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November 6, 2020
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https://www.sciencedaily.com/releases/2020/11/201106113914.htm
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Higher-resolution imaging of living, moving cells using plasmonic metasurfaces
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In the quest to image exceedingly small structures and phenomenon with higher precision, scientists have been pushing the limits of optical microscope resolution, but these advances often come with increased complication and cost.
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Now, researchers in Japan have shown that a glass surface embedded with self-assembled gold nanoparticles can improve resolution with little added cost even using a conventional widefield microscope, facilitating high-resolution fluorescence microscopy capable of high-speed imaging of living cells.Because optical microscopes magnify light to obtain detailed images of a structure, the size of objects that can be distinguished has long been limited by diffraction -- a property of light that causes it to spread when passing through an opening.Researchers have been developing techniques to overcome these limits with highly advanced optical systems, but many of them depend on the use of strong lasers, which can damage or even kill living cells, and scanning of the sample or processing of multiple images, which inhibits real-time imaging."Recent techniques can produce stunning images, but many of them require highly specialized equipment and are incapable of observing the movement of living cells," says Kaoru Tamada, distinguished professor at Kyushu University's Institute for Materials Chemistry and Engineering.Imaging cells using real-time fluorescence microscopy methods, Tamada and her group found that they could improve resolution under a conventional widefield microscope to near the diffraction limit just by changing the surface under the cells.In fluorescence microscopy, cell structures of interest are tagged with molecules that absorb energy from incoming light and, through the process of fluorescence, re-emit it as light of a different color, which is collected to form the image.Though cells are usually imaged on plain glass, Tamada's group coated the glass surface with a self-assembled layer of gold nanoparticles covered with a thin layer of silicon dioxide, creating a so-called metasurface with special optical properties.Only 12 nm in diameter, the organized metal nanoparticles exhibit a phenomenon known as localized surface plasmon resonance, which allows the metasurface to collect energy from nearby light-emitting molecules for highly efficient re-emission, thereby producing enhanced emission confined to the 10-nm thick nanoparticle surface."By introducing the nanoparticles, we have effectively created a light-emitting plane that is only several nanometers thick," explains Tamada. "Because the light of interest is emitted from such a thin layer, we can better focus on it."Additional benefits arise from energy transfer to the metasurface being fast, further localizing emission points by reducing diffusion, and the metasurface's high refractive index, which helps to improve resolution according to Abbe's diffraction limit.Using the metasurface, the researchers imaged in real-time mouse cells known as 3T3 fibroblasts that were genetically engineered to produce a protein called paxillin that is modified to emit green light when excited. Paxillin plays a key role in creating focal adhesions -- points where molecules in the cell membrane interact with the outside world.Illuminating the entire sample with laser light perpendicular to the surface, the researchers were able to image changes in paxillin near the cell membrane with a higher resolution using the metasurface instead of glass.Tilting the illumination light to achieve total internal reflection, the researchers could obtain images with even higher contrast because most of the illumination light is reflected off the surface with only a small amount reaching the cell side, thereby reducing stray emission produced by illumination penetrating deep into the cell.Analysis of images recorded every 500 milliseconds with a super-resolution digital camera revealed clear differences in intensity over spots covering only a few pixels, indicating the resolution was about 200 nm -- close to the diffraction limit.Cells could also be imaged longer on the metasurface because the emission was enhanced despite a lower input energy, thereby reducing cell damage over time."Metasurfaces are a promising option for improving resolution for researchers around the world using conventional optical microscopes that they already have," comments Tamada.In addition to continuing to improve the surfaces for use with conventional microscopes, the researchers are also exploring the advantages they can have for more sophisticated microscope systems.
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Biotechnology
| 2,020 |
November 6, 2020
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https://www.sciencedaily.com/releases/2020/11/201106113919.htm
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Sugar-coated viral proteins hijack and hitch a ride out of cells
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Researchers from the Universities of Melbourne, York, Warwick and Oxford have shed light on how encapsulated viruses like hepatitis B, dengue and SARS-CoV-2 hijack the protein manufacturing and distribution pathways in the cell -- they have also identified a potential broad spectrum anti-viral drug target to stop them in their tracks.
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The findings have been published in Professor Spencer Williams from the School of Chemistry at Bio21 said the research will help define a new 'host-directed' approach for treating infections by encapsulated viruses."One approach to treating viral infections is to make a new drug for each virus that comes along. But it is slow. An alternative and attractive approach is to make a drug against a human target that viruses need to replicate. The same drug can then be used and reused against many different viruses, even ones that have yet to emerge," he said.The findings result from work by Professor Gideon Davies and his UK team who clarified how the structure of the catalytic domain of human enzyme that trims sugar molecules from proteins during their production and Professor Williams' and his Bio21 team, who developed a series of inhibitors to block the enzyme.When tested in human cell lines, these inhibitors where shown to reduce infection in dengue viruses."Encapsulated viruses tend to harness the 'glycosylation' step of protein production, whereby glycans, or sugar molecules coat newly assembled proteins," said Professor Williams."The sugar molecules provide instructions for proteins to fold into their correct 3D structure as well as transport instructions for the protein to be brought to its next destination within the cell. Glycosylation is facilitated by various enzymes that synthesize, trim, check and modify these sugar molecules."Our body's cells contain around 42 million protein molecules. Protein production is a complex, multi-step process within the cell. Like products on a factory assembly-line, all proteins pass through 'quality control' check points where they are inspected before they are transported to their destination, to carry out their functions.Viruses are not living organisms, but biological programs encoded in ribonucleic acid (RNA) or deoxyribonucleic acid (DNA).They come to life when they enter a living cell and hijack the protein production systems. Viruses use the cell's machinery to copy their DNA or RNA (in the case of SARS-CoV2, it's RNA) and to produce the proteins they need to make copies of themselves.The viral proteins produced in an infected cell undergo the 'glycosylation' and then pass through the quality control steps, which involves 'trimming' by an enzyme called 'MANEA'."Trimming is a crucial quality control step and when it does not occur, client proteins are marked for degradation. MANEA represents a key target for broad spectrum drug development against encapsulated viruses, as inhibitors will trigger destruction of their proteins," said Professor Davies.Because viruses hijack this unusual biosynthetic pathway, it makes it a good potential drug target.Researchers at the University of Warwick and University of Oxford studied the effect of the best inhibitors on viral replication.
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Biotechnology
| 2,020 |
November 5, 2020
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https://www.sciencedaily.com/releases/2020/11/201105183801.htm
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Gene in mice controls food cravings, desire to exercise
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National Institutes of Health researchers have discovered a gene in mice that controls the craving for fatty and sugary foods and the desire to exercise. The gene, Prkar2a, is highly expressed in the habenula, a tiny brain region involved in responses to pain, stress, anxiety, sleep and reward. The findings could inform future research to prevent obesity and its accompanying risks for cardiovascular disease and diabetes. The study was conducted by Edra London, Ph.D., a staff scientist in the section on endocrinology and genetics at NIH's Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), and colleagues. It appears in
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Prkar2a contains the information needed to make two subunits -- molecular components -- of the enzyme protein kinase A. Enzymes speed up chemical reactions, either helping to combine smaller molecules into larger molecules, or to break down larger molecules into smaller ones. Protein kinase A is the central enzyme that speeds reactions inside cells in many species. In a previous study, the NICHD team found that despite being fed a high fat diet, mice lacking functioning copies of Prkar2a were less likely to become obese than wild type mice with normally functioning Prkar2a.The researchers determined that Prkar2a-negative mice ate less high-fat food than their counterparts, not only when given unlimited access to the food, but also after a fast. Similarly, the Prkar2a negative mice also drank less of a sugar solution than the wild type mice. The Prkar2a-negative mice were also more inclined to exercise, running 2-3 times longer than wild type mice on a treadmill. Female Prkar2a-negative mice were less inclined to consume high fat foods than Prkar2-negative males, while Prkar2-negative males showed less preference for the sugar solution than Prkar2-negative females.
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Biotechnology
| 2,020 |
November 5, 2020
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https://www.sciencedaily.com/releases/2020/11/201105183757.htm
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Mystery molecule in bacteria is revealed to be a guard
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Peculiar hybrid structures called retrons that are half RNA, half single-strand DNA are found in many species of bacteria. Since their discovery around 35 years ago, researchers have learned how to use retrons for producing single strands of DNA in the lab, but no one knew what their function was in the bacteria, despite much research into the matter. In a paper published today in
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The study, conducted in the lab of Prof. Rotem Sorek of the Institute's Molecular Genetics Department, was led by Adi Millman, Dr. Aude Bernheim and Avigail Stokar-Avihail in his lab. Sorek and his team did not set out to solve the retron mystery; they were seeking new elements of the bacterial immune system, specifically elements that help bacteria to fend off viral infection. Their search was made easier by their recent finding that bacteria's immune system genes tend to cluster together in the genome within so-called defense islands. When they uncovered the unique signature of retron within a bacterial defense island, the team decided to investigate further.Their initial research showed that this retron was definitely involved in protecting bacteria against the viruses known as phages that specialize in infecting bacteria. As the researchers looked more closely at additional retrons located near known defense genes, they found that the retrons were always connected -- physically and functionally -- to one other gene. When either the accompanying gene or the retron was mutated, the bacteria were less successful in fighting off phage infection.The researchers then set out to look for more such complexes in defense islands. Eventually, they identified some 5,000 retrons, many of them new, in different defense islands of numerous bacterial species.To check if these retrons function, generally, as immune mechanisms, the researchers transplanted many retrons, one by one, into laboratory bacterial cells that were lacking retrons. As they suspected, in a great number of these cells they found retrons protecting the bacteria from phage infection.How do retrons do this? Focusing back on one particular kind of retron and tracing its actions in the face of phage infection, the research team discovered that its function is to cause the infected cell to commit suicide. Cell suicide, once thought to belong solely to multicellular organisms, is a last-ditch means of aborting widespread infection -- if the suicide mechanism works fast enough to kill the cell before the virus finishes making copies of itself and spreading out to other cells.Further investigation showed that retrons do not sense the phage invasion itself, but rather keep watch on another part of the immune system known as RecBCD, which is one of the bacterium's first lines of defense. If it realizes that the phage has tampered with the cell's RecBCD, the retron activates its program through the second, linked genes to kill the infected cell and protect the rest of the colony."It's a clever strategy, and we found it works in a similar way to a guard mechanism employed in plant cells," says Sorek. "Just like viruses that infect plants, phages come equipped with a variety of inhibitors to block assorted parts of the cell immune response. The retron, like a guard mechanism known to exist in plants, does not need to be able to identify all possible inhibitors, just to have a handle on the functioning of one particular immune complex. Infected plant cells apply this 'abortive infection' method, killing off a small region of a leaf or root, in an effort to save the plant itself. Since most bacteria live in colonies, this same strategy can promote the survival of the group, even at the expense of individual members."Retrons are so useful to biotechnology because they begin with a piece of RNA, which is the template for the synthesis of the DNA strand. This template in the retron sequence can be swapped out for any desired DNA sequence and used, sometimes in conjunction with another tool borrowed from the bacterial immune toolkit -- CRISPR -- to manipulate genes in various ways. Sorek and his team believe that within the diverse list of retrons they identified may be hiding more than a few that could provide better templates for specific gene editing needs.
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Biotechnology
| 2,020 |
November 5, 2020
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https://www.sciencedaily.com/releases/2020/11/201105134515.htm
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Discovery of shape of the SARS-CoV-2 genome after infection could inform new COVID-19 treatments
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Scientists at the University of Cambridge, in collaboration with Justus-Liebig University, Germany, have uncovered how the genome of SARS-CoV-2 -- the coronavirus that causes COVID-19 -- uses genome origami to infect and replicate successfully inside host cells. This could inform the development of effective drugs that target specific parts of the virus genome, in the fight against COVID-19.
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SARS-CoV-2 is one of many coronaviruses. All share the characteristic of having the largest single-stranded RNA genome in nature. This genome contains all the genetic code the virus needs to produce proteins, evade the immune system and replicate inside the human body. Much of that information is contained in the 3D structure adopted by this RNA genome when it infects cells.The researchers say most current work to find drugs and vaccines for COVID-19 is focused on targeting the proteins of the virus. Because the shape of the RNA molecule is critical to its function, targeting the RNA directly with drugs to disrupt its structure would block the lifecycle and stop the virus replicating.In a study published today in the journal "The RNA genome of coronaviruses is about three times bigger than an average viral RNA genome -- it's huge," said lead author Dr Omer Ziv at the University of Cambridge's Wellcome Trust/Cancer Research UK Gurdon Institute.He added: "Researchers previously proposed that long-distance interactions along coronavirus genomes are critical for their replication and for producing the viral proteins, but until recently we didn't have the right tools to map these interactions in full. Now that we understand this network of connectivity, we can start designing ways to target it effectively with therapeutics."In all cells the genome holds the code for the production of specific proteins, which are made when a molecular machine called a ribosome runs along the RNA reading the code until a 'stop sign' tells it to terminate. In coronaviruses, there is a special spot where the ribosome only stops 50% of the times in front of the stop sign. In the other 50% of cases, a unique RNA shape makes the ribosome jump over the stop sign and produce additional viral proteins. By mapping this RNA structure and the long-range interactions involved, the new research uncovers the strategies by which coronaviruses produce their proteins to manipulate our cells."We show that interactions occur between sections of the SARS-CoV-2 RNA that are very long distances apart, and we can monitor these interactions as they occur during early SARS-CoV-2 replication," said Dr Lyudmila Shalamova, a co-lead investigator at Justus-Liebig University, Germany.Dr Jon Price, a postdoctoral associate at the Gurdon Institute and co-lead of this study, has developed a free, open-access interactive website hosting the entire RNA structure of SARS-CoV-2. This will enable researchers world-wide to use the new data in the development of drugs to target specific regions of the virus's RNA genome.The genome of most human viruses is made of RNA rather than DNA. Ziv developed methods to investigate such long-range interactions across viral RNA genomes inside the host cells, in work to understand the Zika virus genome. This has proved a valuable methodological basis for understanding SARS-CoV-2.This research is a collaborative study between the group of Professor Eric Miska at the University of Cambridge's Gurdon Institute and Department of Genetics, and the group of Professor Friedemann Weber from the Institute for Virology, Justus-Liebig University, Gießen, Germany. The authors are grateful for the support of the Biochemistry Department at the University of Cambridge, who provided specialist laboratory facilities for performing part of this research.
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Biotechnology
| 2,020 |
November 4, 2020
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https://www.sciencedaily.com/releases/2020/11/201104114741.htm
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Biologists create 'atlas' of gene expression in neurons, documenting diversity of brain cells
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New York University researchers have created a "developmental atlas" of gene expression in neurons, using gene sequencing and machine learning to categorize more than 250,000 neurons in the brains of fruit flies. Their study, published in
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"Diversity of the different cell types that make up our brains can only be fully understood in light of their developmental history," said NYU Biology Professor Claude Desplan, the study's senior author.Brains are composed of thousands of different types of neurons. Despite sharing the same genetic information, neurons achieve this diversity by turning on different sets of genes in each neuron type and at each point in their development.To understand the diversity of brain cells, researchers have long studied fruit flies, whose brains, although much simpler than those of humans, can be used as a model system. Researchers previously identified the roughly 60,000 cells and 200 neuronal types that make up fruit flies' optic lobes, the areas of the brain that process visual information, including color vision and detection of objects and motion.In their new study in The researchers created their "atlas" by taking advantage of a form of a recently invented technique known as single-cell mRNA sequencing, which allowed them to capture and sequence mRNA from more than 250,000 single cells. Using a combination of machine learning approaches, they assigned each of these cells to a specific cell type throughout development."Our datasets almost completely account for the known neuronal diversity of the optic lobes and can serve as a paradigm to understand brain development across species," said Neset Özel, a postdoctoral associate at NYU and one of the study's lead authors. "The 'atlas' constitutes an enormous resource for the research community: we can now simply look up whether a particular gene is active or not in any cell type of our choice and at any point during its development."While building their "developmental atlas," the researchers made several discoveries. First, they found a completely new type of neurons in fruit flies, which is present only on the surface of the optic lobe during development but is removed through programmed cell death right before the flies hatch."While we do not yet understand the functions of these previously unknown neurons, neurons with very similar properties -- called Cajal-Retzius cells -- also exist in mammalian brains, and they are critical for proper brain development," said Felix Simon, a biology doctoral student at NYU and the other lead author of the study.In addition, the researchers found that neurons exhibit the highest levels of molecular diversity during development compared to adult neurons, allowing cells during development to form connections with specific partner cells -- and avoid the wrong ones. As a result, neurons can gain distinct features and functions solely due to their developmental history, even though their physiological properties in adult brains might be identical."This has large implications for the studies of neurodevelopmental disorders. Disruptions to neural circuit function could occur entirely due to defects in certain genetic programs that are only transiently active during development and would be impossible to understand by simply looking at the end result," explained Özel.Finally, the study revealed that neurons that look identical in form can express different sets of genes in the upper versus lower part of the brain. These differences can give flies the ability to perform different calculations on the visual information they receive -- for instance, the sky versus the ground.
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Biotechnology
| 2,020 |
November 4, 2020
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https://www.sciencedaily.com/releases/2020/11/201104102205.htm
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Microbial space travel on a molecular scale
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Galactic cosmic and solar UV radiation, extreme vacuum, temperature fluctuations: how can microbes exposed to these challenges in space survive? Scientists investigated how the space-surviving microbes could physically survive the transfer from one celestial body to another.
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Since the dawn of space exploration, humankind has been fascinated by survival of terrestrial life in outer space. Outer space is a hostile environment for any form of life, but some extraordinarily resistant microorganisms can survive. Such extremophiles may migrate between planets and distribute life across the Universe, underlying the panspermia hypothesis or interplanetary transfer of life.The extremophilic bacterium Deinococcus radiodurans withstands the drastic influence of outer space: galactic cosmic and solar UV radiation, extreme vacuum, temperature fluctuations, desiccation, freezing, and microgravity. A recent study examined the influence of outer space on this unique microbe on a molecular level. After 1 year of exposure to low Earth orbit (LEO) outside the International Space Station during the Tanpopo space Mission, researches found that D. radiodurans escaped morphological damage and produced numerous outer membrane vesicles. A multifaceted protein and genomic responses were initiated to alleviate cell stress, helping the bacteria to repair DNA damage and defend against reactive oxygen species. Processes underlying transport and energy status were altered in response to space exposure. D. radiodurans used a primordial stress molecule polyamine putrescine as a reactive oxygen species scavenger during regeneration from space exposure."These investigations help us to understand the mechanisms and processes through which life can exist beyond Earth, expanding our knowledge how to survive and adapt in the hostile environment of outer space. The results suggest that survival of D. radiodurans in LEO for a longer period is possible due to its efficient molecular response system and indicate that even longer, farther journeys are achievable for organisms with such capabilities" says Tetyana Milojevic, a head of Space Biochemistry group at the University of Vienna and a corresponding author of the study.Together with the colleagues from Tokyo University of Pharmacy and Life Science (Japan), Research Group Astrobiology at German Aerospace Center (DLR, Cologne), Vienna Metabolomics Centre (ViMe) at the University of Vienna and Center for Microbiome Research at Medical University Graz, researches answered the question not only till which extend but how extremophilic microbes can tolerate drastic space conditions.
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Biotechnology
| 2,020 |
November 4, 2020
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https://www.sciencedaily.com/releases/2020/11/201104083016.htm
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Scientists identify synthetic mini-antibody to combat COVID-19
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By screening hundreds of synthetic mini-antibodies called sybodies, a group of scientists has identified one that might stop SARS-CoV-2 from infecting human cell.
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The ability of SARS-CoV-2 to infect cells depends on interactions between the viral spike protein and the human cell surface protein ACE2. To enable the virus to hook onto the cell surface, the spike protein binds ACE2 using three finger-like protrusions, called the receptor binding domains (RBDs). Blocking the RBDs therefore has the potential to stop the virus from entering human cells. This can be done using antibodies.Nanobodies, small antibodies found in camels and llamas, are promising as tools against viruses due to their high stability and small size. Although obtaining them from animals is time consuming, technological advances now allow for rapid selection of synthetic nanobodies, called sybodies. A technology platform to select sybodies from large synthetic libraries was recently developed in the lab of Markus Seeger at the University of Zurich, and made available for this study.EMBL Hamburg's Christian Löw group searched through the existing libraries to find sybodies that could block SARS-CoV-2 from infecting human cells. First, they used the viral spike protein's RBDs as bait to select those sybodies that bind to them. Next, they tested the selected sybodies according to their stability, effectiveness, and the precision of binding. Among the best binders, one called sybody 23 turned out to be particularly effective in blocking the RBDs.To learn exactly how sybody 23 interacts with the viral RBDs, researchers in the group of Dmitri Svergun at EMBL Hamburg analysed the binding of sybody 23 to the RBDs by small-angle X-ray scattering. In addition, Martin Hällberg at CSSB and Karolinska Institutet used cryo-EM to determine the structure of the full SARS-CoV-2 spike bound to sybody 23. The RBDs switch between two positions: in the 'up' position the RBDs poke out, ready to bind ACE2; in the 'down' position they are furled to hide from the human immune system. The molecular structures revealed that sybody 23 binds RBDs in both 'up' and 'down' positions, and blocks the areas where ACE2 would normally bind. This ability to block RBDs regardless of their position might explain why sybody 23 is so effective.Finally, to test if sybody 23 can neutralise a virus, the group of Ben Murrell at Karolinska Institutet used a different virus, called a lentivirus, modified such that it carried SARS-CoV-2's spike protein on its surface. They observed that sybody 23 successfully disabled the modified virus in vitro. Additional tests will be necessary to confirm whether this sybody could stop SARS-CoV-2 infection in the human body."The collaborative spirit has been enormous in these times, and everybody was motivated to contribute," says Christian Löw, one of the lead scientists in the study. The researchers started the project as soon as they received approval from EMBL leadership to reopen their laboratories during the COVID-19 lockdown. They managed to select the candidate sybodies and perform the analyses in just a few weeks."Getting the results so quickly was only possible because the methodologies we used had already been established for other research projects unrelated to SARS-CoV-2. Developing these tools would have taken significantly more time and resources," says Löw.The results of this project hold out the promise of a potential way to treat COVID-19. In future work, the scientists will perform further analyses to confirm whether sybody 23 could be an effective COVID-19 treatment.
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Biotechnology
| 2,020 |
November 3, 2020
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https://www.sciencedaily.com/releases/2020/11/201103112524.htm
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The birth of a tRNA gene
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Translation is the process by which genetic information is converted into proteins, the workhorses of the cell. Small molecules called transfer RNAs ("tRNAs") play a crucial role in translation; they are the adapter molecules that match codons (the building blocks of genetic information) with amino acids (the building blocks of proteins). Organisms carry many types of tRNAs, each encoded by one or more genes (the "tRNA gene set").
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Broadly speaking, the function of the tRNA gene set -- to translate 61 types of codons into 20 different kinds of amino acids -- is conserved across organisms. Nevertheless, tRNA gene set composition can vary considerably between organisms. How and why these differences arise has been a question of long-standing interest among scientists.Jenna Gallie (Research Group Leader at the Max Planck Institute for Evolutionary Biology) and her team have investigated how the tRNA gene set of the bacterium Pseudomonas fluorescens can evolve, using a combination of mathematical modelling and lab-based experiments."We started by removing one type of tRNA from the bacterium's genome, resulting in a bacterial strain that grows slowly. We gave this slow-growing strain the opportunity to improve its growth during a real-time evolution experiment. We saw the strain improve repeatedly and rapidly. The improvement was due to the duplication of large chunks of bacterial genetic information, with each duplication containing a compensatory tRNA gene. Ultimately, the elimination of one tRNA type was compensated by an increase in the amount of a second, different tRNA type." Jenna Gallie said. The duplicated tRNA type can compensate because it is able to perform, at a lower rate, the codon-amino acid matching function of the eliminated tRNA type.Comparisons of tRNA genes in related genomes have previously provided evidence for the duplication of some tRNA genes throughout evolutionary history. The experiments described here provide direct, empirical evidence that tRNA gene sets can evolve through duplication events.
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Biotechnology
| 2,020 |
November 3, 2020
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https://www.sciencedaily.com/releases/2020/11/201103104729.htm
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New AI tool provides much-needed help to protein scientists across the world
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Using artificial intelligence, UCPH researchers have solved a problem that until now has been the stumbling block for important protein research into the dynamics behind diseases such as cancer, Alzheimer's and Parkinson's, as well as in the development of sustainable chemistry and new gene-editing technologies.
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It has always been a time-consuming and challenging task to analyse the huge datasets collected by researchers as they used microscopy and the smFRET technique to see how proteins move and interact with their surroundings. At the same time the task required a high level of expertise. Hence, the proliferation of stuffed servers and hard drives. Now researchers at the Department of Chemistry, Nano-Science Center, Novo Nordisk Foundation Center for Protein Research and the Niels Bohr Institute, University of Copenhagen, have developed a machine learning algorithm to do the heavy lifting."We used to sort data until we went loopy. Now our data is analysed at the touch of button. And, the algorithm does it at least as well or better than we can. This frees up resources for us to collect more data than ever before and get faster results," explains Simon Bo Jensen, a biophysicist and PhD student at the Department of Chemistry and the Nano-Science Center.The algorithm has learned to recognize protein movement patterns, allowing it to classify data sets in seconds -- a process that typically takes experts several days to accomplish."Until now, we sat with loads of raw data in the form of thousands of patterns. We used to check through it manually, one at a time. In doing so, we became the bottleneck of our own research. Even for experts, conducting consistent work and reaching the same conclusions time and time again is difficult. After all, we're humans who tire and are prone to error," says Simon Bo Jensen.The studies about the relationship between protein movements and functions conducted by the UCPH researchers is internationally recognized and essential for understanding how the human body functions. For example, diseases including cancer, Alzheimer's and Parkinson's are caused by proteins clumping up or changing their behaviour. The gene-editing technology CRISPR, which won the Nobel Prize in Chemistry this year, also relies on the ability of proteins to cut and splice specific DNA sequences. When UCPH researchers like Guillermo Montoya and Nikos Hatzakis study how these processes take place, they make use of microscopy data."Before we can treat serious diseases or take full advantage of CRISPR, we need to understand how proteins, the smallest building blocks, work. This is where protein movement and dynamics come into play. And this is where our tool is of tremendous help," says Guillermo Montoya, Professor at the Novo Nordisk Foundation Center for Protein Research.It appears that protein researchers from around the world have been missing just such a tool. Several international research groups have already presented themselves and shown an interest in using the algorithm."This AI tool is a huge bonus for the field as a whole because it provides common standards, ones that weren't there before, for when researchers across world need to compare data. Previously, much of the analysis was based on subjective opinions about which patterns were useful. Those can vary from research group to research group. Now, we are equipped with a tool that can ensure we all reach the same conclusions," explains research director Nikos Hatzakis, Associate Professor at the Department of Chemistry and Affiliate Associate Professor at the Novo Nordisk Foundation Center for Protein Research.He adds that the tool offers a different perspective as well:"While analysing the choreography of protein movement remains a niche, it has gained more and more ground as the advanced microscopes needed to do so have become cheaper. Still, analysing data requires a high level of expertise. Our tool makes the method accessible to a greater number of researchers in biology and biophysics, even those without specific expertise, whether it's research into the coronavirus or the development of new drugs or green technologies."
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Biotechnology
| 2,020 |
November 2, 2020
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https://www.sciencedaily.com/releases/2020/11/201102142247.htm
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Microfluidics helps engineers watch viral infection in real time
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Get your popcorn. Engineers and virologists have a new way to watch viral infection go down.
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The technique uses microfluidics -- the submillimeter control of fluids within a precise, geometric structure. On what is basically a tricked-out microscope slide, chemical engineers from Michigan Technological University have been able to manipulate viruses in a microfluidic device using electric fields. The study, published this summer in Viruses carry around an outer shell of proteins called a capsid. The proteins act like a lockpick, attaching to and prying open a cell's membrane. The virus then hijacks the cell's inner workings, forcing it to mass produce the virus's genetic material and construct many, many viral replicas. Much like popcorn kernels pushing away the lid of an overfilled pot, the new viruses explode through the cell wall. And the cycle continues with more virus lockpicks on the loose."When you look at traditional techniques -- fluorescent labeling for different stages, imaging, checking viability -- the point is to know when the membrane is compromised," said Adrienne Minerick, study co-author, dean of the College of Computing and a professor of chemical engineering. "The problem is that these techniques are an indirect measure. Our tools look at charge distribution, so it's heavily focused on what's happening between the cell membrane and virus surface. We discovered with greater resolution when the virus actually goes into the cell."Watching the viral infection cycle and monitoring its stages is crucial for developing new antiviral drugs and gaining better understanding of how a virus spreads. Dielectrophoresis happens when polarizable cells get pushed around in a nonuniform electric field. The movement of these cells is handy for diagnosing diseases, blood typing, studying cancer and many other biomedical applications. When applied to studying viral infection, it's important to note that viruses have a surface charge, so within the confined space in a microfluidic device, dielectrophoresis reveals the electric conversation between the virus capsid and the proteins of a cell membrane."We studied the interaction between the virus and cell in relation to time using microfluidic devices," said Sanaz Habibi, who led the study as a doctoral student in chemical engineering at Michigan Tech. "We showed we could see time-dependent virus-cell interactions in the electric field."Watching a viral infection happen in real time is like a cross between a zombie horror film, paint drying and a Bollywood epic on repeat. The cells in the microfluidic device dance around, shifting into distinct patterns with a dielectric music cue. There needs to be the right ratio of virus to cells to watch infection happen -- and it doesn't happen quickly. Habibi's experiment runs in 10-hour shifts, following the opening scenes of viral attachment, a long interlude of intrusion, and eventually the tragic finale when the new viruses burst out, destroying the cell in the process.Before they burst, cell membranes form structures called blebs, which change the electric signal measured in the microfluidic device. That means the dielectrophoresis measurements grant high-resolution understanding of the electric shifts happening at the surface of the cell through the whole cycle.Viral infections are top of mind right now, but not all viruses are the same. While microfluidic devices that use dielectrophoresis could one day be used for on-site, quick testing for viral diseases like COVID-19, the Michigan Tech team focused on a well-known and closely studied virus, the porcine parvovirus (PPV), which infects kidney cells in pigs.But then the team wanted to push the envelope: They added the osmolyte glycine, an important intervention their collaborators study in viral surface chemistry and vaccine development."Using our system, we could show time-dependent behavior of the virus and cell membrane. Then we added the osmolyte, which can act as an antiviral compound," Habibi explained. "We thought it would stop the interaction. Instead, it looked like the interaction continued to happen at first, but then the new viruses couldn't get out of the cell."That's because glycine likely interrupts the new capsid formation for the replicated viruses within the cell itself. While that specific portion of the viral dance happens behind the curtain of the cell wall, the dielectric measurements show a shift between an infected cycle where capsid formation happens and an infected cell where capsid formation is interrupted by glycine. This difference in electrical charge indicates that glycine prevents the new viruses from forming capsids and stops the would-be viral lockpickers from hitting their targets."When you are working with such small particles and organisms, when you're able to see this process happening in real time, it's rewarding to track those changes," Habibi said.This new view of the interactions between virus capsids and cell membranes could speed up testing and characterizing viruses, cutting out expensive and time-consuming imaging technology. Perhaps in a future pandemic, there will be point-of-care, handheld devices to diagnose viral infections and we can hope medical labs will be outfitted with other microfluidic devices that can quickly screen and reveal the most effective antiviral medications.
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Biotechnology
| 2,020 |
November 2, 2020
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https://www.sciencedaily.com/releases/2020/11/201102120051.htm
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Breakdown of gene coordination during aging suggests a substantial challenge to longevity
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Although all humans share similar changes during aging such as gray hair, wrinkles, and a general decline in function, aging is considered to be the result of a cellular wear-and-tear process due to accumulated random damage, such as genetic mutations or DNA structural damage.
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How is it that random, disorganized damage, which accumulates differently among different humans, and moreover, among different cells of the same individual eventually leads to the same outcomes? Several theories try to address this paradox, and they have great implications for our ability to affect the aging process, making elderly life better and longer. The potential to develop treatments for aging depends on understanding the fundamental process of growing old.A common approach holds that most cells in the human body are barely damaged during aging, while just a few "rotten apples" -- a small fraction of non-functioning cells -- are significantly damaged. Accordingly, a potential treatment for aging could involve removing these few highly-damaged cells.Approximately 15 years ago Prof. Jan Vijg proposed a different approach. He suggested that the proper function of biological tissues may decline during aging because many cells lose their ability to tightly regulate their genes. According to Vijg's theory, there are no single non-functioning cells -- or rotten apples -- on the one hand, but none of the apples is "fresh" on the other. Evidence for Vijg's theory has never been fully presented, until now.In a study published today in the journal To test the consistency of this phenomenon, they analyzed data collected from more than twenty experiments from six different labs around the world. In all cases they found reduced levels of coordination during aging among different organisms: human, mice and fruit flies, and among different cell types: brain cells, Hematopoietic stem cells, pancreatic cells and more."In biology it is very difficult to achieve consistent results for different types of cells, tissues, experiments and organisms due to the high sensitivity of equipment and experimental setup," says Dr. Orr Levy from the group of the study's lead author, Dr. Amir Bashan, of the Department of Physics at Bar-Ilan University. "Our method found the same pattern in more than 20 datasets. Finding evidence for coordination of genes was amazing, but even more outstanding was finding that this property of coordination dramatically declines with age," added Guy Amit from Bashan's team. Bashan and team collaborated on the research with Prof. Haim Cohen and Prof. Sol Efroni from Bar-Ilan University, and Prof. Yang-Yu Liu and Prof. Peter Castadli from Harvard Medical School.The researchers also observed coordination reduction in tissues with an increased level of damage, suggesting a direct link between increased damage level and coordination breakdown. The findings support the theory that during aging, accumulated random damage affects regulation mechanisms and disrupts the ability of genes to coordinate (resulting in a general decrease in tissue function), just like an orchestra without proper coordination between musicians ruins a symphony.This study conclusively demonstrates the long-speculated relationship between aging, gene regulation and somatic damage. The results open up new avenues of research with practical implications. If the same level of coordination reduction between genes is indeed a leading cause for aging phenomena, there may be a need to change course in current efforts to develop aging treatments.This project was primarily funded by the Azrieli Foundation.
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Biotechnology
| 2,020 |
November 2, 2020
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https://www.sciencedaily.com/releases/2020/11/201102120045.htm
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'BAH-code' reader senses gene-silencing tag in cells
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University of North Carolina Lineberger Comprehensive Cancer Center researchers have identified a new and evolutionarily conserved pathway responsible for "closing down" gene activity in the mammalian cell. The finding is closely related to the Polycomb pathway defined decades ago by a set of classic genetic experiments carried out in fruit flies.
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UNC Lineberger's Greg Wang, PhD, associate professor in the UNC School of Medicine Department of Biochemistry and Biophysics, and his colleagues reported in the journal In cells, the Polycomb pathway generates a small chemical tag, H3K27me3, to silence genes. Molecular players related to Polycomb and H3K27me3 are frequently found to be associated with pathogenesis, notably cancer and developmental syndromes."H3K27me3 is sort of like a gene 'tag' for silencing. Our initial interest towards BAHCC1 actually stems from a connection to cancer, in particular, leukemia. And previously, BAHCC1 was little studied," said Wang, the study's senior author. "Work on BAHCC1 turns out to be a very interesting journey leading to fundamental understanding of Polycomb and gene silencing in general."Huitao Fan, PhD, UNC Lineberger and UNC School of Medicine Department of Biochemistry and Biophysics, and Jiuwei Lu, PhD, University of California, Riverside, were the study's first authors.Leukemia is a cancer that affects the blood and bone marrow. The American Cancer Society estimated that more than 60,000 people will be diagnosed with leukemia in the United States this year, and the disease will cause more than 23,000 deaths.In their study, researchers in Wang's lab discovered in retrospective analysis of published data that high expression of the BAHCC1 gene was found in different types of leukemia. Using CRISPR-cas9-based state-of-the-art gene loss-of-function techniques, Wang and his team demonstrated the dependence of various acute leukemia models on BAHCC1 in progression of the disease. They determined BAHCC1 inhibits tumor suppressors to help drive acute leukemia. This function of BAHCC1 relies on an ability harbored within its protein module BAH to scan and directly bind the silencing-related tag, H3K27me3, found on the genes to be silencedThe team led by the other senior author of the paper, Jikui Song, PhD, University of California, Riverside, generated an atomic view of how the BAH module in BAHCC1 binds to the H3K27me3 tag.Wang said this study challenges the current norm that tends to emphasize the previously known CBX proteins as the main effector of H3K27me3 and Polycomb silencing in mammals."We now realize that there exists a previously unexplored chapter of important mechanisms that animal cells use for silencing genes," Wang said. "And based on recent research of others, we believe the BAH module and related pathways are evolutionarily ancient and conserved among fungi, plants and animals. In leukemia, BAHCC1 is co-opted to silence tumor suppressive genes and to promote a cancerous program."The researchers said they will continue to study the underlying mechanisms behind these proteins and their relationship to biology and diseases with the goal of developing therapeutic approaches.
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Biotechnology
| 2,020 |
November 2, 2020
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https://www.sciencedaily.com/releases/2020/11/201102110039.htm
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How the immune system remembers viruses
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When a virus enters the body, it is picked up by certain cells of the immune system. They transport the virus to the lymph nodes where they present its fragments, known as antigens, to CD8+ T cells responsible control of viral infections. Each of these cells carries a unique T cell receptor on the surface that can recognize certain antigens. However, only very few T cell receptors match a given viral the antigen.
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To bring the infection under control and maximize the defenses against the virus, these few antigen-specific T cells start dividing rapidly and develop into effector T cells. These kill virus-infected host cells and then die off themselves once the infection is cleared. Some of these short-lived effector cells -- according to the generally accepted theory -- turn into memory T cells, which persist in the organism long term. In case the same pathogen enters the body again, memory T cells are already present and ready to fight the invader more swiftly and effectively than during the first encounter."Prevailing scientific opinion says that activated T cells first become effector cells and only then gradually develop into memory cells," says Dr. Veit Buchholz, a specialist in microbiology and working group leader at the Institute for Medical Microbiology, Immunology and Hygiene at TUM. "In our view, however, that isn't the case. It would mean that the more effector cells are formed after contact with the pathogen, the more numerous the memory cells would become." However, Buchholz and his colleagues observed a different course of events and have now published their results in the journal "We investigated the antiviral immune responses resulting from individual activated T cells in mice and traced the lineage of the ensuing memory cells using single-cell fate mapping," reports first author Dr. Simon Grassmann. "Based on these experiments, we were able to show that certain 'T cell families' descended from individual cells form up to 1000 times more 'memory' than others. However, these long-term dominating T cell families only contributed little to the magnitude of the initial immune response, which was dominated by effector cells derived from other shorter-lived T cell families."At the level of individual cells, it therefore became evident that development of effector and memory cells segregates at a much earlier stage than previously believed: "Already in the first week after the confrontation with the pathogen, we saw major differences in the transcriptomes of the detected T cell families," says Lorenz Mihatsch, also a first author of the study. "Normally at this time of the immune response CD8+ T cells are enriched in molecules that help to kill virus infected cells. However, we found no indication of these cytolytic molecules in the long-term dominating T cell families. Instead, they were already geared exclusively towards memory development at this early stage."These results could help to improve vaccine development in the future, says Veit Buchholz: "To generate an optimal immune response through vaccination, the body needs to produce as many memory cells as possible. For that purpose, it is important to have a precise understanding of how individual T cells are programmed." Buchholz's study might also prove useful in helping to recognize sooner whether a new vaccine is effective. "To determine the long-term strength of an immune response, it could be helpful to measure the number of memory precursors within a few days of administering a vaccine," says Buchholz.
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Biotechnology
| 2,020 |
November 1, 2020
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https://www.sciencedaily.com/releases/2020/11/201101210349.htm
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Researchers discover mechanism that allows SINEUPs to amplify protein production
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Scientists from an international group led by the RIKEN Center for Integrative Medical Sciences and Yokohama City University have discovered that a pair of proteins play a key role in allowing an important type of functional non-coding RNA, known as SINEUPs, to act to promote their target messenger RNA.SINEUPs are a recently discovered type of RNA that work specifically to amplify the production of proteins by messenger RNAs, and hence could be important for developing therapeutics for diseases where a certain protein is insufficiently synthesized.
