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September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113356.htm
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'Stretching rack' for cells
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The behavior of cells is controlled by their environment. Besides biological factors or chemical substances, physical forces such as pressure or tension are also involved. Researchers from Karlsruhe Institute of Technology (KIT) and Heidelberg University developed a method that enables them to analyze the influence of external forces on individual cells. Using a 3D printing process, they produced micro-scaffolds, each of which has four pillars on which a cell is located. Triggered by an external signal, a hydrogel inside the scaffold swells and pushes the pillars apart, so that the cell must "stretch."
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The work is part of the "3D Matter Made to Order" (3DMM2O) Cluster of Excellence. The researchers report on their results in Many cellular biological processes, such as wound healing or the development of tissue, are strongly influenced by the properties of their environment. Cells react, for example, to biological factors or chemical substances. However, research is increasingly focusing on physical forces acting on the cells: How exactly do the cells adapt to these forces?Within the framework of the German-Japanese University Consortium HeKKSaGOn and in cooperation with Australian scientists, the 3DMM2O team has taken a particularly ingenious approach to this question. For the production of their cell "stretching racks" they used "direct laser writing", a special 3D printing process in which a computer-controlled laser beam is focused into a special printer ink liquid. Its molecules react only at the exposed areas and form a solid material there. All other areas remain liquid and can be washed away. "This is an established method in our Cluster of Excellence for building three-dimensional structures - on the micrometer scale and below," explains Marc Hippler from the KIT Institute of Applied Physics, lead author of the publication.In the current case, the researchers used three different printer inks: The first ink, made of protein-repellent material, was used to form the actual micro-scaffold. Using a second ink of protein-attracting material, they then produced four horizontal bars that are connected to one of the scaffold pillars each. The cell is anchored to these four bars. The real showstopper, however, is the third ink: The scientists used it to "print" a mass inside the scaffold. If they then add a special liquid, the hydrogel swells. It thus develops a force sufficient to move the pillars - and the bars with them. This, in turn, has the effect of stretching the cell that is fixed to the bars.The scientists of the Cluster of Excellence placed two completely different cell types on their micro stretching rack: human bone tu-mor cells and embryonic mouse cells. They found that the cells counteract the external forces with motor proteins and thus greatly increase their tensile forces. When the external stretching force is removed, the cells relax and return to their original state. "This be-havior is an impressive demonstration of the ability to adapt to a dynamic environment. If the cells were unable to recover, they would no longer fulfill their original function - for example wound closure," says Professor Martin Bastmeyer from the Zoological Institute of KIT.As the team further discovered, a protein called NM2A (non-muscle myosin 2A) plays a decisive role in the cells' response to mechani-cal stimulation: Genetically modified bone tumor cells that cannot produce NM2A were barely able to counteract the external defor-mation.Work in the cluster of excellence was carried out by Heidelberg scientists from the field of biophysical chemistry as well as physics and cell- and neurobiology from KIT. Members of the German-Japanese University Consortium HeKKSaGOn include, among oth-ers, Heidelberg University, Karlsruhe Institute of Technology and Osaka University.In the 3D Matter Made to Order (3DMM2O) Cluster of Excellence, scientists of Karlsruhe Institute of Technology and Heidelberg Uni-versity conduct interdisciplinary research into innovative technolo-gies and materials for digital scalable additive manufacture to en-hance the precision, speed, and performance of 3D printing. Work is aimed at completely digitizing 3D manufacture and materials pro-cessing from the molecule to the microstructure. In addition to fund-ing as a cluster of excellence under the Excellence Strategy compe-tition launched by the federation and the federal states, 3DMM3O is financed by Carl Zeiss Foundation.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113348.htm
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RAP tag: A new protein purification approach
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Whether it's our diets, building strength, or as part of medical advancements, it is no secret that proteins form an important part of our lives. Tracking how proteins work and move in cells, and purifying engineered proteins, are important tools for researchers. Traditional approaches to label proteins of interest, called "tagging," have the disadvantage of interfering with protein characteristics, including function and localization. Sometimes, these tags can also cross-react, which makes the information they provide nonspecific. A successful protein tagging system needs to be highly specific and have high affinity.