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While it was once believed that DNA was simply transcribed into RNA which was then translated into proteins, it is now known that RNA plays a more complex role. While nearly all DNA is transcribed into RNA, it turns out that only 30% of RNA is translated into proteins. The remaining 70% play roles such as enhancing gene expression, epigenetic regulation and -- in the case of SINEUPs -- up-regulating the production of proteins by target RNA.The current research, published in According to Hazuki Takahashi of the RIKEN Center for Integrative Medical Sciences, one of the corresponding authors of the paper, "We wanted to figure out the mechanism for the action of SINEUPs. Understanding how these RNAs work would be a tremendous breakthrough, because there are a number of diseases caused by a failure of genes to create sufficient quantities of a certain protein, and knowing how SINEUPs function could provide us with a way to remedy this."The group did have clues from their previous research. They had noted that the SINEUPs only affected the action of their target messenger RNA when they had been transported, together with the messenger RNA, out of the cell nucleus and into the cytosol where the protein production takes place.Through a series of experiments involving both natural SINEUPs and artificial SINEUPs fitted with a fluorescent protein to allow the team to examine their movements, they discovered that a pair of RNA binding proteins, called PTBP1 and HNRNPK, interact with the SINEUPs both to allow their transport and to make it possible for them to act upon the messenger RNA. These two proteins are quite interesting as they have been found to work together in a variety of biological functions such as maintaining the pluripotency of cells. They are also biologically very important, as it has been shown that knocking out the HNRNPK gene in mice is lethal embryonically.According to Piero Carninci of the RIKEN Center for Integrative Medical Sciences, the leader of the research group, "We are very pleased to have discovered the role of these binding proteins in the activities of SINEUPs. Because of the ability of SINEUPs specifically to modulate the translation of targeted mRNAs as needed, they are ideal for future therapies in humans where increasing the level of a specific protein could have a therapeutic effect. There are hundreds of diseases that would benefit from SINEUPs treatments, caused by deficiency of one functional copy of a gene: these diseases are known with the general terms of haploinsufficiencies. In addition, SINEUPs have potential to enhance currently limited antibody drug production. Understanding the mechanism of SINEUPs and other functional long non-coding RNAs mechanism is a very important first step for future applications of these RNAs for improving human health."
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Biotechnology
| 2,020 |
November 2, 2020
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https://www.sciencedaily.com/releases/2020/11/201102090900.htm
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Criss-crossing viruses give rise to peculiar hybrid variants
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For millions of years, viruses have participated in a far-flung, import-export business, exchanging fragments of themselves with both viral and non-viral agents and acquiring new features. What these tiny entities lack in outward complexity, they make up for with their astonishing abilities to swap out modular genomic components and ceaselessly reinvent themselves.
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In new research appearing in the journal Referred to as cruciviruses, these minute forms reveal a fusion of components from both RNA and DNA viruses, proving that these previously distinct genomic domains can, under proper conditions, intermingle, producing a hybrid or chimeric viral variant.Varsani, a virologist at the Arizona State Univeristy Biodesign Center for Fundamental and Applied Microbiomics, is deeply intrigued with these new viruses, which are starting to crop up in greater abundance and diversity in a wide range of environments."It is great to see the research groups that first identified cruciviruses around the same time teaming up for the sharing and mining of metagenomic data with an aim to identify a larger diversity of cruciviruses," says Varsani, an associate professor with ASU School of Life Sciences.Crucivirus sequences were identified by Varsani's colleague and co-author Kenneth M. Stedman and his group at Portland State University. The team detected the viruses flourishing in an extreme environment -- Boiling Springs Lake (BSL) in Lassen Volcanic National Park, Northern California. Around the same time, Varsani and Mya Breitbart's research group identified a crucivirus in a dragonfly sample from Florida.Since their discovery in 2012, cruciviruses have been found in diverse environments around the world, from lakes in upstate New York and Florida, to the Antarctic and deep-sea sediments. Some 80 distinct cruciviruses had been identified, prior to the current study, which expands the number to 461.The first cruciviruses were identified using a technique known as viral metagenomics, in which viral genetic material obtained directly from the environment is sequenced rather than being cultivated or cultured from a host species or natural reservoir.The results of these early investigations revealed peculiar genetic sequences, radically distinct from anything that had been seen before. This sequences clearly displayed the signature of a DNA virus, yet also contained a gene that appeared to be derived from an RNA virus.Using a shotgun approach to trawl through a potentially vast sequence space, viral metagenomics enables researchers to identify all of the genomic patterns present in an environmental sample, then separate out distinct viral sequences, like a fisherman retrieving a variety of sea creatures from his net.The technique has revolutionized the discipline of virology. In addition to identifying a galaxy of previously unknown viruses, metagenomics has offered up exciting clues about genetic diversity and is helping to unlock some of the secrets of viral evolution, all without the need to initially isolate viral species or cultivate viruses in the lab.Cruciviruses belong to a broader class of viruses known as CRESS, (for circular Rep-encoding single-stranded) DNA viruses which have recently been classified into the phylum Cressdnaviricota. The defining characteristic of such viruses is their mode of replication, which relies on a specific component, known as the Rep protein. The Rep protein is important for guiding the replication method of these viruses, known as rolling circle DNA replication. Presence of the Rep protein and rolling circle replication pinpoints a virus as belonging to cressdnaviruses and helps researchers untangle the devilishly complex relationships and lineages found in the viral world.In addition to the Rep found in cressdnaviruses, cruciviruses contain another centrally important feature -- a capsid protein that is similar to that previously found only in RNA viruses. Capsids are vitally important, forming the outer shell or envelope that encloses the virus's identity -- its genetic sequence. The capsid shelters the vital nucleic acids sequestered within from digestion by host cell enzymes, enables virus particles to attach themselves to host cells and allows viruses to evade host cell defenses. Finally, capsids contain specialized features that give the virus its ability to puncture the host cell membrane and inject viral nucleic acid into the cell's cytoplasm.Analysis indicates that the capsid protein of cruciviruses is closely related to the capsid protein of another virus from the family Tombusviridae -- a single-stranded RNA virus known to infect plants. This hybrid viral character, containing both DNA- and RNA virus derived coding components, is what makes cruciviruses so unique.But how did a run-of-the-mill cressdnavirus come to acquire its RNA virus capsid protein coding sequence? This remains an issue of considerable debate, though presumably some form of lateral gene transfer occurred.Viruses can acquire genes from their immediate progenitors, the way genetic traits are passed from human parents to their offspring. Viruses, however, are far more genetically promiscuous, collecting new genes from the cells they infect, from other unrelated viruses and even from bacterial symbionts. (The phenomenon is also common among bacteria, which can use horizontal gene transfer to acquire antibiotic resistance.)Through some such mechanism, a cressdnavirus acquired an RNA virus capsid-like gene, creating the first crucivirus. It also appears that various cruciviruses have actively exchanged functional elements among themselves, further scrambling their evolutionary history.While the HOW of crucivirus DNA-RNA recombination remains mysterious, the WHY may be more straightforward. Clearly, the ability to borrow genetic traits from such distantly related viral sources could provide single-stranded DNA viruses with a considerable adaptive edge.In the current study, researchers explored a vast dataset including 461 cruciviruses and 10 capsid-encoding circular genetic elements identified from varied environments and organisms, making this the most expansive investigation of crucivirus sequences yet undertaken.The samples were found in environments ranging from temperate lakes to permafrost and lurking within organisms including red algae and invertebrates. The study points to the stramenopiles/alveolates/Rhizaria or SAR supergroup, (a diverse assemblage of eukaryotes, including many photosynthetic organisms) as the plausible candidate hosts for these unusual viruses, though this has yet to be verified.After examining the windfall of sequences, the researchers assembled similarity networks of cruciviral proteins with related viruses to try to better understand the twisting evolutionary paths that may have given rise to them, finding a rich cross-pollination of viral traits between many large families of viruses including Geminiviridae, Circoviridae, Nanoviridae, Alphasatellitidae, Genomoviridae, Bacilladnaviridae, Smacoviridae, and Redondoviridae.The findings may provide new insights into the early transition from RNA as the primary hereditary molecule of life to the adoption of more complex DNA genomes that has come to dominate life in the cellular world. The existence and behavior of cruciviruses suggest that viruses may have played a crucial role in this all-important transition, acting as a kind of genomic bridge between the RNA and DNA worlds, during the earliest emergence of life, though much more work is needed to explore these possibilities.Recombining in endless forms, viruses have become the planet's most ubiquitous biological entities, affecting every living organism and occupying every ecological niche. Increasingly, viruses are revealing themselves not only as agents of disease but as drivers of species evolution and vital actors in the molding of ecosystems.The expanded abilities of cruciviruses to borrow genomic elements from the most far-flung regions of viral sequence space suggest that entirely new virus groups may arise though prolific recombination events between distantly related forms.
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Biotechnology
| 2,020 |
October 30, 2020
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https://www.sciencedaily.com/releases/2020/10/201030122552.htm
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New cause of inflammation in people with HIV identified
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While current antiretroviral treatments for HIV are highly effective, data has shown that people living with HIV appear to experience accelerated aging and have shorter lifespans -- by up to five to 10 years -- compared to people without HIV. These outcomes have been associated with chronic inflammation, which could lead to the earlier onset of age-associated diseases, such as atherosclerosis, cancers, or neurocognitive decline. A new study led by researchers at Boston Medical Center examined what factors could be contributing to this inflammation, and they identified the inability to control HIV RNA production from existing HIV DNA as a potential key driver of inflammation. Published in
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After infection, HIV becomes a part of an infected person's DNA forever, and in most cases, infected cells are silent and do not replicate the virus. Occasionally, however, RNA is produced from this HIV DNA, which is a first step towards virus replication. Antiretroviral treatments help prevent HIV and AIDS-related complications, but they do not prevent the chronic inflammation that is common among people with HIV and is associated with mortality."Our study set out to identify a possible association between HIV latently infected cells with chronic inflammation in people with HIV who have suppressed viral loads," said Nina Lin, MD, a physician scientist at Boston Medical Center (BMC) and Boston University School of Medicine (BUSM).For this study, researchers had a cohort of 57 individuals with HIV who were treated with antiretroviral therapy. They compared inflammation in the blood and various virus measurements among younger (age less than 35 years) and older (age greater than 50 years) people living with HIV. They also compared the ability of the inflammation present in the blood to activate HIV production from the silent cells with the HIV genome. Their results suggest that an inability to control HIV RNA production even with antiretroviral drugs correlates with inflammation."Our findings suggest that novel treatments are needed to target the inflammation persistent in people living with HIV," said Manish Sagar, MD, an infectious diseases physician and researcher at BMC and the study's corresponding author. 'Current antiretroviral drugs prevent new infection, but they do not prevent HIV RNA production, which our results point as a potential key factor driving inflammation in people living with HIV."According to the Centers for Disease Control and Prevention, it is estimated that 1.2 million Americans are living with HIV; however, approximately 14 percent of these individuals are not aware that they are infected. Another CDC reporter found that of those diagnosed and undiagnosed with HIV in 2018, 76 percent had received some form of HIV care; 58 percent were retained in care; and 65 percent had undetectable or suppressed HIV viral loads. Antiretroviral therapy prevents HIV progression and puts the risk of transmission almost to zero.The authors note that these results need to be replicated in larger cohorts. "We hope that our study results will serve as a springboard for examining drugs that stop HIV RNA production as a way to reduce inflammation," added Sagar, also an associate professor of medicine and microbiology at BUSM.This study was supported in part by the National Institutes of Health (grant award numbers AG060890 and AI145661, the Boston University Genomic Science Institute and was facilitated by the Providence/Boston Center for AIDS Research.
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Biotechnology
| 2,020 |
October 30, 2020
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https://www.sciencedaily.com/releases/2020/10/201030111809.htm
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Beetroot peptide as potential drug candidate for treating diseases
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In a recent study, a research group led by Christian Gruber at MedUni Vienna's Institute of Pharmacology isolated a peptide (small protein molecule) from beetroot. The peptide is able to inhibit a particular enzyme that is responsible for the breakdown of messenger molecules in the body. Due to its particularly stable molecular structure and pharmacological properties, the beetroot peptide may be a good candidate for development of a drug to treat certain inflammatory diseases, such as e.g. neurodegenerative and autoimmune diseases.
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The peptide that occurs in the roots of beetroot plants belongs to a group of molecules that plants use inter alia as a chemical defence against pests such as e.g. bacteria, viruses or insects. "By analysing thousands of genomic data, our team was able to define a number of new cysteine-rich peptides and assign them phylogenetically in the plant kingdom. In this process, our attention was drawn to a possible function as so-called 'protease inhibitors'. The beetroot peptide can therefore inhibit enzymes that digest proteins," explains Gruber.The beetroot peptide specifically inhibits prolyl oligopeptidase (POP), which is involved in the breakdown of protein hormones in the body and is therefore able to regulate inflammatory reactions. POP is a much-discussed drug target for neurodegenerative and inflammatory diseases, such as Alzheimer's and multiple sclerosis, for example. "This means that, in future studies, this group of plant peptides called 'knottins', such as those found in beetroot, could potentially provide a drug candidate for treating these diseases."The peptide not only occurs in the root vegetables but can also be detected in commercially available beetroot juice -- albeit in very low concentrations. "Although beetroot counts as a very healthy vegetable, it would be unreasonable to hope that dementia could be prevented by regular consumption of beetroot," stresses the MedUni Vienna pharmacologist. "The peptide only occurs in very small quantities and it is not clear whether it can as such be absorbed via the gastrointestinal tract."The research work being conducted by Gruber's laboratory utilized the idea of harnessing Nature's blueprint to develop drug candidates. "We are searching through large databases containing genetic information of plants and animals, decoding new types of peptide molecules and studying their structure, aiming to test them pharmacologically on enzymes or cellular receptors (such as one of the prominent drug target classes, the so-called G protein-coupled receptors) and finally analysing them in the disease models," explains Gruber. Potential drug candidates are chemically synthesised in a slightly modified form based on the natural product, in order to obtain optimised pharmacological properties. This concept appears to be successful: a few years ago the research team generated a drug candidate T20K for MS with a synthesised plant peptide (cyclotide), which has recently been tested successfully in a Phase 1 trial by the Swedish firm Cyxone under a MedUni Vienna licence, and is now being prepared for a Phase 2 clinical trial.
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Biotechnology
| 2,020 |
October 30, 2020
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https://www.sciencedaily.com/releases/2020/10/201030111741.htm
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Microbes in the gut could be protective against hazardous radiation exposure
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A new study by scientists at UNC Lineberger Comprehensive Cancer Center and colleagues published Oct. 30, 2020, in
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The researchers noted that only an 'elite' set of mice had a high abundance of two types of bacteria, Lachnospiraceae and Enterococcaceae, in their guts that strongly countered the effects of the intense radiation. Importantly for humans, these two types of bacteria were found to be abundant in leukemia patients with mild GI symptoms who underwent radiotherapy.The study showed that the presence of the two bacteria led to an increased production of small molecules known as propionate and tryptophan. These metabolites provided long-term protection from radiation, lessened damage to bone marrow stem cell production, mitigated the development of severe gastrointestinal problems and reduced damage to DNA. Both metabolites can be purchased in some countries as health supplements but there is currently no evidence that the supplements could aid people exposed to intense forms of radiation."This truly trans-UNC collaborative effort showed that these beneficial bacteria caused a profound change in gut metabolites," said corresponding author Jenny P.Y. Ting, PhD, William Rand Kenan Professor of Genetics in the UNC School of Medicine and a UNC Lineberger immunology program co-leader.Damage to bodily organs from high levels of radiation, either from accidental exposure, cancer radiotherapy, targeted radiation attacks, among other forms of exposure, can lead to serious illness and even death. Blood cells in the body as well as tissues in the GI tract renew quickly and therefore remain particularly susceptible to radiation damage. On the protective side, however, the GI tract hosts over 10 trillion microbial microorganisms that could play an important role in limiting radiation-induced damage."Substantial federal efforts have been made to mitigate acute radiation symptoms -- however, it remains a long-standing and unresolved problem," said first study author Hao Guo, PhD, a postdoctoral fellow in Ting's lab. "Our work produced a comprehensive dataset of bacteria and metabolites that can serve as a powerful resource to identify actionable therapeutic targets in future microbiome studies."Because radiotherapy that is widely used to treat cancer often leads to GI side-effects, the investigators wanted to understand how their experiments in mice could translate to people. They worked with colleagues at Duke University, Memorial Sloan Kettering and Weill Cornell Medical College, and studied fecal samples from 21 leukemia patients due to receive radiation therapy as part of an arduous stem cell transplant conditioning. The scientists found that patients with shorter periods of diarrhea had significantly higher abundances of Lachnospiraceae and Enterococcaceae than patients with longer periods of diarrhea. These findings correlated closely with the researcher's findings in mice although Ting cautions that much larger studies are needed to verify these conclusions.Importantly for potential human use, in mice that were supplemented with Lachnospiraceae, the benefits of cancer radiotherapy were not lessened."Granulocyte-colony stimulating factor is the only drug that has been approved by the FDA as an effective countermeasure for high-dose radiation exposure, but it is expensive and has potential adverse side-effects," said Ting. "However, bacteria that we can cultivate, and especially metabolites that are relatively inexpensive and already elements in the food we eat, may be a good alternative."The researchers are hoping to launch a clinical trial soon in people to test the benefits of giving these metabolites to patients receiving radiation.
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Biotechnology
| 2,020 |
October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029171649.htm
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Comparing sensitivity of all genes to chemical exposure
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A University of Massachusetts Amherst environmental health scientist has used an unprecedented objective approach to identify which molecular mechanisms in mammals are the most sensitive to chemical exposures.
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The study, published in the journal "When we identified all the sensitive genes, we were very much surprised that almost every well-known molecular pathway is sensitive to chemicals to a certain degree," says lead author Alexander Suvorov, associate professor in the School of Public Health and Health Sciences.The study identified genes and molecular pathways most sensitive to chemical exposures, including mechanisms involving aging, lipid metabolism and autoimmune disease. "These findings for the first time prove that current epidemics in metabolic and autoimmune disorders may be partly due to a very broad range of chemical exposures," Suvorov says.To carry out their analysis, Suvorov and five students -- undergraduates Victoria Salemme, Joseph McGaunn and Menna Teffera, and graduate students Anthony Poluyanoff and Saira Amir -- extracted data on chemical-gene interactions from the Comparative Toxicogenomics Database, which includes human, rat and mouse genes.The UMass Amherst team created a database of 591,084 chemical-gene interactions reported in 2,169 studies that used high-throughput gene expression analysis, which means they looked at multiple genes. Low-throughout analysis focuses only on a single gene."In the recent past, everything that we knew about molecular mechanisms affected by chemicals was coming from low-throughput experiments," Suvorov says, which led toxicology researchers to focus on those already identified genes, rather than looking for chemical sensitivity among a fuller range of genes."I wanted to find some approach that would tell us in a completely unbiased way which mechanisms are sensitive and which are not. I wondered if we were missing a significant toxic response just because no one ever looked for it," Suvorov says. "By overlaying many high-throughput studies, we can see changes in the expression of all genes all at once. And that is unbiased because we are not cherry-picking any particular molecular mechanisms."The interactions analyzed encompassed 17,338 unique genes and 1,239 unique chemicals. The researchers split their database of chemicals into two parts -- pharmaceutical chemicals, which are designed to target known molecular cascades; and other chemicals such as industrial, agricultural, cosmetics and pollutants. When the sensitivity of genes to pharmaceutical chemicals was compared to the sensitivity of genes to the other chemicals, the results were the same. "That proves that when analysis is done on really big numbers of chemicals, their composition does not matter," Suvorov says.The study confirmed the molecular mechanisms that were previously recognized as being sensitive to chemical exposure, such as oxidative stress. The study's new findings that the pathways involving aging, lipid metabolism and autoimmune disease are also highly sensitive suggest that chemical exposures may have a role in such conditions as diabetes, fatty liver disease, lupus and rheumatoid arthritis, among others."This study represents a significant step forward in the use of genomic data for the improvement of public health policies and decisions," Suvorov says, "and the public health field will benefit from a future focus of toxicological research on these identified sensitive mechanisms."
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Biotechnology
| 2,020 |
October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029171646.htm
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Tuning biomolecular receptors for affinity and cooperativity
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Our biological processes rely on a system of communications -- cellular signals -- that set off chain reactions in and between target cells to produce a response. The first step in these often complex communications is the moment a molecule binds to a receptor on or in a cell, prompting changes that can trigger further signals that propagate across systems. From food tasting and blood oxygenation during breathing to drug therapy, receptor binding is the fundamental mechanism that unlocks a multitude of biological functions and responses.
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UC Santa Barbara researchers in chemist Kevin Plaxco's lab are deeply interested in the mechanics of biomolecular receptors, which have great potential biotechnology applications, including the design of biosensors. In a paper in the "There is a trade-off between cooperativity and affinity," said Gabriel Ortega, lead author of the study. This type of balancing act is common in nature, he added. "If you improve one property of a system, you're most likely making another property worse."And so it is with cooperativity, a property related to the ability of multibinding site receptors to respond to small changes in the concentration of their target molecule. Same goes for affinity, the concentration of the target molecule that is required for it to bind its receptor -- related to the receptors' sensitivity to the smallest concentrations of target molecule."Nature wants to achieve very tight regulation of all processes that occur in the body," Ortega explained. For that to happen, our bodies need to be able to distinguish between small changes in the concentration of target molecules, he said, and, in the case of cooperative binding, mount a more dramatic, more "all-or-none" response to changes in concentration."The most typical example is hemoglobin binding oxygen," Ortega said. Carried by blood cells, these proteins have four binding sites for oxygen, which they gather as blood flows through the lungs.The first binding event has the lowest affinity. "It acts like a gatekeeper, and it absorbs a lot of the signal," he said, "but once you occupy that lower affinity regime, the other sites, which have a higher affinity, bind more readily." He likens it to a system of connected pools where the first is the deepest and acts like a reservoir. Once it saturates, the rest fill up almost instantaneously."You want the hemoglobin to be able to completely capture the oxygen when it's in the lungs, and then completely release the oxygen when it's in the tissues," Ortega said, adding that many biological processes require such a digital-like response, in which receptors shuttle between nearly fully activated or nearly completely shut down in response to small changes in a signaling cue. Signals between brain and nerve cells operate this way, as do muscle cells.Cooperative receptors are also of interest in biological engineering. For example, they can be used to improve the precision with which biosensors measure their targets (by steepening the curve relating output to target concentration), which can be very beneficial for pharmaceutical applications where some drugs, such as chemotherapies, feature a very narrow range between ineffective and toxic.Here's the rub: To create that steeper, cooperative receptor, the first binding event has to have a low affinity, which means the overall affinity of the receptor site is lower than it would be if it were comprised only of its highest-affinity receptor. In areas like biosensing, this means the improved precision that comes with cooperativity is linked to an inability to detect the lowest concentrations of the target."If you make cooperativity better but it comes at the expense of pushing your detection capacity outside the window that you want to detect, then it's useless," Ortega said.The researchers explored a way to sidestep this seesaw with a method that can increase both affinity and cooperativity in their aptamer-based biosensors, and allow biosensor designers to fine tune between cooperativity and affinity."If you add more high-affinity binding sites, then you're still improving your responsiveness because you're still improving cooperativity, but now your overall affinity is going to be closer to the highest affinity site, thus improving your sensitivity," Ortega said.In the Plaxco Lab, aptamers -- single strands of DNA -- act as their multisite receptors, changing shape as they come into contact with target molecules (In this case, the chemotherapy drug doxorubicin). Two binding sites, one with low affinity and one with high affinity, produce a cooperative response with the overall affinity being the average of the two; a third high-affinity site pushes the average affinity higher while increasing cooperativity. The result? A sensor that can detect not only low doxorubicin concentrations but also minute changes in those concentrations.Meanwhile, added Ortega, adding another low-affinity receptor can increase cooperativity even further, albeit at the cost of reducing affinity a bit more."You're always going to get progressively more cooperativity and more affinity (relative to fewer binding sites)," Ortega said. "And by playing with the affinity of each individual binding site you can tailor your system to any affinity-cooperativity combination in between."The researchers plan to put their design to work to improve the aptamer-based sensors that they have already developed to detect clinically relevant molecules. They are moving toward in-vitro and in-vivo studies in which they deploy these sensors to detect the presence and concentration of target molecules in real-time. In addition, Ortega plans to use these new design principles to work in much more delicate and complex protein systems."I think that now that we have proof that these fundamental principles work, we can try to use them in proteins," he said.Research in this paper was conducted also by Frederick W. Dahlquist at UCSB, and Davide Mariottini, Alessandra Troina and Francesco Ricci at the University of Rome.
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Biotechnology
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October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029141949.htm
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Molecular compass for cell orientation
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Plants have veins that transport nutrients throughout their whole body. These veins are organized in a highly ordered manner. The plant hormone auxin travels directionally from cell-to-cell and provides cells with positional information, coordinating them during vein formation and regeneration. Until now, it remained a mystery how cells translate auxin signal into a formation of a complex system of veins. Scientists at the Institute of Science and Technology (IST) Austria discovered a molecular machinery that perceives a local auxin concentration and allows cells to synchronize their behavior to coordinate veins formation and regeneration. The scientists published their study in the journal
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The human body uses veins and blood to transport nutrients and oxygen throughout the body. Plants use a similar approach, the vascular systems. These veins transport nutrients for survival and define the size, structure, and position of new leaves and allow long-range communication between distant organs. Now, scientists from the group of Prof Jiri Friml at IST Austria, discovered how the plant hormone auxin dictates the newly formed veins' position. "Auxin decides which cells will differentiate into vascular tissue and orchestrates them to form intricate vein patterns," explains Jakub Hajný, who led the study. When cells have no ability to sense auxin signal, plant forms disorganized veins with disconnections that limit nutrients distribution. In case of mechanical damage, it also decreases regeneration after wounding.Already decades ago, scientists suspected that auxin is the vein-inducing signal organizing tissue into the formation of conserved vein patterns. However, scientists could not understand how the cells decrypt this chemical signal into a cellular response so far. The Friml group managed to identify the responsible proteins, called CAMEL and CANAR which serves as auxin sensor. The CAMEL/CANAR complex most likely perceives the auxin concentration in the neighborhood and allows cells to synchronize their orientations to create continuous veins. "It is basically a molecular compass for cell orientation, only instead of a magnetic field, it detects auxin concentration," explains Jakub Hajný. Thus, the team discovered molecular machinery underlying auxin-mediated vein formation and regeneration.
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Biotechnology
| 2,020 |
October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029115822.htm
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Identifying biomolecule fragments in ionizing radiation
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When living cells are bombarded with fast, heavy ions, their interactions with water molecules can produce randomly scattered 'secondary' electrons with a wide range of energies. These electrons can then go on to trigger potentially damaging reactions in nearby biological molecules, producing electrically charged fragments. So far, however, researchers have yet to determine the precise energies at which secondary electrons produce certain fragments. In a new study published in
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Through a better understanding of how biomolecules such as DNA are damaged by ionising radiation, researchers could make important new advances towards more effective cancer therapies. Like molecular bullets, heavy ions will leave behind nanometre-scale tracks as they pass through water; scattering secondary electrons as they deposit their energy. These electrons may then either attach themselves to nearby molecules if they have lower energies, potentially causing them to fragment afterwards; or they may trigger more direct fragmentation if they have higher energies. Since water comprises 70% of all molecules in living cells, this effect is particularly pronounced in biological tissues.In their previous research, Tsuchida's team bombarded liquid droplets containing the amino acid glycine with fast, heavy carbon ions, then identified the resulting fragments using mass spectrometry. Drawing on these results, the researchers have now used computer models incorporating random sampling methods to simulate secondary electron scattering along a carbon ion's water track. This allowed them to calculate the precise energy spectra of secondary electrons produced during ion bombardment; revealing how they related to the different types of glycine fragment produced. Through this approach, Tsuchida and colleagues showed that while electrons with energies lower 13 electronvolts (eV) went on to produce negatively charged fragments including ionised cyanide and formate, those in the range between 13eV and 100eV created positive fragments such as methylene amine.
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Biotechnology
| 2,020 |
October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029105032.htm
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new gelatin microcarrier for cell production
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Researchers from Singapore-MIT Alliance for Research and Technology (SMART), MIT's research enterprise in Singapore, have developed a novel microcarrier for large-scale cell production and expansion that offers higher yield and cost-effectiveness compared to traditional methods, and reduces steps required in the cell retrieval process. Microcarriers are particles used in bioreactor-based cell manufacturing of anchorage-dependent cells.
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SMART's newly developed dissolvable gelatin-based microcarrier has proven useful for expansion of mesenchymal stromal cells (MSCs), a cell type of great current interest as they can be isolated from adult tissues and further expanded to treat various ailments such as bone and cartilage defects and the body's rejection of foreign bone-marrow and cells (called graft vs. host disease). This dissolvability of the microcarriers also eliminates an additional separation step to retrieve the cells from the microcarriers. This reduces the complexity of cell manufacturing and increasing the ease with which the therapeutic cells can be harvested to make the product for patients.SMART's Critical Analytics for Manufacturing Personalized-Medicine (CAMP) Interdisciplinary Research Group (IRG) discovered that gelatin microcarriers, which fully dissolves in enzymatic treatment, can be useful in the cell recovery step -- one of the current bottlenecks faced in 3D microcarrier culture. The novel gelatin microcarrier showed higher yield and reduced cell loss at the cell harvesting step compared to commercial microcarriers, with comparable cell attachment efficiency and proliferation rate.Their discovery is explained in a paper titled "Dissolvable gelatin-based microcarriers generated through droplet microfluidics for expansion and culture of mesenchymal stromal cells" published in Biotechnology Journal and co-authored by researchers from SMART CAMP, Massachusetts Institute of Technology (MIT), National University of Singapore (NUS) and City University of Hong Kong (CityU)."Our study achieved over 90% harvest rate of cells grown on the gelatin microcarriers, which is significantly higher than the 50-60% harvest rate seen in current standards," said Dr. Ee Xien Ng, lead author of the paper and CAMP alumnus. "Using gelatin microcarriers also achieved tight control over microcarrier dimensions (for example, microcarrier diameter and stiffness) that facilitate uniform environmental conditions for controlling consistent cell numbers per microcarrier."The research also showed that MSCs cultured by gelatin microcarriers retain critical quality attributes of retrieved cells, such as a higher degree of trilineage multipotency with more balanced differentiation performance compared to commercial microcarriers. Most commercial microcarriers showed similar trends in adipogenic differentiation efficiency, while losing some degrees of chondrogenic and osteogenic differentiation capability."Innovations in microcarriers will aid in the scalability of certain cell types such as mesenchymal stromal cells for cell-based therapy, including for regenerative medicine applications," says Professor Krystyn J. Van Vliet, co-author of the paper as well as Lead Principal Investigator at CAMP and Professor of Materials Science and Engineering and Biological Engineering at MIT. "Developing a microcarrier platform for MSC culture has been a key part for SMART CAMP's understanding and managing the critical quality attributes of these cell therapy products. We hope our findings help bring about better, more efficient and scalable cell therapies with predictable therapeutic outcomes for multiple patient needs, and high harvesting efficiency of those potent cells."While the study focused on whether gelatin microcarriers are suitable for MSC culture and expansion, the team's research could potentially be extended for other types of anchorage-dependent cells.The research is carried out by SMART and supported by the National Research Foundation (NRF) Singapore under its Campus for Research Excellence And Technological Enterprise (CREATE) programme.
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Biotechnology
| 2,020 |
October 29, 2020
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https://www.sciencedaily.com/releases/2020/10/201029105022.htm
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Mouse studies link some autism to brain cells that guide sociability and platonic love
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Johns Hopkins Medicine researchers report that new experiments with genetically engineered mice have found clear connections among a range of autism types and abnormalities in brain cells whose chemical output forges platonic (non-sexual) feelings of love and sociability.
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The findings, the researchers say, could eventually fuel the development of autism therapies that target disease symptoms spurred on by abnormalities in parvocellular oxytocin neurons, which are brain cells in the hypothalamus of mammals.A report on the experiments was published online Oct. 27 in The investigators pursued evidence of the connections because of long-known variations in forms and symptoms of autism spectrum disorders, and because those with Fragile X -- an inherited disorder that occurs in one in 4,000 males and one in 6,000 females -- frequently is characterized by the inability to form close social bonds."Autism is defined by impaired social behaviors, but not all social behaviors are the same," says Gül Dölen, M.D., Ph.D., associate professor of neuroscience at the Johns Hopkins University School of Medicine. "People with autism generally have less difficulty with developing very close, family bonds than with friendships. Our experiments provide evidence that these two types of affection are encoded by different types of oxytocin neurons, and that disruption of one of these types of neurons is responsible for the characteristic social impairments seen in autism."For more than a century, Dölen says, scientists have known there are two types of neurons in the hypothalamus. The neurons release the so-called "love hormone" oxytocin, which induces contractions during childbirth, reduces stress and fosters bonding among animals across mammalian species, including humans.A magnocellular oxytocin neuron, which is one type of oxytocin-releasing neuron, releases huge quantities of oxytocin to the brain and body -- as much as 500 times or more than is released by parvocellular oxytocin neurons, which limit their scope and avoid flooding the body with all-consuming feelings of love.As their name suggests, magnocellular oxytocin neurons are larger than other neurons and can send their arm-like axons beyond the blood-brain barrier. Among their functions, magnocellular oxytocin neurons stir filial love -- what Dölen calls "mad love" -- and bonding between infants and mothers, and between sexual partners.Dölen's research shows that parvocellular oxytocin neurons, which comes from the Greek word "parvo" or "small" -- also encode social behaviors, but a different kind than the magnocellular neurons encode. While magnocellular oxytocin neurons encode social behaviors related to reproduction (pair bonding and parental bonding), parvocellular oxytocin neurons encode social behaviors related to what Dölen calls "love in moderation," or the platonic love that is important to communities (friends and colleagues).To study if and how autism symptoms are associated with disruptions in either or both of magnocellular and parvocellular neurons, Dölen and her team first genetically engineered mice to glow a fluorescent light in all oxytocin neurons, magno and parvo. Then, knowing that magnocellular neurons project their axons and chemicals beyond the blood/brain barrier, the research team used dyes that stay within the barrier to mark only the parvocellular neurons -- which are rarer and harder to detect, as well as smaller in size.Next, Dölen enlisted the help of Johns Hopkins scientist Loyal Goff, Ph.D., an expert in charting the genetic profile of individual cells. The technique, called single cell sequencing, specifically reads an individual cell's RNA -- a genetic cousin to DNA -- which indicates how the cell's genetic code is being read and which proteins are being produced. The way our genetic code is read makes one cell type different from another."This study is a comprehensive characterization of two types of closely-related neurons involved in the regulation of social behavior," says Goff, assistant professor of genetic medicine at the Johns Hopkins University School of Medicine. "One of the things that makes this study so unique is the multi-modal aspect of this characterization; relating anatomical, morphological, electrophysiological, transcriptional, genetic, and behavioral features to fully define the relevant and important differences between these two types of neurons."The research team used single cell sequencing and other gene-tracking tools and techniques to ensure that the subpopulations of magnocellular and parvocellular neurons were, indeed, distinct, so that they could genetically alter each group to determine if a change would induce autism-like behaviors in mice. What the researchers measured included how much the mice liked their social interactions and how much they preferred things associated with those social interactions (such as bedding).To re-create a model of autism in mice, the scientists turned to the FMR1 gene, which is linked to Fragile X, an inherited disorder characterized by intellectual disability, but also one of the most commonly identified causes of autism, occurring in about five percent of people with the condition.In humans, the FMR1 gene is silenced through a cellular process that adds chemicals called methyl groups to the gene. This same process does not occur in mice, so to replicate the FMR1 gene abnormality, the scientists genetically engineered the mice to have no functioning FMR1 gene either throughout the brain or only in parvocellular neurons.The researchers studied how mice without FMR1 valued the rewards from forming a social bond with an adult female mouse serving as a surrogate parent. These mice learned to like bedding associated with the surrogate parent, but not bedding associated with social interactions with peer mice -- evidence that mutations in genes that cause autism selectively disrupt platonic love, but spare filial love.When the scientists deleted the FMR1 gene in parvocellular cells only, not magnocellular cells, the mice had the same reaction: intact affinity for things associated with their surrogate parent, compared with things associated with peer mice. The scientists found no such preference in mice lacking FMR1 in oxytocin magnocellular cells.In a further set of experiments to pin down the specificity of their findings with the oxytocin-producing neurons, the scientists studied how certain genes linked to risk for autism were turned on or off, or expressed, among the two types of oxytocin neurons. They found that significantly more autism risk genes had higher expression levels in parvocellular neurons compared with magnocellular neurons. However, when the scientists looked at genes for schizophrenia, Alzheimer's disease and diabetes, there were no such differences in gene expression between the two oxytocin neuron types."This tells us that the difference we are seeing between the two types of oxytocin neurons relates to the disease that is characterized by impaired social behaviors, but not diseases where this behavior is not a defining symptom," says Dölen.She also notes, "What may be happening in the brain is that even though all brain cells may carry a particular mutation associated with autism, some neurons are more vulnerable to the symptoms related to social bonding."Dölen plans to conduct similar studies on genes associated with other types of autism. She says her work may indicate that drugs currently being tested for autism -- such as intranasal oxytocin -- could prove ineffective because the treatments target magnocellular neurons, which the new study indicates is not central to the disease. Instead, she says, their evidence suggests that parvocellular oxytocin neurons should be the focus of drug development for autism.Other scientists who conducted the research include Eastman Lewis, Genevieve Stein-O'Brien, Alejandra Patino, Romain Nardou, Cooper Grossman, Matthew Brown, Bidii Bangamwabo, Ndeye Ndiaye and Daniel Giovinazzo from Johns Hopkins, and Ian Dardani and Connie Jiang from the University of Pennsylvania.This research was funded by the National Institutes of Health's National Institute of Mental Health (R56MH115177 and R01MH117127), the Chan-Zuckerberg Initiative and the National Science Foundation.
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Biotechnology
| 2,020 |
October 28, 2020
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https://www.sciencedaily.com/releases/2020/10/201028134036.htm
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Learning the language of sugars
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We're told from a young age not to eat too much sugar, but in reality, our bodies are full of the stuff. The surface of every living cell, and even viruses, is covered in a mess of glycans: long, branching chains of simple sugars linked together by covalent bonds. These cell-surface sugars are crucial for regulating cell-cell contact, including the attachment of bacteria to healthy host cells. Glycans are also found on all other biological polymers, including proteins and RNA, and their presence impacts the polymers' stability and function.
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Despite their ubiquity and importance, glycans remain poorly understood because of their complexity. Rather than just the four nucleotide "letters" that make up DNA and RNA molecules, glycans have an "alphabet" of hundreds of different monosaccharides that can be strung together into sequences with a seemingly infinite array of lengths and branches. In addition, an individual glycan sequence can be changed due to the interplay of multiple enzymes and conditions both within and outside a cell, without the need for genetic mutations.Now, a team of scientists from the Wyss Institute for Biologically Inspired Engineering at Harvard University and the Massachusetts Institute of Technology (MIT) has cracked the glycan code by developing new machine learning and bioinformatics methods that enable researchers to systematically study glycans and identify sequences that play a role in the interactions of microbes and their host cells, as well as other still-unknown functions. The tools are presented in a new paper published today in "The language-based models that we have created can be used to predict whether and how a given glycan will be detected by the human immune system, thus helping us determine whether a strain of bacteria that harbors that glycan on its surface is likely to be pathogenic," said first author Daniel Bojar, Ph.D., a Postdoctoral Fellow at the Wyss Institute and MIT. "These resources also enable the study of glycan sequences involved in molecular mimicry and immune evasion, expanding our understanding of host-microbe interactions."Because glycans are the outermost layer of all living cell types, they are necessarily involved in the process of infection, both in the interaction of a prokaryotic bacterium binding to a eukaryotic host cell, and the interactions between the cells of the immune system. This has created an evolutionary arms race, in which bacterial glycans evolve to mimic those found on their hosts' cells to evade immune detection, and hosts' glycans are modified so that pathogens can no longer use them to gain access. To trace this history of glycan sequence development and identify meaningful trends and patterns, the research team turned to machine learning algorithms -- specifically, natural language processing (NLP), which has previously demonstrated success in analyzing other biopolymers, like RNA and proteins."Languages are actually quite similar to molecular sequences: the order of the elements matters, elements that are not next to each other can still influence each other, and their structures evolve over time," said co-author Rani Powers, Ph.D., a Senior Staff Scientist at the Wyss Institute.First, the team needed to assemble a large database of glycan sequences on which an NLP-based algorithm could be trained. They combed through existing datasets both online and in the academic literature to create a database of 19,299 unique glycan sequences, which they dubbed SugarBase. Within SugarBase they identified 1,027 unique glycan molecules or bonds they termed "glycoletters" making up the glycan alphabet, which could theoretically be combined into "glycowords" that the team defined as three glycoletters and two bonds.To develop an NLP-based model that could analyze sequences of glycoletters and pick out distinct glycowords, the team chose to use a bidirectional recurrent neural network (RNN). RNNs, which also underlie the "autocomplete" feature of text messaging and email software, predict the next word in a sequence given the preceding words, enabling them to learn complex, order-dependent interactions. They trained their glycoletter-based language model, dubbed SweetTalk, on sequences from SugarBase, and used it to predict the next most probable glycoletter in a glycan sequence based on the preceding glycoletters, in the context of glycowords.SweetTalk revealed that from the close to 1.2 trillion theoretically possible glycowords, only 19,866 distinct glycowords (~0.0000016%) were present in the database of existing glycans. The observed glycowords also tended to be clustered together in groups with highly similar sequences, partly indicating the taxonomic groups in which the glycowords are found, rather than distributed evenly among all possible sequence combinations. These outcomes likely reflect the high "cost" to an organism of evolving dedicated enzymes to construct specific glycan substructures -- in that scenario, it is more evolutionarily efficient to tweak existing glycowords rather than generate completely new ones.Given the important role glycans play in human immunity, the researchers fine-tuned SweetTalk using a smaller, curated list of glycans that are known from the literature to cause an immune response. When predicting the immunogenicity of glycan sequences from SugarBase, the SweetTalk model achieved an accuracy of ~92%, compared to an accuracy of ~51% for a model trained on scrambled glycan sequences. For example, glycans that are rich in a simple sugar called rhamnose, which is found in bacteria but not in mammals, were unambiguously labeled as immunogenic by SweetTalk. The model's excellent performance indicated that language-based models could be used to study characteristics of glycans on a large scale and with many potential applications, such as the exploration of glycan-immune system interactions.Based on the success of their first glycan-focused deep learning model, the team had a hunch that deep learning could also illuminate the "family tree" of glycan sequences. To achieve this, they constructed a language model-based classifier called SweetOrigins. They first pre-trained SweetOrigins with a SweetTalk model, then used the language-like properties of glycans to fine-tune the new model on a different task: predicting the taxonomic group of glycans by learning species-specific features of glycans that indicate their evolutionary history. They replicated this structure for each level of classification, from individual species all the way up to domains (e.g., Bacteria, Eukarya), creating eight SweetOrigins models that were able to classify the taxonomic group of a glycan with high accuracy. For example, the model accurately predicted glycans from the kingdoms Animalia (91.1%) and Bacteria (97.2%), allowing a glycan of unknown origin to be quickly classified as either animal-associated, microbe-associated, or found on both cell types.The researchers then used SweetOrigins to investigate host-pathogen interactions, reasoning that differences in the glycans associated with various strains of E. coli bacteria could be used to predict how infectious the strains are. They trained a deep learning-based classifier with the same language model architecture as SweetOrigins on E. coli-specific glycan sequences, and were able to predict E. coli strain pathogenicity with an accuracy of ~89%. It also placed the majority of glycans that are associated with E. coli strains of unknown pathogenicity at various places along the spectrum of infectiousness, helping to identify strains that are likely to be pathogenic to humans."Interestingly, the glycans that our model predicts are most associated with infection bear a striking resemblance to glycans found on the cells that form the mucosal barriers in animals' bodies, which keep pathogens out," said Diogo Camacho, Ph.D., a co-corresponding author of the paper and Senior Bioinformatics Scientist at the Wyss Institute. "This suggests that the glycans on pathogenic bacteria have evolved to mimic those found on the hosts' cells, facilitating their entry and evasion of the immune system."To more deeply probe how glycans function in host-microbe interactions, the team developed a glycan sequence alignment method, which compares individual glycan sequences to determine regions that are conserved between glycans and, therefore, likely serve a similar function. They chose a specific polysaccharide sequence from the pathogen Staphylococcus aureus that is known to increase bacterial virulence and hypothesized that this glycan helped the bacterium escape immune detection. When they compared that polysaccharide to similar glycan sequences in the dataset, they found the best alignment result with the enterobacterial common antigen (ECA), a glycan found on the Enterobacteriaceae family of symbiotic and pathogenic bacteria.The team also found ECA-like sequences associated with bacteria in the Staphylococcus, Acinetobacter, and Haemophilus genera, which are not part of the Enterobacteriaceae family that typically carries the ECA. This insight suggests that, in addition to mimicking the glycans found on their hosts, bacterial glycans can also evolve to mimic those found on other bacteria such as those in our microbiome, and that pathogenicity can arise via glycans on microbes that are not traditionally thought to be dangerous."The resources we developed here -- SugarBase, SweetTalk, and SweetOrigins -- enable the rapid discovery, understanding, and utilization of glycan sequences, and can predict the pathogenic potential of bacterial strains based on their glycans," said co-corresponding author Jim Collins, Ph.D., a Wyss Core Faculty member who is also the Termeer Professor of Medical Engineering & Science at MIT. "As glycobiology progresses, these tools can be readily expanded and updated, eventually allowing for the precise classification of glycans and facilitating the glycan-based study of host-microbe interactions at unprecedented resolution, potentially leading to new antimicrobial therapeutics.""This achievement is yet another example of the power of applying computational approaches to biological problems that have so far defied resolution because of their complexity. I am also very impressed with this team for making their tools openly available to researchers around the world, which promises to accelerate the pace of our collective understanding of glycans and their impact on human health," said Wyss Institute Founding Director Don Ingber, M.D., Ph.D. Ingber is also the Judah Folkman Professor of Vascular Biology at Harvard Medical School and the Vascular Biology Program at Boston Children's Hospital, as well as Professor of Bioengineering at Harvard's John A. Paulson School of Engineering and Applied Sciences.This research was supported by the Predictive BioAnalytics Initiative at the Wyss Institute for Biologically Inspired Engineering.
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Biotechnology
| 2,020 |
October 28, 2020
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https://www.sciencedaily.com/releases/2020/10/201028124532.htm
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Models for potential precursors of cells endure simulated early-Earth conditions
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Membraneless compartments -- models for a potential step in the early evolution of cells -- have been shown to persist or form, disappear, and reform in predictable ways through multiple cycles of dehydration and rehydration. Such wet-dry cycles were likely common conditions during the early development of life on Earth and could be a driving force for reactions important for the evolution of life.
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Understanding how the compartments -- known as complex coacervates -- respond to wet-dry cycling also informs current applications of the droplets, which are found in many household items, such as adhesives, cosmetics, fragrances, and food, and could be used in drug delivery systems. A paper describing the research, led by Penn State scientists, appears October 27, 2020 in the journal "Wet-dry cycling has gotten attention recently in attempts to produce molecules that could be the precursors to life, things like the building blocks of RNA, DNA, and proteins," said Hadi Fares, a NASA Postdoctoral Program Fellow at Penn State and the first author of the paper. "We are looking into a possible step further in the evolution of life. If these building blocks form compartments -- the precursors of cells -- what happens if they undergo the same type of wet-dry cycling?"The researchers make membraneless compartments, which form through liquid-liquid phase separation in a manner akin to oil droplets forming as a salad dressing separates, by controlling the concentrations of reagents in a solution. When the conditions -- pH, temperature, salt and polymer concentrations -- are right, droplets form that contain higher concentrations of the polymers than the surrounding solution. Like oil drops in water, there is no physical barrier or membrane that separates the droplets from their surroundings.Dehydrating the solution, like what could happen during dry periods on a pre-life Earth where small ponds or puddles might regularly dry up, changes all of these factors. The researchers, therefore, wanted to know what would happen to the membraneless compartments in their experimental system if they recreated these wet-dry cycles."We first mapped out how the compartments form when we alter the concentrations of the polymers and the salt," said Fares. "This 'phase diagram' is experimentally determined and represents the physical chemistry of the system. So, we know whether or not droplets will form for different concentrations of polymers and salt. We can then start with a solution with concentrations at any point on this phase diagram and see what happens when we dehydrate the sample."If the researchers start with a solution with concentrations that favor the formation of droplets, dehydration can change the concentrations such that the droplets disappear. The droplets then reappear when the sample is rehydrated. They can also start with a solution in which no droplets form and dehydration could bring the concentrations into the range that droplets begin to form. The behavior of the droplets during dehydration and rehydration match the predictions based on the experimentally derived phase diagram and they continue to do so through several iterations of the wet-dry cycle.Next, the researchers addressed the ability of droplets to incorporate RNA molecules inside of the membraneless compartments. The "RNA world" hypothesis suggests that RNA may have played an important role in the early evolution of life on Earth and previous experimental work has shown that RNA in these solutions becomes concentrated inside of the droplets."As we dry droplets that contain RNA, the overall concentration of RNA in the solution increases but the concentration of RNA inside the droplets remains fairly stable," said Fares. "The preference of RNA molecules to be inside the droplets seems to decrease. We believe that this is because as they dry the composition inside the droplets is changing to look more like the composition outside the droplets."The research team also looked at the ability of RNA to move into and within the droplets during dehydration. As they dry the sample the movement of RNA into and out of the droplets increases massively, but movement within the droplets increases only modestly. This difference in RNA mobility could have implications for the exchange of RNA among droplets during dehydration, which could in turn be functionally important in protocells."What we are showing is that as the membraneless compartments dry, they are able to preserve, at least to some extent, their internal environment," said Fares. "Importantly, the behavior of the coacervates, or protocells, whether they persist or disappear and reappear through the wet-dry cycle, is predicable from the physical chemistry of the system. We can therefore use this model system to think about the chemistry that might have been important for the early evolution of life."Beyond early life scenarios, the research has implications much closer to home."People underestimate how important coacervates are beyond their role as a model for protocells," said Christine Keating, Distinguished Professor of Chemistry at Penn State and leader of the research team. "Many of the things that you have in your house that appear cloudy have coacervates in them. Any time you want to compartmentalize something, whether it's for drug delivery, a fragrance, a nutrient, or food product, coacervates may be involved. Understanding something new about the physical chemistry of the process of droplet formation will be important for all of these things."
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Biotechnology
| 2,020 |
October 28, 2020
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https://www.sciencedaily.com/releases/2020/10/201028110631.htm
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Genetic analysis system yields new insights into bacterial pneumonia
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A team of infectious disease researchers has developed a new method to identify virulence genes in Streptococcus pneumoniae, the leading cause of bacterial pneumonia. Using this technique in a mouse model of pneumonia, they were able to gain new insights into the progression of the disease and its interaction with the flu virus.
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"Bacterial pneumonia is a lot more common, and more deadly, after a viral infection. Historically, a lot of the deaths during flu outbreaks such as the 1918 pandemic have been attributed to pneumococcal pneumonia," said Jacqueline Kimmey, assistant professor of microbiology and environmental toxicology at UC Santa Cruz and co-first author of a paper on the new findings published October 28 in Kimmey and her colleagues developed a new method for performing functional gene analysis to identify the genes that drive virulence in S. pneumoniae. Their method builds on the powerful gene editing technology known as CRISPR, which can be modified to selectively silence targeted genes with a technique called CRISPR interference. The researchers created a pooled library of S. pneumoniae strains in which each of the bacteria's genes was targeted by CRISPR interference in one of the bacterial strains.The CRISPR interference system was inducible by the antibiotic doxycycline, so the genes were not silenced until the bacteria (which were resistant to the antibiotic) were introduced into mice given doxycycline-containing feed. In addition, a genetic "barcode" on the guide RNAs used to target the silenced genes enabled the researchers to easily track each strain after infection. With a single sequencing step, they could identify which strains had survived and caused infections in the mice."It's a very efficient way to shut off individual genes and find out which ones are important," Kimmey explained.The system also enabled the researchers to assess a crucial phase of the infection when most of the bacteria die off. Only a small number of bacteria survive this "bottleneck" and go on to cause invasive disease."The lungs are actually really good at clearing infection," Kimmey said. "Even when we gave mice quite a high load of bacteria, there was a huge bottleneck, and very few bacteria made it into the blood."The researchers estimated that as few as 25 bacterial cells could survive the bottleneck and cause disease. They also found a surprising amount of variation in the outcome of the bottleneck, even though the mice were genetically identical and were infected through a carefully controlled protocol. The effects of the bottleneck overshadowed the gene silencing effects, resulting in little difference between the control mice and those in which bacterial genes were silenced."There was no consistency in terms of which strains survived, and there was huge variability in the size of the bottleneck," Kimmey said. "We know there is a lot of variability in the clinical progression of the disease in humans, so it is very exciting to see so much variation in this highly controlled system."The researchers then added flu to the system, infecting the mice with type A influenza prior to introducing S. pneumoniae. In mice pre-infected with influenza, there was no bottleneck, and a relatively small dose of bacteria caused rampant infection in the lungs. This enabled the researchers to assess the effects of gene silencing on the virulence of the bacteria.The results pointed to several genes as having important roles in pneumococcal infections, including genes identified as virulence factors in previous studies, such as the bacterial capsule genes. Surprisingly, the gene for the bacteria's main toxin, pneumolysin, did not appear to be necessary for the development of infections. Together with other recent findings, this suggests that pneumolysin may be more important for transmission than for survival in the host, the researchers said.A mysterious aspect of S. pneumoniae infections is that it is a very common colonizer of the upper respiratory tract without causing disease in most people."We really don't know what controls that," Kimmey said. "There seems to be a large population of people who are colonized, and normally that's okay. But a viral infection may predispose them and increase the risk of bacterial pneumonia."To get a better understanding of the variable outcomes seen in this study, Kimmey said she plans to use the CRISPR interference system to study the progression of infections in greater detail. In clinical settings, variability in the progression of disease can be attributed to a wide range of factors. In this controlled study, the process of infection itself seemed to be highly variable."The system we developed gave us a very elegant way of showing the variability of outcomes and what seems like random variation in the course of infections, even in a controlled system," she said.
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Biotechnology
| 2,020 |
October 28, 2020
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https://www.sciencedaily.com/releases/2020/10/201028082954.htm
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Scientists discover new organic compounds that could have helped form the first cells
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Chemists studying how life started often focus on how modern biopolymers like peptides and nucleic acids contributed, but modern biopolymers don't form easily without help from living organisms. A possible solution to this paradox is that life started using different components, and many non-biological chemicals were likely abundant in the environment. A new survey conducted by an international team of chemists from the Earth-Life Science Institute (ELSI) at Tokyo Institute of Technology and other institutes from Malaysia, the Czech Republic, the US and India, has found that a diverse set of such compounds easily form polymers under primitive environmental conditions, and some even spontaneously form cell-like structures.
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Understanding how life started on Earth is one of the most challenging questions modern science attempts to explain. Scientists presently study modern organisms and try to see what aspects of their biochemistry are universal, and thus were probably present in the organisms from which they descended. The best guess is that life has thrived on Earth for at least 3.5 billion of Earth's 4.5 billion year history since the planet formed, and most scientists would say life likely began before there is good evidence for its existence. Problematically, since Earth's surface is dynamic, the earliest traces of life on Earth have not been preserved in the geological record. However, the earliest evidence for life on Earth tells us little about what the earliest organisms were made of, or what was going on inside their cells. "There is clearly a lot left to learn from prebiotic chemistry about how life may have arisen," says the study's co-author Jim Cleaves.A hallmark of life is evolution, and the mechanisms of evolution suggest that common traits can suddenly be displaced by rare and novel mutations which allow mutant organisms to survive better and proliferate, often replacing previously common organisms very rapidly. Paleontological, ecological and laboratory evidence suggests this occurs commonly and quickly. One example is an invasive organism like the dandelion, which was introduced to the Americas from Europe and is now a commo weed causing lawn-concerned homeowners to spend countless hours of effort and dollars to eradicate. Another less whimsical example is COVID-19, a virus (technically not living, but technically an organism) which was probably confined to a small population of bats for years, but suddenly spread among humans around the world. Organisms which reproduce faster than their competitors, even only slightly faster, quickly send their competitors to what Leon Trotsky termed the "ash heap of history." As most organisms which have ever existed are extinct, co-author Tony Z. Jia suggests that "to understand how modern biology emerged, it is important to study plausible non-biological chemistries or structures not currently present in modern biology which potentially went extinct as life complexified."This idea of evolutionary replacement is pushed to an extreme when scientists try to understand the origins of life. All modern organisms have a few core commonalities: all life is cellular, life uses DNA as an information storage molecule, and uses DNA to make ribonucleic RNA as an intermediary way to make proteins. Proteins perform most of the catalysis in modern biochemistry, and they are created using a very nearly universal "code" to make them from RNA. How this code came to be is in itself enigmatic, but these deep questions point to their possibly having been a very murky period in early biological evolution ~ 4 billion years ago during which almost none of the molecular features observed in modern biochemistry were present, and few if any of the ones that were present have been carried forward.Proteins are linear polymers of amino acids. These floppy strings of polymerised amino acids fold into unique three-dimensional shapes, forming extremely efficient catalysts which foster precise chemical reactions. In principle, many types of polymerised molecules could form similar strings and fold to form similar catalytic shapes, and synthetic chemists have already discovered many examples. "The point of this kind of study is finding functional polymers in plausibly prebiotic systems without the assistance of biology, including grad students," says co-author Irena Mamajanov.Scientists have found many ways to make biological organic compounds without the intervention of biology, and these mechanisms help explain these compounds' presence in samples like carbonaceous meteorites, which are relics of the early solar system, and which scientists don't think ever hosted life. These primordial meteorite samples also contain many other types of molecules which could have formed complex folded polymers like proteins, which could have helped steer primitive chemistry. Proteins, by virtue of their folding and catalysis mediate much of the complex biochemical evolution observed in living systems. The ELSI team reasoned that alternative polymers could have helped this occur before the coding between DNA and protein evolved. "Perhaps we cannot reverse-engineer the origin of life; it may be more productive to try and build it from scratch, and not necessarily using modern biomolecules. There were large reservoirs of non-biological chemicals that existed on the primeval Earth. How they helped in the formation of life-as-we-know-it is what we are interested in," says co-author Kuhan Chandru.The ELSI team did something simple yet profound: they took a large set of structurally diverse small organic molecules which could plausibly be made by prebiotic processes and tried to see if they could form polymers when evaporated from dilute solution. To their surprise, they found many of the primitive compounds could, though they also found some of them decomposed rapidly. This simple criterion, whether a compound is able to be dried without decomposing, may have been one of the earliest evolutionary selection pressures for primordial molecules.The team conducted one further simple test. They took these dried reactions, added water and looked at them under a microscope. To their surprise, some of the products of these reaction formed cell-sized compartments. That simple starting materials containing 10 to 20 atoms can be converted to self-organised cell-like aggregates containing millions of atoms provides startling insight into how simple chemistry may have led to complex chemistry bordering on the kind of complexity associated with living systems, while not using modern biochemicals."We didn't test every possible compound, but we tested a lot of possible compounds. The diversity of chemical behaviors we found was surprising, and suggests this kind of small-molecule to functional-aggregate behavior is a common feature of organic chemistry, which may make the origin of life a more common phenomenon than previously thought," concludes co-author Niraja Bapat.
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Biotechnology
| 2,020 |
October 27, 2020
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https://www.sciencedaily.com/releases/2020/10/201027143551.htm
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Scientists map structure of potent antibody against coronavirus
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Scientists at Fred Hutchinson Cancer Research Center in Seattle have shown that a potent antibody from a COVID-19 survivor interferes with a key feature on the surface of the coronavirus's distinctive spikes and induces critical pieces of those spikes to break off in the process.
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The antibody -- a tiny, Y-shaped protein that is one of the body's premier weapons against pathogens including viruses -- was isolated by the Fred Hutch team from a blood sample received from a Washington state patient in the early days of the pandemic.The team led by Drs. Leo Stamatatos, Andrew McGuire and Marie Pancera previously reported that, among dozens of different antibodies generated naturally by the patient, this one -- dubbed CV30 -- was 530 times more potent than any of its competitors.Using tools derived from high-energy physics, Hutch structural biologist Pancera and her postdoctoral fellow Dr. Nicholas Hurlburt have now mapped the molecular structure of CV30. They and their colleagues published their results online today in the journal The product of their research is a set of computer-generated 3D images that look to the untrained eye as an unruly mass of noodles. But to scientists they show the precise shapes of proteins comprising critical surface structures of antibodies, the coronavirus spike and the spike's binding site on human cells. The models depict how these structures can fit together like pieces of a 3D puzzle."Our study shows that this antibody neutralizes the virus with two mechanisms. One is that it overlaps the virus's target site on human cells, the other is that it induces shedding or dissociation of part of the spike from the rest," Pancera said.On the surface of the complex structure of the antibody is a spot on the tips of each of its floppy, Y-shaped arms. This infinitesimally small patch of molecules can neatly stretch across a spot on the coronavirus spike, a site that otherwise works like a grappling hook to grab onto a docking site on human cells.The target for those hooks is the ACE2 receptor, a protein found on the surfaces of cells that line human lung tissues and blood vessels. But if CV30 antibodies cover those hooks, the coronavirus cannot dock easily with the ACE2 receptor. Its ability to infect cells is blunted.This very effective antibody not only jams the business end of the coronavirus spike, it apparently causes a section of that spike, known as S1, to shear off. Hutch researcher McGuire and his laboratory team performed an experiment showing that, in the presence of this antibody, there is reduction of antibody binding over time, suggesting the S1 section was shed from the spike surface.The S1 protein plays a crucial role in helping the coronavirus to enter cells. Research indicates that after the spike makes initial contact with the ACE2 receptor, the S1 protein swings like a gate to help the virus fuse with the captured cell surface and slip inside. Once within a cell, the virus hijacks components of its gene and protein-making machinery to make multiple copies of itself that are ultimately released to infect other target cells.The incredibly small size of antibodies is difficult to comprehend. These proteins are so small they would appear to swarm like mosquitos around a virus whose structure can only be seen using the most powerful of microscopes. The tiny molecular features Pancera's team focused on the tips of the antibody protein are measured in nanometers -- billionths of a meter.Yet structural biologists equipped with the right tools can now build accurate 3D images of these proteins, deduce how parts of these structures fit like puzzle pieces, and even animate their interactions.Fred Hutch structural biologists developed 3D images of an antibody fished from the blood of an early COVID-19 survivor that efficiently neutralized the coronavirus.Dr. Nicholas Hurlburt, who helped develop the images, narrates this short video showing how that antibody interacts with the notorious spikes of the coronavirus, blocking their ability to bind to a receptor on human cells that otherwise presents a doorway to infection.Key to building models of these nanoscale proteins is the use of X-ray crystallography. Structural biologists determine the shapes of proteins by illuminating frozen, crystalized samples of these molecules with extremely powerful X-rays. The most powerful X-rays come from a gigantic instrument known as a synchrotron light source. Born from atom-smashing experiments dating back to the 1930s, a synchrotron is a ring of massively powerful magnets that are used to accelerate a stream of electrons around a circular track at close to the speed of light. Synchrotrons are so costly that only governments can build and operate them. There are only 40 of them in the world.Pancera's work used the Advanced Photon Source, a synchrotron at Argonne National Laboratory near Chicago, which is run by the University of Chicago and the U.S. Department of Energy. Argonne's ring is 1,200 feet in diameter and sits on an 80-acre site.As the electrons whiz around the synchrotron ring, they give off enormously powerful X-rays -- far brighter than the sun but delivered in flashes of beams smaller than a pinpoint.Structural biologists from around the world rely on these brilliant X-ray beamlines to illuminate frozen crystals of proteins. They reveal their structure in the way these bright beams are bent as they pass though the molecules. It takes powerful computers to translate the data readout from these synchrotron experiments into the images of proteins that are eventually completed by structural biologists.The Fred Hutch team's work on CV30 builds on that of other structural biologists who are studying a growing family of potent neutralizing antibodies against the coronavirus. The goal of most coronavirus vaccine candidates is to stimulate and train the immune system to make similar neutralizing antibodies, which can recognize the virus as an invader and stop COVID-19 infections before they can take hold.Neutralizing antibodies from the blood of recovered COVID-19 patients may also be infused into infected patients -- an experimental approach known as convalescent plasma therapy. The donated plasma contains a wide variety of different antibodies of varying potency. Although once thought promising, recent studies have cast doubt on its effectiveness.However, pharmaceutical companies are experimenting with combinations of potent neutralizing antibodies that can be grown in a laboratory. These "monoclonal antibody cocktails" can be produced at industrial scale for delivery by infusion to infected patients or given as prophylactic drugs to prevent infection. After coming down with COVID-19, President Trump received an experimental monoclonal antibody drug being tested in clinical trials by the biotech company Regeneron, and he attributes his apparently quick recovery to the advanced medical treatment he received.The Fred Hutch research team holds out hope that the protein they discovered, CV30, may prove to be useful in the prevention or treatment of COVID-19. To find out, this antibody, along with other candidate proteins their team is studying, need to be tested preclinically and then in human trials."It is too early to tell how good they might be," Pancera said.This work was supported by donations to the Fred Hutch COVID-19 Research Fund.
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Biotechnology
| 2,020 |
October 27, 2020
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https://www.sciencedaily.com/releases/2020/10/201027133723.htm
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Scientists uncover prophage defense mechanisms against phage attacks in mycobacteria
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A phage is a virus that invades a bacterial cell. While harmless to human cells, phages are potentially deadly to bacteria since many phages enter a cell in order to hijack its machinery in order to reproduce itself, thus destroying the cell.
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While this is bad news for bacteria, it may be good news for humans. There is a growing need to develop new treatments that effectively attack deadly strains of bacteria that have become resistant to other medicines. Already used with success in some parts of the world, phage therapy is gaining traction as a more widespread way to fight antibiotic-resistant bacterial infections and even, at some point, some viral infections including, according to a recent article, possibly COVID-19.Among the challenges: a virus type known as a prophage. A phage enters a bacterial cell and, instead of destroying it, takes up residence. Called a "prophage," it fights off other viruses' attempts to invade. According to Vassie Ware, a professor in Lehigh University's Department of Biological Sciences, many bacterial strains contain prophages. These prophages, she says, may provide defense systems that would make therapeutic uses of phages more challenging. In order to eradicate a pathogen, phages may need to overcome an already-in-residence prophage's defense systems.Ware and her team (former PhD student Catherine Mageeney, current PhD student Hamidu Mohammed and former undergraduate student Netta Cudkevich), collaborating with former Lehigh Chemical and Biomolecular Engineering and Bioengineering faculty member Javier Buceta and his team (former postdoctoral associate Marta Dies, recent PhD students Samira Anbari and Yanyan Chen), recently conducted a study that focused on a phage called Butters (discovered by Lena Ma in Lehigh's SEA-PHAGES Program in 2012) that attacks a bacterial strain related to mycobacteria that cause tuberculosis or other human infections.The group uncovered a two-component system of Butters prophage genes that encode proteins that "collaborate" to block entry and subsequent infection of some phages, but not others. While the Butters prophage cannot protect the bacterial cell against all phage attacks, they discovered that more than one defense system is present in the Butters prophage defense repertoire. These weapons, they discovered, are specific for different types of phages. These findings were published in an article earlier this month in "Previous findings by several members of our research team working with other collaborators showed that prophages express genes that defend their bacterial host from infection by some specific groups of phages. For Butters, no genes involved in defense against specific phages had been previously identified," says Ware. "With our experimental approach, we expected to identify genes involved in defense against infection by several phages, but were not expecting to uncover interactions between the two proteins that affected how one of the proteins functions in defense."The Ware/Buceta team used a multidisciplinary approach to identify the genes and interactions. They utilized bioinformatics tools to predict structural features of proteins encoded by genes expressed by the Butters prophage and to probe databases for the presence of Butters genes within known bacterial strains. Molecular biology techniques were used to engineer mycobacterial strains to express phage genes from the prophage. Microbiology experiments included immunity plating efficiency assays for each engineered bacterial strain to determine if the gene in question would protect the engineered bacterial strain from infection by a particular phage type.This strategy, says Ware, allowed identification of specific genes as part of the defense mechanism against specific viral attack.They also conducted microscopy experiments for live-cell imaging to visualize the cellular location of phage proteins within engineered bacterial cells and to show a functional interaction between the phage proteins in question. Biochemical experiments determined that the phage proteins likely interact physically as part of the defense mechanism."Collectively, these approaches provided data that allowed the team to construct a model for how the Butters prophage two-component system may function in defense against specific viral attack," says Ware.Adds Ware: "The diversity of defense systems that exists demonstrates that efforts to establish generic sets of phage cocktails for phage therapy to kill pathogenic bacteria will likely be more challenging."In addition to advancing phage therapy development, the team's discovery may also be important for engineering phage-resistant bacteria that could be used in the food industry and in some biotechnology applications.
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Biotechnology
| 2,020 |
October 27, 2020
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https://www.sciencedaily.com/releases/2020/10/201027105411.htm
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Not all mutations are bad: Researchers identify differences between benign and pathogenic variants
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An international team of researchers led by Cleveland Clinic's Lerner Research Institute has performed for the first time a wide-scale characterization of missense variants from 1,330 disease-associated genes. Published in
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"Our study serves as a powerful resource for the translation of personal genomics to personal diagnostics and precision medicine, and can aid variant interpretation, inform experiments and help accelerate personalized drug discovery," said Dennis Lal, PhD, assistant staff, Genomic Medicine, and the study's lead author. Recent large-scale DNA sequencing efforts have detected millions of missense variants, where mistakes in the DNA code change the amino acid (molecular building block of a protein) makeup of proteins. Some of these variants are pathogenic, meaning they alter the structure and function of a protein in a way that leads to disease, while others are benign with no impact on health. The vast majority, however, are considered variants of uncertain significance because their effects remain unknown.While methods to predict variant pathogenicity exist, they do not elucidate why some variants are more or less likely to cause disease than others or establish their functional impact. Additionally, pathogenic and benign variants can co-exist in almost every disease-associated gene. As such, gaining a better understanding of the mechanistic differences between benign and pathogenic variants will be a critical next step in the development of novel therapies for genetic disorders.Considering that a protein's function is closely linked to its three-dimensional structure, in this study the research team identified and compared the protein features of amino acids affected by pathogenic versus benign missense variants. Features that are more frequently mutated in pathogenic variants compared to benign variants (3D mutational hotspots) are likely crucial to protein fitness and thus could help explain the molecular determinants of pathogenicity.Looking at 1,330 disease-associated genes, the researchers analyzed a set of 40 features and found that 18 were significantly associated with pathogenic variants, 14 were significantly associated with benign variants and the remaining eight had no significant association with any variant type."By considering genetic variation in the context of proteins' three-dimensional organization, we present for the first time an atlas of molecular properties of pathogenic mutations that addresses the differences between benign and disease-causing mutations," said Lal. "This study focused on 1,330 genes associated with rare types of genetic disorders, so we are currently extending our project to look at more genes and milder disorders." Data from this study (including precomputed P3DFiDAGS1330 and P3DFiProteinclass values for every possible amino acid exchange in proteins encoded by 1,330 disease-associated genes, along with the explicit listing of the 3D features of the altered site as the rationale for the index) is available through the dedicated web server MISCAST.Sumaiya Iqbala, PhD, is first author on the study, which was supported by the Stanley Center for Psychiatric Research. Iqbala is a post-doctoral research associate at the Broad Institute of MIT and Harvard and the Analytical and Translational Genetics Unit at Massachusetts General Hospital.
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Biotechnology
| 2,020 |
October 27, 2020
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https://www.sciencedaily.com/releases/2020/10/201027105352.htm
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Glowing mice shine light on night vision
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Publishing in
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These 'PKAchu' mice were developed to provide researchers with a way of visualizing the activation of one of the body's most essential and widely studied proteins."PKA is found in many cells and is involved in a wide variety of biological processes. It's natural that researchers would find a way to observe its activities," explains first author Shinya Sato of the Graduate School of Biostudies."PKAchu mice were developed in 2012 -- 'chu' being Japanese for 'squeak' -- to allow us to closely monitor how PKA acts during specific biological processes. I decided to apply this to my work in retina biology."The team first developed a method for recording high resolution, microscopic images of living retinal tissue. They then observed how PKA reacts to light stimulation. Knowing the pathways involved, the team hypothesized that light would deactivate PKA.But to their surprise, the exact opposite happened."We started with a six-second illumination of the tissue. Incredibly, this activated PKA in the selected area for nearly 15 minutes," continues Sato. "We then did a ten-minute illumination, during which PKA was inactive. But when the lights were turned off, PKA kicked into gear. It was as if the darkness had activated it."Single-cell level analysis revealed that this lights-off PKA activation occurred only in rod cells, which are indispensable for our night vision.Sato hypothesizes that this previously unknown mechanism of rod-specific PKA activation may be a key in boosting light sensitivity in our eyes, contributing to our night vision. Rod-type photoreceptor cells are thought to have evolved from color-sensing cone cells. PKA activation, it now appears, is rod-specific.Michiyuki Matsuda, the study's senior author, concludes: "We have not only uncovered many interesting aspects of retinal cells, but the further utility of PKAchu mice as well. We are excited to uncover the mechanisms and purpose behind these new findings, and perhaps illuminate our understanding of conditions such as night blindness."
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Biotechnology
| 2,020 |
October 26, 2020
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https://www.sciencedaily.com/releases/2020/10/201026135800.htm
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How cells use mechanical tension sensors to interact with their environment
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Actin is among the most abundant proteins in cells, and it has many jobs -- from giving the cell its very shape and structure to managing networks of proteins crucial to numerous cellular functions. Without it, the fragile fundamental unit of life would crumble.
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A good deal of actin's activity is based on mechanical signaling; it appears to somehow sense physical feedback from the environment and respond accordingly. Now, a new study in eLife describes how this mechanical signaling works. By literally stretching individual actin filaments, the researchers determined a process by which actin transmits cellular mechanical messages to other proteins. Because hundreds of different proteins bind actin filaments, the implications of this discovery are far-reaching and may ultimately explain how cells mechanically control movement -- including how cardiac cells contract or how motile cells move."The idea that actin filaments could potentially be tiny stretchy tension sensors in the cell has been banging around in the literature for a while, but I think we really proved it here," says Gregory M. Alushin, assistant professor and head of the Laboratory of Structural Biophysics and Mechanobiology. "We're just now beginning to understand the nuances of mechanical signaling in the cell."The lifecycle of a cell is intimately tied to chemical signaling, in which well-studied networks of ligands and receptors feed information to the cell and determine whether it grows, divides, migrates, or dies. But physical forces also pass crucial signals onto cells through an entirely different phenomenon in which molecules push and pull on each other, perpetually coming together and dissociating.While decades of research have shed light on the chemical signaling process, the particulars of mechanical signaling are still poorly understood. It is clear, for instance, that when a cell clings to the bottom of a petri dish or makes contact with neighboring cells, its interaction with its surroundings is driven by actin, which binds to so-called adhesion proteins in the cell's outer rim.But it remained unclear how cells pass a mechanical signal from their environment along to actin, and how actin then relays that signal to either beckon adhesion proteins or push them away."We've known about actin-binding adhesion proteins for many years," says Lin Mei, a graduate fellow in chemical biology at Rockefeller. "But before our study, there was no research proving that stretching actin conveys a mechanical signal to proteins that can sense this mechanical force."To further investigate, Alushin and Mei stretched actin -- quite literally.In collaboration with Rockefeller's Shixin Liu, the researchers undertook the painstaking task of suspending a single actin filament, which measures about 1/15,000th the width of a human hair, between two microscopic beads that anchor each end of the filament. They then exposed the filament to an adhesion protein known as α-catenin, and used an innovative technology called laser tweezers to pull on the actin protein just enough to mimic the minute tension it might experience in a cell.They observed that actin bound α-catenin better while they were pulling on it, implying that actin was transmitting a mechanical signal to α-catenin, and that α-catenin had the capacity to receive this signal.But a similar adhesion protein, vinculin, proved to be signal-deaf. With the help of advanced electron microscopy techniques, the researchers homed in on the crucial difference between α-catenin and vinculin. "There was this one peptide in α-catenin protein, a floppy little tail that partially folds only when it is bound to actin directly," Alushin says. "All other parts of the two proteins were essentially the same."He and his co-workers suspected that actin was relaying a perfectly good mechanical signal, but that only the floppy tail of α-catenin was prepared to receive it. Vinculin, bereft of floppy tail, was simply missing actin's call. To test this theory, they engineered a version of vinculin with an α-catenin tail transplanted onto it -- and the floppy-tailed vinculin began binding better when actin was stretched. Actin was the signal transmitter; the floppy tail, the receiver.Alushin notes that, while α-catenin and its floppy tail may eventually become an appealing target for clinical therapies, the new findings are first and foremost a coup for the burgeoning field of mechanobiology, which studies how mechanical forces drive crucial processes at the cellular level. "We know that α-catenin is critical in brain development and frequently mutated in cancer, but most of what we know about it is that, if you get rid of it, everything else in the cell breaks," he says. "By precisely defining the force-detector in α-catenin, we will enable researchers to figure out exactly what its function is in mechanical signaling.""We suspect that there are hundreds of other proteins that directly sense force transmitted by actin," adds Mei. "Our work provides the foundation and the molecular details to begin searching for all of the other force-sensitive proteins."
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Biotechnology
| 2,020 |
October 26, 2020
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https://www.sciencedaily.com/releases/2020/10/201026135750.htm
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Can scientists take the STING out of common respiratory viruses?
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University of North Carolina School of Medicine scientists have made a curious discovery about a well-known human protein that helps the immune system fight viral infections. The lab of Stan Lemon MD, and colleagues found that one class of viruses actually requires this protein to infect cells and replicate.
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Published in the "We found that a large proportion of these rhinoviruses, particularly the ones that cause severe disease, need a human protein called STING to make copies of its RNA," said Lemon, professor of medicine and microbiology and immunology at the UNC School of Medicine. "We don't know how or why; we'll have to study this further. But our work opens the door to a new strategy for controlling infection of these pesky and at times very dangerous pathogens."Viruses are relatively simple bugs able to infect human cells and then replicate to cause diseases from the common cold to COVID-19 and more dangerous pathogens, such as HIV and Ebola. Humans have developed some defenses against these invasions, and one part of the defense is called the 'stimulator of interferon gene' protein, or STING, so named for its ability to sense invaders and enhance our immune response to many viruses, including herpes viruses and cytomegalovirus, a common bug that infects half of adults by age 40 and causes symptoms similar to many other viral infections. Rhinoviruses turn STING against us, and use it to promote their own growth. The genomes of many viruses are made up of DNA, whereas the genomes of rhinoviruses are composed of RNA, a similar kind of genetic code at the foundation of all living things. STING helps us defend against DNA viruses, but is instead helping this RNA virus.Human rhinoviruses comprise a large group of common respiratory tract pathogens -- dozens of different viruses -- that are associated with asthma, pneumonia and exacerbations of chronic lung disease in both children and adults. There are no vaccines available to prevent these common infections because the viruses are very diverse and different from each other in terms of how they are 'seen' by the human immune system."If you are immune to one, you can easily catch another," Lemon said. "There is also no effective antiviral therapy for any of them."Lemon's lab studies how viruses like rhinoviruses interact with human proteins with the hope of finding a way to block or limit the interaction and, as a result, limit symptoms and disease progression. While studying the human proteins involved in this interaction, they found something completely unexpected. Using cell cultures, they employed experimental techniques to disable STING to see what happened. To their surprise, the virus could not infect cells and replication was stopped."It could be possible to target this protein with a small molecule in way that would benefit people with rhinoviruses, especially children and others who can become severely ill," Lemon said.
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Biotechnology
| 2,020 |
October 23, 2020
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https://www.sciencedaily.com/releases/2020/10/201023141012.htm
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SPOTlight supercharges cell studies
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Researchers at Rice University and Baylor College of Medicine have developed a new method to isolate specific cells, and in the process found a more robust fluorescent protein.