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In a study published in September 2020 in Frontiers in Plant Science, researchers from the University of Tsukuba, led by Professor Kenji Miura, have described a new tagging system for detecting and purifying proteins in plant cells. This approach uses a short sequence called a "RAP tag" to label proteins. An antibody, PMab-2, is then able to specifically recognize the RAP tag and can be used to purify the proteins of interest.In describing this approach, Professor Miura says, "The high affinity and specificity of immunoaffinity chromatography using monoclonal antibodies makes it a very powerful tool, especially for the purification of proteins expressed at low levels." A hurdle to applying this approach, however, is the high cost of reagents, especially that of antibodies.To get around this, Professor Miura and colleagues explored whether they could produce the PMab-2 antibody in the plant model Nicotiana benthamiana, a relative of the tobacco plant. Not only could they successfully produce PMab-2, they went on to show that the plant-produced PMab-2 behaved similarly to that produced in animal cells. This discovery opens the door to reducing the cost of antibody production, and could be applied more widely across scientific fields.Testing the feasibility of a RAP-tagged/ PMab-2 affinity purification approach, the researchers then expressed RAP-tagged proteins in plant cells. They found that these tagged proteins could be specifically identified using the PMab-2 antibody. Moreover, RAP-tagged recombinant proteins, involving the fusion of sequences from more than one protein, and protein complexes were also expressed in these cells and identified by PMab-2. These proteins could also be purified from plant cells using the PMab-2 antibody, indicating that the RAP tag can be used for both protein detection and purification from soluble plant extracts."Plants are an extremely valuable resource for molecular biology," explains Professor Miura. "They can be used as bioreactors to produce large amounts of proteins because they are unlikely to suffer from contamination issues faced by bacterial and mammalian cell systems."The results presented by the team show that this approach has the potential to be widely applied across the molecular sciences.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113339.htm
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Secure nano-carrier delivers medications directly to cells
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Medications often have unwanted side-effects. One reason is that they reach not only the unhealthy cells for which they are intended, but also reach and have an impact on healthy cells. Researchers at the Technical University of Munich (TUM), working together with the KTH Royal Institute of Technology in Stockholm, have developed a stable nano-carrier for medications. A special mechanism makes sure the drugs are only released in diseased cells.
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The human body is made up of billions of cells. In the case of cancer, the genome of several of these cells is changed pathologically so that the cells divide in an uncontrolled manner. The cause of virus infections is also found within the affected cells. During chemotherapy for example, drugs are used to try to destroy these cells. However, the therapy impacts the entire body, damaging healthy cells as well and resulting in side effects which are sometimes quite serious.A team of researchers led by Prof. Oliver Lieleg, Professor of Biomechanics and a member of the TUM Munich School of BioEngineering, and Prof. Thomas Crouzier of the KTH has developed a transport system which releases the active agents of medications in affected cells only. "The drug carriers are accepted by all the cells," Lieleg explains. "But only the diseased cells should be able to trigger the release of the active agent."The scientists have now shown that the mechanism functions in tumor model systems based on cell cultures. First they packaged the active ingredients. For this purpose, they used so-called mucins, the main ingredient of the mucus found for example on the mucus membranes of the mouth, stomach and intestines. Mucins consist of a protein background to which sugar molecules are docked. "Since mucins occur naturally in the body, opened mucin particles can later be broken down by the cells," Lieleg says.Another important part of the package also occurs naturally in the body: deoxyribonucleic acid (DNA), the carrier of our genetic information. The researchers synthetically created DNA structures with the properties they desired and chemically bonded these structures to the mucins. If glycerol is now added to the solution containing the mucin DNA molecules and the active ingredient, the solubility of the mucins decreases, they fold up and enclose the active agent. The DNA strands bond to one another and thus stabilize the structure so that the mucins can no longer unfold themselves.The DNA-stabilized particles can only be opened by the right "key" in order to once again release the encapsulated active agent molecules. Here the researchers use what are called microRNA molecules. RNA or ribonucleic acid has a structure very similar to that of DNA and plays a major role in the body's synthesis of proteins; it can also regulate other cell processes."Cancer cells contain microRNA strands whose structure we know precisely," explains Ceren Kimna, lead author of the study. "In order to use them as keys, we modified the lock accordingly by meticulously designing the synthetic DNA strands which stabilize our medication carrier particles." The DNA strands are structured in such a way that the microRNA can bind to them and as a result break down the existing bonds which are stabilizing the structure. The synthetic DNA strands in the particles can also be adapted to microRNA structures which occur with other diseases such as diabetes or hepatitis.The clinical application of the new mechanism has not yet been tested; additional laboratory investigations with more complex tumor model systems are necessary first. The researchers also plan to investigate further modifying this mechanism to release active agents in order to improve existing cancer therapies.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113327.htm
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New method allows precise gene control by light
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A novel optical switch makes it possible to precisely control the lifespan of genetic "copies." These are used by the cell as building instructions for the production of proteins. The method was developed by researchers from the universities of Bonn and Bayreuth. It may significantly advance the investigation of dynamic processes in living cells. The study is published in the journal
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Metaphorically speaking, every human cell contains in its nucleus a huge library of tens of thousands of books, the genes. Each of these books in turn contains the building instructions for a protein. When the cell needs a certain protein, a transcription of the corresponding instructions is made. These transcriptions are called mRNAs (RNA is a slightly modified form of DNA).A cellular mechanism ensures that the mRNA transcriptions are "shredded" again after a short time. This ensures that the protein is only produced as long as it is actually needed. Several decades ago, researchers came up with the idea of using this shredder for their own purposes: By specifically attaching a marker to certain mRNAs, they ensure that the transcriptions are not used as building instructions at all, but are destroyed immediately: a process also known as RNA silencing. The cell then lacks the corresponding protein. This makes it possible to find out which function it would actually be responsible for.The approach that the groups from Bonn and Bayreuth have now published is based on this method. However, it is nowhere near as crude, but allows a far more differentiated control over the lifespan of the mRNA copies. "We use a bacterial molecule to control the shredding of mRNA transcriptions with the help of light," explains Prof. Dr. Günter Mayer, who heads the Chemical Biology & Medicinal Chemistry Research Group at the LIMES Institute of the University of Bonn.The bacterial molecule with the abbreviation PAL acts as a kind of switch. It changes its shape under the influence of blue light. In the process, a pocket is exposed that can bind to certain molecules. "We searched a huge library of artificially produced short RNA molecules called aptamers," says Mayer. "Eventually we came across an aptamer that's a good match for the pocket in the PAL molecule."The researchers have now coupled this aptamer to one of the molecular markers that can attach to mRNAs and thereby release them for degradation. "When we irradiate the cell with blue light, PAL binds to the marker via the aptamer and thus puts it out of action," explains Mayer's colleague Sebastian Pilsl. "The mRNA is then not destroyed, but translated into the corresponding protein." As soon as the researchers switch off the blue light, PAL releases the label again. Now it can attach itself to the mRNA, which is then shredded.This will in future enable researchers to investigate exactly where and when a protein is needed in a cell, simply by immersing an area of the cell in blue light at a certain time and then looking at the consequences. In the current study they applied this to proteins that play an important role in the regulation of the cell cycle and cell division. The combination of aptamer and degradation marker is introduced into the cell by genetic engineering. This means that it generates the light-dependent degradation signal itself; it does not have to be supplied from outside.The aptamer can be combined with any markers, each of which in turn serves as a shredder signal for a specific mRNA. "This method can therefore be used to switch off practically every mRNA molecule in the cell in a controlled manner," emphasizes Prof. Dr. Andreas Möglich from the University of Bayreuth. In the recently published pilot study, it all worked both simply and reliably. The researchers therefore see great potential in their method for the investigation of dynamic processes in living cells and organisms.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200924141527.htm