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Both the platform and the protein could be highly useful to synthetic biologists and biomedical researchers. They often need to single out cells with specific visual phenotypes like shape or activity determined by their genetic or epigenetic makeup or their developmental history.Rice graduate student Jihwan (James) Lee and François St-Pierre, an assistant professor of neuroscience at Baylor College of Medicine and an adjunct assistant professor of electrical and computer engineering at Rice, and their team reported their results in Science Advances.Lee and his colleagues dubbed their platform SPOTlight, short for Single-cell Phenotypic Observation and Tagging with Light. It addresses the limitations of existing sorting techniques to isolate single live cells with unique profiles from heterogenous populations.They then leveraged the method for protein engineering to develop the most photostable yellow fluorescent protein reported to date."We basically developed a platform that allows one to screen for spatial and temporal properties of individual cells," said Lee, the first author and a student in Rice's Systems, Synthetic and Physical Biology program working in St-Pierre's Baylor lab."This is done by first observing the cells under a microscope," he said. "The cells express a special protein so that shining a spot of light on desired cells make them go red. We can then easily separate red cells from the rest using a common device called a flow cytometer."That "special" photoactivatable fluorescent protein irreversibly transitions from dark to bright after being zapped by violet light. Photoactivatable dyes can also be used instead of proteins. In effect, cells are left with a long-lasting tag.To only tag cells of interest, the team used a digital micromirror device, an array of tiny motor-driven mirrors also used in digital projectors, to give it the ability to light up single cells. "These micromirrors rotate and turn to define a region of your sample, down to single cells," Lee said. "This is all automated. There's a motorized microscope stage that moves the cells on an imaging plate around a predefined zone, and the DMD will shine light only on a particular cell."Through SPOTlight, a researcher can observe a population of hundreds of thousands of human or yeast cells over time to find those with desirable cellular dynamics, subcellular structures or shapes. Custom software can then be used to identify all cells with the desired profile, and instruct the light source and the DMD to photoactivate them with violet light."Then we use a flow cytometer or cell-sorting machine that can detect and recover the cells we tagged while throwing away the rest," Lee said. "After we've recovered our cells of interest, we can send them for sequencing or conduct further studies."Lee said the prototype tags individual cells in 45 seconds to a minute. "That depends on the power of the light," he said. "With a stronger light source, we should be able to do this even faster, maybe down to a few seconds per cell."To demonstrate the utility of SPOTlight, Lee and his colleagues used it to screen 3 million mutant cells expressing a library of fluorescent proteins, ultimately identifying and refining a yellow fluorescent protein they call mGold."It's a variant of an existing fluorescent probe called mVenus," Lee said. "The problem with mVenus is that it photobleaches very fast. It becomes dimmer and dimmer as you keep shining light on it. If you're monitoring cells expressing mVenus for a long time, there comes a time where the fluorescent protein is no longer detectable. So we decided to screen for mVenus mutants with better fluorescent stability."He said researchers typically engineer fluorescent proteins by shining light on bacterial colonies expressing the proteins to see which one is brightest. With SPOTlight, "we can screen for brightness and photostability at the same time," Lee said. "This isn't something people commonly did, but biology isn't static. It's moving in time and space, so it's important to have these temporal properties as well."Compared with commonly used yellow fluorescent proteins, mGold was four to five times more stable," he said."Important developmental events and behaviors require monitoring for many minutes, hours or days and it's frustrating when the probes we use to image these processes go dark before we've been able to capture the whole story," St-Pierre said."It's like having a power outage in the middle of watching a good movie," he said. "Building on our work with mGold, we now want to use SPOTlight to develop probes that will enable us to watch full movies."Similarly, SPOTlight can enable synthetic biologists to engineer new proteins, nucleic acids or cells," St-Pierre said. "More broadly, this method can help any researcher seeking to unravel the genetic or epigenetic determinants of an interesting cellular phenotype, including such clinically relevant properties as resistance to disease or treatment."
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Biotechnology
| 2,020 |
October 23, 2020
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https://www.sciencedaily.com/releases/2020/10/201023135323.htm
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Protective shield: Membrane-attached protein protects bacteria and chloroplasts from stress
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Stress is present everywhere, even bacteria and plant cells have to cope with it. They express various specific stress proteins, but how exactly this line of defense works is often not clear. A group of scientists headed by Professor Dirk Schneider of Johannes Gutenberg University Mainz (JGU) has now discovered a protective mechanism in cyanobacteria as well as in chloroplasts of plant cells: Complex ring structures formed by a protein attach to cell membranes and dissociate. Thereafter, the individual proteins spread out on the membrane surface and form a carpet structure. "Via formation of such a shield, bacteria and chloroplasts protect their membranes under certain stress conditions," stated Professor Dirk Schneider, head of the Membrane Biochemistry section at the JGU Department of Chemistry.
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The biochemist and his team have examined the protein IM30, i.e., the inner membrane-associated protein having a mass of approximately 30 kilodaltons. Previous studies have already shown that the IM30 protein is involved in the formation and preservation of membranes in photosynthetic cells. Without IM30, the amount of thylakoid membranes, in which the photosynthetic light reaction occurs, decreases, ultimately resulting in cell death. The hitherto unknown molecular mechanism of membrane stabilization has now been observed and revealed in detail. The results of this collaborative research project have recently been published in the Nature journal "For quite some time now, we were well aware that IM30 is somehow related to stress. However, we did not know how exactly these proteins manage to protect the cells on a molecular level," explained Schneider. Employing biochemical and biophysical methods in cooperation especially with Professor Stefan Weber of the Max Planck Institute of Polymer Research in Mainz and Professor Eva Wolf of the Institute of Molecular Biology (IMB), the mystery was finally solved. Using atomic force microscopy, the scientists were able to observe how the ring structures disassemble and form carpets on membrane surfaces. "For the very first time we were able to visualize the neat IM30 structure on the surface of membranes," said Schneider.Intrinsically disordered proteins have important functionsIM30 belongs to the group of intrinsically disordered proteins, which have shifted into the focus of science in recent years. When IM30 binds to the membrane, it unfolds in half -- which makes it particularly complicated to study. The traditional understanding of proteins has been based on the assumption that their function is associated with its structure and that disordered structures more or less take over no function. "It is now becoming increasingly clear that disordered protein regions can be involved in defined interactions," stated Schneider as to the classification of the results in a large-scale context.The study defines the thus far enigmatic structural basis for the physiological function of IM30 and related proteins, including the phage shock protein A (PspA), the main representative of the protein family to which IM30 belongs. It also "highlights a hitherto unrecognized concept of membrane stabilization by intrinsically disordered proteins," stated the authors in the "Our discovery now answers the long-standing question as to how exactly the protein protects the membrane. This, however, raises new questions, for example how the individual proteins exactly interact on the membrane surface and form the carpet," said Schneider about the research now planned.
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Biotechnology
| 2,020 |
October 23, 2020
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https://www.sciencedaily.com/releases/2020/10/201023095855.htm
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Researchers reveal why heat stress damages sperm
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University of Oregon biologists have used the model organism Caenorhabditis elegans to identify molecular mechanisms that produce DNA damage in sperm and contribute to male infertility following exposure to heat.
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In humans, the optimal temperature for sperm production is just below body temperature, in a range of about 90-95 degrees F. Human studies have found that exposure to temperatures as little as 1 degree C (1.8 F) above this normal range adversely affects male fertility, said Diana Libuda, a professor in the Department of Biology and Institute of Molecular Biology.The phenomenon of heat-induced male infertility is well known, and the effects of modern exposures to heat such as hot tubs, tight clothing and excessive drive times have been extensively studied. The underlying mechanisms that damage sperm and impair fertilization are not completely understood."In both humans and C. elegans, relatively small increases in temperature are sufficient to reduce male fertility," said Libuda.An increase of 2 C (3.6 F) above normal in C. elegans, a type of roundworm, led to a 25-fold increase in DNA damage in developing sperm compared to unexposed sperm. Eggs fertilized by these damaged sperm failed to produce offspring.This basic research discovery is detailed in a paper published online Oct. 15 in the journal The study provides a roadmap for scientists to pursue studies in mammals and humans to confirm if the same mechanisms contribute to male infertility, said R. Scott Hawley, a meiosis research expert who was not involved in the research.Hawley, a member of the National Academy of Sciences and dean emeritus of the graduate school of the Stowers Institute for Medical Research in Kansas City, Missouri, had heard about preliminary findings at an academic conference."I think this is a hallmark paper because it shows an environmental effect that alters specific DNA sequences and the presumably the proteins that control their activity," Hawley said. "What Diana and Nicole's work has done is to clearly say what goes wrong, at the level of molecules, when sperm-making is altered by heat, at least in worms."The paper also helps to understand how meiosis, the process that produces sex cells, differs between sperm and eggs.Sperm, the smallest cell in a person's body, form by the billions at temperatures below body temperature and are produced throughout the entire adult lifespan. Eggs, the largest cells in a person's body, are formed internally, where a consistent temperature is maintained, and are produced only for a limited time during fetal development."We know that sperm development is very sensitive to increased temperature, while egg development is not affected," Kurhanewicz said. "The data presented in this paper suggest that another way egg and sperm develop differently is in how tightly they control the ability of mobile DNA elements, which are also known as 'jumping genes' or transposons, to move in the genome, and how sensitive to heat stress those mechanisms are in preventing that movement."Transposons are DNA segments that move around and alter genetic information by inserting themselves in new positions. They also leave DNA damage in their wake. Movement of these "jumping genes" is normally repressed in developing sperm and eggs. However, this study found that with exposure to heat transposons are moving specifically in developing sperm.The research team used microscopy to observe developing sperm and eggs under both normal and heat-stressed conditions. In the latter, the researchers saw higher amounts of DNA damage in sperm, but not eggs. Using next-generation genome sequencing, they also identified the locations of transposons across the whole genome with and without exposure to heat."We found that after heat shock, certain transposons are found in new and more variable locations in the male genome," Kurhanewicz said.The study, Hawley said, not only shows that a small rise in temperature affects meiotic divisions but she also identifies a mechanism -- not only where the error occurs but what the error is."This is where it gets exciting," he said. "If we can determine how much of a change is bad, and if you are really concerned about the environmental matters such as hot tubs or 'boxers versus briefs', this type molecular understanding may allow us to reframe the debate on solid scientific grounds."
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Biotechnology
| 2,020 |
October 22, 2020
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https://www.sciencedaily.com/releases/2020/10/201022144544.htm
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Discoveries reshape understanding of gut microbiome
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The human gut is home to microorganisms that outnumber our cells by a factor of 10 to 1. Now, discoveries by scientists at the Oklahoma Medical Research Foundation have redefined how the so-called gut microbiome operates and how our bodies coexist with some of the 100 trillion bacteria that make it up.
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The new findings appear in the journal Using research models, OMRF's Lijun Xia, M.D., Ph.D., led a team of scientists who found the microbiome controls the creation of a sticky layer of special forms of sugar-enriched mucus that encapsulates and travels with fecal matter. The mucus -- which the researchers showed not to be static as previously thought -- acts as a barrier between bacteria in feces and the thousands of immune cells in the colon. Without the mucus, the whole system gets thrown out of balance."The colon is not just a digestive organ, but an immune organ," said Xia, who holds the Merrick Foundation Chair in Biomedical Research at OMRF. "Our microbiome begins to develop at the moment of birth and evolves throughout our lives. It's essential for the growth and maturation of the acquired immune system in our body. When it's not well developed or cared for, it doesn't operate as it should, which can lead to diseases."The overall health of the gut microbiome is dependent on the presence of its mucus. And although mucus production can be interrupted, OMRF researchers showed that it can be restored.In the study, the researchers found that the fecal matter of mice treated with a broad-spectrum antibiotic had no trace of the mucus coating. And when mice without this protective barrier received a transplant of fecal matter with microbiome, their mucus production jump-started.This may have significant treatment implications for patients whose microbiome is out of balance, Xia explained."Whether because of antibiotics interrupting mucus production or a total colon removal due to ulcerative colitis, painful inflammation can result," said Xia. "Now that we better understand the role and origin of this mucus, we will study how we can supplement it or restore its production."OMRF President Stephen Prescott, M.D., noted that the gut microbiome has taken on increasing importance in medical research as scientists have recognized the roles its microorganisms play in our overall health."Researchers have found these tiny organisms living in our digestive tracts may be key players in obesity, diabetes and a variety of autoimmune and digestive diseases. Dr. Xia's work proves a strong connection between the protective coating driven by the microbiome and the development of a disease like colitis," said Prescott.The findings may open the door to alternatives to colonoscopies for monitoring conditions like inflammatory bowel disease. "Rather than repeated invasive procedures to track the progression of IBD, we may be able to measure the presence of the mucus in a fecal sample and assess a patient's gut health," Xia said.The ramifications of this research are far-reaching and reinforce the importance of "basic" science, Prescott added."Our researchers had to understand the origin and function of this mucus and how it relates to the microbiome to learn how it connects with disease. We now understand that the long-term health implications when this pair gets out of balance can be devastating, and Dr. Xia's work can guide us to answers," Prescott said.Kirk Bergstrom, Ph.D., made essential contributions to this project when he was a senior scientist in Xia's lab. He is now new faculty at the University of British Columbia. Xindi Shan, Ph.D., a postdoctoral researcher in Xia's lab, also contributed significantly to this work. Other scientists from OMRF and institutions including Cedars Sinai Medical Center, University of British Columbia and the University of Oklahoma Health Sciences Center also contributed to the project.This research was funded by National Institutes of Health grant nos. R01DK085691 and GM103441, as well as the Oklahoma Center for Adult Stem Cell Research, a program of TSET, and the Stephenson Cancer Center.
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Biotechnology
| 2,020 |
October 21, 2020
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https://www.sciencedaily.com/releases/2020/10/201021112356.htm
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Transcription factors may inadvertently lock in DNA mistakes
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Transcription factor proteins are the light switches of the human genome. By binding to DNA, they help turn genes "on" or "off" and start the important process of copying DNA into an RNA template that acts as a blueprint for a new protein.
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By being choosy about which genes they turn on, transcription factors determine which rooms in the house are lighted and which aren't, or rather, which components of a person's genome are activated.A team of Duke researchers has found that transcription factors have a tendency to bind strongly to "mismatched" sections of DNA, sections of the code that were not copied correctly. The strong binding of transcription factors to mismatched sections of regulatory DNA might be a way in which random mutations become a problem that leads to disease, including cancer.The findings appear Oct. 21 in the journal Most of the time, DNA replication in the body goes smoothly, with nucleotides locking arms with their complementary base pair and marching through the cycle together in intended A-T and C-G fashion. However, as Gordan describes it, "no polymerase is perfect" and every now and then, a nucleotide will be paired with the wrong partner, resulting in a mismatch.Pipetting transcription factor proteins on slides pre-blotted with thousands of DNA molecule samples, a research team led by Duke computational biologist Raluca Gordan Ph.D., showed that the proteins had a stronger bond with the sections of DNA with the mismatched base pairs than with those with perfectly matched base pairs, or "normal" DNA structure.But what makes these 'mistakes' an attractive binding site for transcription factor proteins? For insight, Gordan, an associate professor in the Department of Biostatistics and Bioinformatics and the Department of Computer Science, reached out to Hashim Al-Hashimi, Ph.D., a James B. Duke Professor of Biochemistry, and expert in DNA structure and dynamics who works just across the street.Al-Hashimi studies nucleic acids (DNA and RNA) and their interactions with proteins and small molecules, with the idea that how these biomolecules look and move is as important for their function as their chemical properties.Looking at the experimental results, Gordan and Al-Hashimi came to the conclusion that the strong interaction between transcription factor proteins and mismatched DNA has a lot to do with laziness. When a transcription factor protein binds to DNA, it must spend energy distorting the site, for example by bending the DNA to its will. However, mismatched sections of DNA are already distorted, so the transcription factor protein has to do less work."That's when the transcription factor doesn't need to pay that energetic penalty" to get the job done, Gordan said."If we are ever to attain a deep and predictive understanding of how DNA is recognized by proteins in cells, we need to go beyond the conventional description in terms of static structures and move towards describing both DNA and the protein molecules that bind to them in terms of dynamic structures that have different preferences to adopt a wide range of shapes," Al-Hashimi said.Gordan said that going forward, the team hopes to understand how this interaction relates to disease development. If a mismatched base pair, bound strongly by a transcription factor, makes it through the DNA replication cycle without being repaired by another type of protein -- known as a repair enzyme -- it can become a mutation, and mutations can lead to genetic diseases like cancer and neurodegeneration."We are now convinced that the interactions between transcription factors and mismatches are really strong," she said. "So the next step is to understand what this means for the cell.""We already know that regulatory regions of the genome harbor more cancer mutations than expected by chance. We just do not know why. The strong interactions between transcription factors and DNA mismatches, which could interfere with repair of the mismatches, provide a novel mechanism for the accumulation of mutations in regulatory DNA."
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Biotechnology
| 2,020 |
October 21, 2020
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https://www.sciencedaily.com/releases/2020/10/201021112404.htm
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The consequences of mating at the molecular level
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While it is known that stem cells have the ability to develop into all tissues in a precisely regulated process, the way environmental cues affect stem cell behavior has remained poorly understood. In a new study, researchers from the University of Tsukuba discovered that neurons producing the neurotransmitter octopamine regulate the behavior of germline stem cells (GSCs) in response to environmental cues, such as mating.
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The ovaries of the fruit fly Drosophila melanogaster have been a robust model system for studying the relationship between environmental cues and stem cell biology. In fruit flies, GSCs give rise to eggs and exist in close proximity to somatic cells. Somatic cells comprise several types of cells in support of the budding eggs. As with other stem cells, when GSCs divide, one daughter cell retains its stem cell identity, while the other differentiates into multiple progeny cells. The balance between self-renewal and differentiation is tightly regulated, both by cues within and outside the environment in which GSCs reside (also called a niche). Mating is one such external cue known to increase GSCs."It is well known that a molecule called sex peptide from the male seminal fluid activates neurons located in the uterine lumen. We have previously shown that these neurons are essential for stimulating the biosynthesis of ovarian steroid hormones to increase the number of GSCs," says corresponding author of the study Professor Ryusuke Niwa. "The goal of our study was to investigate how the information from mating is transmitted from these neurons to GSCs at the molecular and cellular levels."To achieve their goal, the researchers took a genetic approach to investigate which gene is responsible for the increase of GSCs upon mating, and found that the octopamine receptor Oamb is the one through which octopamine exerts its effect on GSCs. Through a series of experiments, the researchers then found that Oamb in escort cells, one type of somatic cell adjacent to GSCs, modulates GSC increase after mating and the subsequent release of octopamine by neurons. At the molecular level, Oamb activation by octopamine resulted in an increase in calcium signaling in escort cells. Calcium is a potent biomolecule and changes in cellular calcium levels strongly affect cell behavior.Because it had previously been shown that ovarian steroid hormones were involved in the increase of GSCs, the researchers next investigated the relationship between ovarian steroid hormones and the calcium-dependent GSC increase. Their results showed that ovarian steroid hormones are indeed required to increase the number of GSCs. Next, the researchers asked which molecules play a role in stimulation of escort cells by octopamine and found that the protein matrix metalloproteinase 2 is required upon the calcium-dependent GSC increase. Finally, the researchers showed that the neurons projecting to the ovaries to increase GSCs do so via specialized proteins, called nicotinic acetylcholine receptors. These results provide a complete picture as to how neuronal activation results in increased ovarian stem cells."These are striking results that show the molecular mechanism underlying the coupling of the nervous system with stem cell behavior in response to environmental cues, such as mating," says Professor Niwa. "Our results could help unravel the conserved systemic and neuronal regulatory mechanisms for stem cell homeostasis in animals."
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Biotechnology
| 2,020 |
October 20, 2020
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https://www.sciencedaily.com/releases/2020/10/201020150512.htm
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Repairing the photosynthetic enzyme Rubisco
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Manajit Hayer-Hartl, head of the research group "Chaperonin-assisted Protein Folding," has a long-standing interest in the central enzyme of photosynthesis called Rubisco. Her team has already reported on many of the interacting partners of Rubisco that are required for the folding and assembly of this highly abundant protein. In the current study, they have elucidated how Rubisco activase works. As the name indicates, this enzyme is critical for repairing Rubisco once it has lost its activity. The study was published in
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The enzyme Rubisco catalyzes the assimilation of COThe enzyme Rubisco activase, Rca, is present in plants, algae and certain cyanobacteria. Rca is a ring-shaped complex of six subunits with a central pore. How exactly Rca interacts with the inhibited Rubisco and releases the bound sugar from the active site pocket of Rubisco, restoring its COThey discovered that the Rca grabs the N-terminal tail of Rubisco and by pulling and pushing actions, using the energy of ATP, opens the active site pocket. This results in the release of the inhibitory sugar molecule. In cyanobacteria Rubisco is packaged into specialized micro-compartments called carboxysomes, in which a high concentration of COIn an earlier study, Hayer-Hartl showed how Rubisco is recruited into carboxysomes via interactions with the SSUL domains of the scaffolding protein CcmM. Interestingly, the researchers now found that Rca is recruited into carboxysomes using a very similar trick. The Rca hexamer also contains SSUL domains that dock onto Rubisco during carboxysome formation. This makes sure that enough Rca is present inside carboxysomes to perform its essential repair function. Thus, Rca not only functions in Rubisco activation but also mediates its own recruitment into carboxysomes.Manajit Hayer-Hartl concludes: "Rca is absolutely required for Rubisco to function optimally. Deciphering its mechanism and dual function in cyanobacteria will further help us to make photosynthesis more effective in the future. Hopefully, this will get us closer to our ultimate goal, to increase agricultural productivity."
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Biotechnology
| 2,020 |
October 20, 2020
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https://www.sciencedaily.com/releases/2020/10/201020131355.htm
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Targeting the shell of the Ebola virus
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As the world grapples with the coronavirus (COVID-19) pandemic, another virus has been raging again in the Democratic Republic of the Congo in recent months: Ebola. Since the first terrifying outbreak in 2013, the Ebola virus has periodically emerged in Africa, causing horrific bleeding in its victims and, in many cases, death.
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How can we battle these infectious agents that reproduce by hijacking cells and reprogramming them into virus-replicating machines? Science at the molecular level is critical to gaining the upper hand -- research you'll find underway in the laboratory of Professor Juan Perilla at the University of Delaware.Perilla and his team of graduate and undergraduate students in UD's Department of Chemistry and Biochemistry are using supercomputers to simulate the inner workings of Ebola, observing the way molecules move, atom by atom, to carry out their functions. In the team's latest work, they reveal structural features of the virus's coiled protein shell, or nucleocapsid, that may be promising therapeutic targets, more easily destabilized and knocked out by an antiviral treatment.The research is highlighted in the Tuesday, Oct. 20 issue of the "The Ebola nucleocapsid looks like a Slinky walking spring, whose neighboring rings are connected," Perilla said. "We tried to find what factors control the stability of this spring in our computer simulations."The life cycle of Ebola is highly dependent on this coiled nucleocapsid, which surrounds the virus's genetic material consisting of a single strand of ribonucleic acid (ssRNA). Nucleoproteins protect this RNA from being recognized by cellular defense mechanisms. Through interactions with different viral proteins, such as VP24 and VP30, these nucleoproteins form a minimal functional unit -- a copy machine -- for viral transcription and replication.While nucleoproteins are important to the nucleocapsid's stability, the team's most surprising finding, Perilla said, is that in the absence of single-stranded RNA, the nucleocapsid quickly becomes disordered. But RNA alone is not sufficient to stabilize it. The team also observed charged ions binding to the nucleocapsid, which may reveal where other important cellular factors bind and stabilize the structure during the virus's life cycle.Perilla compared the team's work to a search for molecular "knobs" that control the nucleocapsid's stability like volume control knobs that can be turned up to hinder virus replication.The UD team built two molecular dynamics systems of the Ebola nucleocapsid for their study. One included single-stranded RNA; the other contained only the nucleoprotein. The systems were then simulated using the Texas Advanced Computing Center's Frontera supercomputer -- the largest academic supercomputer in the world. The simulations took about two months to complete.Graduate research assistant Chaoyi Xu ran the molecular simulations, while the entire team was involved in developing the analytical framework and conducting the analysis. Writing the manuscript was a learning experience for Xu and undergraduate research assistant Tanya Nesterova, who had not been directly involved in this work before. She also received training as a next-generation computational scientist with support from UD's Undergraduate Research Scholars program and NSF's XSEDE-EMPOWER program. The latter has allowed her to perform the highest-level research using the nation's top supercomputers. Postdoctoral researcher Nidhi Katyal's expertise also was essential to bringing the project to completion, Perilla said.While a vaccine exists for Ebola, it must be kept extremely cold, which is difficult in remote African regions where outbreaks have occurred. Will the team's work help advance new treatments?"As basic scientists we are excited to understand the fundamental principles of Ebola," Perilla said. "The nucleocapsid is the most abundant protein in the virus and it's highly immunogenic -- able to produce an immune response. Thus, our new findings may facilitate the development of new antiviral treatments."Currently, Perilla and Jodi Hadden-Perilla are using supercomputer simulations to study the novel coronavirus that causes COVID-19. Although the structures of the nucleocapsid in Ebola and COVID-19 share some similarities -- both are rod-like helical protofilaments and both are involved in the replication, transcription and packing of viral genomes -- that is where the similarities end."We now are refining the methodology we used for Ebola to examine SARS-CoV-2," Perilla said.
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Biotechnology
| 2,020 |
October 20, 2020
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https://www.sciencedaily.com/releases/2020/10/201020131336.htm
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The road to uncovering a novel mechanism for disposing of misfolded proteins
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About 30 years ago, Dr. Richard Sifers set out on a journey to discover why people with a rare condition known as alpha1-antitrypsin (AAT) deficiency present with high variation in the severity of liver disease. His journey led him to the discovery of fundamental underpinnings of this condition and, unexpectedly, to uncovering a novel cellular mechanism for disposing of misfolded proteins. The latter has implications not just for AAT-deficiency, but also for other more common conditions associated with accumulation of defective proteins, including neurological disorders, such as Alzheimer's disease.
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AAT deficiency can develop in people who carry the AAT gene with a mutation called Z."I began studying AAT deficiency because I was intrigued by the wide range of severity of the condition. Some of the people carrying two copies of the Z mutation developed lung disease late in life and some developed liver disease. Interestingly, the condition also could appear very early in life. Some newborns and infants developed severe liver disease and needed to have a transplant to live," said Sifers, professor of pathology & immunology and member of the Dan L Duncan Comprehensive Cancer Center at Baylor College of Medicine.Other groups had shown that about 1 in 1,700 people carry two copies of the AAT-Z gene. However, only about 17 percent of these newborns with AAT-Z had clinically significant liver disease, and less than 3 percent of those progressed to life-threatening, end-stage disease as infants.One of Sifers's first contributions was to help develop the first screening test to determine whether a newborn was at risk of developing severe liver disease."Developing the screening method made me realize that I could tell whether a child was at a high risk of having liver disease, but still did not know what was causing the condition," Sifers said.AAT is a protein produced by the liver and transported through the blood to the lungs, where it protects them from damage caused by other enzymes that breakdown proteins in the lung. The Z mutation produces a defective AAT protein that cannot fold into an appropriate 3-D conformation. Inappropriately folded AAT-Z proteins cannot exit the liver, so they do not travel to the lungs to protect them from destruction. This can lead to lung damage contributing to emphysema and other lung conditions."As I studied the disease, I noticed that AAT-Z, which should be released from the liver, was actually accumulating," Sifers said. "This suggested that the naturally disposing mechanism of the cell might not be working."Sifers and others dug deeper into how cells dispose of misfolded proteins. They discovered that cells shuttle defective proteins from their place of synthesis, the endoplasmic reticulum (ER), to the cytosol, where they are degraded in a cellular structure called a proteasome. Key to this process is to tag the proteins for destruction."Specifically, we found that the human enzyme mannosidase Man1b1 acted like a quality-control factor that mediated the removal of the sugar mannose from misfolded AAT-Z proteins, promoting their degradation."The AAT deficiency model has been used by many other researchers studying conditions also linked to toxic accumulation of misfolded proteins in cells, altogether called conformational diseases. This approach has accelerated the understanding of the underlying causes of these conditions, offering novel opportunities for potential treatments.Although researchers knew that liver injury associated with AAT deficiency was linked to accumulation of misfolded AAT-Z proteins in the liver, there was still no explanation for the severe liver disease in infants.In a 2009 paper, Sifers and his colleagues studied liver tissue samples from unrelated infants or children older than 2 years who had undergone liver transplantation for end-stage liver disease. They also conducted genetic linkage and functional laboratory experiments with other cells cultured in the lab.They showed that certain genetic modification, a single nucleotide polymorphism, that leads to changes in the expression of the Man1b1 gene results in lower levels of Man1b1protein in the endoplasmic reticulum of liver cells.Sifers and colleagues proposed that lower levels of Man1b1 impair the liver's capacity to deal with the accumulation of misfolded AAT-Z. This likely accelerates reaching the tolerable threshold for protein accumulation, resulting in earlier liver failure."I was delighted that after years of research, we had found an explanation for the mystery of AAT deficiency-associated liver disease in infants and wondered whether my lab would make other major contributions in the future," Sifers said.As Sifers and colleagues continued studying Man1b1, they unexpectedly came across a role for this protein that had not been described before."We found that, in addition to tagging misfolded proteins for degradation by enzymatically removing mannose groups, Man1b1 also promotes protein degradation by another mechanism that is independent from the first," Sifers said.Sifers and his co-authors, Dr. Ashlee H. Sun, now at Polypus-transfection Biotechnology, and Dr. John R. Collette, postdocs in his lab, reported in the The researchers propose that the new unconventional system might be involved in the elimination of soluble protein aggregates that have been associated with conformational diseases. For instance, human Man1b1 has been linked to the causes of multiple congenital disorders of intellectual disability and HIV infection, and to poor prognosis in patients with bladder cancer."Our work is a clear example that studying rare diseases can bring solutions for more common conditions."This study was supported by research grants from Alpha-1 Foundation.
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Biotechnology
| 2,020 |
October 20, 2020
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https://www.sciencedaily.com/releases/2020/10/201020105551.htm
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CRISPR meets Pac-Man: New DNA cut-and-paste tool enables bigger gene edits
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Gene editing for the development of new treatments, and for studying disease as well as normal function in humans and other organisms, may advance more quickly with a new tool for cutting larger pieces of DNA out of a cell's genome, according to a new study by UC San Francisco scientists.
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Publication of the UCSF study on Oct. 19, 2020 in the journal Though now employed as a research tool in laboratories around the world, CRISPR evolved eons ago in bacteria as a means to fight their ancient nemeses, a whole host of viruses known as bacteriophages. When bacteria encounter a phage, they incorporate a bit of the viral DNA into their own DNA, and it then serves as a template to make RNA that binds to the corresponding viral DNA in the phage itself. The CRISPR enzymes then target, disable and kill the phage.In his latest work exploring this ancient and strange arms race, principal investigator Joseph Bondy-Denomy, PhD, associate professor in the UCSF Department of Microbiology and Immunology, joined scientists Bálint Csörg?, PhD, and Lina León to develop and test a new CRISPR tool.The already renowned CRISPR-Cas9 ensemble is like a molecular chisel that can be used to rapidly and precisely excise a small bit of DNA at a targeted site. Other methods can then be used to insert new DNA. But the new CRISPR-Cas3 system adapted by the UCSF scientists employs a different bacterial immune system. The key enzyme in this system, Cas3, acts more like a molecular wood chipper to remove much longer stretches of DNA quickly and accurately."Cas3 is like Cas9 with a motor -- after finding its specific DNA target, it runs on DNA and chews it up like a Pac-Man," Bondy-Denomy said.This new capability to delete or replace long stretches of DNA will enable researchers to more efficiently assess the importance of genomic regions that contain DNA sequences of indeterminate function, according to Bondy-Denomy, an important consideration in understanding humans and the pathogens that plague them."Previously, there was no easy and reliable way to delete very large regions of DNA in bacteria for research or therapeutic purposes," he said. "Now, instead of making 100 different small DNA deletions we can just make one deletion and ask, 'What changed?'"Because bacteria and other types of cells are commonly used to produce small molecule or protein-based pharmaceuticals, CRISPR-Cas3 will enable biotechnology industry scientists to more easily remove potentially pathogenic or useless DNA from these cells, according to Bondy-Denomy."Large swathes of bacterial DNA are poorly understood, with unknown functions that in some cases are not necessary for survival," Bondy-Denomy said. "In addition, bacterial DNA contains large stretches of DNA imported from other sources, which can cause disease in the bacterium's human host, or divert bacterial metabolism."CRISPR-Cas3 also should also allow entire genes to be inserted into the genome in industrial, agricultural or even in human gene therapy applications, Bondy-Denomy said.The UCSF researchers selected and modified the CRISPR-Cas3 system used by the bacterium Pseudomonas aeruginosa, and demonstrated in this species and in three others, including bacteria that cause disease in humans and plants, that their more compact version functions well to remove selected DNA in all four species. Other CRISPR-Cas3 systems have been made to work in human and other mammalian cells, and that also should be achievable for the modified P. aeruginosa system, Bondy-Denomy said.Bondy-Denomy studies a range of bacteria, bacteriophage, and CRISPR systems to learn more about how they work and to find useful molecular tools. "CRISPR-Cas3 is by far the most common CRISPR system in nature," he said. "About 10 times as many bacterial species use a Cas3 system as use a Cas9 system. It may be that Cas3 is a better bacterial immune system because it shreds phage DNA."Unlike Cas9, when Cas3 binds to its precise DNA target it begins chewing up one strand of the double-stranded DNA in both directions, leaving a single strand exposed. The deletions obtained in the UCSF experiments ranged in size, in many cases encompassing as many as 100 bacterial genes. The CRISPR-Cas3 mechanism should also allow for easier replacement of deleted DNA with a new DNA sequence, the researchers found.For DNA deletion and editing in the lab, scientists program CRISPR systems to target specific DNA in the genome of an organism of interest using any guide sequence they choose.In the new CRISPR-Cas3 study, by manipulating the sequences of DNA provided to the bacteria for repairing the deletions, the researchers were able to precisely set the boundaries of these large DNA repairs, something they were unable to accomplish with CRISPR-Cas9.Bondy-Denomy previously discovered anti-CRISPR strategies that phage evolved to fight back against bacteria, and these might prove useful for stopping the gene editing reactions driven by Cas enzymes used as human therapeutics before side effects arise, or in using phage to remove unwanted bacteria that have populated the gut, he said. Apart from E. coli and a couple of other species, relatively little is known about the 1,000 or so bacterial species that normally reside there."Non-model microbes have largely been left behind in the genetics world, and there is a huge need for new tools to study them," he said.
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Biotechnology
| 2,020 |
October 19, 2020
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https://www.sciencedaily.com/releases/2020/10/201019125516.htm
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Magnetic field and hydrogels could be used to grow new cartilage
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Using a magnetic field and hydrogels, a team of researchers in the Perelman School of Medicine at the University of Pennsylvania have demonstrated a new possible way to rebuild complex body tissues, which could result in more lasting fixes to common injuries, such as cartilage degeneration. This research was published today in
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"We found that we were able to arrange objects, such as cells, in ways that could generate new, complex tissues without having to alter the cells themselves," said the study's first author, Hannah Zlotnick, a graduate student in Bioengineering who works in the McKay Orthopaedic Research Laboratory at Penn Medicine. "Others have had to add magnetic particles to the cells so that they respond to a magnetic field, but that approach can have unwanted long-term effects on cell health. Instead, we manipulated the magnetic character of the environment surrounding the cells, allowing us to arrange the objects with magnets."In humans, tissues like cartilage can often break down, causing joint instability or pain. Often, the breakdown isn't in total, but covers an area, forming a hole. Current fixes are to fill those holes in with synthetic or biologic materials, which can work but often wear away because they are not the same exact material as what was there before. It's similar to fixing a pothole in a road by filling it with gravel and making a tar patch: the hole will be smoothed out but eventually wear away with use because it's not the same material and can't bond the same way.What complicates fixing cartilage or other similar tissues is that their make-up is complex."There is a natural gradient from the top of cartilage to the bottom, where it contacts the bone," Zlotnick explained. "Superficially, or at the surface, cartilage has a high cellularity, meaning there is a higher number of cells. But where cartilage attaches to the bone, deeper inside, its cellularity is low."So the researchers, which included senior author Robert Mauck, PhD, director of the McKay Lab and a professor of Orthopaedic Surgery and Bioengineering, sought to find a way to fix the potholes by repaving them instead of filling them in. With that in mind, the research team found that if they added a magnetic liquid to a three-dimensional hydrogel solution, cells, and other non-magnetic objects including drug delivery microcapsules, could be arranged into specific patterns that mimicked natural tissue through the use of an external magnetic field.After brief contact with the magnetic field, the hydrogel solution (and the objects in it) was exposed to ultraviolet light in a process called "photo crosslinking" to lock everything in place, and the magnetic solution subsequently was diffused out. After this, the engineered tissues maintained the necessary cellular gradient.With this magneto-patterning technique, the team was able to recreate articular cartilage, the tissue that covers the ends of bones."These magneto-patterned engineered tissues better resemble the native tissue, in terms of their cell disposition and mechanical properties, compared to standard uniform synthetic materials or biologics that have been produced," said Mauck. "By locking cells and other drug delivering agents in place via magneto-patterning, we are able to start tissues on the appropriate trajectory to produce better implants for cartilage repair."While the technique was restricted to in vitro studies, it's the first step toward potential longer-lasting, more efficient fixes in living subjects."This new approach can be used to generate living tissues for implantation to fix localized cartilage defects, and may one day be extended to generate living joint surfaces," Mauck explained.Co-authors on this study included Sarah Gullbrand and James Carey of the University of Pennsylvania and Andy Clark and Xuemei Cheng of Bryn Mawr College.Financial support was provided by the National Institutes of Health (grant numbers R01 AR056624, R01 AR071340, T32 AR007132, and P30 AR069619), the Department of Veteran Affairs (IK6 RX003416) and the National Sciences Foundation for Engineering Mechanobiology (CMMI: 15-48571).
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Biotechnology
| 2,020 |
October 19, 2020
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https://www.sciencedaily.com/releases/2020/10/201019125356.htm
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Prebiotic chemistry: In the beginning, there was sugar
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Organic molecules formed the basis for the evolution of life. But how could inorganic precursors have given rise to them? Ludwig-Maximilians-Universitaet (LMU) in Munich chemist Oliver Trapp now reports a reaction pathway in which minerals catalyze the formation of sugars in the absence of water.
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More than 4 billion years ago, the Earth was very far from being the Blue Planet it would later become. At that point it had just begun to cool and, in the course of that process, the concentric structural zones that lie ever deeper beneath our feet were formed. The early Earth was dominated by volcanism, and the atmosphere was made up of carbon dioxide, nitrogen, methane, ammonia, hydrogen sulfide and water vapor. In this decidedly inhospitable environment the building blocks of life were formed. How then might this have come about?Researchers have puzzled over the question for decades. The first breakthrough was made in 1953 by two chemists, named Stanley Miller and Harold C. Urey, at the University of Chicago. In their experiments, they simulated the atmosphere of the primordial Earth in a closed reaction system that contained the gases mentioned above. A miniature 'ocean' was heated to provide water vapor, and electrical discharges were passed through the system to mimic the effects of lightning. When they analyzed the chemicals produced under these conditions, Miller and Urey detected amino acids -- the basic constituents of proteins -- as well as a number of other organic acids.It is now known that the conditions employed in these experiments did not reflect those that prevailed on the early Earth. Nevertheless, the Miller-Urey experiment initiated the field of prebiotic chemical evolution. However, it not throw much light on how other classes of molecules found in all biological cells -- such as sugars, fats and nucleic acids -- might have been generated. These compounds are however indispensable ingredients of the process that led to the first bacteria and subsequently to photosynthetic cyanobacteria that produced oxygen. This is why Oliver Trapp, Professor of Organic Chemistry at LMU, decided to focus his research on the prebiotic synthesis of these substances.The story of synthetic routes from smaller precursors to sugars goes back almost a century prior to the Miller-Urey experiment. In 1861, the Russian chemist Alexander Butlerov showed that formaldehyde could give rise to various sugars via what became known as the formose reaction. Miller und Urey in fact found formic acid in their experiments, and it can be readily reduced to yield formaldehyde. Butlerov also discovered that the formose reaction is promoted by a number of metal oxides and hydroxides, including those of calcium, barium, thallium and lead. Notably calcium is abundantly available on and below the Earth's surface.However, the hypothesis that sugars could have been produced via the formose reaction runs into two difficulties. The 'classical' formose reaction produces a diverse mixture of compounds, and it takes place only in aqueous media. These requirements are at odds with the fact that sugars have been detected in meteorites.Together with colleagues at LMU and the Max Planck Institute for Astronomy in Heidelberg, Trapp therefore decided to explore whether formaldehyde could give rise to sugars in a solid-phase system. With a view to simulating the kinds of mechanical forces to which solid minerals would have been subjected, all the reaction components were combined in a ball mill -- in the absence of solvents, but adding enough formaldehyde to saturate the powdered solidsAnd indeed, the formose reaction was observed and several different minerals were found to catalyze it. The formaldehyde was adsorbed onto the solid particles, and the interaction resulted in the formation of the formaldehyde dimer (glycolaldehyde) -- and ribose, the 5-carbon sugar that is an essential constituent of ribonucleic acid (RNA). RNA is thought to have merged prior to DNA, and it serves as the repository of genetic information in many viruses, as well as providing the templates for protein synthesis in all cellular organisms. More complex sugars were also obtained in the experiments, together with a few byproducts, such as lactic acid and methanol."Our results provide a plausible explanation for the formation of sugars in the solid phase, even under extraterrestrial settings in the absence of water," says Trapp. They also prompt new questions that may point to new and unexpected prebiotic routes to the basic components of life as we know it, as Trapp affirms. "We are convinced that these new insights will open up entirely new perspectives for research on prebiotic, chemical evolution," he says.