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Finding the Achilles' heel of a killer parasite
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Two studies led by UT Southwestern researchers shed light on the biology and potential vulnerabilities of schistosomes -- parasitic flatworms that cause the little-known tropical disease schistosomiasis. The findings, published online today in
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About 240 million people around the world have schistosomiasis -- mostly children in Africa, Asia, and South America in populations that represent "the poorest of the poor," says study leader James J. Collins III, Ph.D., associate professor in UTSW's department of pharmacology.Most of those infected survive, but those who die often suffer organ failure or parasite-induced cancer. Symptoms can be serious enough to keep people from living productive lives, Collins says.The parasite that causes this disease has a complicated life cycle that involves stages in both freshwater snails and mammals. Dwelling in mammalian hosts' circulatory systems, schistosomes feed on blood and lay copious numbers of eggs, all while causing an array of symptoms including abdominal pain, diarrhea, bloody stool, or blood in the urine. Larval worms are released from snails into water, where the flatworms then may infect humans by penetrating the skin. Schistosomiasis may become a chronic disease that affects the person for years.Only one drug, praziquantel, is available to treat this condition. However, Collins explains, it is of limited use -- it doesn't kill all intramammalian stages of the schistosome life cycle, and it has a variable cure rate in some endemic settings. There's been little interest by pharmaceutical companies in developing new drugs for this disease, he adds, because there is no monetary incentive to do so. Consequently, relatively few studies have been devoted to understanding schistosomes' basic biology, which might reveal inherent weaknesses that could serve as targets for new drugs.To that end, Collins and his colleagues embarked upon two separate studies -- one at the cellular level and another at the molecular level -- to better understand these organisms.In the first study, the researchers delved into the cell types that make up these flatworms. Although the worms are multicellular organisms composed of a variety of unique tissue types, researchers knew little about the different cell populations in these parasites.With a goal to create an atlas of cell types in In the second study, the researchers used RNAi to sort out the function of about 20 percent of By deactivating the genes one by one, Collins and his colleagues identified more than 250 genes crucial for survival. Using a database of pharmacological compounds, the researchers then searched for drugs that had the potential to act on proteins produced by these genes, identifying several compounds with activity on worms. The team also uncovered two protein kinases -- a group of proteins renowned for their ability to be targeted by drugs -- that are essential for muscle function. When these kinases were inhibited, the worms became paralyzed and eventually died, suggesting that drugs targeting these proteins could eventually treat people with schistosomiasis. A next step in the research will be to search for inhibitors of these proteins.Collins notes that these strides in understanding the basic biology of schistosomes could eventually lead to new treatments to save untold lives in places where schistosomiasis is endemic."This is a very important disease that most people have never heard of," he says. "We need to invest and invigorate research on these parasites."UTSW researchers who contributed to the first study include George Wendt, Lu Zhao, Rui Chen, and Michael L. Reese. UTSW researchers who contributed to the second study include Jipeng Wang, Carlos Paz, Irina Gradinaru, and Julie N. R. Collins.The first study was supported by grants from the National Institutes of Health (R01 R01AI121037, R01 R01AI150715, R21 R21AI133393, and F30 1F30AI131509-01A1, the Welch Foundation (I-1948-20180324 and I-1936-20170325), the National Science Foundation (MCB1553334), the Burroughs Wellcome Fund, the Wellcome Trust (107475/Z/15/Z), and the Bill and Melinda Gates Foundation (OPP1171488).The second study was supported by grants from the National Institutes of Health (R01AI121037), the Welch Foundation (I-1948-20180324), the Burroughs Wellcome Fund, and the Wellcome Trust (107475/Z/15/Z and 206194).James Collins is Rita C. and William P. Clements, Jr. Scholar in Biomedical Research.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113636.htm
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Color-coded biosensor illuminates in real time how viruses attack hosts
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Infectious viruses come in many shapes and sizes and use slightly different attack mechanisms to make humans and animals sick. But all viruses share something in common: They can only do damage by replicating inside the cells of another organism -- their host.
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This broad, fundamental process of how viruses trick host cells into making copies of the virus has had a team of Colorado State University scientists captivated for several years. A collaboration between the labs of Monfort Professor Tim Stasevich, in the Department of Biochemistry and Molecular Biology, and Associate Professor Brian Munsky, in the Department of Chemical and Biological Engineering, is on a mission to understand, in visual detail and with mathematical precision, all aspects of viral attack strategies, including how viruses invade host cell protein-making machinery. Their work, supported by grants from the National Institute of General Medicine and the W. M. Keck Foundation, could provide insight into predicting and fighting back against all manner of viral diseases.For the first time ever, the team has shown an important mechanism in this host-attacking process, at the single-molecule level in living cells, and they have reproduced these behaviors in computational models. Their new experiments and models, published in Nature Structural and Molecular Biology, reveal in unprecedented detail how viruses initiate translation of genetic material into proteins.Since viruses do not encode their own replication machinery, they hijack that of their host cells by stealing cellular machines called ribosomes, which are essential for making proteins from the genetic material found in RNA. Many viral genomes contain special RNA structures called Internal Ribosome Entry Sites, or IRES, that capture ribosomes from the host, forcing those ribosomes to make viral proteins.Researchers know that when IRES-related RNA translation takes place, the virus has succeeded in commandeering the host's ribosomes. The CSU researchers invented a biosensor that lights up blue when viral translation is happening, and green when normal host translation is happening, in single living cells. This design allows them to differentiate between normal host processes and viral processes, in real time.The sensor combines the relevant bits of virus (not the whole virus) that interact with and steal host ribosomes, along with two distinct protein tags that glow the moment RNA is translated. First author and graduate student Amanda Koch spent more than a year developing the sensor, with the goal of looking at host protein RNA translation, and virus-related RNA translation, at the same time.Luis Aguilera, a postdoctoral researcher in the Munsky group, built a detailed computational model to reproduce Koch's fluorescence microscopy videos. By analyzing Koch's data through the lens of dozens of hypotheses and millions of possible combinations, Aguilera discovered complex biochemical mechanisms that the biochemists couldn't directly see. His models showed that both healthy human RNA and viral RNA fluctuate between states that actively express proteins and those that are silent.In addition to examining viral translation in normal cells, Koch's biosensor allows the researchers to visualize the effects of different types of stress that cells undergo when being attacked by a virus, and how, where and when normal versus viral translation increase or decrease. The integration of Koch's microscopy data and Aguilera's computational models revealed that the relationship between normal and IRES-mediated translation is largely one-sided -- in healthy cells, normal translation dominates, but in cells under stress, IRES translation dominates.The Stasevich and Munsky teams envision that the combination of their unique biochemical sensors and detailed computational analyses will provide powerful tools to understand, predict, and control how future drugs might work to inhibit viral translation without affecting host translation.As the researchers look ahead to the future, they have their sights next set on COVID-19. Although SARS-CoV-2 does not contain an IRES, according to Koch "our biosensor is modular and can easily incorporate pieces of SARS-CoV-2 to explore how it uniquely hijacks host replication machinery during infection.""We are proving, more and more, that we can look at these nuanced dynamics of how viruses are sneaking past their hosts to infect a lot of cells and make us sick," Koch said.
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Biotechnology
| 2,020 |
September 25, 2020
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https://www.sciencedaily.com/releases/2020/09/200925113623.htm
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Lending color to dead cells -- A novel natural dye for screening cell viability
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Conducting studies
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Researchers have elucidated several methods to tell live and dead cells apart, and one popular approach is the "dye exclusion test (DET)" using synthetic dyes. In conventional DET, a dye such as trypan blue or methylene blue selectively permeates and stains dead cells, distinguishing them from live cells. This seems simple enough, but these synthetic dyes have been known to damage living cells in the culture as well. This renders them unusable for long-term studies with a single culture.Fortunately, as is described in their study published in MDPI Biology, a team of scientists from the Tokyo University of Science, Japan?comprising Assistant Professor Ryoma Tagawa, Professor Yoshikazu Higami, Professor Eiji Tokunaga, and Assistant Professor Kyohei Yamashita?recently discovered an alternative to DET with synthetic dyes: DET using a natural pigment made from Dr Yamashita and a colleague were working alongside Dr Koji Yamada and Dr Kengo Suzuki from euglena Co., Ltd. to find effective ways of culturing Dr Yamashita then went on to lead another study demonstrating the applicability of MP in DET for another single-cell organism species with a vastly different structure, In their most recent study, the one published in MDPI Biology, Dr Yamashita and colleagues proved that MP can be used to ascertain the viability of breast cancer cells. They found that, unlike trypan blue, MP does not damage living cells and is robust against a typical chemotherapy drug cisplatin. Moreover, MP took only ten minutes to stain dead cells and costs a tenth of what trypan blue does. Considering all this, Dr Yamashita remarks: "The proposed natural pigment enables the long-term monitoring of the life and death of cells, which may bring about improvements in the efficiency of biomass production, basic research on metabolic mechanisms, and applied research in fields such as breeding."In addition to its use as a reagent to monitor the life and death of cells, Dr Yamashita notes that the pigment is also nutritious to living cells and has antioxidative characteristics, which is useful for boosting culture efficiency and performing quality control in the food industry, where safe fermentation is critical. It is also safe to humans and the environment.This applicability of MP to completely different kinds of cells -- breast cancer, There is certainly a bright and colorful future ahead for this promising natural dye!