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Biotechnology
| 2,020 |
October 19, 2020
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https://www.sciencedaily.com/releases/2020/10/201019111918.htm
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AI methods of analyzing social networks find new cell types in tissue
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In situ sequencing enables gene activity inside body tissues to be depicted in microscope images. To facilitate interpretation of the vast quantities of information generated, Uppsala University researchers have now developed an entirely new method of image analysis. Based on algorithms used in artificial intelligence, the method was originally devised to enhance understanding of social networks. The researchers' study is published in
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The tissue composing our organs consists of trillions of cells with various functions. All the cells in an individual contain the same genes (DNA) in their nuclei. Gene expression occurs by means of "messenger RNA" (mRNA) -- molecules that carry messages from the nucleus to the rest of the cell, to direct its activities. The mRNA combination thus defines the function and identity of every cell.RNA transcripts are obtainable through in situ sequencing. The researchers behind the new study had previously been involved in developing this method, which shows millions of detected mRNA sequences as dots in microscope images of the tissue. The problem is that distinguishing all the important details may be difficult. This is where the new AI-based method may come in useful, since it allows unsupervised detection of cell types as well as detection of functions within an individual cell and of interactions among cells."We're using the latest AI methods -- specifically, graph neural networks, developed to analyse social networks; and adapting them to understand biological patterns and successive variation in tissue samples. The cells are comparable to social groupings that can be defined according to the activities they share in their social networks like Twitter, sharing their Google search results or TV recommendations," says Carolina Wählby, professor of quantitative microscopy at the Department of Information Technology, Uppsala University.Earlier analytical methods of this type of data depend on knowing which cell types the tissue contains, and identifying the cell nuclei in it, in advance. The method conventionally used, known as "single-cell analysis," may lose some mRNA and miss certain cell types. Even with advanced automated image analysis, it is often difficult to find the various cell nuclei if, for example, the cells are packed densely together."With our analysis, which we call 'spage2vec', we can now get corresponding results without any previous knowledge of expected cell types. And what's more, we can find new cell types and intra- or intercellular functions in tissue," Wählby says.The research group are now working further on its analytical method by investigating differentiation and organisation of various types of cells during the early development of the heart. This is pure basic research, intended to provide more knowledge of the mechanisms that govern development, both when everything is functioning as it should and when a disease is present. In another project, a collaboration with cancer researchers, the Uppsala group are hoping to be able to apply the new methods to gain a better understanding of how tumour tissue interacts, at molecular level, with surrounding healthy tissue. The aim is that, in the long term, this will culminate in better treatments that can be adapted to individual patients.
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Biotechnology
| 2,020 |
October 19, 2020
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https://www.sciencedaily.com/releases/2020/10/201019103441.htm
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Advancing wildlife genomics through the development of molecular methods
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A team of scientists from the Leibniz Institute for Zoo and Wildlife Research (Leibniz-IZW), the Australian Museum and the Max Delbrück Center for Molecular Medicine (MDC) report a new method for identifying any genome sequence located next to a known sequence. It is often difficult to precisely determine unknown sequences close to small known fragments. Whole genome sequencing can be a solution, but it's a very cost intensive approach. In order to find a more efficient technique, the scientists developed Sonication Inverse PCR (SIP): First, DNA is cut into random pieces using ultrasound waves. After DNA fragmentation, long-range inverse PCR is performed followed by long-fragment high-throughput sequencing. SIP can be used to characterise any DNA sequence (near a known sequence) and can be applied across genomics applications within a clinical setting as well as molecular evolutionary analyses. The results are reported in the scientific journal
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Many methods have been developed to identify sequences next to a determined sequence of interest. Inverse PCR based methods are among the most common methods and have been used for decades but suffer from bias because of the way DNA is cut apart by enzymes: They need to find specific sequence motifs that are not evenly spread across the DNA. Therefore, many neighbouring sequences to a target cannot be characterised without technical difficulty or without the expense and effort of whole genome sequencing. "Sonication Inverse PCR (SIP) circumvents this problem by using high-frequency sound waves to randomly cut the DNA, eliminating the bias resulting from the use of enzymes," Prof Alex Greenwood from Leibniz-IZW explains. "The fragments are then turned into circles and the so-called inverse PCR is applied." With the development of long-fragment sequencing, the authors were able to target 4-6 thousand base long inverse PCR fragments and sequenced them at high-throughput on the PacBio RS II sequencing platform.The new method was tested on a complex model, the koala retrovirus (KoRV), a high copy retrovirus found in the koala (Phascolarctos cinereus) genome. Targeting the ends of the integrated virus, the full spectrum of viral integrations in the genome could be determined using a small 'known' piece of viral DNA. Mapping the integrations against reference genomes provided precise genomic locations for each integration at a resolution that would otherwise require a large sequencing effort. "Applying this method allowed us to discover a koala specific defense mechanism against KoRV," says Dr Ulrike Löber from the MDC."SIP is economical and can be simultaneously applied to many samples by including barcodes to the PCR primers, making the method cost efficient," adds Dr David Alquezar, former member of the Leibniz-IZW team and now manager of the Australian Centre for Wildlife Genomics at the Australian Museum. The authors continue to apply SIP to address different problems, such as how viruses become integrated into genomes and how they cause diseases. In conclusion, SIP provides a new protocol for high-throughput profiling of flanking sequences next to any region of interest coupled with long-range sequencing, allowing scientists to study complex biological systems such as mobile genetic elements.
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Biotechnology
| 2,020 |
October 19, 2020
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https://www.sciencedaily.com/releases/2020/10/201014114646.htm
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DNA-peptide interactions create complex behaviors which may have helped shape biology
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Deoxyribonucleic acid (DNA)-protein interactions are extremely important in biology. For example, each human cell contains about 2 meters worth of DNA, but this is packaged into a space about one million times smaller. The information in this DNA allows the cell to copy itself. This extreme packaging is mainly accomplished in cells by wrapping the DNA around proteins. Thus, how DNA and proteins interact is of extreme interest to scientists trying to understand how biology organises itself. New research by scientists at the Earth-Life Science Institute (ELSI) at Tokyo Institute of Technology and the Institut Pierre-Gilles de Gennes, ESPCI Paris, Université PSL suggests that the interactions of DNA and proteins have deep-seated propensities to form higher-ordered structures such as those which allow the extreme packaging of DNA in cells.
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Modern living cells are principally composed of a few classes of large molecules. DNA gets the lion's share of attention as it is the repository of the information cells use to build themselves generation after generation. This information-rich DNA is normally present as a double-stranded caduceus of two polymers wrapped around each other, with much of what makes the information DNA contains obscured to the external environment because the information-bearing parts of the molecules are engaged with their complementary strand. When DNA is copied into ribonucleic acid (RNA), its strands are pulled apart to allow its more complex surfaces to interact, which enables it to be copied into single-stranded RNA polymers. These RNA polymers are finally read out by biological processes into proteins, which are polymers of a variety of amino acids with extremely complicated surface properties. Thus, DNA and RNA are somewhat predictable in terms of their chemical behaviour as polymers, while proteins are not.Polymeric molecules, those composed or repeated types of subunits, can display complex behaviours when mixed with other chemicals, especially when dissolved in a solvent like water. Chemists have developed a complex set of terms for how compounds behave when they are mixed. For example, the proteins in cow's milk are considered a colloidal (or homogeneous noncrystalline suspended mixture which does not settle and cannot be separated by physical means) suspension in water. When lemon juice is added to milk, the suspended proteins reorganise themselves to produce the visible self-organisation of curds, which do separate into a new phase. There are other types of this phenomenon chemists have discovered over the years, for example, liquid crystals (LC). LCs are formed when a molecules have an elongated shape or the tendency to make linear aggregates (like stacks of molecules one on top of each other): the resulting material presents a mixture of the properties of a crystal and a liquid: the material thus has a certain degree of order like a solid (for example, parallel orientation of the molecules) but still retains its fluidity (molecules can easily slip on and by each other). We all experience liquid crystals in the various screens we interact with every day "LCDs," or liquid crystal displays, which use these variable properties to make the images we see on our device screens. In their work, Fraccia and Jia, showed that double-stranded DNA and peptides can generate many different LC phases in a very peculiar way: the LCs actually form in membraneless droplets, called coacervates, where DNA and peptides are spontaneously co-assembled and ordered. This process brings DNA and peptides to very high concentrations, comparable to that of a cell's nucleus, which is 100-1000 times greater of that of the very diluted initial solution (which is the maximum concentration that can likely be achieved on early Earth). Thus, such spontaneous behaviour can in principle favour the formation of the first cell-like structures on early Earth, which would take advantage of the ordered, but fluid, LC matrix in order to gain stability and functionality, and to favour the growth and the evolution of primitive biomolecules.The cut-off between when these higher-order properties begin to present themselves is not always clear cut. When molecules interact at the molecular level, they often "self-organise." One can think of the process of adding sand to a sandpile: as one sprinkles more and more sand to a pile, it tends to form a "low energy" final state -- a pile. Though the addition of sand grains may cause some new structures to form locally, at some point, the addition of one more grain causes a landslide in the pile which reinforces the conical shape of the pile.Though we all benefit from the existence of these phenomena, the scientific community may be missing important aspects of the implications of this type of self-organisation, Jia and Fraccia argue. The combination of these collective material self-organising effects may be relevant at many scales of biology and may be important for biomolecular structure transitions in cell physiology and disease. In particular, the researchers discovered that various liquid crystalline structures could be accessed continuously simply by changes in environmental conditions, even as simple as changes in salinity or temperature; given the numerous unexplored conditions, this work suggests many more novel self-organised LC mesophases with potential biological function could be discovered in the near future.This new understanding of biopolymeric self-organisation may also be important for understanding how life self-organised to become living in the first place. Understanding how primitive collections of molecules could have structured themselves into collectively behaving aggregates is a significant avenue of future research."When the general public hears about liquid crystals, they might think of TV screens and engineering applications. However, very few would immediately think of basic science. Most researchers would not even make the connection between LCs and the origins of life. We hope this work will help increase the public's understanding of LCs in the context of the origins of life," says co-author Jia.Finally, this work may also be relevant to disease. For example, recent discoveries regarding diseases including Alzheimer's, Parkinson's, Huntington's Disease, and ALS (Lou Gehrig's Disease) have pointed to intracellular phase transitions and separation leading to membraneless droplets as potential major causes.The researchers noted that though their work was heavily impacted by the pandemic, they did their best to keep working under the global shutdowns and travel restrictions.
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Biotechnology
| 2,020 |
October 16, 2020
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https://www.sciencedaily.com/releases/2020/10/201016143050.htm
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Fats fighting back against bacteria
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Droplets of fat inside our cells are helping the body's own defence system fight back against infection, University of Queensland researchers have discovered.
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The international collaboration between UQ Institute for Molecular Bioscience researchers Professor Robert Parton and Professor Matt Sweet, and the University of Barcelona's Professor Albert Pol found that these fat droplets are both a food source and weapon against bacterial invaders."It was previously thought that bacteria were merely using the lipid droplets to feed on, but we have discovered these fatty droplets are involved in the battle between the pathogens and our cells," Professor Parton said."Fat is part of the cell's arsenal -- cells manufacture toxic proteins, package them into the lipid droplets, then fire them at the intruders."This is a new way that cells are protecting themselves, using fats as a covert weapon, and giving us new insights into ways of fighting infection."With antibiotic-resistant superbugs on the rise, researchers are determined to find alternative ways to fight infection.One possibility is ramping up the body's natural defences."We showed that upon infection of white blood cells called macrophages, lipid droplets move to the part of the macrophage where the bacteria are present," Professor Sweet said.The bacterial infection also changed the way that white blood cells used energy."Lipid droplets can be used as a fuel source for mitochondria when there aren't enough other nutrients," Professor Sweet said."During an infection, lipid droplets move away from the mitochondria and attack the bacteria instead, altering metabolism of the cell."Cell biologist Professor Parton was inspired to continue this research after the phenomenon was seen in fruit flies."Most people thought the lipid droplets were 'blobs of fat', only useful for energy storage but now we are seeing that they act as metabolic switches in the cell, defend against infection and much more -- there are now entire scientific conferences of researchers working on them," he said."Our next step is to find out how the lipid droplets target the bacteria."By understanding the body's natural defences, we can develop new therapies that don't rely on antibiotics to fight drug-resistant infections."VIDEO --
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Biotechnology
| 2,020 |
October 16, 2020
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https://www.sciencedaily.com/releases/2020/10/201016122403.htm
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Pinpointing the 'silent' mutations that gave the coronavirus an evolutionary edge
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We know that the coronavirus behind the COVID-19 crisis lived harmlessly in bats and other wildlife before it jumped the species barrier and spilled over to humans.
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Now, researchers at Duke University have identified a number of "silent" mutations in the roughly 30,000 letters of the virus's genetic code that helped it thrive once it made the leap -- and possibly helped set the stage for the global pandemic. The subtle changes involved how the virus folded its RNA molecules within human cells.For the study, published Oct. 16 in the journal "We're trying to figure out what made this virus so unique," said lead author Alejandro Berrio, a postdoctoral associate in biologist Greg Wray's lab at Duke.Previous research detected fingerprints of positive selection within a gene that encodes the "spike" proteins studding the coronavirus's surface, which play a key role in its ability to infect new cells.The new study likewise flagged mutations that altered the spike proteins, suggesting that viral strains carrying these mutations were more likely to thrive. But with their approach, study authors Berrio, Wray and Duke Ph.D. student Valerie Gartner also identified additional culprits that previous studies failed to detect.The researchers report that so-called silent mutations in two other regions of the SARS-CoV-2 genome, dubbed Nsp4 and Nsp16, appear to have given the virus a biological edge over previous strains without altering the proteins they encode.Instead of affecting proteins, Berrio said, the changes likely affected how the virus's genetic material -- which is made of RNA -- folds up into 3-D shapes and functions inside human cells.What these changes in RNA structure might have done to set the SARS-CoV-2 virus in humans apart from other coronaviruses is still unknown, Berrio said. But they may have contributed to the virus's ability to spread before people even know they have it -- a crucial difference that made the current situation so much more difficult to control than the SARS coronavirus outbreak of 2003.The research could lead to new molecular targets for treating or preventing COVID-19, Berrio said."Nsp4 and Nsp16 are among the first RNA molecules that are produced when the virus infects a new person," Berrio said. "The spike protein doesn't get expressed until later. So they could make a better therapeutic target because they appear earlier in the viral life cycle."More generally, by pinpointing the genetic changes that enabled the new coronavirus to thrive in human hosts, scientists hope to better predict future zoonotic disease outbreaks before they happen."Viruses are constantly mutating and evolving," Berrio said. "So it's possible that a new strain of coronavirus capable of infecting other animals may come along that also has the potential to spread to people, like SARS-CoV-2 did. We'll need to be able to recognize it and make efforts to contain it early."
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Biotechnology
| 2,020 |
October 16, 2020
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https://www.sciencedaily.com/releases/2020/10/201016090213.htm
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Gel instrumental in 3D bioprinting biological tissues
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The eventual creation of replacement biological parts requires fully three-dimensional capabilities that two-dimensional and three-dimensional thin-film bioprinting cannot supply. Now, using a yield stress gel, Penn State engineers can place tiny aggregates of cells exactly where they want to build the complex shapes that will be necessary to replace bone, cartilage and other tissues.
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"The reason why this is important is that the current cell aggregate bioprinting techniques can't make complicated configurations and is mostly in 2D and 3D thin films or simple configurations," said Ibrahim T. Ozbolat, Hartz Family Career Development Associate Professor of Engineering Science and Mechanics. "If we want complicated 3D, we need a supportive field."That supportive field, the researchers report today (Oct. 16) in The researchers are using an aspiration-assisted bioprinting system that they demonstrated earlier this year to pick up aggregates of cells and place them precisely inside of the gel. The stress of the aspiration nozzle against the gel liquefies it, but once the aspiration nozzle releases cell aggregates and withdraws, the gel returns to solid again, self-healing. The tiny balls of cells rest upon each other and self-assemble, creating a solid tissue sample within the gel.The researchers can place different types of cells, in small aggregates, together to form the required shape with the required function. Geometric shapes like the cartilage rings that support the trachea, could be suspended within the gel."We tried two different types of gels, but the first one was a little tricky to remove," said Ozbolat. "We had to do it through washing. For the second gel, we used an enzyme that liquefied the gel and removed it easily.""What we are doing is very important because we are trying to recreate nature," said Dishary Banerjee, postdoctoral researcher in engineering science and mechanics. "In this technology it is very important to be able to make free-form, complex shapes from spheroids."The researchers used a variety of approaches, creating theoretical models to get a physical understanding of what was happening. They then used experiments to test if this method could produce complex shapes.
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Biotechnology
| 2,020 |
October 15, 2020
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https://www.sciencedaily.com/releases/2020/10/201015111731.htm
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Machine learning uncovers potential new TB drugs
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Machine learning is a computational tool used by many biologists to analyze huge amounts of data, helping them to identify potential new drugs. MIT researchers have now incorporated a new feature into these types of machine-learning algorithms, improving their prediction-making ability.
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Using this new approach, which allows computer models to account for uncertainty in the data they're analyzing, the MIT team identified several promising compounds that target a protein required by the bacteria that cause tuberculosis.This method, which has previously been used by computer scientists but has not taken off in biology, could also prove useful in protein design and many other fields of biology, says Bonnie Berger, the Simons Professor of Mathematics and head of the Computation and Biology group in MIT's Computer Science and Artificial Intelligence Laboratory (CSAIL)."This technique is part of a known subfield of machine learning, but people have not brought it to biology," Berger says. "This is a paradigm shift, and is absolutely how biological exploration should be done."Berger and Bryan Bryson, an assistant professor of biological engineering at MIT and a member of the Ragon Institute of MGH, MIT, and Harvard, are the senior authors of the study, which appears today in Machine learning is a type of computer modeling in which an algorithm learns to make predictions based on data that it has already seen. In recent years, biologists have begun using machine learning to scour huge databases of potential drug compounds to find molecules that interact with particular targets.One limitation of this method is that while the algorithms perform well when the data they're analyzing are similar to the data they were trained on, they're not very good at evaluating molecules that are very different from the ones they have already seen.To overcome that, the researchers used a technique called Gaussian process to assign uncertainty values to the data that the algorithms are trained on. That way, when the models are analyzing the training data, they also take into account how reliable those predictions are.For example, if the data going into the model predict how strongly a particular molecule binds to a target protein, as well as the uncertainty of those predictions, the model can use that information to make predictions for protein-target interactions that it hasn't seen before. The model also estimates the certainty of its own predictions. When analyzing new data, the model's predictions may have lower certainty for molecules that are very different from the training data. Researchers can use that information to help them decide which molecules to test experimentally.Another advantage of this approach is that the algorithm requires only a small amount of training data. In this study, the MIT team trained the model with a dataset of 72 small molecules and their interactions with more than 400 proteins called protein kinases. They were then able to use this algorithm to analyze nearly 11,000 small molecules, which they took from the ZINC database, a publicly available repository that contains millions of chemical compounds. Many of these molecules were very different from those in the training data.Using this approach, the researchers were able to identify molecules with very strong predicted binding affinities for the protein kinases they put into the model. These included three human kinases, as well as one kinase found in Mycobacterium tuberculosis. That kinase, PknB, is critical for the bacteria to survive, but is not targeted by any frontline TB antibiotics.The researchers then experimentally tested some of their top hits to see how well they actually bind to their targets, and found that the model's predictions were very accurate. Among the molecules that the model assigned the highest certainty, about 90 percent proved to be true hits -- much higher than the 30 to 40 percent hit rate of existing machine learning models used for drug screens.The researchers also used the same training data to train a traditional machine-learning algorithm, which does not incorporate uncertainty, and then had it analyze the same 11,000 molecule library. "Without uncertainty, the model just gets horribly confused and it proposes very weird chemical structures as interacting with the kinases," Hie says.The researchers then took some of their most promising PknB inhibitors and tested them against Mycobacterium tuberculosis grown in bacterial culture media, and found that they inhibited bacterial growth. The inhibitors also worked in human immune cells infected with the bacterium.Another important element of this approach is that once the researchers get additional experimental data, they can add it to the model and retrain it, further improving the predictions. Even a small amount of data can help the model get better, the researchers say."You don't really need very large data sets on each iteration," Hie says. "You can just retrain the model with maybe 10 new examples, which is something that a biologist can easily generate."This study is the first in many years to propose new molecules that can target PknB, and should give drug developers a good starting point to try to develop drugs that target the kinase, Bryson says. "We've now provided them with some new leads beyond what has been already published," he says.The researchers also showed that they could use this same type of machine learning to boost the fluorescent output of a green fluorescent protein, which is commonly used to label molecules inside living cells. It could also be applied to many other types of biological studies, says Berger, who is now using it to analyze mutations that drive tumor development.The research was funded by the U.S. Department of Defense through the National Defense Science and Engineering Graduate Fellowship; the National Institutes of Health; the Ragon Institute of MGH, MIT, and Harvard' and MIT's Department of Biological Engineering.
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Biotechnology
| 2,020 |
October 15, 2020
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https://www.sciencedaily.com/releases/2020/10/201015101807.htm
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Novel antiviral strategy for treatment of COVID-19
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A research team led by Professor Hongzhe SUN, Norman & Cecilia Yip Professor in Bioinorganic Chemistry, Department of Chemistry, Faculty of Science, and Professor Kwok Yung YUEN, Henry Fok Professor in Infectious Diseases, Department of Microbiology, Li Ka Shing Faculty of Medicine of the University of Hong Kong (HKU), has discovered a novel antiviral strategy for treatment of COVID-19.
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They discovered that a class of metallodrugs currently used in the treatment of other infectious diseases is showing efficacy to potently suppress SARS-CoV-2 replication and relieve viral-associated symptoms in an animal model.The findings provide a new and readily available therapeutic option with high clinical potential for infection with SARS-CoV-2. This ground-breaking work has been published online in a top-class scientific journal SARS-CoV-2 is an emerging coronavirus that has caused over 30 million laboratory-confirmed cases and more than 1 million deaths globally of COVID-19 since December 2019. As the process of developing an effective vaccine is still ongoing, another approach for prevention and treatment of the disease is to identify anti-COVID-19 agents from existing virus-specific antiviral drugs to repurpose their uses to target the new virus. Remdesivir, a broad-spectrum antiviral drug, has been reported to show efficacy towards SARS-CoV-2. However, global shortage of the drug, its relatively high price and lack of significant clinical benefits in severe cases, are factors that have limited its wider applications. Clinical trials on a series of antiviral agents are still ongoing which have yet to demonstrate therapeutic efficacies. Therefore, greater efforts are needed to extend the evaluation to cover a wider spectrum of clinically approved drugs, which hopefully could open the way to alternative treatment strategies against the disease through some readily available channels.Generally, metal compounds are used as anti-microbial agents; their antiviral activities have rarely been explored. After screening a series of metallodrugs and related compounds, the research team identified ranitidine bismuth citrate (RBC), a commonly used anti-ulcer drug which contains the metal Bismuth for treatment of Helicobacter pylori-associated infection, as a potent anti-SARS-CoV-2 agent, both in vitro and in vivo.RBC targets the vital non-structural protein 13 (Nsp13), a viral helicase essential for SARS-CoV-2 to replicate, by displacing the crucial zinc(II) ions in the zinc-binding with Bismuth-ions, to potently suppress the activity of the helicase.RBC has been demonstrated to greatly reduce viral loads by over 1,000-folds in SARS-CoV-2-infected cells. In particular, in a golden Syrian hamster model, RBC suppresses SARS-CoV-2 replications to reduce viral loads by ~100 folds in both the upper and lower respiratory tracts, and mitigates virus-associated pneumonia. RBC remarkably diminishes the level of prognostic markers and other major pro-inflammatory cytokines and chemokines in severe COVID-19 cases of infected hamsters, compared to the Remdesivir-treated group and control group.RBC exhibits a low cytotoxicity with a high selectivity index at 975 (the larger the number the safer the drug), as compared to Remdesivir which has a low selectivity index at 129. The finding indicates a wide window between the drug's cytotoxicity and antiviral activity, which allows a great flexibility in adjusting its dosages for treatment.The team investigated the mechanisms of RBC on SARS-CoV-2 and revealed for the first time the vital Nsp13 helicase as a druggable target by RBC. It irreversibly kicks out the crucial zinc(II) ions in the zinc-binding domain to change it to bismuth-bound via a distinct metal displacement route. RBC and its Bi(III) compounds dysfuntionalised the Nsp13 helicase and potently inhibited both the ATPase (IC50=0.69 µM) and DNA-unwinding (IC50=0.70 µM) activities of this enzyme.The research findings highlight viral helicases as a druggable target, and the high clinical potential of bismuth(III) drugs and other metallodrugs for treatment of SARS-CoV-2 infections. Hopefully, following this important breakthrough, more antiviral agents from readily available clinically approved drugs could be identified for potential treatment of COVID-19 infections. They can be in the form of combination regimens (cocktails) with drugs that exhibit anti-SARS-CoV-2 activities including RBC, dexamethasone and interferon-β1b.
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Biotechnology
| 2,020 |
October 15, 2020
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https://www.sciencedaily.com/releases/2020/10/201015101830.htm
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Boost to develop microalgae into health foods
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Dietary supplementation of fatty acids produced from microalgae have wide-reaching health benefits for humans, including the ability to reduce obesity, diabetes and fatty liver disease, preventing hair loss, and assisting wounds to heal.
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However, its widespread development has been hampered by the current limits of bioimaging tools needed to allow easy, rapid and non-invasive evaluation of lipid conditions within microalgae.A novel protocol to detect lipid production in microalgae has been discovered at Flinders University by Mohsinul Reza, a PhD student under the supervision of Professors Jian Qin and Youhong Tang.The study details the development of a novel protocol by using a novel aggregation induced emission (AIE) fluorescent bioprobe to detect the production of lipid drops from microalgae.Mr Reza has discovered the optimal condition to maximise the production of long-chain polyunsaturated fatty acid such as omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in Euglena gracilis -- a species of lipid-producing microalgae that can biosynthesise multiple beneficiary compounds as food supplements for human health."This technique enables us to visualise the distribution and quantity of lipid drops in live algae on a confocal microscope," says Professor Jian Qin. "This new method could screen the capacity of lipid droplet production in other algal species that have the potential as a source to produce healthy food for humans."The new technique improves on traditional fluorescent probes currently used for lipid imaging, which often suffer from reduced photostability and difficulties in dye acquisition techniques that limits their usage for microscopic imaging.The new bioprobe DPAS (a lipid-specific AIE fluorogen that is synthesised from very cheap materials) could surpass the performances of the traditional fluorophore for lipid droplets staining in terms of photostability, rapid and easy sample preparation techniques.This new technique significantly eases the lipid study in this algal cell type. This fluorescent probe is also biocompatible and suitable for multicolour imaging that broadens the horizon of this dye for biological studies.The researchers also observed cultural conditions that can produce higher amount of health beneficiary fatty acids, which suggests that promising bio-functional compounds could be available from culturing Euglena gracilis microalgae in the applied conditions.In details, the researchers tested five different treatments and analysed the results using DPAS and BODIPY (a well-known staining probe) to compare the results. They found that the presence of organic carbon in the form of glucose and deprivation of nitrogen and calcium from the algal culture enhanced lipid production in a dark condition.
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Biotechnology
| 2,020 |
October 14, 2020
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https://www.sciencedaily.com/releases/2020/10/201014141141.htm
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Virus-mimicking drug helps immune system target cunning cancer cells
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Researchers at the UCLA Jonsson Comprehensive Cancer Center have found that a drug that activates the body's natural defenses by behaving like a virus may also make certain stealthy melanoma tumors visible to the immune system, allowing them to be better targeted by immunotherapy.
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The findings, published today in the journal "Most immunotherapy approaches rely on the ability of T cells to recognize and kill tumor cells," said lead and corresponding author Dr. Anusha Kalbasi, an assistant professor of radiation oncology at the David Geffen School of Medicine at UCLA and member of the Jonsson Cancer Center. "But in some patients, tumors escape the immune system through mutations in genes involved in the interferon signaling pathway. This is a critical pathway because it normally allows tumors to increase their antigen presentation, an intricate machinery that makes tumors visible to T cells."Interferons are proteins in cells that respond to viral infection by impeding the virus's ability to replicate and alerting the immune system to marshal its forces. Activating interferon signaling in tumors helps slow down tumor division and can lead to the release of molecules that recruit more immune cells to the tumor."This coordinated effort as a result of interferon signaling can help the immune system better identify and kill tumor cells," Kalbasi said.The team first attempted to overcome defective interferon signaling by using adoptive T cell therapy, a type of immunotherapy that involves extracting T cells from a patient and engineering them in the laboratory to recognize and kill cancer cells. The researchers found that these T cells remained ineffective against tumors with defective interferon signaling.The authors then engineered mouse melanoma tumor cells with a gene called NLRC5. NLRC5 increased antigen presentation even in the absence of interferon signaling and restored the effectiveness of the T cells. While this approach was effective in mice, engineering tumor cells in humans was not as simple.Instead, Kalbasi and his colleagues turned to a virus-mimicking drug called BO-112 that activates virus-sensing pathways in tumors. When the drug was injected directly into the tumor in the laboratory, the team discovered that the activation of virus-sensing pathways increased antigen presentation even when interferon signaling was defective. As a result, these tumors could be recognized and killed by T cells."This study helps us understand the interdependence between interferon signaling and antigen presentation, which gives us important insights into how tumor cells are recognized by the immune system," said the study's senior author, Dr. Antoni Ribas, a professor of medicine at the Geffen School of Medicine and director of the tumor immunology program at the Jonsson Cancer Center. "New strategies to promote antigen presentation to make tumors more visible to the immune system will allow immunotherapy to be effective for even more tumor types."The findings also highlight the potential of other promising clinical approaches that bypass tumor interferon signaling and antigen presentation, like CAR, or chimeric antigen receptor-based T cell therapy, which can recognize and kill tumor cells even in absence of antigen presentation.Kalbasi is now leading a human clinical trial of the combination therapy of nivolumab, an immune checkpoint blockade drug, and BO-112 in people with certain types of sarcoma who are undergoing radiation followed by surgery. The idea is to activate the immune system against the patient's tumor while the tumor is still in the body.
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Biotechnology
| 2,020 |
October 14, 2020
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https://www.sciencedaily.com/releases/2020/10/201014082758.htm
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Molecular dance keeps your heart beating
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It might look like a little game at the molecular scale.
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Filament-like proteins in heart muscle cells have to be exactly the same length so that they can coordinate perfectly to make the heart beat.Another protein decides when the filament is the right size and puts a wee little cap on it. But, if that protein makes a mistake and puts the cap on too early, another protein, leiomodin, comes along and knocks the cap out of the way.This little dance at the molecular scale might sound insignificant, but it plays a critical role in the development of healthy heart and other muscles. Reporting in the journal, The finding could someday lead to improved diagnostics and medical treatments for serious and sometimes devastating hereditary heart conditions that come about from genetic mutations in the proteins. One of these conditions, cardiomyopathy, affects as many as one in 500 people around the world and can often be fatal or have lifetime health consequences. A similar condition called nemaline myopathy affects skeletal muscles throughout the body with often devastating consequences."Mutations in these proteins are found in patients with myopathy," said Alla Kostyukova, associate professor in the Gene and Linda Voiland School of Chemical Engineering and Bioengineering and leader of the project. "Our work is to prove that these mutations cause these problems and to propose strategies for treatment."Heart muscle is made of tiny thick and thin filaments of proteins. With the help of electrical signals, the rope-like filaments bind and unbind in an intricate and precise architecture, allowing heart muscle to contract and beat.The thin filaments are made of actin, the most abundant protein in the human body. Tropomysin, another protein, wraps itself around the actin filaments. Tropomyosin together with two other proteins, tropomodulin and leiomodin, at the end of the actin filaments act as a sort of cap and determine the filament length."It's beautifully designed," said Kostyukova, whose research is focused on understanding protein structures.And, tightly regulated.To keep heart muscle healthy, the actin filaments, which are about a micron long, all have to be the exact same length. In families with cardiomyopathy, genetic mutations result in formation of filaments that are either too short or too long. Those affected can have significant heart problems that cause disability, illness and death.In a project that spanned seven years, the researchers proved that leiomodin attaches to the end of the actin filament and kicks out the other protein, tropomodulin, to assure the actin filament's proper length."This is the first time that this has been shown with the atomic-level precision," said Dmitri Tolkatchev, research assistant professor in the Voiland School and lead author on the paper. "Previously, several laboratories attempted to solve this problem with very little success. With our data we finally have a direct proof."The researchers used state-of-the-art approaches to make the key proteins and study them at the molecular and cellular level. The work entailed designing the molecules, constructing them at the gene level in a plasmid, and then producing them into bacterial or cardiac cells. The researchers used nuclear magnetic resonance, which works on the same physical principle as Magnetic Resonance Imaging (MRIs), to understand the proteins' binding at the atomic level. They also used molecular dynamic simulation to model them."The probability of being able to show this mechanism was not high, but the impact of the discovery is," said Tolkatchev, an expert in nuclear magnetic resonance. "This was a very important problem to study and could have a significant impact in the field of muscle mechanics."The researchers hope to continue the work, identifying additional components and molecular mechanisms that regulate thin filament architecture, whether diseased or healthy.
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Biotechnology
| 2,020 |
October 13, 2020
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https://www.sciencedaily.com/releases/2020/10/201013134304.htm
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Bacterial toxin with healing effect
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A bacterial toxin promoting tissue healing has been discovered. The compound, found in Staphylococcus aureus, does not just damage cells, but also stimulates tissue regeneration.
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Normally they are among the many harmless organisms found in and on the human body: one in four people have millions of The Professor for Pharmaceutical Chemistry and his team have studied the molecular defence mechanisms of the human immune system in the fight against such In their latest study, the researchers from the University of Jena, Jena University Hospital and the Leibniz Institute on Aging -- Fritz Lipmann Institute (FLI), together with colleagues from Harvard Medical School and the University of Naples, have studied in particular the bacterial toxin "?-Hemolysin" and examined its effect on M2 macrophages. M2 macrophages are immune cells which, in the later stages of an inflammatory reaction, ensure that bacteria that have been killed, and damaged cell components, are removed, and that the tissue regenerates. "They are therefore a kind of cellular waste disposal," says Paul Jordan, doctoral candidate in Werz's team and lead author of the publication, describing the function of these cells.The researchers showed that ?-hemolysin binds to specific receptor proteins on the surface of M2 macrophages and thus triggers the production of anti-inflammatory messenger substances in the cells, which then cause the inflammation to resolve. In the study, the scientists were also able to show that these transmitters promote tissue regeneration in an animal model. The anti-inflammatory messenger substances include resolvins, maresins and protectins that are formed from omega-3 fatty acids.
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Biotechnology
| 2,020 |
October 13, 2020
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https://www.sciencedaily.com/releases/2020/10/201013124105.htm
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Mechanism that restores cell function after genome damage
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A research team from Cologne has discovered that a change in the DNA structure -- more precisely in the chromatin -- plays a decisive role in the recovery phase after DNA damage. The key is a double occupation by two methyl groups on the DNA packaging protein histone H3 (H3K4me2). The discovery was made by scientists under the direction of Prof. Björn Schumacher of the Cluster of Excellence for Aging Research CECAD, the Center for Molecular Medicine Cologne (CMMC), and the Institute for Genome Stability in Aging and Disease at the University of Cologne. The specific change enables genes to be reactivated and proteins to be produced after damage: The cells regain their balance and the organism recovers. The protective role of H3K4me2 was identified in experiments with the nematode Caenorhabditis elegans. The study has now been published in the journal
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The genome in every human cell is damaged on a daily basis, for example in the skin by UV radiation from the sun. Damage to the DNA causes diseases such as cancer, influences development, and accelerates aging. Congenital malfunctions in DNA repair can lead to extremely accelerated aging in rare hereditary diseases. Therefore, preservation and reconstruction processes are particularly important to ensure development and to maintain tissue function. DNA, which is rolled up on packaging proteins -- the histones -- like on cable drums, is regulated by methyl groups. Various proteins are responsible for placing methyl groups on histones or removing them. The number of groups on the packaging proteins affects the activity of genes and thus the protein production of the cell.In experiments with the nematode, the research team showed that after repairing damaged DNA, two methyl groups were increasingly found on the DNA packages. Furthermore, they found that errors in placing these two methyl groups on the histones (H3K4me2) accelerated the damage-induced aging process, while increased position of this histone alteration prolongs the lifespan after DNA damage. By controlling the proteins that either set or remove these methyl groups, the resistance to DNA damage -- and thus the aging process of the animals -- could be influenced.Further analysis of the role of these two methyl groups showed that the enrichment of H3K4 after genome damage with two methyl groups supports the cells in restoring the balance after DNA damage.'Now that we know the exact changes in chromatin, we can use this to precisely limit the consequences of DNA damage,' said Schumacher. 'I hope that these findings will enable us to develop therapies for hereditary diseases characterized by developmental disorders and premature aging. Due to the fundamental importance of DNA damage in the aging process, such approaches could also counteract normal aging and prevent age-related diseases.'
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Biotechnology
| 2,020 |
October 14, 2020
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https://www.sciencedaily.com/releases/2020/10/201014161648.htm
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Why do identical cells act differently? Team unravels sources of cellular 'noise'
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University of Texas at Dallas researchers have taken an important step toward explaining why genetically identical cells can produce varying amounts of the same protein associated with the same gene.
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In a study published Aug. 18 and appearing in the Sept. 18 print edition of the journal Understanding why and how such fluctuations, or cellular noise, occur is a fundamental bioengineering problem, said Dr. Leonidas Bleris, associate professor of bioengineering in the Erik Jonsson School of Engineering and Computer Science and Fellow, Cecil H. and Ida Green Professor in Systems Biology Science."The quest to understand cellular noise is driven primarily by our interest in applying engineering to biology. To paraphrase [physicist] Richard Feynman, by understanding, we will be able to create," said Bleris, the corresponding author of the study. "We are interested in applying control in cells to achieve desirable objectives. The applications range from sophisticated gene therapy to engineering pathways in order to produce valuable compounds."Nearly every cell in a person's body contains the same DNA, the master set of genetic instructions for making the complex proteins that do most of the biological work. DNA segments called genes encode specific proteins. But the amount of protein produced by a given gene -- referred to as gene expression -- can vary not only between people, but also among identical cells in the same person. That fluctuation in gene expression between identical cells is called cellular noise.Using a combination of experiments and theory, Bleris and his colleagues analyzed each stage of the process by which information in DNA is converted to proteins, a process referred to as the central dogma of molecular biology.The process begins with transcription of a gene, in which the information in DNA is copied into a related kind of genetic material called RNA. The cell then uses the information in the RNA to build proteins.To understand the source of cellular noise, researchers in the Bleris Lab engineered custom genetic circuits, a synthetic biology approach that allowed the team to isolate each step of the standard protein-making process.Then, the team used the gene-editing tool CRISPR to insert single copies of these circuits surgically in a predetermined genomic location in human cells. This combination of synthetic biology and genome editing made it possible for the team to analyze fluctuations in cells at different stages of the protein-making process and pinpoint transcription as the source of noise.The 2020 Nobel Prize in chemistry was awarded Oct. 7 to two scientists who developed the CRISPR/Cas9 genetic scissors -- Dr. Emmanuelle Charpentier, director of the Max Planck Unit for the Science of Pathogens in Berlin, and Dr. Jennifer Doudna, a biochemist at the University of California, Berkeley and a Howard Hughes Medical Institute investigator.Understanding differences in how genetically identical cells behave can help scientists develop more effective, targeted therapies, Bleris said."Eventually, once we have a better understanding of how our genes operate in their intrinsically fluctuating environment, we will be able to engineer a more sophisticated class of gene therapies that can more appropriately address the diseases that ail humanity," said Tyler Quarton PhD'19, one of the study's lead authors.The research also raises questions for further study."Understanding the sources of noise opens the path for asking new questions: What is the biological function of noise? Is noise used by cells to introduce diversity, or is it simply a nuisance?" explained Taek Kang, a biomedical engineering doctoral student, Eugene McDermott Graduate Fellow and co-lead author.The team also included Vasileios Papakis BS'20; Khai Nguyen, biomedical engineering senior; Chance Nowak, a molecular and cell biology doctoral student; and Dr. Yi Li, a research scientist in bioengineering.The research was supported by the National Science Foundation, including Bleris' NSF Faculty Early Career Development (CAREER) Award.