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Biotechnology
| 2,020 |
September 24, 2020
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https://www.sciencedaily.com/releases/2020/09/200924141550.htm
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Twinkling, star-shaped brain cells may hold the key to why, how we sleep
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A new study published today in the journal
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Led by researchers at Washington State University's Elson S. Floyd College of Medicine, the study builds new momentum toward ultimately solving the mystery of why we sleep and how sleep works in the brain. The discovery may also set the stage for potential future treatment strategies for sleep disorders and neurological diseases and other conditions associated with troubled sleep, such as PTSD, depression, Alzheimer's disease, and autism spectrum disorder."What we know about sleep has been based largely on neurons," said lead author and postdoctoral research associate Ashley Ingiosi. Neurons, she explained, communicate through electrical signals that can be readily captured through electroencephalography (EEG). Astrocytes -- a type of glial (or "glue") cell that interacts with neurons -- do not use electrical signals and instead use a process known as calcium signaling to control their activity.It was long thought that astrocytes -- which can outnumber neurons by five to one -- merely served a supportive role, without any direct involvement in behaviors and processes. Neuroscientists have only recently started to take a closer look at their potential role in various processes. And while a few studies have hinted that astrocytes may play a role in sleep, solid scientific tools to study their calcium activity have not been available until recently, Ingiosi said.To delve deeper into astrocytes' role in sleep, she and her coauthors used a rodent model to record astrocytes' calcium activity throughout sleep and wake, as well as after sleep deprivation. They used a fluorescent calcium indicator that was imaged via tiny head-mounted microscopes that looked directly into the brains of mice as they moved around and behaved as they normally would. This indicator allowed the team to see calcium-driven fluorescent activity twinkling on and off in astrocytes during sleep and waking behaviors. Their one-of-a-kind methodology using these miniature microscopes allowed the team to conduct the first-ever study of astrocytes' calcium activity in sleep in freely behaving animals.The research team set out to answer two main questions: do astrocytes change dynamically across sleep and wake states like neurons do? And do astrocytes play a role in regulating sleep need, our natural drive to sleep?Looking at astrocytes in the frontal cortex, an area of the brain associated with measurable EEG changes in sleep need, they found that astrocytes' activity changes dynamically across the sleep-wake cycle, as is true for neurons. They also observed the most calcium activity at the beginning of the rest phase -- when sleep need is greatest -- and the least calcium activity at the end of the test phase, when the need for sleep has dissipated.Next, they kept mice awake for the first 6 hours of their normal rest phase and watched calcium activity change in parallel with EEG slow wave activity in sleep, a key indicator of sleep need. That is, they found that sleep deprivation caused an increase in astrocyte calcium activity that decreased after mice were allowed to sleep.Their next question was whether genetically manipulating astrocyte calcium activity would impact sleep regulation. To find out, they studied mice that lacked a protein known as STIM1 selectively in astrocytes, which reduced the amount of available calcium. After being sleep deprived, these mice did not sleep as long or get as sleepy as normal mice once allowed to sleep, which further confirmed earlier findings that suggest that astrocytes play an essential role in regulating the need for sleep.Finally, they tested the hypothesis that perhaps astrocyte calcium activity merely mirrors the electrical activity of neurons. Studies have shown that the electrical activity of neurons becomes more synchronized during non-REM sleep and after sleep deprivation, but the researchers found the opposite to be true for astrocytes, with calcium activity becoming less synchronized in non-REM sleep and after sleep deprivation."This indicates to us that astrocytes are not just passively following the lead of neurons," said Ingiosi. "And because they don't necessarily display the same activity patterns as neurons, this might actually implicate a more direct role for astrocytes in regulating sleep and sleep need."More research is needed to further unravel the role of astrocytes in sleep and sleep regulation, Ingiosi said. She plans to study astrocytes' calcium activity in other parts of the brain that have been shown to be important for sleep and wake. In addition, she would like to look at astrocytes' interactions with different neurotransmitters in the brain to start to tease out the mechanism by which astrocytes might drive sleep and sleep need."The findings of our study suggest that we may have been looking in the wrong place for more than 100 years," said senior author and professor of biomedical sciences Marcos Frank. "It provides strong evidence that we should be targeting astrocytes to understand why and how we sleep, as well as for the development of therapies that could help people with sleep disorders and other health conditions that involve abnormal sleep."Support for the study came from the National Institutes of Health.