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Biotechnology
| 2,020 |
October 13, 2020
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https://www.sciencedaily.com/releases/2020/10/201013105750.htm
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Stopping lethal lung damage from the flu with a natural human protein
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The raging lung inflammation that can contribute to death from the flu can be stopped in its tracks by a drug derived from a naturally occurring human protein, a new animal study suggests.
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In mouse studies, all untreated animals given a lethal dose of influenza died within days. All but one of the infected mice treated with the experimental therapy not only survived, but remained energetic and kept weight on -- despite having high levels of the flu virus in their lungs.The experimental treatment is a heavy dose of MG53, part of a family of proteins that plays an essential role in cell membrane repair. Already identified as a potential therapy for conditions ranging from Alzheimer's disease to persistent skin wounds, MG53 was found in this study to prevent death from a lethal flu infection by blocking excessive inflammation -- without having any effect on the virus itself.The researchers are currently testing the effects of the therapy in mice infected with SARS-CoV-2, the coronavirus that causes COVID-19."I haven't ever seen anything like this before," said Jacob Yount, associate professor of microbial infection and immunity at The Ohio State University and co-lead author of the study. "Even though these mice had the same viral load as the untreated mice, they didn't get very sick -- with the lethal dose of the flu."Yount, whose lab studies the immune response against viral infections, co-led the work with Jianjie Ma, professor of cardiac surgery at Ohio State, who discovered MG53 and its role in cell repair and has been developing the protein as a therapeutic agent.The paper was published online Oct. 8 in the The collaboration on this work grew out of a proposal by Matthew Sermersheim, a graduate student in Ma's lab, to expand on the investigation of MG53's links to inflammation. In the July 17 issue of Nature Communications, Sermersheim was the first author of a study showing that the lungs of mice lacking the MG53 gene and infected with flu responded with extensive inflammation compared to normal mice -- indicating that MG53 has a protective role in the immune response.For this new work, the scientists put MG53 to the test against influenza, which, along with other respiratory viruses, is a top-10 cause of death worldwide.The researchers infected mice with a dose of an H1N1 strain of influenza and treated half with a placebo. Using recombinant human MG53, a molecule Ma's lab has been developing as a drug, the researchers treated the other half of mice with seven daily injections beginning 24 hours after infection. The untreated mice showed an aggressive loss of weight and died within nine days, but 92% of the treated mice lost very little weight, remained active and returned to their normal weight by two weeks after infection."The protein has a way to recognize tissue that's been injured and it can go there directly," Ma said. "We are basically enhancing a natural anti-inflammatory mechanism in the body so that when you face the crisis of an aggressive virus infection, the body can better defend itself."Despite the strikingly different outcomes, the viral loads in both sets of mice were similar -- meaning an MG53-based agent is not an anti-viral drug. Even teeming with the flu virus, the airways of treated mice showed little tissue damage.Though the team is still working to fully identify how this protection occurs, the researchers determined that MG53 stops an immune response mishap called a "cytokine storm," which leads to tissue damage. The research also showed that MG53 mitigates an infection-related cell-death process called pyroptosis, which also promotes inflammation and lung dysfunction."A lot of the lung damage with the flu virus is actually caused by excessive inflammation from our own immune response," Yount said. "If you can dampen that hyperactive immune response, you'll have less tissue damage, even though the virus is still replicating at really high levels."Lung tissue damaged by inflammation is deadly because it allows fluid and cells to build up in airways, preventing the lungs from absorbing oxygen.Ma's previous work in animal models suggests driving up levels of MG53 in the body for therapeutic purposes is safe: Mice his lab has genetically engineered to over-produce the protein live longer and healthier lives than normal mice. Though the scientists envision MG53 as part of a cocktail of drugs targeting deadly viral infections, they caution that much more research is needed before a therapy is available for humans."We need better anti-inflammatory tissue repair therapies," Ma said. "We don't have COVID-19 data yet, but even with influenza, which hits us on a seasonal basis, this application could make quite a bit of difference."
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Biotechnology
| 2,020 |
October 12, 2020
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https://www.sciencedaily.com/releases/2020/10/201012164223.htm
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Scientists engineer bacteria-killing molecules from wasp venom
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A team led by scientists in the Perelman School of Medicine at the University of Pennsylvania has engineered powerful new antimicrobial molecules from toxic proteins found in wasp venom. The team hopes to develop the molecules into new bacteria-killing drugs, an important advancement considering increasing numbers of antibiotic-resistant bacteria which can cause illness such as sepsis and tuberculosis.
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In the study, published today in the There is an urgent need for new drug treatments for bacterial infections, as many circulating bacterial species have developed a resistance to older drugs. The U.S. Centers for Disease Control & Prevention has estimated that each year nearly three million Americans are infected with antibiotic-resistant microbes and more than 35,000 die of them. Globally the problem is even worse: Sepsis, an often-fatal inflammatory syndrome triggered by extensive bacterial infection, is thought to have accounted for about one in five deaths around the world as recently as 2017."New antibiotics are urgently needed to treat the ever-increasing number of drug-resistant infections, and venoms are an untapped source of novel potential drugs. We think that venom-derived molecules such as the ones we engineered in this study are going to be a valuable source of new antibiotics," said study senior author César de la Fuente, PhD, a Presidential Assistant Professor in Psychiatry, Microbiology, and Bioengineering at Penn.De la Fuente and his team started with a small protein, or "peptide," called mastoparan-L, a key ingredient in the venom of Vespula lewisii wasps. Mastoparan-L-containing venom is usually not dangerous to humans in the small doses delivered by wasp stings, but it is quite toxic. It destroys red blood cells, and triggers a type of allergic/inflammatory reaction that in susceptible individuals can lead to a fatal syndrome called anaphylaxis -- in which blood pressure drops and breathing becomes difficult or impossible.Mastoparan-L (mast-L) also is known for its moderate toxicity to bacterial species, making it a potential starting point for engineering new antibiotics. But there are still some unknowns, including how to enhance its anti-bacterial properties, and how to make it safe for humans.The team searched a database of hundreds of known antimicrobial peptides and found a small region, the so-called pentapeptide motif, that was associated with strong activity against bacteria. The researchers then used this motif to replace a section at one end of mast-L that is thought to be the chief source of toxicity to human cells.In a key set of experiments, the researchers treated mice with mast-MO several hours after infecting them with otherwise lethal, sepsis-inducing strains of the bacteria E. coli or Staphylococcus aureus. In each test the antimicrobial peptide kept 80 percent of treated mice alive. By contrast, mice treated with mast-L were less likely to survive, and showed severe toxic side-effects when treated with higher doses -- doses at which mast-MO caused no evident toxicity.The potency of mast-MO in these tests also appeared to be comparable to existing antibiotics such as gentamicin and imipenem -- for which alternatives are needed due to the spread of resistant bacterial strains.De la Fuente and his colleagues found evidence in the study that mast-MO kills bacterial cells by making their outer membranes more porous -- which can also improve the ability of co-administered antibiotics to penetrate the cells -- and by summoning antimicrobial white blood cells. At the same time, mast-MO appears to damp down the kind of harmful immune-overreaction that can lead to severe disease in some bacterial infections.The researchers created dozens of variants of mast-MO and found several that appeared to have significantly enhanced antimicrobial potency with no toxicity to human cells. They hope to develop one or more of these molecules into new antibiotics -- and they expect to take a similar approach in future to turn other venom toxins into promising antibiotic candidates."The principles and approaches we used in this study can be applied more broadly to better understand the antimicrobial and immune-modulating properties of peptide molecules, and to harness that understanding to make valuable new treatments," de la Fuente said.
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Biotechnology
| 2,020 |
October 12, 2020
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https://www.sciencedaily.com/releases/2020/10/201012103129.htm
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Dueling proteins give shape to plants
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In an elegant choreography, plants take cues from their environment and channel them into flowers, roots, or branches. In a new paper in
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Long interested in the translation of these environmental inputs to physical traits, Wagner and her team have been studying two key groups of proteins that influence plant form and timing of developmental transitions. Terminal Flower 1 (TLF1) proteins promote branch formation. When it is repressed, flowers grow. Flowering Locus T (FT) proteins, on the other hand, promote flowering in response to seasonal cues like day length. Strangely enough, the two proteins are almost identical."These two elements have significance galore," Wagner says. "Besides flowering, they're involved in tuberization in potatoes, bulb formation in onions, tendril formation in grapes, growth cessation in trees, lots of things."Manipulating these genes, some have argued, could lead to the next "green revolution" as one could theoretically "trick" a plant that normally only flowers in the long days of summer to flower quickly, and thus produce fruit or seed, in the short days of winter with a deft genetic edit of TFL1. Or, in an area with a longer growing season, a clever manipulation of growth architecture via FT could encourage increased branching and then a later, and more abundant, flowering and fruit development on the many branches.Plants have already been bred to have reduced TFL1 activity. Tomato gardeners may know these as determinant plants, which set all their flowers at the same time, as opposed to the indeterminant variety, which continue to branch, flower, and fruit over a period of months. Determinant plants make commercial agriculture more efficient, as fruits can be harvested in one go as opposed to repeated passes.It was known that TFL1 and FT1 acted in opposing directions, each "tuning" the activity of the other, but the mechanism of their antagonism has remained fuzzy. In part this was because studying them has presented a technical challenge: They are only present in low levels in a limited number of cells.Wagner's groups, however, had overcome similar challenges in studying a regulator of plant chromatin in earlier work, so they were undeterred in taking on these two dueling proteins.In the current study, to flesh out TFL1 and FT1's molecular mechanism, they first looked to see where TFL1 was found in the nuclei of plant cells, using the model species Arabidopsis thaliana. They found thousands of sites to which it bound, acting through the transcription factor FD, as neither TFL1 nor FT otherwise can bind directly to DNA. The sites to which TFL1 was recruited were consistent with its role in suppressing flowering and in suppressing gene expression.The researchers next examined the relationship of both TFL1 and FT with LEAFY, a gene that is known to give rise to flowers. When they mutated the sites where TFL1 regulates the LEAFY gene, LEAFY protein was now found in parts of the plant where TFL1 is present."We also saw something that we didn't expect," Wagner says. "LEAFY was gone from all the regions that should make flowers."That suggested to the team that an unknown factor may be activating the LEAFY gene specifically in the flowering portion of plants via the same site through which TFL1 acts. So, they looked to FT, because of its importance in promoting flowering. By experimentally augmenting FT expression, they found that FT, also binding to the FD transcription factor, was required to act upon LEAFY to promote flower formation.Seeing that FT and TFL1 both required FD in order to act, they sought to confirm how this competition played in plants. "We wanted to really test its biological contribution: What does it mean to the plant to lose this?" Wagner says.They saw that under conditions that would normally induce the plant to flower, plants that still had normal FT failed to. "It was a strong phenotype and made it very clear to us that FT and TFL1 compete for this FD factor binding site," she says.TFL1 and FT are mobile and easily convert from one to another. Wagner is very interested in learning more about their mechanism of action to understand how plants can "tune" their growth to best adapt to their environment. In future work, she and her lab plan to continue working out the details, including how both proteins respond to cues from the environment like day length or shade under a tree. She also wants to know how they can control such different processes like flower, bulb, or tendril formation.And Wagner believes the findings have ample potential for applications in plant breeding and agriculture. "You can imagine locally adapting crops, maybe for a high-altitude site in the Himalayas or maybe a location in the far north where days are short," she says. "These elements could even play a role in rational solutions for climate change, breeding plants that are specifically adapted for new conditions."The study was supported by the National Science Foundation (grants 1557529 and 1905062).
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008142154.htm
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Quality control mechanism closes the protein production 'on-ramps'
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Recent work led by Carnegie's Kamena Kostova revealed a new quality control system in the protein production assembly line with possible implications for understanding neurogenerative disease.
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The DNA that comprises the chromosomes housed in each cell's nucleus encodes the recipes for how to make proteins, which are responsible for the majority of the physiological actions that sustain life. Individual recipes are transcribed using messenger RNA, which carries this piece of code to a piece of cellular machinery called the ribosome. The ribosome translates the message into amino acids -- the building blocks of proteins.But sometimes messages get garbled. The resulting incomplete protein products can be toxic to cells. So how do cells clean up in the aftermath of a botched translation?Some quality assurance mechanisms were already known -- including systems that degrade the half-finished protein product and the messenger RNA that led to its creation. But Kostova led a team that identified a new tool in the cell's kit for preventing damage when protein assembly goes awry. Their work was published by Using CRISPR-Cas9-based genetic screening, the researchers discovered a separate, and much needed, device by which the cell prevents that particular faulty message from being translated again. They found two factors, called GIGYF2 and 4EHP, which prevent translation from being initiated on problematic messenger RNA fragments."Imagine that the protein assembly process is a highway and the ribosomes are cars traveling on it," Kostova explained. "If there's a bad message producing incomplete protein products, it's like having a stalled car or two on the road, clogging traffic. Think of GIGYF2 and 4EHP as closing the on-ramp, so that there is time to clear everything away and additional cars don't get stalled, exacerbating the problem."Loss of GIGYF2 has previously been associated with neurodegenerative and neurodevelopmental problems. It is possible that these issues are caused by the buildup of defective proteins that occurs without the ability to prevent translation on faulty messenger RNAs.
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008142112.htm
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Moles: Intersexual and genetically doped
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Moles are special creatures that roam in an extreme habitat. As mammals that burrow deep into the earth, they have forepaws with an extra finger and exceptionally strong muscles. What's more, female moles are intersexual while retaining their fertility. Typical for mammals, they are equipped with two X chromosomes, but they simultaneously develop functional ovarian and testicular tissues. In female moles, both tissue types are united in one organ, the ovotestis -- something that is unique among mammals.
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The testicular tissue of the female mole does not produce sperm, but large amounts of the sex hormone testosterone, meaning the females have similarly high levels as the males. Presumably this natural "doping" makes the female moles aggressive and muscular, an advantage for life underground, where they have to dig burrows and fight for resources.In a study in the journal The study was conducted by an international team co-led by Professor Stefan Mundlos, Research Group leader at the Max Planck Institute for Molecular Genetics (MPIMG) and Director at the Institute for Medical Genetics and Human Genetics at Charité -- Universitätsmedizin Berlin and by Dr. Darío Lupiáñez, Research Group Leader at the Berlin Institute for Medical Systems Biology (BIMSB), which is part of the Max Delbrück Center for Molecular Medicine in the Helmholtz Association (MDC)."Since Darwin, it has been generally accepted that the different appearances of living organisms are the result of gradual changes in genetic makeup that have been passed onto subsequent generations," says Mundlos. "But how are DNA changes and their manifestations in the appearance of an organism related in concrete terms, and how can we uncover such changes?"To pursue this question, the researchers have completely sequenced the genome of the Iberian mole (Talpa occidentalis) for the first time. Moreover, they examined the three-dimensional structure of the genome within the cell. In the nucleus, genes and their associated control sequences form regulatory domains -- relatively isolated "neighborhoods" consisting of large regions where DNA sections interact frequently with each other."We hypothesized that in moles, there are not only changes in the genes themselves, but particularly in the regulatory regions belonging to these genes," says Mundlos.In the course of the moles' evolution, then not only would individual letters of the DNA have changed, also larger pieces of the genome would have shifted, says the researcher. If segments of DNA move from one location to another, completely new or reorganized regulatory domains can emerge and thus activate new genes and enhance or attenuate their expression."The sexual development of mammals is complex, although we have a reasonably good idea on how this process takes place," says Darío Lupiáñez. "At a certain point, sexual development usually progresses in one direction or another, male or female. We wanted to know how evolution modulates this sequence of developmental events, enabling the intersexual features that we see in moles."In fact, when comparing the genome to that of other animals and humans, the team discovered an inversion -- i.e., an inverted genomic segment -- in a region known to be involved in testicular development. The inversion causes additional DNA segments to get included in the regulatory domain of the gene FGF9, which reorganizes the control and regulation of the gene. "This change is associated with the development of testicular tissue in addition to ovarian tissue in female moles," explains Dr. Francisca Martinez Real, lead author of the study and scientist at the MPIMG as well as the Institute for Medical Genetics and Human Genetics at Charité.The team also discovered a triplication of a genomic region responsible for the production of male sex hormones (androgens), more specifically the androgen production gene CYP17A1. "The triplication appends additional regulatory sequences to the gene -- which ultimately leads to an increased production of male sex hormones in the ovotestes of female moles, especially more testosterone," says Real.The highly territorial moles cannot be kept in the laboratory, which particularly challenged the work of the researchers. "We had to do all our research on wild moles," says Lupiáñez. He and Real spent months in southern Spain collecting samples for their experiments. "However, this drawback also became a strength in our study. Our results are not limited to laboratory animals, but extend our knowledge to wild animals."The research group proved that the two genome mutations actually contribute to the special sexuality of female moles by creating a mouse model in which they mimic the genomic changes observed in moles. Of the altered animals, the female mice had androgen levels that were as high as in normal male mice. They were also significantly stronger than their unaltered conspecifics.With moles, the sexes are not that clearly delimited from one another; instead, females move on a spectrum between typically female and typically male phenotypes, i.e., they are intersexual."Our findings are a good example of how important the three-dimensional organization of the genome is for evolution," says Lupiáñez. "Nature makes use of the existing toolbox of developmental genes and merely rearranges them to create a characteristic such as intersexuality. In the process, other organ systems and development are not affected.""Historically, the term intersexuality has caused considerable controversy," says Mundlos. "There was and continues to be a tendency to characterize intersexual phenotypes as pathological conditions. Our study highlights the complexity of sexual development and how this process can result in a wide range of intermediate manifestations that are a representation of natural variation."
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008142104.htm
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HIV up close: Unprecedented view of virus reveals essential steps for causing AIDS
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Accomplishing a feat that had been a pipe dream for decades, scientists have recreated in a test tube the first steps of infection by HIV (human immunodeficiency virus), the virus that causes AIDS (acquired immunodeficiency syndrome). Doing so has provided up-close access to the virus -- which is otherwise obstructed from view deep within the cell -- and enabled identification of essential components that HIV needs to replicate within its human host.
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Specifically, the scientists were able to monitor the virus as it replicated its genome and inserted it into target DNA, mirroring steps that ordinarily take place within the host. Published in "This is teaching us how HIV infects," says Wesley I. Sundquist, Ph.D., distinguished professor of biochemistry at University of Utah Health. He is co-senior author of the study with his former trainee, Owen Pornillos, Ph.D., now an associate professor at the University of Virginia. The co-first authors are Devin Christensen, Ph.D., and Barbie Ganser-Pornillos, Ph.D. "We are learning new things about one of the most significant pathogens that humans have ever encountered, and that is important."For all its danger, HIV is deceptively simple in appearance. The virus resembles a rounded ice cream cone, where an outer shell encapsulates the virus' genetic material inside. Previously, it had been thought that the main role of the shell, called the capsid, was to protect its precious cargo. But the investigations by Sundquist and Pornillos' team show that the capsid also plays an active role in infection.Carrying out initial steps of infection in a test tube allowed the research team to precisely manipulate HIV in ways that had not been possible before. They found that when they used genetic and biochemical methods to destabilize the capsid, HIV could not effectively replicate its genetic material. It was the first direct demonstration that, rather than serving merely as packaging, the capsid is an essential component of the HIV infection process itself.If seeing is believing, then watching the HIV molecule in action gave credence to the experimental finding. Recent advances in cryo-electron microscopy and molecular modeling have made it possible to see the virus -- which, at 130nm, is about 60 times smaller than a red blood cell -- in exquisite detail. Using these techniques, the team visualized each of the 240 tiny protein "tiles" that fit together to make the cone-shaped outer shell. With the up-close view, the scientists could literally see that the capsid remained largely intact throughout the replication process, called reverse transcription."This is different than in the textbooks," Sundquist says. "Our data indicate that the viral capsid plays an active and indispensable role in supporting efficient reverse transcription."Sundquist says that the discovery may help explain why an investigative HIV drug developed by Gilead, the first to target the capsid, is a potent inhibitor of the virus. Previous work by Sundquist, Pornillos, and others elucidating the structure and function of the HIV capsid informed the design of the drug, which has performed well in phase 1 clinical trials. Additional insights gained through the test tube system could improve drug design even further.Advances in microscope technology, coupled with dogged persistence, led to the new view of HIV, which was first discovered as the cause of AIDS more than 35 years ago. It took years of trial and error to determine the minimum components required for recapitulating the process in a test tube, outside the cell. Now that the simplified system is up and running, Pornillos says, it opens doors to learning more fundamental truths about a familiar foe."For me, there is both the fundamental knowledge aspect of it, but also the translational aspect that could help us come up with better ways to stop HIV," Pornillos says. "That's why it's great research."
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008142046.htm
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Setting a TRAP for pandemic-causing viruses
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A research team led by Nagoya University scientists in Japan has developed an approach that can quickly find synthetic proteins that specifically bind to important targets, such as components of the SARS-CoV-2 virus. The method was published in the journal
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"We developed a laboratory technique for rapid selection of synthetic proteins that strongly bind to SARS-CoV-2," says Nagoya University biomolecular engineer Hiroshi Murakami. "High-affinity synthetic proteins can be used to develop sensitive antigen tests for SARS-CoV-2 and for future use as neutralization antibodies in infected patients."Murakami and his colleagues had previously developed a protein selection lab test called TRAP display, which stands for 'transcription-translation coupled with association of puromycin linker.' Their approach skips two time-consuming steps in another commonly used technique for searching through synthetic protein libraries. But their investigations indicated there was a problem with the puromycin linker.In the current study, the team improved their technique by modifying the puromycin linker. Ultimately, they were able to use their TRAP display to identify nine synthetic proteins that bind to the spike protein on SARS-CoV-2's outer membrane. The approach took only four days compared to the weeks it would take using the commonly used messenger RNA display technology.TRAP display involves using a large number of DNA templates that code for and synthesize trillions of proteins carrying random peptide sequences. The synthetic proteins are linked to DNA with the help of the modified puromycin linker and then exposed to a target protein. When the whole sample is washed, only the synthetic proteins that bind to the target remain. These are then placed back into the TRAP display for further rounds until only a small number of very specific target-binding synthetic proteins are left.The researchers investigated the nine synthetic proteins that were found to bind to SARS-CoV-2. Some were specifically able to detect SARS-CoV-2 in nasal swabs from COVID-19 patients, indicating they could be used in test kits. One also attaches to the virus to prevent it from binding to the receptors it uses to gain access to human cells. This suggests this protein could be used as a treatment strategy."Our high-speed, improved TRAP display could be useful for implementing rapid responses to subspecies of SARS-CoV-2 and to other potential new viruses causing future pandemics," says Murakami.
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008142036.htm
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Understanding the progress of viral infections
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A team of researchers at the Institute for the Genetics of Heart Diseases at Münster University created a viral expression model that can be used to simulate and analyse a large number of viral infections -- including the one with SARS-CoV-2. The results are published in the journal
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It is only 120 millionths of a millimetre in size but can bring entire countries to a standstill: the coronavirus. Even if it were to disappear one day, viral infections will still be among the most frequent and difficult-to-treat diseases in humans. Even decades of research have only produced a few standardized vaccines and strategies for treatment to combat just a small number of viruses. Nor has there been much research into viral mechanisms of action -- which was a reason for Prof. Guiscard Seebohm and his team at the Institute for the Genetics of Heart Diseases of Münster University to focus their attention on precisely this topic. And the team has now succeeded in making a groundbreaking development: it has created a viral expression model which can be used to simulate and analyse a large number of viral infections -- including the one with SARS-CoV-2. The results can be read in the current issue of One virus that is much less well-known than SARS-CoV-2, but which can be transmitted in the same way, is the Coxsackie virus B3 (CVB3). "Its symptoms," explains Guiscard Seebohm, "are mostly similar to those for flu, as is the recovery time: after two or three weeks, any patients with a CVB3 infection are, as a rule, well again, and don't have any obvious long-term impairment." But -- not always, says Guiscard Seebohm, who heads the Cellular Electrophysiology and Molecular Biology department. Apart from any acute infections, he explains, a viral infection also contains the risk of a chronic infection, with the consequence of continuous damage to certain internal organs -- which can lead to death. This means that months or even years later, an inflammation of the heart muscle, or type 1 diabetes, can occur in some patients who had CVB3 in the past. Histological examinations of patients showed, in some cases, serious damage to the tissue structure. Also: even years after the acute infection occurred, tissue analyses prove the existence of a virus in the genes.So far there has been an insufficient amount of study devoted to the question of how a CVB3 infection becomes chronic and how exactly an acute infection progresses. In this respect, the Guiscard Seebohm team has succeeded in taking a great step forward. It developed an expression model for CVB3, based on stem cells, in order to get to the bottom of the mechanisms of action in this virus -- as a prototype for the effects of viruses in general. In a study, the model was tested for its controllability in heart muscle cells cultured from stem cells. In the process, the team of researchers was able to achieve a stable integration of the genetic information from a non-infectious variant of CVB3 into the genetic material in human stem cells. The latter can be converted into any kind of human tissue in the laboratory, making it possible to precisely investigate viral mechanisms. The CVB3 expression can be specifically activated by means of a chemical signal.Guiscard Seebohm is delighted at this success, because, as he says, "As a result of this unique human viral expression system based on stem cells, it will now be possible to simulate a large number of diseases in their progression and, for the first time, analyse them with the utmost precision." In Guiscard Seebohm's view there is something else that is just as important: the system is completely controllable. The team of researchers managed to steer the CVB3 expression in the expression model time-wise, both in stem cells and in differentiated heart muscle cells. At the same time, the researchers were able to vary the quantity of viral proteins produced as well as their localization. In other words, the extent of the viral infection, the infection pattern and the time progression can all be adapted to whatever topics researchers are working on.The production of the first fully controllable viral expression model in human cells, its proven functionality, and the transferability to patients all open up numerous new approaches for research. It is not only that any infection with CVB3 and other viruses such as coronavirus and influenza can be examined with a very high resolution; this new method also means that the borders of what can be researched can be extended. Follow-up studies on the controlled expression of CVB3 in hiPSC are already underway and showing promising results. Last but not least, Dr. Stefan Peischard, the lead author of the study now published, and his colleagues in the Seebohm team hope that their work will lead to significant benefits for patients.
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008124429.htm
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How an egg cell's 'operating manual' sets the stage for fertility
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Recently published work from Carnegie's Allan Spradling and Wanbao Niu revealed in unprecedented detail the genetic instructions immature egg cells go through step by step as they mature into functionality. Their findings improve our understanding of how ovaries maintain a female's fertility.
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The general outline of how immature egg cells are assisted by specific ovarian helper cells starting even before a female is born is well understood. But Spradling and Niu mapped the gene activity of thousands of immature egg cells and helper cells to learn how the stage is set for fertility later in life.Even before birth, "germ" cells assemble a finite number of cell clusters called follicles in a female's ovaries. Follicles consist of an immature egg cell and some "helper" cells, which guide the egg through its maturation process. It is from a follicle that a mature egg cell bursts during ovulation."Follicles are slowly used up during a female's reproductive lifespan and menopause ensues when they run out. Understanding what it takes for follicles to form and develop successfully, helps us learn how damaged genes or adverse environmental factors, including a poor diet, can interfere with fertility," explained Spradling. "By documenting the follicle's genetic operating manual, problems in egg development that might lead to birth defects -- as a result of mutations or due to bad nutrition -- can be better understood and reduced."Spradling and Niu sequenced 52,500 mouse ovarian cells at seven stages of follicle development to determine the relative expression of thousands of genes and to characterize their roles.The study also illuminated how mammalian ovaries produce two distinct types of follicles and Spradling and Niu were able to identify many differences in gene activity between them.The first, called wave 1 follicles, are present in the ovary even before puberty. In mice, they generate the first fertile eggs; their function in humans is poorly understood, but they may produce useful hormones. The second type, called wave 2 follicles, are stored in a resting state but small groups are activated to mature during a female's hormonal cycle, ending in ovulation. The findings help clarify each type's roles.Spradling and Niu's work and all its underlying data were published by Proceedings of the National Academy of Sciences."We hope our work will serve as a genetic resource for all researchers who study reproduction and fertility," concluded Spradling.This work was funded by the Howard Hughes Medical Institute.
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008121304.htm
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Taking sides: Factors that influence patterns in protein distribution
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In plants, many proteins are found at only one end of a cell, giving them a polarity like heads and tails on a coin.
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Often, cells next to each other have these proteins at the same end, like a stack of coins with heads all facing up. This protein patterning is critical for how plant cells orient and coordinate themselves to produce the leaves, flowers, stems and roots that adorn our gardens and provide us with all our food and the oxygen we breathe.Previously it's been unclear how this head-to-tail protein patterning is produced: can it arise within each cell, or does it depend on a collective effort of many cells working together?A new paper, published in The team, from the John Innes Centre, studied a protein called BASL that is normally found at only one end of the cells giving rise to leaf pores. By tagging the BASL protein with fluorescence and introducing it into cultured plant cells they could see where the protein went.They showed that even if the cells were stripped of their walls, to create membrane-enclosed spheres, the BASL protein went to only one end of the cell, forming a cap. Time-lapse movies showed that position of BASL labelling changed over time, like a polar ice cap wandering over the earth's surface. However, when cells reformed their walls, the BASL cap could become fixed, and cells elongated into sausage shapes, with the cap remaining at one of the rounded ends.Lead author Dr Jordi Chan says, "It was incredibly exciting to see polarity in isolated plant cells for the first time. It was like seeing a boring-looking planet suddenly light up to reveal a cap, and then elongating while keeping the cap at one end."The results show that cell polarity can arise within cells and likely orients their growth. Signalling between cells may then coordinate polarity, aligning the heads and tails of different cells in a tissue, guiding how they grow collectively and develop into a plant.
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Biotechnology
| 2,020 |
October 8, 2020
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https://www.sciencedaily.com/releases/2020/10/201008104234.htm
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Stem cell sheets harvested in just two days
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Stem cells are cell factories that constantly divide themselves to create new cells. Implanting stem cells in damaged organs can regenerate new tissues. Cell sheet engineering, which allows stem cells to be transplanted into damaged areas in the form of sheets made up of only cells, completely eliminates immune rejection caused by external substances and encourages tissue regeneration. A research team led by POSTECH recently succeeded in drastically reducing the harvest period of such stem cell sheets.
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A joint research team comprised of Professor Dong Sung Kim and researcher Andrew Choi of POSTECH's Department of Mechanical Engineering and Dr. InHyeok Rhyou and Dr. Ji-Ho Lee of the Department of Orthopedic Surgery at Pohang Semyung Christianity Hospital has significantly reduced the total harvest period of a stem cell sheet to two days. The nanotopography of poly(N-isopropylacrylamide) (PNIPAAm), which abruptly changes its roughness depending on temperature, allows harvesting of cell sheets that consist of mesenchymal stem cells derived from human bone marrow. Considering that it takes one week on average to make stem cells into sheets using the existing techniques developed so far, this is the shortest harvest time on record. These research findings were published as a cover paper in the latest issue of Professor Kim's research team focused on PNIPAAm, a polymer that either combines with water or averts it depending on the temperature. In previous studies, PNIPAAm has been introduced as a coating material for cell culture platform to harvest cell sheets, but the range of utilization had been hampered due to the limited types of cells that can be made into sheets. For the first time in 2019, the research team developed a technology of easily regulating the roughness of 3D bulk PNIPAAm and has stably produced various types of cells into sheets.The study conducted this time focused on making stem cells -- that are effective in tissue regeneration -- into sheets in a short time in order to increase their direct utility. The team achieved this by applying an isotropic pattern of nanopores measuring 400 nanometers (nm, 1 billionth of a meter) on the surface of a 3D bulk PNIPAAm. As a result, not only did the formation and maturity of human bone marrow-derived mesenchymal stem cells on the nanotopography of bulk PNIPAAm accelerate, but the surface roughness of bulk PNIPAAm at room temperature below the lower critical solution temperature (LCST) was also rapidly increased, effectively inducing the detachment of cell sheets. This in turn enabled the rapid harvesting of human bone marrow-derived mesenchymal stem cell sheets."At least five days are needed to harvest stem cell sheets reported through previous researches," commented Andrew Choi, the " author of the paper. "We can now harvest them in just two days with the PNIPAAm nanotopography developed this time.""We have significantly shortened the harvest time by introducing nanotopography on the surface of the 3D bulk PNIPAAm to produce mature stem cell sheets for the first time in the world," remarked Professor Dong Sung Kim who led the study. He added, "We have opened up the possibility of applying the sheets directly to patients in the future."The research was conducted with the support from Basic Research Program (Mid-career Researcher Program) and the Biomedical Technology Development Program of the National Research Foundation and the Ministry of Science and ICT of Korea.
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Biotechnology
| 2,020 |
October 7, 2020
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https://www.sciencedaily.com/releases/2020/10/201007145424.htm
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Researchers develop tools to sharpen 3D view of large RNA molecules
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University of Maryland scientists have developed a method to determine the structures of large RNA molecules at high resolution. The method overcomes a challenge that has limited 3D analysis and imaging of RNA to only small molecules and pieces of RNA for the past 50 years.
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The new method, which expands the scope of nuclear magnetic resonance (NMR) spectroscopy, will enable researchers to understand the shape and structure of RNA molecules and learn how they interact with other molecules. The insights provided by this technology could lead to targeted RNA therapeutic treatments for disease. The research paper on this work was published in the journal "The field of nuclear magnetic resonance spectroscopy has been stuck looking at things that are small, say 35 RNA building blocks or nucleotides. But most of the interesting things that are biologically and medically relevant are much bigger, 100 nucleotides or more," said Kwaku Dayie, a professor of chemistry and biochemistry at UMD and senior author of the paper. "So, being able to break down the log jam and look at things that are big is very exciting. It will allow us to peek into these molecules and see what is going on in a way we haven't been able to do before."In NMR spectroscopy, scientists direct radio waves at a molecule, exciting the atoms and "lighting up" the molecule. By measuring changes in the magnetic field around the excited atoms -- the nuclear magnetic resonance -- scientists can reconstruct characteristics such as the shape, structure and motion of the molecule. The data this produces can then be used to generate images, much like MRI images seen in medicine.Ordinarily, NMR signals from the many atoms in a biological molecule such as RNA overlap with each other, making analysis very difficult. However, in the 1970s, scientists learned to biochemically engineer RNA molecules to work better with NMR by replacing the hydrogen atoms with magnetically active fluorine atoms. In relatively small molecules of RNA consisting of 35 or fewer nucleotides, the fluorine atoms light up readily when hit with radio waves and remain excited long enough for high-resolution analysis. But as RNA molecules get larger, the fluorine atoms light up only briefly, then quickly lose their signal. This has prevented high-resolution 3D analysis of larger RNA molecules.Previous work by others had shown that fluorine continued to produce a strong signal when it was next to a carbon atom containing six protons and seven neutrons (C-13). So, Dayie and his team developed a relatively easy method to change the naturally occurring C-12 in RNA (which has 6 protons and 6 neutrons) to C-13 and install a fluorine atom (F-19) directly next to it.Dayie and his team first demonstrated that their method could produce data and images equal to current methods by applying it to pieces of RNA from HIV containing 30 nucleotides, which had been previously imaged. They then applied their method to pieces of Hepatitis B RNA containing 61 nucleotides -- nearly double the size of previous NMR spectroscopy possible for RNA.Their method enabled the researchers to identify sites on the hepatitis B RNA where small molecules bind and interact with the RNA. That could be useful for understanding the effect of potential therapeutic drugs. The next step for the researchers is to analyze even larger RNA molecules."This work allows us to expand what can be brought into focus," Dayie said. "Our calculations tell us that, in theory, we can look at really big things, like a part of the ribosome, which is the molecular machine that synthesizes proteins inside cells."By understanding the shape and structure of a molecule, scientists can better understand its function and how it interacts with its environment. What's more, this technology will enable scientists to see the 3D structure as it changes, because RNA molecules in particular change shape frequently. This knowledge is key to developing therapeutics that narrowly target disease-specific molecules without affecting healthy cell functions."The hope is that if researchers know the nooks and crannies in a molecule that is dysfunctional, then they can design drugs that fill the nooks and crannies to take it out of commission," Dayie said. "And if we can follow these molecules as they change shape and structure, then their response to potential drugs will be a little bit more predictable, and designing drugs that are effective can be more efficient."
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Biotechnology
| 2,020 |
October 7, 2020
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https://www.sciencedaily.com/releases/2020/10/201007123119.htm
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Fighting intestinal infections with the body's own endocannabinoids
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Endocannabinoids, signaling molecules produced in the body that share features with chemicals found in marijuana, can shut down genes needed for some pathogenic intestinal bacteria to colonize, multiply, and cause disease, new research led by UT Southwestern scientists shows.
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The findings, published online today in Discovered in 1992, endocannabinoids are lipid-based neurotransmitters that play a variety of roles in the body, including regulating immunity, appetite, and mood. Cannabis and its derivatives have long been used to relieve chronic gastrointestinal conditions, including irritable bowel syndrome and inflammatory bowel disease. Studies have shown that dysregulation of the body's endocannabinoid system can lead to intestinal inflammation and affect the makeup of gut microbiota, the population of different bacterial species that inhabit the digestive tract.However, study leader Vanessa Sperandio, Ph.D., professor of microbiology and biochemistry at UTSW, says it's been unknown whether endocannabinoids affect susceptibility to pathogenic gastrointestinal infections.To help answer this question, Sperandio and her colleagues worked with mice genetically altered to overproduce the potent mammalian endocannabinoid 2-arachidonoyl glycerol (2-AG) in various organs, including the intestines. When the researchers infected these animals and their unmodified littermates with Citrobacter rodentium, a bacterial pathogen that attacks the colon and causes marked inflammation and diarrhea, the mutant mice developed only mild symptoms compared with the more extreme gastrointestinal distress exhibited by their littermates. Examination of the mutant animals' colons showed far lower inflammation and signs of infection. These mice also had significantly lower fecal loads of C. rodentium bacteria and cleared their infection days faster than their unmodified littermates. Treating genetically unmodified animals with a drug that raised levels of 2-AG in the intestines produced similar positive effects.Sperandio's team found that increased levels of 2-AG could also attenuate Salmonella typhimurium infections in mice and impede enterohemorrhagic Escherichia coli -- a particularly dangerous gastrointestinal bacteria that infects humans -- in order to express the virulence traits needed for a successful infection.Conversely, when the researchers treated mammalian cells in petri dishes with tetrahydrolipstatin, a Food and Drug Administration-approved compound sold commercially as Alli that inhibits 2-AG production, they became more susceptible to the bacterial pathogens.Further experiments showed that 2-AG exerted these effects on C. rodentium, S. typhimurium, and E. coli by blocking a bacterial receptor known as QseC. When this receptor senses the host signaling molecules epinephrine and norepinephrine, it triggers a molecular cascade necessary to establish infection. Plugging this receptor with 2-AG prevents this virulence program from activating, Sperandio explains, helping to protect against infection.Sperandio notes that these findings could help explain some of the effects of cannabis use on inflammatory bowel conditions. Although studies have shown that cannabis can lower inflammation, recent research has shown that these conditions also tend to have a bacterial component that might be positively affected by plant cannabinoids.In addition, cannabis compounds or synthetic derivatives could eventually help patients kick intestinal bacterial infections without antibiotics. This could be particularly useful for infections caused by enterohemorrhagic Escherichia coli, Sperandio says, which produces a deadly toxin when it's treated with antibiotics, rendering these drugs not only counterproductive but extremely dangerous. Because many virulent bacteria that colonize areas elsewhere in the body also have the QseC receptor, she adds, this strategy could be used more broadly to fight a variety of infections."By harnessing the power of natural compounds produced in the body and in plants," she says, "we may eventually treat infections in a whole new way."
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Biotechnology
| 2,020 |
October 7, 2020
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https://www.sciencedaily.com/releases/2020/10/201007085615.htm
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Deep learning takes on synthetic biology
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DNA and RNA have been compared to "instruction manuals" containing the information needed for living "machines" to operate. But while electronic machines like computers and robots are designed from the ground up to serve a specific purpose, biological organisms are governed by a much messier, more complex set of functions that lack the predictability of binary code. Inventing new solutions to biological problems requires teasing apart seemingly intractable variables -- a task that is daunting to even the most intrepid human brains.
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Two teams of scientists from the Wyss Institute at Harvard University and the Massachusetts Institute of Technology have devised pathways around this roadblock by going beyond human brains; they developed a set of machine learning algorithms that can analyze reams of RNA-based "toehold" sequences and predict which ones will be most effective at sensing and responding to a desired target sequence. As reported in two papers published concurrently today in "These achievements are exciting because they mark the starting point of our ability to ask better questions about the fundamental principles of RNA folding, which we need to know in order to achieve meaningful discoveries and build useful biological technologies," said Luis Soenksen, Ph.D., a Postdoctoral Fellow at the Wyss Institute and Venture Builder at MIT's Jameel Clinic who is a co-first author of the first of the two papers.The collaboration between data scientists from the Wyss Institute's Predictive BioAnalytics Initiative and synthetic biologists in Wyss Core Faculty member Jim Collins' lab at MIT was created to apply the computational power of machine learning, neural networks, and other algorithmic architectures to complex problems in biology that have so far defied resolution. As a proving ground for their approach, the two teams focused on a specific class of engineered RNA molecules: toehold switches, which are folded into a hairpin-like shape in their "off" state. When a complementary RNA strand binds to a "trigger" sequence trailing from one end of the hairpin, the toehold switch unfolds into its "on" state and exposes sequences that were previously hidden within the hairpin, allowing ribosomes to bind to and translate a downstream gene into protein molecules. This precise control over the expression of genes in response to the presence of a given molecule makes toehold switches very powerful components for sensing substances in the environment, detecting disease, and other purposes.However, many toehold switches do not work very well when tested experimentally, even though they have been engineered to produce a desired output in response to a given input based on known RNA folding rules. Recognizing this problem, the teams decided to use machine learning to analyze a large volume of toehold switch sequences and use insights from that analysis to more accurately predict which toeholds reliably perform their intended tasks, which would allow researchers to quickly identify high-quality toeholds for various experiments.The first hurdle they faced was that there was no dataset of toehold switch sequences large enough for deep learning techniques to analyze effectively. The authors took it upon themselves to generate a dataset that would be useful to train such models. "We designed and synthesized a massive library of toehold switches, nearly 100,000 in total, by systematically sampling short trigger regions along the entire genomes of 23 viruses and 906 human transcription factors," said Alex Garruss, a Harvard graduate student working at the Wyss Institute who is a co-first author of the first paper. "The unprecedented scale of this dataset enables the use of advanced machine learning techniques for identifying and understanding useful switches for immediate downstream applications and future design."Armed with enough data, the teams first employed tools traditionally used for analyzing synthetic RNA molecules to see if they could accurately predict the behavior of toehold switches now that there were manifold more examples available. However, none of the methods they tried -- including mechanistic modeling based on thermodynamics and physical features -- were able to predict with sufficient accuracy which toeholds functioned better.The researchers then explored various machine learning techniques to see if they could create models with better predictive abilities. The authors of the first paper decided to analyze toehold switches not as sequences of bases, but rather as two-dimensional "images" of base-pair possibilities. "We know the baseline rules for how an RNA molecule's base pairs bond with each other, but molecules are wiggly -- they never have a single perfect shape, but rather a probability of different shapes they could be in," said Nicolaas Angenent-Mari, a MIT graduate student working at the Wyss Institute and co-first author of the first paper. "Computer vision algorithms have become very good at analyzing images, so we created a picture-like representation of all the possible folding states of each toehold switch, and trained a machine learning algorithm on those pictures so it could recognize the subtle patterns indicating whether a given picture would be a good or a bad toehold."Another benefit of their visually-based approach is that the team was able to "see" which parts of a toehold switch sequence the algorithm "paid attention" to the most when determining whether a given sequence was "good" or "bad." They named this interpretation approach Visualizing Secondary Structure Saliency Maps, or VIS4Map, and applied it to their entire toehold switch dataset. VIS4Map successfully identified physical elements of the toehold switches that influenced their performance, and allowed the researchers to conclude that toeholds with more potentially competing internal structures were "leakier" and thus of lower quality than those with fewer such structures, providing insight into RNA folding mechanisms that had not been discovered using traditional analysis techniques."Being able to understand and explain why certain tools work or don't work has been a secondary goal within the artificial intelligence community for some time, but interpretability needs to be at the forefront of our concerns when studying biology because the underlying reasons for those systems' behaviors often cannot be intuited," said Jim Collins, Ph.D., the senior author of the first paper. "Meaningful discoveries and disruptions are the result of deep understanding of how nature works, and this project demonstrates that machine learning, when properly designed and applied, can greatly enhance our ability to gain important insights about biological systems." Collins is also the Termeer Professor of Medical Engineering and Science at MIT.While the first team analyzed toehold switch sequences as 2D images to predict their quality, the second team created two different deep learning architectures that approached the challenge using orthogonal techniques. They then went beyond predicting toehold quality and used their models to optimize and redesign poorly performing toehold switches for different purposes, which they report in the second paper.The first model, based on a convolutional neural network (CNN) and multi-layer perceptron (MLP), treats toehold sequences as 1D images, or lines of nucleotide bases, and identifies patterns of bases and potential interactions between those bases to predict good and bad toeholds. The team used this model to create an optimization method called STORM (Sequence-based Toehold Optimization and Redesign Model), which allows for complete redesign of a toehold sequence from the ground up. This "blank slate" tool is optimal for generating novel toehold switches to perform a specific function as part of a synthetic genetic circuit, enabling the creation of complex biological tools."The really cool part about STORM and the model underlying it is that after seeding it with input data from the first paper, we were able to fine-tune the model with only 168 samples and use the improved model to optimize toehold switches. That calls into question the prevailing assumption that you need to generate massive datasets every time you want to apply a machine learning algorithm to a new problem, and suggests that deep learning is potentially more applicable for synthetic biologists than we thought," said co-first author Jackie Valeri, a graduate student at MIT and the Wyss Institute.The second model is based on natural language processing (NLP), and treats each toehold sequence as a "phrase" consisting of patterns of "words," eventually learning how certain words are put together to make a coherent phrase. "I like to think of each toehold switch as a haiku poem: like a haiku, it's a very specific arrangement of phrases within its parent language -- in this case, RNA. We are essentially training this model to learn how to write a good haiku by feeding it lots and lots of examples," said co-first author Pradeep Ramesh, Ph.D., a Visiting Postdoctoral Fellow at the Wyss Institute and Machine Learning Scientist at Sherlock Biosciences.Ramesh and his co-authors integrated this NLP-based model with the CNN-based model to create NuSpeak (Nucleic Acid Speech), an optimization approach that allowed them to redesign the last 9 nucleotides of a given toehold switch while keeping the remaining 21 nucleotides intact. This technique allows for the creation of toeholds that are designed to detect the presence of specific pathogenic RNA sequences, and could be used to develop new diagnostic tests.The team experimentally validated both of these platforms by optimizing toehold switches designed to sense fragments from the SARS-CoV-2 viral genome. NuSpeak improved the sensors' performances by an average of 160%, while STORM created better versions of four "bad" SARS-CoV-2 viral RNA sensors whose performances improved by up to 28 times."A real benefit of the STORM and NuSpeak platforms is that they enable you to rapidly design and optimize synthetic biology components, as we showed with the development of toehold sensors for a COVID-19 diagnostic," said co-first author Katie Collins, an undergraduate MIT student at the Wyss Institute who worked with MIT Associate Professor Timothy Lu, M.D., Ph.D., a corresponding author of the second paper."The data-driven approaches enabled by machine learning open the door to really valuable synergies between computer science and synthetic biology, and we're just beginning to scratch the surface," said Diogo Camacho, Ph.D., a corresponding author of the second paper who is a Senior Bioinformatics Scientist and co-lead of the Predictive BioAnalytics Initiative at the Wyss Institute. "Perhaps the most important aspect of the tools we developed in these papers is that they are generalizable to other types of RNA-based sequences such as inducible promoters and naturally occurring riboswitches, and therefore can be applied to a wide range of problems and opportunities in biotechnology and medicine."Additional authors of the papers include Wyss Core Faculty member and Professor of Genetics at HMS George Church, Ph.D.; and Wyss and MIT Graduate Students Miguel Alcantar and Bianca Lepe."Artificial intelligence is wave that is just beginning to impact science and industry, and has incredible potential for helping to solve intractable problems. The breakthroughs described in these studies demonstrate the power of melding computation with synthetic biology at the bench to develop new and more powerful bioinspired technologies, in addition to leading to new insights into fundamental mechanisms of biological control," said Don Ingber, M.D., Ph.D., the Wyss Institute's Founding Director. Ingber is also the Judah Folkman Professor of Vascular Biology at Harvard Medical School and the Vascular Biology Program at Boston Children's Hospital, as well as Professor of Bioengineering at Harvard's John A. Paulson School of Engineering and Applied Sciences.This work was supported by the DARPA Synergistic Discovery and Design program, the Defense Threat Reduction Agency, the Paul G. Allen Frontiers Group, the Wyss Institute for Biologically Inspired Engineering, Harvard University, the Institute for Medical Engineering and Science, the Massachusetts Institute of Technology, the National Science Foundation, the National Human Genome Research Institute, the Department of Energy, the National Institutes of Health, and a CONACyT grant.
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Biotechnology
| 2,020 |
October 7, 2020
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https://www.sciencedaily.com/releases/2020/10/201007083443.htm
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Nobel Prize in Chemistry 2020: CRISPR/Cas9 method for genome editing
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The Royal Swedish Academy of Sciences has decided to award the Nobel Prize in Chemistry 2020 to Emmanuelle Charpentier, Max Planck Unit for the Science of Pathogens, Berlin, Germany, and Jennifer A. Doudna, University of California, Berkeley, USA "for the development of a method for genome editing."
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Emmanuelle Charpentier and Jennifer A. Doudna have discovered one of gene technology's sharpest tools: the CRISPR/Cas9 genetic scissors. Using these, researchers can change the DNA of animals, plants and microorganisms with extremely high precision. This technology has had a revolutionary impact on the life sciences, is contributing to new cancer therapies and may make the dream of curing inherited diseases come true.Researchers need to modify genes in cells if they are to find out about life's inner workings. This used to be time-consuming, difficult and sometimes impossible work. Using the CRISPR/Cas9 genetic scissors, it is now possible to change the code of life over the course of a few weeks."There is enormous power in this genetic tool, which affects us all. It has not only revolutionised basic science, but also resulted in innovative crops and will lead to ground-breaking new medical treatments," says Claes Gustafsson, chair of the Nobel Committee for Chemistry.As so often in science, the discovery of these genetic scissors was unexpected. During Emmanuelle Charpentier's studies of Charpentier published her discovery in 2011. The same year, she initiated a collaboration with Jennifer Doudna, an experienced biochemist with vast knowledge of RNA. Together, they succeeded in recreating the bacteria's genetic scissors in a test tube and simplifying the scissors' molecular components so they were easier to use.In an epoch-making experiment, they then reprogrammed the genetic scissors. In their natural form, the scissors recognise DNA from viruses, but Charpentier and Doudna proved that they could be controlled so that they can cut any DNA molecule at a predetermined site. Where the DNA is cut it is then easy to rewrite the code of life.Since Charpentier and Doudna discovered the CRISPR/Cas9 genetic scissors in 2012 their use has exploded. This tool has contributed to many important discoveries in basic research, and plant researchers have been able to develop crops that withstand mould, pests and drought. In medicine, clinical trials of new cancer therapies are underway, and the dream of being able to cure inherited diseases is about to come true. These genetic scissors have taken the life sciences into a new epoch and, in many ways, are bringing the greatest benefit to humankind.
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Biotechnology
| 2,020 |
October 6, 2020
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https://www.sciencedaily.com/releases/2020/10/201006114305.htm
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Watch how cells squeeze through channels
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Observations of cells moving through small channels shed new light on cell migration in 3D environments, researchers report October 6 in
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"Our results describe how cells can migrate and deform through confined spaces, providing potentially new ways to envision cell motility in small blood capillaries in vivo," says senior study author Daniel Riveline of the University of Strasbourg in France.Cell migration plays a key role in a variety of biological phenomena, ranging from early development to disease processes. But cell motility has mainly been studied on flat surfaces rather than in 3D environments similar to blood vessels and other structures commonly found in the body. To address this gap, Riveline and his collaborators studied cell motion in microfabricated channels that had either open or closed configurations (i.e., confined by three or four walls, respectively). In addition, some channels were straight, whereas others had various bottlenecks to mimic cell blockage in small veins.As expected, fibroblasts moved freely in straight channels. But in the presence of bottlenecks, the nucleus sometimes prevented cell passage, causing pauses in cell motion. Other times, the cells anchored and pulled locally to deform the nucleus and allow cell passage. Additional results suggested that cells would not be able to change their direction of motion when entering a sufficiently small capillary, and that chemical gradients can induce directional motion.The researchers also studied the movements of oral squamous epithelial cells, including some with mutant keratin protein implicated in squamous cancers. In normal cells, keratin accumulated at the rear of the nucleus during passage through bottlenecks, potentially to facilitate deformation of the organelle. By contrast, the mutant cells could not pass through bottlenecks, indicating that defects in keratin impair motion in confined spaces, possibly by preventing the nucleus from deforming. The findings also suggest that squamous cancer cells could be blocked within small capillaries, potentially allowing them to penetrate tissues."Because initial arrest in the capillary is critical for tumor cells to metastasize to secondary sites in distant organs, blockage by mutant keratin may provide advantages for tumor seeding, survival, and proliferation," Riveline says. "Future studies could take this channel strategy to identify signaling networks that are modified in the context of cancer."
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Biotechnology
| 2,020 |
October 6, 2020
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https://www.sciencedaily.com/releases/2020/10/201006114300.htm
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New techniques probe vital and elusive proteins
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The number of proteins in the human body, collectively known as the proteome, is vast. Somewhere between 80,000 and 400,000 proteins circulate in our cells, tissues and organs, carrying out a broad range of duties essential for life. When proteins go awry, they are responsible for a myriad of serious diseases.
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Now, researchers at the Biodesign Center for Applied Structural Discovery and ASU's School of Molecular Sciences, along with their colleagues, investigate a critically important class of proteins, which adorn the outer membranes of cells. Such membrane proteins often act as receptors for binding molecules, initiating signals that can alter cell behavior in a variety of ways.A new approach to acquiring structural data of membrane proteins in startling detail is described in the new study. Cryogenic electron microscopy (or cryo-EM) methods, a groundbreaking suite of tools, is used. Further, use of so-called LCP crystallization and Microcrystal electron diffraction (MicroED) help unveil structural details of proteins that have been largely inaccessible through conventional approaches like X-ray crystallography.The findings describe the first use of LCP-embedded microcrystals to reveal high-resolution protein structural details using MicroED. The new research graces the cover of the current issue of the Cell Press journal "LCP was a great success in membrane protein crystallization, according to Wei Liu, a corresponding author of the new study. "The new extensive application of LCP-MicroED offers promise for improved approaches for structural determination from challenging protein targets. These structural blueprints can be used to facilitate new therapeutic drug design from more precise insights."One class of membrane proteins of particular interest are the G-protein-coupled receptors (GPCRs), which form the largest and most varied group of membrane receptors found in eukaryotic organisms, including humans.The physiological activities of GPCRs are so important that they are a major target for a wide range of therapeutic drugs. This is where problems arise however, as determining the detailed structure of membrane proteins -- an essential precursor to accurate drug design -- often poses enormous challenges.The technique of X-ray crystallography has been used to investigate the atomic-scale structures and even dynamic behavior of many proteins. Here, crystallized samples of the protein under study are struck with an X-ray beam, causing diffraction patterns, which appear on a screen. Assembling thousands of diffraction snapshots allows a high-resolution 3D structural image to be assembled with the aid of computers.Yet many membrane proteins, including GPCRs, don't form large, well-ordered crystals appropriate for X-ray crystallography. Further, such proteins are delicate and easily damaged by X-radiation. Getting around the problem has required the use of special devices known as X-ray free electron lasers or XFELS, which can deliver a brilliant burst of X-ray light lasting mere femtoseconds, (a femtosecond is equal to one quadrillionth of a second or about the time it takes a light ray to traverse the diamere of a virus). The technique of serial femtosecond X-ray crystallography allows researchers to obtain a refraction image before the crystalized sample is destroyed.Nevertheless, crystallization of many membrane proteins remains an extremely difficult and imprecise art and only a handful of these gargantuan XFEL machines exist in the world.Enter cryogenic electron microscopy and MicroED. This ground-breaking technique involves flash-freezing protein crystals in a thin veneer of ice, then subjecting them to a beam of electrons. As in the case of X-ray crystallography, the method uses diffraction patterns, this time from electrons rather than X-rays, to assemble final detailed structures.MicroED excels in collecting data from crystals too small and irregular to be used for conventional X-ray crystallography. In the new study, researchers used two advanced techniques in tandem in order to produce high-resolution diffraction images of two important model proteins: Proteinase K and the A2A adenosine receptor, whose functions include modulation of neurotransmitters in the brain, cardiac vasodilation and T-cell immune response.The proteins were embedded in a special type of crystal known as a lipidic cubic phase or LCP crystal, which mimics the native environment such proteins naturally occur in. The LCP samples were then subjected to electron microscopy, using the MicroED method, which permits the imaging of extremely thin, sub-micron-sized crystals. Further, continuous rotation of LCP crystals under the electron microscope allows multiple diffraction patterns to be acquired from a single crystal with an extremely low, damage-free electron dose.The ability to examine proteins that can only form micro- or nanocrystals opens the door to the structural determination of many vitally important membrane proteins that have eluded conventional means of investigation, particularly GPCRs.
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Biotechnology
| 2,020 |
October 6, 2020
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https://www.sciencedaily.com/releases/2020/10/201006114241.htm
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RTL1 gene a likely culprit behind temple and Kagami-Ogata syndromes
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Temple and Kagami-Ogata syndromes are serious genetic conditions that can lead to a variety of health problems with neonatal lethality, and in the case of Temple syndrome, severe growth problems occur. However, the genetic mechanisms of these illnesses are not well understood. But now, researchers from Tokyo Medical and Dental University (TMDU) have identified a gene that appears to be responsible for symptoms of both conditions, with important implications for human evolutionary genetics.
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In a study published last month in Development the research team has revealed that deficiency and overproduction of Retrotransposon Gag like 1 (Rtl1), which is a mouse ortholog of the human RTL1 gene, is significantly associated with muscle symptoms in models of Temple and Kagami-Ogata syndromes, respectively.Temple and Kagami-Ogata syndromes are characterized by unique postnatal muscle-related symptoms and prenatal placental problems. Although Rtl1 has previously been found to be responsible for prenatal placental malformations in mouse models of these conditions, the causative gene for the muscle-related symptoms has not been identified. The researchers at Tokyo Medical and Dental University (TMDU) aimed to address this in their recent study."Although Rtl1 is essential for maintaining placental fetal capillaries," says lead author of the study, Moe Kitazawa, "Little is known about the role of Rtl1 in other forms of muscular abnormalities."The researchers extensively examined the role of Rtl1 in fetal muscle development at both the cellular and tissue level. They used two previously generated mouse models, one with a loss of Rtl1 to model Temple syndrome, and one with an overproduction of Rtl1 to model Kagami-Ogata syndrome."Our data clearly demonstrate that RTL1 is of critical physiological significance," explains Fumitoshi Ishino, senior author. "In the mouse models of both Temple and Kagami-Ogata syndrome, we detected severe but distinct abnormalities in the neonatal muscles that are necessary for respiration, such as the intercostal, abdominal, and diaphragm muscles."Particularly, the overproduction of Rtl1 in the mouse model of Kagami-Ogata syndrome led to respiratory problems and muscle damage, resulting in neonatal death and the loss of Rtl1 was a major cause of abnormal muscle tone, such as hypotonia, and likely to be associated with feeding difficulty/poor sucking function in Temple syndrome patients.Thus, RTL1 appears to be the major gene responsible for muscle and placental defects in these two genetic conditions."This is the first demonstration that a form of RTL1 specific to eutherians (placental mammals) plays an important role in fetal and neonatal muscle development," says Kitazawa."This work demonstrates that LTR retrotransposons, which are a specific type of mobile DNA from which RTL1 originated, have influenced a variety of eutherian-specific traits, including the skeletal muscles, placenta, and the brain. Thus, these findings have important implications for understanding the acquisition of genes throughout the process of eutherian evolution," say senior authors Kaneko-Ishino and Ishino.
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Biotechnology
| 2,020 |
October 5, 2020
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https://www.sciencedaily.com/releases/2020/10/201005170839.htm
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How malaria parasites withstand a fever's heat
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Even when a person suffering from malaria is burning up with fever and too sick to function, the tiny blood-eating parasites lurking inside them continue to flourish, relentlessly growing and multiplying as they gobble up the host's red blood cells.
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The single-celled Plasmodium parasites that cause 200 million cases of malaria each year can withstand feverish temperatures that make their human hosts miserable. And now, a Duke University-led team is beginning to understand how they do it.Assistant professor of chemistry Emily Derbyshire and colleagues have identified a lipid-protein combo that springs into action to gird the parasite's innards against heat shock.Understanding how the malaria parasite protects its cells against heat stress and other onslaughts could lead to new ways to fight resistant strains, which have evolved ways to survive the drugs traditionally used to kill them, the researchers say.Nearly half of the world's population is at risk of contracting malaria. The disease kills 400,000 people a year, most of them children.Long before the cause of malaria was identified, the disease's harrowing fevers were well known. References to them have been found on 5,000-year-old clay tablets from ancient Mesopotamia. The Greek poet Homer wrote about their misery. Hippocrates too.The Duke team, collaborating with professor of biological engineering Jacquin Niles at the Massachusetts Institute of Technology, wanted to know how the malaria parasites inside a person's body make it through these fevers unscathed.When the parasites enter a person's bloodstream through the bite of an infected mosquito, the temperature around them jumps from the balmy mid-70s of the mosquito to 98.6 degrees in the human. The human host's body temperature can then rocket to 105 degrees or higher before dropping back down to normal two to six hours later, a roller coaster pattern that repeats itself every two to three days."It's like going from room temperature water to a hot tub," said first author Kuan-Yi Lu, who earned his Ph.D. in molecular genetics and microbiology in Derbyshire's lab at Duke.For the paper, published Sept. 25 in the journal To mimic malarial fever in the lab, the researchers placed malaria-infected red blood cells in an incubator heated to 104 degrees Fahrenheit for six hours before bringing them back down to normal body temperature, 98.6 degrees.They found that when temperatures rise, the parasites produce more of a lipid molecule called phosphatidylinositol 3-phosphate, or PI(3)P.This substance builds up in the outer wall of a tiny sac inside the parasite's cells called the food vacuole -- the protist's version of a gut. There, it recruits and binds to another molecule, a heat shock protein called Hsp70, and together they help shore up the food vacuole's outer walls.Without this lipid-protein boost, the team found that heat can make the food vacuole start to leak, unleashing its acidic contents into the gel-like fluid that fills the cell and possibly even digesting the parasite from the inside.The findings are important because they could help researchers make the most of existing malaria drugs.Previous research has shown that malaria parasites with higher-than-normal PI(3)P levels are more resistant to artemisinins, the leading class of antimalarials. Since artemisinins were first introduced in the 1970s, partial resistance has been increasingly reported in parts of Southeast Asia, raising fears that we may be losing one of our best weapons against the disease.But the Duke-led study raises the possibility that new combination therapies for malaria -- artemisinins combined with other drugs that reduce the parasite's PI(3)P lipid levels and disrupt the food vacuole's membrane -- could be a way to re-sensitize resistant parasites, breaking down their defenses so the malaria treatments we already have are effective again."If there is an alternative way to increase the permeability of the digestive vacuole, it could make the digestive vacuole more accessible to those drugs again," Lu said.The findings also suggest caution in giving malaria patients ibuprofen for fever if they're already taking artemisinin-based compounds, Derbyshire said. That's because artemisinins kill malaria parasites by damaging their cell's survival machinery, including the machinery that makes PI(3)P. If artemisinins suppress PI(3)P levels, and thereby make malaria parasites more vulnerable to heat stress, then fever reducers could prolong the time it takes for artemisinin-based drugs to kill the parasites, as some reports have suggested.Much remains to be learned, Derbyshire said. "There's more work to do to establish the mode of action. But you could imagine designing new combination therapies to try and extend the life of artemisinin and prolong its effectiveness," Derbyshire said.This work was supported by the National Institutes of Health (DP2AI138239) and the Bill & Melinda Gates Foundation (OPP1132312, OPP1162467).
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Biotechnology
| 2,020 |
October 5, 2020
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https://www.sciencedaily.com/releases/2020/10/201005101532.htm
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Cyanobacteria as 'green' catalysts in biotechnology
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Cyanobacteria, despite staining water green through their special pigments, are colloquially known as "blue-green algae," and convert light energy into chemical energy particularly effectively thanks to their highly active photosynthetic cells. This makes them attractive for biotechnological application, where they could be used as environmentally friendly and readily available biocatalysts for the production of new chemicals using specifically introduced enzymes.
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What sounds good in theory, is still facing obstacles in the practical large-scale technological implementation. A decisive limiting factor is currently the availability of light, as Robert Kourist from the Institute of Molecular Biotechnology at Graz University of Technology explains: "When cyanobacteria are densely grown, i.e. in high concentrations, only the cells located on the outside receive enough light. Inside it's pretty dark. This means that the amount of catalyst cannot be increased at will. After a cell density of a few grams per litre, the photosynthetic activity and thus the productivity of the cells decreases sharply. This is of course a considerable disadvantage for large-scale biotechnological production."By comparison, previously established biocatalysts such as yeasts can be used with cell densities of 50 grams per litre and more. The established production organisms have the major disadvantage that they depend on agricultural products as a basis for growth and thus consume many resources. "Algae-based catalysts can be grown from water and CO2, so they are 'green' in a two-fold sense. For this reason, intensive efforts are under way to increase the catalytic performance of cyanobacteria," said Kourist.Together with Ruhr University Bochum and the Finnish University of Turku, the algae working group at TU Graz has now succeeded in increasing precisely this catalytic performance by specifically redirecting the photosynthetic electron flow to the desired catalytic function. "For the first time, we were able to measure the supply of photosynthetic energy directly in the cells in a time-resolved manner so that we were able to identify bottlenecks in the metabolism," explains Marc Nowaczyk from the Chair of Plant Biochemistry at the Ruhr University Bochum."We have switched off a system in the genome of the cyanobacterium that is supposed to protect the cell from fluctuating light. This system is not necessary under controlled cultivation conditions, but consumes photosynthetic energy. Energy that we prefer to put into the target reaction," explains Hanna Büchsenschütz, doctoral student at TU Graz and first author of the study. In this way, the problem of low productivity of cyanobacteria due to high cell densities can be solved. "To put it another way, we can only use a certain amount of cells. That's why we have to make the cells go faster. We have developed a method using so-called metabolic engineering that makes cyanobacteria a great deal more mature for biotechnological application," said Kourist.In addition to increasing the productivity of the cell itself through targeted interventions at the gene level, the Graz researchers are also working on new concepts for the algae cultivation process. One approach is to introduce light sources directly into the cell suspension, for example via mini LEDs. New geometries are also being experimented with. Thus, cyanobacteria in the form of encapsulated small spheres, so-called "beads," can absorb more light overall. Robert Kourist comments: "It is very important to develop all measures on the way to large-scale industrial application of algae-based biocatalysts in an integrated way. This is only possible with interdisciplinary research that looks at the function of an enzyme in the same way as we look at engineering in the photosynthetic cell."
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Biotechnology
| 2,020 |
October 5, 2020
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https://www.sciencedaily.com/releases/2020/10/201005005927.htm
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A tale of two cesspits: DNA reveals intestinal health in Medieval Europe and Middle East
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A new study published this week demonstrates a first attempt at using the methods of ancient bacterial detection, pioneered in studies of past epidemics, to characterize the microbial diversity of ancient gut contents from two medieval latrines. The findings provide insights into the microbiomes of pre-industrial agricultural populations, which may provide much-needed context for interpreting the health of modern microbiomes.
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Over the years, scientists have noted that those living in industrialized societies have a notably different microbiome compared to hunter-gatherer communities around the world. From this, a growing body of evidence has linked changes in our microbiome to many of the diseases of the modern industrialized world, such as inflammatory bowel disease, allergies, and obesity. The current study helps to characterize the change in gut microbiomes and highlights the value of ancient latrines as sources of bio-molecular information.Piers Mitchell of Cambridge University specializes in the gut contents of past people through analysis of unusual substrates. By looking at the contents of archaeological latrines and desiccated faeces under the microscope, he and his team have learned volumes about the intestinal parasites that plagued our ancestors."Microscopic analysis can show the eggs of parasitic worms that lived in the intestines, but many microbes in the gut are simply too small to see," comments Mitchell. "If we are to determine what constitutes a healthy microbiome for modern people, we should start looking at the microbiomes of our ancestors who lived before antibiotic use, fast food, and the other trappings of industrialization."Kirsten Bos, a specialist in ancient bacterial DNA from the Max Planck Institute for the Science of Human History and co-leader the study, was first skeptical about the feasibility of investigating the contents of latrines that had long been out of order."At the outset we weren't sure if molecular signatures of gut contents would survive in the latrines over hundreds of years. Many of our successes in ancient bacterial retrieval thus far have come from calcified tissues like bones and dental calculus, which offer very different preservation conditions. Nevertheless," says Bos, "I was really hoping the data here would change my perspective."The team analyzed sediment from medieval latrines in Jerusalem and Riga, Latvia dating from the 14th-15th century CE. The first challenge was distinguishing bacteria that once formed the ancient gut from those that were introduced by the environment, an unavoidable consequence of working with archaeological material.The researchers identified a wide range of bacteria, archaea, protozoa, parasitic worms, fungi and other organisms, including many taxa known to inhabit the intestines of modern humans."It seems latrines are indeed valuable sources for both microscopic and molecular information," concludes Bos.Susanna Sabin, a doctoral alumna of the MPI-SHH who co-led the study, compared the latrine DNA to those from other sources, including microbiomes from industrial and foraging populations, as well as waste water and soil."We found that the microbiome at Jerusalem and Riga had some common characteristics -- they did show similarity to modern hunter gatherer microbiomes and modern industrial microbiomes, but were different enough that they formed their own unique group. We don't know of a modern source that harbors the microbial content we see here."The use of latrines, where the faeces of many people are mixed together, allowed the researchers unprecedented insight into the microbiomes of entire communities."These latrines gave us much more representative information about the wider pre-industrial population of these regions than an individual faecal sample would have," explains Mitchell. "Combining evidence from light microscopy and ancient DNA analysis allows us to identify the amazing variety of organisms present in the intestines of our ancestors who lived centuries ago."Despite the promise of this new approach for investigating the microbiome, challenges remain."We'll need many more studies at other archaeological sites and time periods to fully understand how the microbiome changed in human groups over time," says Bos. "However, we have taken a key step in showing that DNA recovery of ancient intestinal contents from past latrines can work."
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Biotechnology
| 2,020 |
October 2, 2020
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https://www.sciencedaily.com/releases/2020/10/201002141910.htm
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Biomedical sciences researchers find new way to prevent and cure rotavirus, other viral infections
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A combination of two substances secreted by the immune system can cure and prevent rotavirus infection, as well as potentially treat other viral infections that target epithelial cells, which cover body surfaces such as skin, blood vessels, organs and the urinary tract, according to researchers in the Institute for Biomedical Sciences at Georgia State University.
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Rotavirus, which causes severe, life-threatening diarrhea in young children and moderate gastrointestinal distress in adults, leads to thousands of deaths in children annually, particularly in developing countries where rotavirus vaccines are only moderately effective. Rotavirus is an RNA virus that primarily infects intestinal epithelial cells.The substances identified in the study, officially known as cytokines, are interleukin 18 (IL-18) and interleukin 22 (IL-22). IL-18 and IL-22 are produced when the body detects a protein in the whip-like appendage of bacteria.The study, which investigated how these cytokines inhibit rotavirus infection, found when mice were treated with both IL-18 and IL-22, the cytokines promoted each other's expression, but also impeded rotavirus by independent, distinct mechanisms that involved activating receptors in intestinal epithelial cells. These actions resulted in rapid and complete expulsion of rotavirus, even in hosts with severely compromised immune systems. The therapy was also found to be effective for norovirus, a contagious virus that causes vomiting and diarrhea. The findings are published in the journal "Our study reports a novel means of eradicating a viral infection, particularly viruses that infect epithelial cells," said Dr. Andrew Gewirtz, senior author of the study and a professor in the Institute for Biomedical Sciences at Georgia State. "The results suggest that a cocktail that combines IL-18 and IL-22 could be a means of treating viral infections that target short-lived epithelial cells with high turnover rates."The study is funded by grants from the National Institutes of Health.
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Biotechnology
| 2,020 |
October 2, 2020
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https://www.sciencedaily.com/releases/2020/10/201002111727.htm
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Cell perturbation system could have medical applications
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Cell lines injected with free nucleic acid are widely used for drug discovery and disease modeling. To avoid genetically mixed cell populations, investigators use dilution techniques to select single cells that will then generate identical lines. However, the route of limiting dilutions is tedious and time consuming.
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A new study by Northwestern researchers shows how Nanofountain Probe Electroporation (NFP-E), a tool that delivers molecules into single-cells, could solve that issue, and could lead to new applications for drug screening and designing patient-specific courses of treatment.The team, led by Northwestern Engineering's Horacio Espinosa and including Joshua Leonard, demonstrates the versatility of NFP-E -- which introduces DNA or RNA into cells using electricity. It can also deliver both proteins and plasmids in a variety of animal and human cell types with dosage control. The team included John Kessler, the Ken and Ruth Davee Professor of Stem Cell Biology and professor of neurology and pharmacology at the Northwestern University Feinberg School of Medicine.The new method can be used to study disease or for cell therapy. In the former, the genome is manipulated. In the latter, gene-editing occurs in cells such as T-cells to treat cancer with immunotherapies.By employing single-cell electroporation, the process of introducing DNA or RNA into single cells using a pulse of electricity, which briefly open pores in the cell membrane, their work shows how NFP-E achieves fine control over the relative expression of two co-transfected plasmids. Moreover, by pairing single-cell electroporation with time-lapse fluorescent imaging, their investigation reveals characteristic times for electro-pore closure."We demonstrated the potential of the NFP-E technology in manipulating a variety of cell types with stoichiometric control of molecular cargo that can be used for conducting a wide range of studies in drug screening, cell therapies, and synthetic biology," said Espinosa, James N. and Nancy J. Farley Professor in Manufacturing and Entrepreneurship and professor of mechanical engineering and (by courtesy) biomedical engineering and civil and environmental engineering.Currently, biomolecules can be delivered into cells in numerous ways: viral vectors; chemical carriers, such as cell-penetrating peptides and polymer nano-capsules; lipofectamine, and bulk electroporation."There exist a number of strategies for delivering biomolecules into cells, but each has its limitations," said Leonard, associate professor of chemical and biological engineering and Charles Deering McCormick Professor of Teaching Excellence. "For instance, chemical carriers confer relatively slow delivery and can be toxic to the cell; viral vectors are often efficient but can induce adverse immune responses and insertional genotoxicity. Use of any traditional method often requires substantial effort to optimize the protocol depending on the cell type and molecule to be delivered, and, therefore, a readily generalizable biomolecule delivery strategy would offer some meaningful advantages."The new NFP-E system enables single-cell delivery of DNA, RNA, and proteins into different immortalized cell lines as well as primary cells with more than 95 percent efficiency and more than 90 percent cell viability."The results indicate that the cell membrane resealing time scales non-linearly with the pulse voltage and the number of electroporation pulses, reaching a maximum at intermediate values," Espinosa said. "That means long pulsing times or high voltages appear not to be necessary for efficient molecular transport across cell membranes. That feature is important in obtaining high transport efficiency while keeping cell toxicity to a minimum."Using single-cell electroporation technology, the researchers were able to understand transport mechanisms involved in localized electroporation-based cell sampling. One obstacle to nondestructive temporal single-cell sampling is the small amounts of cytosol -- the fluid inside cells -- that are extracted, which makes it challenging to test or detect RNA sequences or proteins.Research showed that the scaling of membrane resealing time is a function of various electroporation parameters, providing insight into post-pulse electro-pore dynamics."The work addresses the need to understand ways to increase the cytosol-sampled amount, without adversely affecting cells," Espinosa said. "That can guide the research community in designing experiments aimed at electroporation-based sampling of intracellular molecules for temporal cell analysis."This research is related to
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Biotechnology
| 2,020 |
October 2, 2020
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https://www.sciencedaily.com/releases/2020/10/201002105756.htm
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Hidden DNA fragment the 'trigger switch' for male development
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Biology textbooks may need to be re-written, with scientists finding a new piece of DNA essential to forming male sex organs in mice.
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An international research collaboration with The University of Queensland found the Y-chromosome gene that makes mice male is made up of two different DNA parts, not one, as scientists had previously assumed.UQ's Institute of Molecular Biosciences Emeritus Professor Peter Koopman said the critical DNA fragment had been hidden from researchers for more than 30 years."Expression of the Y chromosomal gene Sry is required for male development in mammals and since its discovery in 1990 has been considered a one-piece gene," he said."Sry turns out to have a cryptic second part, which nobody suspected was there, that is essential for determining the sex of male mice. We have called the two-piece gene Sry-T."The scientists tested their theory and found that male mice (XY) lacking in Sry-T developed as female, while female mice (XX) carrying a Sry-T transgene developed as male.The success rate for the experiments was almost 100 per cent.Emeritus Professor Koopman said the discovery would change how basic biology and evolution was taught around the world."For the last 30 years, we've been trying to figure out how this works," he said."Sry is a master switch gene because it flicks the switch for male development, it gets the ball rolling for a whole series of genetic events that result in a baby being born as a male instead of female."This new piece of the gene is absolutely essential for its function; without that piece, the gene simply doesn't work."We've discovered something massively important in biology here, because without Sry there can be no sexual reproduction and hence no propagation and survival of mammalian species."The discovery may apply to efforts to manipulate sex ratios in agriculture or for biological pest management. But Emeritus Professor Koopman was quick to point out that, for ethical and practical reasons, the discovery cannot be utilised on human embryos."Once we understand better how males and females are specified in non-human species of mammals, then it offers the opportunity to influence that process," he said."The ability to select for the desired sex could dramatically increase efficiencies for agricultural industries such as the dairy industry (females) or the beef industry (males)."People have been trying to figure out ways to skew to the desired sex in these industries for a long time, and now that we understand more about the fundamental mechanism of Sry it may be possible through genetic means."
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Biotechnology
| 2,020 |
October 2, 2020
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https://www.sciencedaily.com/releases/2020/10/201002091057.htm
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Searching for the chemistry of life
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In the search for the chemical origins of life, researchers have found a possible alternative path for the emergence of the characteristic DNA pattern: According to the experiments, the characteristic DNA base pairs can form by dry heating, without water or other solvents. The team led by Ivan Halasz from the Rudjer Boskovic Institute and Ernest Mestrovic from the pharmaceutical company Xellia presents its observations from DESY's X-ray source PETRA III in the journal
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"One of the most intriguing questions in the search for the origin of life is how the chemical selection occurred and how the first biomolecules formed," says Tomislav Stolar from the Rudjer Boskovic Institute in Zagreb, the first author on the paper. While living cells control the production of biomolecules with their sophisticated machinery, the first molecular and supramolecular building blocks of life were likely created by pure chemistry and without enzyme catalysis. For their study, the scientists investigated the formation of nucleobase pairs that act as molecular recognition units in the Deoxyribonucleic Acid (DNA).Our genetic code is stored in the DNA as a specific sequence spelled by the nucleobases adenine (A), cytosine (C), guanine (G) and thymine (T). The code is arranged in two long, complementary strands wound in a double-helix structure. In the strands, each nucleobase pairs with a complementary partner in the other strand: adenine with thymine and cytosine with guanine."Only specific pairing combinations occur in the DNA, but when nucleobases are isolated they do not like to bind to each other at all. So why did nature choose these base pairs?" says Stolar. Investigations of pairing of nucleobases surged after the discovery of the DNA double helix structure by James Watson and Francis Crick in 1953. However, it was quite surprising that there has been little success in achieving specific nucleobase pairing in conditions that could be considered as prebiotically plausible."We have explored a different path," reports co-author Martin Etter from DESY. "We have tried to find out whether the base pairs can be generated by mechanical energy or simply by heating." To this end, the team studied methylated nucleobases. Having a methyl group (-CH3) attached to the respective nucleobases in principle allows them to form hydrogen bonds at the Watson-Crick side of the molecule. Methylated nucleobases occur naturally in many living organisms where they fulfil a variety of biological functions.In the lab, the scientists tried to produce nucleobase pairs by grinding. Powders of two nucleobases were loaded into a milling jar along with steel balls, which served as the grinding media, while the jars were shaken in a controlled manner. The experiment produced A:T pairs which had also been observed by other scientists before. Grinding however, could not achieve formation of G:C pairs.In a second step, the researchers heated the ground cytosine and guanine powders. "At about 200 degrees Celsius, we could indeed observe the formation of cytosine-guanine pairs," reports Stolar. In order to test whether the bases only form the known pairs under thermal conditions, the team repeated the experiments with mixtures of three and four nucleobases at the P02.1 measuring station of DESY's X-ray source PETRA III. Here, the detailed crystal structure of the mixtures could be monitored during heating and formation of new phases could be observed."At about 100 degrees Celsius, we were able to observe the formation of the adenine-thymine pairs, and at about 200 degrees Celsius the formation of Watson-Crick pairs of guanine and cytosine," says Etter, head of the measuring station. "Any other base pair did not form even when heated further until melting." This proves that the thermal reaction of nucleobase pairing has the same selectivity as in the DNA."Our results show a possible alternative route as to how the molecular recognition patterns that we observe in the DNA could have been formed," adds Stolar. "The conditions of the experiment are plausible for the young Earth that was a hot, seething cauldron with volcanoes, earthquakes, meteorite impacts and all sorts of other events. Our results open up many new paths in the search for the chemical origins of life." The team plans to investigate this route further with follow-up experiments at P02.1.DESY is one of the world's leading particle accelerator centres and investigates the structure and function of matter -- from the interaction of tiny elementary particles and the behaviour of novel nanomaterials and vital biomolecules to the great mysteries of the universe. The particle accelerators and detectors that DESY develops and builds at its locations in Hamburg and Zeuthen are unique research tools. They generate the most intense X-ray radiation in the world, accelerate particles to record energies and open up new windows onto the universe. DESY is a member of the Helmholtz Association, Germany's largest scientific association, and receives its funding from the German Federal Ministry of Education and Research (BMBF) (90 per cent) and the German federal states of Hamburg and Brandenburg (10 per cent).
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Biotechnology
| 2,020 |
October 2, 2020
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https://www.sciencedaily.com/releases/2020/10/201002091035.htm
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Laundry lint can cause significant tissue damage within marine mussels
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Microscopic fibres created during the laundry cycle can cause damage to the gills, liver and DNA of marine species, according to new research.
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Scientists at the University of Plymouth exposed the Mediterranean mussel (They demonstrated that increasing the amount of lint resulted in significant abnormality within the mussels' gills, specifically leading to damage of tissues including deformity, extensive swelling and loss of cilia. In the liver, the presence of lint led to atrophy or deformities leading to loss of definition in digestive tubules.The increasing concentration of fibres also led to a reduction in the mussels' ability to filter food particles from the seawater and a significant increase in DNA strand breaks in the blood cells.Scientists say the precise causes of the effects are not wholly clear, but are likely to arise from the fibres themselves and chemicals present within them.They say the findings are unlikely to solely apply to lint, as its properties are consistent with other textiles and fibres found commonly in waste water and throughout the marine environment.The study, published in the journal Dr Andrew Turner, Associate Professor of Environmental Sciences, was the study's senior author and has previously conducted research highlighting the chemicals -- including bromine, iron and zinc -- found within lint.He said: "The laundering of clothes and other textiles is among the most significant sources of synthetic microfibers within the environment. However, despite their known presence in a range of species, there have been very few studies looking in detail at their impact. This study shows for the first time what harm they can cause, and it is particularly interesting to consider that it is not just the fibres themselves which create issues but also the cocktail of more harmful chemicals which they can mobilise."Co-author Awadhesh Jha, Professor in Genetic Toxicology and Ecotoxicology, added: "Mytilus species are commonly used to monitor water quality in coastal areas, and the damage shown to them in this study is a cause for significant concern. Given their genetic similarity to other species and the fact they are found all over the world, we can also assume these effects will be replicated in other shellfish and marine species. Damage to DNA and impairment of the filter feeding abilities would have potential impact on the health of the organisms and the ecosystem. That is particularly significant as we look in the future to increase our reliance on aquaculture as a global source of food."This study is the latest research by the University in the field, with it being awarded a Queen's Anniversary Prize for Higher and Further Education in 2020 for its ground-breaking research and policy impact on microplastics pollution in the oceans.That research has included work showing that washing clothes releases thousands of microplastic particles into environment, and that devices fitted to washing machines can reduce the fibres produced in laundry cycle by up to 80%. Scientists from the University have also showed that wearing clothes could release more microfibres to the environment than washing them.
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Biotechnology
| 2,020 |
October 1, 2020
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https://www.sciencedaily.com/releases/2020/10/201001113702.htm
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Gene expression altered by direction of forces acting on cell
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Tissues and cells in the human body are subjected to a constant push and pull -- strained by other cells, blood pressure and fluid flow, to name a few. The type and direction of the force on a cell alters gene expression by stretching different regions of DNA, researchers at University of Illinois Urbana-Champaign and collaborators in China found in a new study.
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The findings could provide insights into physiology and diseases such as fibrosis, cardiovascular disease and malignant cancer, the researchers said."Force is everywhere in the human body, and both external and internal forces can influence your body far more than you may have thought," said study leader Ning Wang, a professor of mechanical science and engineering at Illinois. "These strains profoundly influence cellular behaviors and physiological functions, which are initiated at the level of gene expression."The effects of physical forces and signals on cells, tissues and organs have been less studied than those of chemical signals and responses, yet physical forces play an important role in how cells function and respond to their environment, Wang said.Most studies seeking to understand the mechanics of cells apply force using a microscope cantilever probe to tap a cell's surface or a focused laser beam to move a tiny particle across the surface. However, these techniques can only move in one dimension. This incomplete picture leaves fundamental questions unanswered, Wang said -- for example, the difference in the responses to shear stress from blood flow and stretching from blood pressure.Wang and his collaborators developed a method that allows them to move a magnetic bead in any direction, giving them a picture of the ways forces act on a cell in 3D. They call it three-dimensional magnetic twisting cytometry.They found that the force from the magnetic bead caused a rapid increase in expression for certain genes, but the amount of the increase depended on the direction the bead moved. When the bead rolled along the long axis of the cell, the increase was the lowest, but when the force was applied perpendicularly -- across the short axis of the cell -- gene activity increased the most. When the bead was moved at a 45-degree angle or rotated in the same plane as the cell to induce shear stress, the response was intermediate."These observations show that gene upregulation and activation are very sensitive to the mode of the applied force, when the magnitude of the force remains unchanged," Wang said.In further experiments, the researchers found that the reason for the difference lies in the method that the forces are relayed to the cell's nucleus, where DNA is housed. Cells have a network of support structures called the cytoskeleton, and the main force-bearing elements are long fibers of the protein actin. When they bend due to a force, they relay that force to the nucleus and stretch the chromosomes.These actin fibers run lengthwise along the cell. So when the force strains them widthwise, they deform more, stretching the chromosomes more and causing greater gene activity, the researchers found. They published their findings in the journal "A stress fiber is like a tense violin string. When a stress is applied across the short axis of the cell, it's just like when a person plucks a violin string vertically from the string's direction to produce a louder, more forceful sound," Wang said.The researchers' next step will be to create disease models to see how different forces might help explain the mechanism of certain diseases, and to identify possible therapeutic targets or applications."In certain diseases, such as aortic valve calcification, arterial atherosclerosis, liver fibrosis or malignant tumors, these cellular responses and adaptation go awry, causing the tissues and organs to function abnormally," Wang said. "This is the first time that the mechanism of living cells' different biological responses to the direction of forces at the level of genes has been revealed, so perhaps with our three-dimensional approach we can understand these diseases better."
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Biotechnology
| 2,020 |
October 1, 2020
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https://www.sciencedaily.com/releases/2020/10/201001113652.htm
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Repurposed anti-malarial compounds kill diarrheal parasite, study finds
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A class of compounds used for malaria treatment also kill the intestinal parasite Cryptosporidium, a leading cause of diarrheal disease and death in children that has no cure, a multi-institution collaboration of researchers found in a new study.
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The compounds, called bicyclic azetidines, specifically target an enzyme responsible for protein production within the parasite, the authors report in the journal "There's an urgent need because young children are dying of this diarrheal pathogen, and there's no effective medicine to treat the infection nor vaccine to prevent the disease," said the study's lead author, Sumiti Vinayak, a pathobiology professor at the University of Illinois Urbana-Champaign. "Immunocompromised patients and agricultural animals, especially young calves, are also very susceptible to Cryptosporidium. This is the first time we have had validation of a compound working on a specific target in this parasite."The researchers began by performing a broad analytical study of existing drugs, looking for any with the potential for repurposing as a Cryptosporidium treatment. After looking at many classes of anti-microbial compounds, they determined that the anti-malarial bicyclic azetidines was a candidate and tested them against Cryptosporidium.After the compounds proved very effective at killing the parasite in cell cultures, the researchers tested them in immunocompromised mice with Cryptosporidium infections. They found that one oral dose a day for four days rid the mice of infection."This study provides a new way of targeting Cryptosporidium. Significantly, because we are repurposing compounds from an anti-malarial program in development, it allows us to apply insights from that program to the treatment of cryptosporidiosis," said Eamon Comer, who led the study at the Broad Institute in Cambridge, Massachusetts. Professors Boris Striepen of the University of Pennsylvania and Christopher D. Huston of the University of Vermont also co-led the study.The researchers then performed comprehensive biochemical and genetic studies to determine how the compounds attacked the parasite. They found that the bicyclic azetidines targeted an enzyme that makes transfer RNA, the molecule that carries amino acids when the cell makes proteins. The Cryptosporidium enzyme is very similar to that of the parasite that causes malaria, but different from the analogous enzyme in humans, Vinayak said, making it an effective drug target.Using CRISPR-Cas9 gene-editing technology, the researchers changed one letter in the DNA of the Cryptosporidium gene for the target enzyme, making it just different enough that the drug would not attack it. That one change made the parasite resistant to the drugs, further confirming that blocking this enzyme is the mechanism by which the drug kills Cryptosporidium, Vinayak said."This is the first time that the mechanism of action of an anti-Cryptosporidium drug candidate has been confirmed," Vinayak said. "It's a good steppingstone to find these compounds that we can feed into the drug-development pipeline. Future research will further evaluate safety and clinical effectiveness, but the discovery of a new and potent series of compounds with a known target puts us on a promising path forward in this important effort to develop urgently needed treatments."
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Biotechnology
| 2,020 |
September 30, 2020
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https://www.sciencedaily.com/releases/2020/09/200930094754.htm
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'Immortal' in tree resin
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The phenomenon of using DNA from old fossils preserved in amber already inspired Hollywood -- in the film Jurassic Park, scientists reproduce the DNA of dinosaurs extracted from a fossil mosquito embedded in a piece of amber and thereby resurrect them. In reality, however, the undertaking is much more difficult: all previous studies in which researchers took DNA samples from insects enclosed in tree resin were the results of modern environmental contamination and, in addition, were unreproducible, subsequently useless under the scientific method.
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An international team led by researchers at the University of Bonn now detected DNA from ambrosia beetles that were trapped in recent tree resin for less than seven years. The study was published in the journal Using so-called ancient DNA, scientists can draw conclusions about long gone times and the organisms living there. The use of organisms trapped in amber (fossil tree resin) with this finality was thought not to be possible after relatively recent fails in looking for DNA in a few thousand-year-old samples."Our new results show that it is indeed possible to genetically study organisms that were embedded in resin, although we do not know the time limit yet" emphasizes study leader Dr. David Peris of the Institute for Geosciences and Meteorology at the University of Bonn. The superordinate aim of the researchers is to dissolve step by step fundamental aspects of the DNA preservation in the resin and to determine the real temporal border, until when DNA in resins remains preserved."Instead of looking for DNA in amber of 100 million years old or more, to dream about the resurrection of dinosaurs, we should start by detecting it in insects trapped a few years ago in resin," highlights David Peris. The resin samples used were six and two years old and came from Madagascar. To detect the DNA, the scientists established a method based on the technique called polymerase chain reaction, which makes it possible to multiply genetic material in a test tube. This method is well-known in criminology and recently became famous, as the basic technology for the detection of the SARS-CoV-2. "This method allowed us to perform several authenticity checks, so that we could say certain that the detected DNA in our experiments was indeed from the beetles preserved in the resin," explains Kathrin Janssen of the Institute of Medical Microbiology, Immunology and Parasitology, also part of University of Bonn, the second lead author.The researchers found that water is stored in the embedded samples longer than previously thought, which has a negative effect on the stability of the DNA. In the future, the scientists plan to gradually analyze older samples with more sensitive "next generation sequencing" methods. "Investigating the time limit of DNA conservation and many other related issues is the aim of future experiments," summarizes Kathrin Janssen.Besides the University of Bonn, the Universitat de Barcelona, the Instituto Geológico y Minero de España (both Spain), the Senckenberg Research Institute in Frankfurt am Main and the University of Bergen (Norway) were involved in the study.
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Biotechnology
| 2,020 |
September 30, 2020
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https://www.sciencedaily.com/releases/2020/09/200930085151.htm
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Breaking COVID-19's 'clutch' to stop its spread
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Scripps Research chemist Matthew Disney, PhD, and colleagues have created drug-like compounds that, in human cell studies, bind and destroy the pandemic coronavirus' so-called "frameshifting element" to stop the virus from replicating. The frameshifter is a clutch-like device the virus needs to generate new copies of itself after infecting cells.
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"Our concept was to develop lead medicines capable of breaking COVID-19's clutch," Disney says. "It doesn't allow the shifting of gears."Viruses spread by entering cells and then using the cells' protein-building machinery to churn out new infectious copies. Their genetic material must be compact and efficient to make it into the cells.The pandemic coronavirus stays small by having one string of genetic material encode multiple proteins needed to assemble new virus. A clutch-like frameshifting element forces the cells' protein-building engines, called ribosomes, to pause, slip to a different gear, or reading frame, and then restart protein assembly anew, thus producing different protein from the same sequence.But making a medicine able to stop the process is far from simple. The virus that causes COVID-19 encodes its genetic sequence in RNA, chemical cousin of DNA. It has historically been very difficult to bind RNA with orally administered medicines, but Disney's group has been developing and refining tools to do so over more than a decade.The scientists' report, titled "Targeting the SARS-CoV-2 RNA Genome with Small Molecule Binders and Ribonuclease Targeting Chimera (RIBOTAC) Degraders," appears Sept. 30 in the journal Disney emphasizes this is a first step in a long process of refinement and research that lies ahead. Even so, the results demonstrate the feasibility of directly targeting viral RNA with small-molecule drugs, Disney says. Their study suggests other RNA viral diseases may eventually be treated through this strategy, he adds."This is a proof-of-concept study," Disney says. "We put the frameshifting element into cells and showed that our compound binds the element and degrades it. The next step will be to do this with the whole COVID virus, and then optimize the compound."Disney's team collaborated with Iowa State University Assistant Professor Walter Moss, PhD, to analyze and predict the structure of molecules encoded by the viral genome, in search of its vulnerabilities."By coupling our predictive modeling approaches to the tools and technologies developed in the Disney lab, we can rapidly discover druggable elements in RNA," Moss says. "We're using these tools not only to accelerate progress toward treatments for COVID-19, but a host of other diseases, as well."The scientists zeroed in on the virus' frameshifting element, in part, because it features a stable hairpin-shaped segment, one that acts like a joystick to control protein-building. Binding the joystick with a drug-like compound should disable its ability to control frameshifting, they predicted. The virus needs all of its proteins to make complete copies, so disturbing the shifter and distorting even one of the proteins should, in theory, stop the virus altogether.Using a database of RNA-binding chemical entities developed by Disney, they found 26 candidate compounds. Further testing with different variants of the frameshifting structure revealed three candidates that bound them all well, Disney says.Disney's team in Jupiter, Florida quickly set about testing the compounds in human cells carrying COVID-19's frameshifting element. Those tests revealed that one, C5, had the most pronounced effect, in a dose-dependent manner, and did not bind unintended RNA.They then went further, engineering the C5 compound to carry an RNA editing signal that causes the cell to specifically destroy the viral RNA. With the addition of the RNA editor, "these compounds are designed to basically remove the virus," Disney says.Cells need RNA to read DNA and build proteins. Cells have natural process to rid cells of RNA after they are done using them. Disney has chemically harnessed this waste-disposal system to chew up COVID-19 RNA. His system is called RIBOTAC, short for "Ribonuclease Targeting Chimera."Adding a RIBOTAC to the C5 anti-COVID compound increases its potency by tenfold, Disney says. Much more work lies ahead for this to become a medicine that makes it to clinical trials. Because it's a totally new way of attacking a virus, there remains much to learn, he says."We wanted to publish it as soon as possible to show the scientific community that the COVID RNA genome is a druggable target. We have encountered many skeptics who thought one cannot target any RNA with a small molecule," Disney says. "This is another example that we hope puts RNA at the forefront of modern medicinal science as a drug target."
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Biotechnology
| 2,020 |
September 29, 2020
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https://www.sciencedaily.com/releases/2020/09/200929123544.htm
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App analyzes coronavirus genome on a smartphone
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A new mobile app has made it possible to analyse the genome of the SARS-CoV-2 virus on a smartphone in less than half an hour.
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Cutting-edge nanopore devices have enabled scientists to read or 'sequence' the genetic material in a biological sample outside a laboratory, however analysing the raw data has still required access to high-end computing power -- until now.The app Genopo, developed by the Garvan Institute of Medical Research, in collaboration with the University of Peradeniya in Sri Lanka, makes genomics more accessible to remote or under-resourced regions, as well as the hospital bedside."Not everyone has access to the high-power computing resources that are required for DNA and RNA analysis, but most people have access to a smartphone," says co-senior author Dr Ira Deveson, who heads the Genomic Technologies Group at Garvan's Kinghorn Centre for Clinical Genomics."Fast, real-time genomic analysis is more crucial today than ever, as a central method for tracking the spread of coronavirus. Our app makes genomic analysis more accessible, literally placing the technology into the pockets of scientists around the world."The researchers report the app Genopo in the journal Genomic sequencing no longer requires a sophisticated lab setup.At the size of a USB stick, portable devices such as the Oxford Nanopore Technologies MinION sequencer can rapidly generate genomic sequences from a sample in the field or the clinic. The technology has been used for Ebola surveillance in West Africa, to profile microbial communities in the Arctic and determine coronavirus evolution during the current pandemic.However, analysing genome sequencing data requires powerful computation. Scientists need to piece the many strings of genetic letters from the raw data into a single sequence and pinpoint the instances of genetic variation that shed light on how a virus evolves."Until now, genomic analysis has required the processing power of high-end server computers or cloud services. We set out to change that," explains co-senior author Hasindu Gamaarachchi, Genomics Computing Systems Engineer at the Garvan Institute."To enable in situ genomic sequencing and analysis, in real time and without major laboratory infrastructure, we developed an app that could execute bioinformatics workflows on nanopore sequencing datasets that are downloaded to a smartphone. The reengineering process, spearheaded by first author Hiruna Samarakoon, required overcoming a number of technical challenges due to various resource constraints in smartphones. The app Genopo combines a number of available bioinformatics tools into a single Android application, 'miniaturised' to work on the processing power of a consumer Android device."The researchers tested Genopo on the raw sequencing data of virus samples isolated from nine Sydney patients infected with SARS-CoV-2, which involved extracting and amplifying the virus RNA from a swab sample, sequencing the amplified DNA with a MinION device and analysing the data on a smartphone. The researchers tested their app on different Android devices, including models from Nokia, Huawei, LG and Sony.The Genopo app took an average 27 minutes to determine the complete SARS-CoV-2 genome sequence from the raw data, which the researchers say opens the possibility to do genomic analysis at the point of care, in real time. The researchers also showed that Genopo can be used to profile DNA methylation -- a modification which changes gene activity -- in a sample of the human genome."This illustrates a flexible, efficient architecture that is suitable to run many popular bioinformatics tools and accommodate small or large genomes," says Dr Deveson. "We hope this will make genomics much more accessible to researchers to unlock the information in DNA or RNA to the benefit of human health, including in the current pandemic."Genopo is a free, open-source application available through the Google Play store (This project was supported by a Medical Research Future Fund (grant APP1173594), a Cancer Institute NSW Early Career Fellowship and The Kinghorn Foundation. Garvan is affiliated with St Vincent's Hospital Sydney and UNSW Sydney.
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Biotechnology
| 2,020 |
September 29, 2020
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https://www.sciencedaily.com/releases/2020/09/200929123518.htm
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Common antioxidant enzyme may provide potential treatment for COVID-19
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Researchers from UCLA and China have found that catalase, a naturally occurring enzyme, holds potential as a low-cost therapeutic drug to treat COVID-19 symptoms and suppress the replication of coronavirus inside the body. A study detailing the research was published in
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Catalase is produced naturally and used by humans, animals and plants. Inside cells, the antioxidant enzyme kick starts the breakdown of hydrogen peroxide, which can be toxic, into water and oxygen. The enzyme is also commonly used worldwide in food production and as a dietary supplement."There is a lot of focus on vaccines and antiviral drugs, and rightly so," said Yunfeng Lu, a UCLA Samueli School of Engineering professor of chemical and biomolecular engineering and a senior author on the study. "In the meantime, our research suggests this enzyme could offer a very effective therapeutic solution for treatment of hyperinflammation that occurs due to SARS-CoV-2 virus, as well as hyperinflammation generally."Lu's group developed the drug-delivery technology used in the experiments. Three types of tests were conducted, each addressing a different symptom of COVID-19.First, they demonstrated the enzyme's anti-inflammatory effects and its ability to regulate the production of cytokines, a protein that is produced in white blood cells. Cytokines are an important part of the human immune system, but they can also signal the immune system to attack the body's own cells if too many are made -- a so-called "cytokine storm" that is reported in some patients diagnosed with COVID-19.Second, the team showed that catalase can protect alveolar cells, which line the human lungs, from damage due to oxidation.Finally, the experiments showed that catalase can repress the replication of SARS?CoV?2 virus in rhesus macaques, a type of monkey, without noticeable toxicity."This work has far-reaching implications beyond the treatment of COVID-19. Cytokine storm is a lethal condition that can complicate other infections, such as influenza, as well as non-infectious conditions, like autoimmune disease," said Dr. Gregory Fishbein, an author on the study and a pathologist at the UCLA David Geffen School of Medicine.The co-lead authors are Zheng Cao, a graduate student involved in Lu's UCLA laboratory; Jing Wen, an assistant professor of UCLA School of Medicine; and Meng Qin from Beijing University of Chemical Technology.In addition to Fishbein, Bin Xu -- a visiting professor with Lu's laboratory -- is another author from UCLA.Researchers on the study are also from the Institute of Medical Biology at the Chinese Academy of Medical Sciences; Jinan University, China; and Vivibaba, a UCLA startup company.The study was supported in part by Beijing Science Sun Pharmaceutical Co. Ltd.
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Biotechnology
| 2,020 |
September 29, 2020
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https://www.sciencedaily.com/releases/2020/09/200929123506.htm
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Mussels connect antibodies to treat cancer
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Antibody-based immunotherapy is one of treatment options for cancer immunotherapy by modulating our immune system with therapeutic antibodies. Among many strategies for cancer therapy, it has been considered as an advanced cancer treatment due to its favorable clinical benefits by effectively activating our immune cells to attack cancer cells in the body. The development of new cancer immunotherapy drugs has gained great attention as the next-generation research area with more than 1,600 clinical trials currently underway worldwide.
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However, conventional cancer treatments that require intravenous administration throughout the whole body can cause side effects on normal cells or tissues by continuously supplying large amounts of antibodies. In particular, if large amounts of antibodies are released at once, there is a risk of excessive immune response that can lead to autoimmune diseases. The common localized treatment, such as local injection to tumor, still had a problem of rapidly spreading antibodies out of the target area by blood flow, leaving only a few, resulting in a significant reduction in treatment efficacy.The POSTECH research team led by professors Hyung Joon Cha, Kye Il Joo, and Dr. Yeonsu Jung of the Department of Chemical Engineering along with Professor Sin-Hyeog Im and Dr. Sung-Min Hwang of the Department of Life Sciences have together developed a novel immunotherapy platform called imuGlue. This new platform can effectively connect mussel adhesive proteins (MPAs) -- which has strong adhesion in even underwater conditions -- to the antibodies used as immune checkpoint inhibitors (ICIs) in order to deliver the antibodies to the target areas.ImuGlue can significantly enhance the efficiency of cancer immunotherapy and reduce the side effects by allowing therapeutic antibodies to stay in the target area for long periods of time even in moisture-rich environment and release antibodies on-demand at cancer sites. It was also demonstrated that imuGlue could be utilized for combination therapy with other immunomodulatory drugs often used in cancer immunotherapy.This new treatment platform for localized cancer immunotherapy has the advantage of not only being able to easily connect various therapeutic antibodies, but also does not mix or lose its property in body fluid- or blood-rich environments and at mucous surfaces. For this reason, it is anticipated to lead in the cancer immunotherapy market as it can be used not just through injections but also by spraying or other unique treatment methods."This study is the first immunotherapy that uses the mussel adhesion protein," commented Professor Hyung Joon Cha who led the study. He added, "As an innovative antibody delivery platform, it should be useful in various forms of immunotherapy."The study was published in
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Biotechnology
| 2,020 |
September 29, 2020
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https://www.sciencedaily.com/releases/2020/09/200929123619.htm
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E. coli engineered to grow on CO2 and formic acid as sole carbon sources
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Most biorefinery processes have relied on the use of biomass as a raw material for the production of chemicals and materials. Even though the use of CO
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Now, a metabolic engineering research group at KAIST has developed a strategy to grow an With support from the C1 Gas Refinery R&D Center and the Ministry of Science and ICT, a research team led by Distinguished Professor Sang Yup Lee stepped up their work to develop an engineered Despite the recent reports by several research groups on the development of The team previously reported the reconstruction of the tetrahydrofolate cycle and reverse glycine cleavage pathway to construct an engineered Metabolic fluxes were also fine-tuned, the gluconeogenic flux enhanced, and the levels of cytochrome bo3 and bd-I ubiquinol oxidase for ATP generation were optimized. This engineered Professor Lee said, "We engineered Professor Lee's team is continuing to develop such a strain. "In the future, we would be delighted to see the production of chemicals from an engineered
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Biotechnology
| 2,020 |
September 28, 2020
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https://www.sciencedaily.com/releases/2020/09/200928133145.htm
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Looking at evolution's genealogy from home
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Evolution leaves its traces -- in particular -- in genomes. Pinpointing its influence is a laborious process -- but one in which Dr. Jürgen Schmitz and his team at the University of Münster are at home. Five years ago, the team made public a web app which can compare the genomes of humans and animals and thus help to provide an understanding of evolutionary developments. The Münster researchers are now going one step further: their new software -- "2-n-way" -- can compare any genomes from and for anyone and systematically search for regions which are characterized by the presence or absence of certain sequences -- or, to put it simply, what is missing and where in the genome and when it got lost or when it newly emerged. This makes it possible to recognize relationships among species or individuals. The Münster researchers have now published details of their new development -- which, like its predecessor -- is freely available on the internet -- in the journal
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Jürgen Schmitz, a biologist and zoologist at the Institute of Experimental Pathology at Münster University's Faculty of Medicine, led the study together with Dr. Gennady Churakov and for him it represents a "unique, forward-pointing opportunity to take a close look at the mutability of multiple genomes." It means that not only genome evolutions can be analysed, but also the occurrence of genetic diseases as a result of deletions or insertions -- i.e. the loss of a DNA segment or the insertion of a new one. The decisive difference between the new model and its predecessor, GPAC -- which has been used hundreds of thousands of times since it was activated -- is that 2-n-way can sequence any number of genomes. "The tool is a response to the modern genomic era -- and it is a piece of software which, despite the complexity behind it, can be used by anyone, whether a non-medical layperson, a student or a professor. Last but not least, the tool links up a very wide range of areas, such as evolution, population genetics and medicine" says Schmitz.The term "2-n-way" is derived from two abbreviations used by specialists: "2-way" stands for the linear alignment of sequences to be compared; and "n-way" means the combination of individual components and the subsequent multiple comparison. But users do not need to know such background information. "They only have to download on the internet the genomes they want to compare. One source, for example, is the website of the National Center for Biotechnology Information (NCBI) in the USA. The genomes fed into 2-n-way are then aligned with each other by the software. The genes which are of particular interest are selected from the genomic "coordinates" -- or "loci," to give them their proper technical term. "The search can be geared for example to some or all of the so-called jumping genes, i.e. those genes which have changed their position in the genome," Schmitz explains.If for example a search is made for certain jumping genes in humans, chimpanzees or rhesus monkeys, the results are given in a table with "plus" and "minus." If the evaluation contains two finds and an exact gap in the rhesus monkey, then the conclusion is that humans and chimpanzees have inherited the jumping gene from a common ancestor and are therefore closely related -- while the rhesus monkey still displays the original locus without any insertion and is therefore only distantly related. However, the software not only indicates whether a certain insertion is present, but also the region in which it is to be found. In addition to the table of correlations, 2-n-way also provides the user with a list of DNA sequences for all loci.Although the new tool has only just been made public, the team of researchers is already looking ahead -- working on the simplification of the individual creation of 2-ways, i.e. the alignment. "This is a process which, at the moment, can take up a lot of time -- which is a bit annoying," says Schmitz. Otherwise, he comments, the tool "has been tested intensively and is absolutely perfect."
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Biotechnology
| 2,020 |
September 28, 2020
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https://www.sciencedaily.com/releases/2020/09/200928125031.htm
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Fungal compound inhibits important group of proteins
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Researchers in the group of Jeroen den Hertog, in collaboration with researchers in Leiden, have found that a compound inhibits a group of proteins called BMP receptors. This compound, called cercosporamide, was previously only known to inhibit a different group of proteins. When overactive, BMP receptors can induce several diseases. Studying compounds that may counteract this overactivity may lead to more treatment options in the future. Their results were published in the scientific journal
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We constantly need new therapeutic compounds for use in the clinic for various reasons, including our increasing age, corresponding illnesses and resistance to existing drugs. Fungi are an excellent, but underexplored source of these kinds of compounds. Researcher Jelmer Hoeksma explains: "Every year new compounds produced by fungi are identified, but so far we have only investigated a very small subset of all existing fungi. This suggests that many more biologically active compounds remain to be discovered."Together with the Westerdijk Fungal Biodiversity Institute, home to the largest collection of live fungi in the world, the researchers set up a large library of filtrates derived from more than ten thousand different fungi. A filtrate contains all the products that the fungus excretes. To search for therapeutic compounds, the researchers investigate the effects of fungal products present in this large library on zebrafish embryos. This enables them to study effects on the whole body during development.Using this approach, the researchers identified a compound, called cercosporamide, that had an effect in zebrafish. This effect is known for a certain type of molecules that inhibit a group of proteins called BMP receptors. When these BMP receptors are overactive, they can induce several diseases, such as Fibrodysplasia ossificans progressiva. In people that suffer from this disease, muscle tissue is progressively replaced with bone tissue, leading to a severe loss of mobility over time. Therefore, finding new compounds that may counteract overactive BMP receptors may provide new options for treatment of such diseases.Although the compound cercosporamide had been identified before, its effect on BMP receptors was unknown until now. The researchers discovered this additional effect because they tested the effects of compounds on whole zebrafish embryos. Additional tests in both zebrafish and human cells confirmed the results.Surprisingly, the molecules of cercosporamide have a completely different structure compared to other common BMP receptor inhibitors. So, even if cercosporamide itself turns out unusable as therapeutic drug, there may be a completely different class of structurally related chemicals that may have BMP receptor inhibiting effects.Currently, the researchers are looking for other bioactive compounds. Hoeksma: "For now, we are continuing to look for (new) compounds and understand their effects. Thus far, we only investigated a small subset of all these fungal products -- we have only scratched the surface."
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Biotechnology
| 2,020 |
September 28, 2020
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https://www.sciencedaily.com/releases/2020/09/200928125016.htm
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How methanogens are able to render oxygen molecules harmless
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Methane is a powerful greenhouse gas that plays a central role in the global carbon cycle. At the same time, it is an important energy source for us humans. About half of its annual production is made by microorganisms known as methanogens that decompose organic material such as dead plants. This normally takes place in a habitat without oxygen as this gas is lethal to methanogens. But even in actually oxygen-free habitats, oxygen molecules occasionally appear. To render these intruders harmless, methanogens possess a special enzyme that is able to convert oxygen into water.
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"Enzymes are vital components of the metabolism of all living organisms and the goal of our laboratory is to understand how these nanomachines are working at the molecular level," says Tristan Wagner from the Max Planck Institute for Marine Microbiology and first author of the study, published in the scientific journal "It was already known that F420-oxidase can convert oxygen into water," says Wagner. "But we succeeded to decrypt the mechanism." The study is a cooperation of scientists from the Max Planck Institute for Marine Microbiology, the Max Planck Institute for Terrestrial Microbiology, the Paul Scherrer Institute, the Interdisciplinary Research Institute of Grenoble and the European Synchrotron Radiation Facility.The mechanism, the researchers revealed, has an important requirement: Oxygen is very reactive, so it is crucial that the reaction is controlled correctly by the enzyme and no solvents are floating around. Otherwise the oxygen could accidentally be transformed in superoxide and kill the anaerobe. The trick of the enzyme F420-oxidase is to use a gas channel and a gating system. The oxygen molecule is first funneled in the specific channel to an appropriate anhydrous catalytic cavity containing iron. Then iron transforms the oxygen in water that will be released by a gating mechanism. For that the cavity begins to move and opens a small "door." Thanks to the movement, the newly generated water is transported outside. The empty cavity closes again and is available for the next oxygen molecule.To gain insights into this mechanism the scientists used X-ray crystallography. They first obtained the crystal structure without oxygen, where they could see the anhydrous catalytic cavity isolated from the solvent. Then, they gassed the enzyme crystals with the inert gas krypton, which, unlike oxygen, can be made visible by X-rays. Afterwards they irradiated the enzyme crystals and were able to detect krypton atoms showing the gas channel leading to the catalytic cavity. The flavodiiron protein and its channel is conserved not only in methanogens, but also in other microorganisms like clostridia (who live mainly in soil or in the digestive tract), in the sulfur bacteria Desulfovibrio gigas or even in the intestinal parasite Giardia intestinalis."This reaction is really fast," says Sylvain Engilberge from the Paul Scherrer Institute and first author of the study next to Tristan Wagner. "This velocity is also the high importance of our investigation." Similar enzymes like laccase are much slower. "For future application of bio-inspired electrochemical processes, we need to learn more from the chemical reaction, structure and function of different groups of oxygen-reducing enzymes," says Engilberge. It would also pave the way of protein engineering to convert a high-rate O2-detoxifier into an electron sink for industrial processes."Our next step would be to understand the diversity of flavodiiron protein," says Tristan Wagner. Some homologues are not targeting oxygen but the poisonous nitric oxide, their enzymes can discriminate between both gases with high specificity. But what is the selective filter? The gas channel? The environment of the catalytic cavity? "More studies have to be carried out to understand how the protein discriminates oxygen and nitric oxide," adds Wagner. With such knowledge, it would be for instance possible to predict from genomic information if a flavodiiron protein would be an oxygen or a nitric oxide scavenger.
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Biotechnology
| 2,020 |
September 28, 2020
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https://www.sciencedaily.com/releases/2020/09/200928125008.htm
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Fine-tuning stem cell metabolism prevents hair loss
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A team of researchers from Cologne and Helsinki has discovered a mechanism that prevents hair loss: hair follicle stem cells, essential for hair to regrow, can prolong their life by switching their metabolic state in response to low oxygen concentration in the tissue. The team was led by Associate Professor Sara Wickström (University of Helsinki and Max Planck Institute for the Biology of Ageing) and the dermatologist Professor Sabine Eming (University of Cologne), and included researchers from the University of Cologne's Cluster of Excellence in Aging Research CECAD, the Max Planck Institute for the Biology of Ageing, Collaborative Research Centre 829 'Molecular Mechanisms Regulating Skin Homeostasis', the Center for Molecular Medicine (CMMC) (all in Cologne), and the University of Helsinki. The paper 'Glutamine Metabolism Controls Stem Cell Fate Reversibility and Long-Term Maintenance in the Hair Follicle' has been published in
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Every day, tissues such as the skin and its hair follicles are exposed to environmental damage like ultraviolet radiation. Damaged material is continuously removed and renewed. On average, 500 million cells and 100 hairs are shed every day, amounting to 1.5 gram of material. The dead material is replaced by stem cells, which are specialized, highly proliferative and long-lived. Tissue function relies on the activity and health of these stem cells; compromised function or reduced number leads to aging. 'Although the critical role of stem cells in aging is established, little is known about the mechanisms that regulate the long-term maintenance of these important cells. The hair follicle with its well understood functions and clearly identifiable stem cells was a perfect model system to study this important question', said Sara Wickström.To understand what made stem cells functionally distinct from their differentiated daughter cells, the team investigated the transcriptional and metabolic profiles of the two cell populations. 'Intriguingly, these studies showed that stem cells and daughter cells have distinct metabolic characteristics', said Dr. Christine Kim, co-leading scientist of the study. 'Our analyses further predicted that Rictor, an important but relatively poorly understood molecular component of the metabolic master regulator mTOR pathway, would be involved.' The mTOR signal transduction regulates processes like growth, energy, and oxygen consumption of cells.In more detailed analyses, the team showed that stem cell depletion was due to the loss of metabolic flexibility. At the end of each regenerative cycle, during which a new hair is made, the stem cells will return to their specific location and resume a quiescent state. Dr. Xiaolei Ding, the other co-leading scientist, explained: 'The key finding of this study is that this so called "fate reversibility" requires a shift from glutamine metabolism and cellular respiration to glycolysis. The stem cells reside in an environment with low oxygen availability and thus use glucose rather than glutamine as a carbon source for energy and protein synthesis. This shift is triggered by the low oxygen concentration and Rictor signaling. The removal of Rictor impaired the ability of this stem cell fate reversal, triggering slow, age-dependent exhaustion of the stem cells and age-induced hair loss.' Ding and Eming had recently generated a genetic mouse model to study Rictor function and observed that mice lacking Rictor had significantly delayed hair follicle regeneration and cycling, which indicated impaired stem cell regulation. 'Interestingly, with aging these mice showed hair loss and reduction in stem cell numbers', said Ding.'A major future goal will be to understand how these preclinical findings might translate into stem cell biology in humans and potentially could be pharmaceutically harnessed to protect from hair follicle aging', said Eming. 'We are particularly excited about the observation that the application of a glutaminase inhibitor was able to restore stem cell function in the Rictor-deficient mice, proving the principle that modifying metabolic pathways could be a powerful way to boost the regenerative capacity of our tissues.'
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113438.htm
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The male Y chromosome does more than we thought
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New light is being shed on a little-known role of Y chromosome genes, specific to males, that could explain why men suffer differently than women from various diseases, including Covid-19.
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The findings were published this month in "Our discovery provides a better understanding of how male genes on the Y chromosome allow male cells to function differently from female cells," said Deschepper, the study's lead author, who is also an associate professor at McGill University."In the future, these results could help to shed some light on why some diseases occur differently in men and women."Humans each have 23 pairs of chromosomes, including one pair of sex chromosomes. While females carry two X sex chromosomes, males carry one X and one Y chromosome. This male chromosome carries genes that females lack. Although these male genes are expressed in all cells of the body, their only confirmed role to date has been essentially limited to the functions of the sex organs.In his study, Deschepper performed a genetic manipulation that inactivated two male genes on the Y chromosome, altering several signalling pathways that play important roles in certain functions of non-sex organ cells. For example, under stress, some of the affected mechanisms could influence the way in which cells in human hearts defend themselves against aggressions such as ischemia (reduced blood supply) or mechanical stress.In addition, the study showed that these male genes performed their regulatory functions in a way that was unusual compared to the mechanisms generally used by most other genes on the non-sex chromosomes. Thus, instead of specifically activating certain genes by direct action at the genome level, the Y chromosome seems to affect cellular functions by acting on protein production.The discovery of these differences in function may explain in part why the functions of male Y chromosome genes have so far been poorly understood, said Deschepper.Males differ from females in the manifestation, severity and consequences of most diseases. A recent example of this duality is Covid-19, which has a mortality rate twice as high in men than in women.
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Biotechnology
| 2,020 |
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