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Biotechnology
| 2,020 |
September 24, 2020
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https://www.sciencedaily.com/releases/2020/09/200924141507.htm
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Mystery of giant proton pump solved
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Mitochondria are the powerhouses of our cells, generating energy that supports life. A giant molecular proton pump, called complex I, is crucial: It sets in motion a chain of reactions, creating a proton gradient that powers the generation of ATP, the cell's fuel. Despite complex I's central role, the mechanism by which it transports protons across the membrane has so far been unknown. Now, Leonid Sazanov and his group at the Institute of Science and Technology Austria (IST Austria) have solved the mystery of how complex I works: Conformational changes in the protein combined with electrostatic waves move protons into the mitochondrial matrix. This is the result of a study published today in
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Complex I is the first enzyme in the respiratory chain, a series of protein complexes in the inner mitochondrial membrane. The respiratory chain is responsible for most of the cell's energy production. In this chain, three membrane proteins set up a gradient of protons, moving them from the cell's cytoplasm into the mitochondrial inner space, called the matrix. The energy for this process comes mostly from the electron transfer between NADH molecules, derived from the food we eat, and oxygen that we breathe. ATP synthase, the last protein in the chain, then uses this proton gradient to generate ATP. Complex I is remarkable not only because of its central role in life, but also for its sheer size: with a molecular weight of 1 Megadalton, the eukaryotic complex I is one of the biggest membrane proteins. Its size also makes complex I hard to study. In 2016, Sazanov and his group were the first to solve the structure of mammalian complex I, following on their 2013 structure of a simpler bacterial enzyme. But the mechanism by which complex I moves protons across the membrane has remained controversial. "One idea was that a part of complex I works like a piston, to open and close channels through which protons travel," explains Sazanov. "Another idea was that residues at the center of complex I act as a driver. It turns out that an even more unusual mechanism is at work."Previously, Sazanov and his group have shown that L-shaped complex I consists of hydrophilic and membrane arms. In the hydrophilic arm, electrons tunnel from NADH to quinone, the hydrophobic electron carrier. The membrane arm, where proton translocation happens, has three similar subunits with structures related to antiporters, and one subunit containing a quinone binding cavity. In this cavity, complex I transfers two electrons per catalytic cycle to quinone, which delivers the electrons further to complexes III and IV. But mystery surrounded how the interaction between electrons and quinone can move four protons per cycle across the membrane, since the antiporter-like subunits are far away from quinone cavity. To solve this puzzle, Sazanov and his team performed cryo-EM on sheep complex I. In a tour-de-force effort, PhD student Domen Kampjut solved 23 different structures of complex I, obtained under different conditions. By adding NADH and quinone, the researchers could capture images of complex I at work, changing conformation between the two main states. Due to high-resolution achieved, they could resolve the water molecules inside the protein, which are essential to allow proton transfer. They found that many water molecules in the central axis of the membrane arm provide a way for protons to hop between polar residues and waters, forming pathways along and across the membrane.But only in one subunit, furthest away from quinone, do protons hop across the membrane. The other two subunits rather provide a coupling between the farthest subunit and quinone. When the binding cavity "waits" for quinone, a helix blocks the water wire in the central axis. When quinone binds in the binding cavity, the protein conformation around this area changes dramatically and this helix rotates. Now, the water wire connects all membrane subunits of complex I and two protons are delivered to quinone, to complete its reduction. This key part of the mechanism creates a charge near the first antiporter and starts an electrostatic wave of interactions between charged residues, which propagates along the antiporters, resulting in the translocation of four protons in total. "We show that a new and unexpected mechanism is at work in complex I. A mixture of both conformational changes and an electrostatic wave pumps protons across the membrane," explains Sazanov. "This mechanism is highly unusual, as it involves the rotation of an entire helix inside the protein. It seems a bit excessive, but probably helps the mechanism to be robust."The new research complements studies from Sazanov group published in the last two months, on the mechanism of proton pumping in bacterial complex I (
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Biotechnology
| 2,020 |
September 24, 2020
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https://www.sciencedaily.com/releases/2020/09/200924101929.htm
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Bioelectronic device achieves unprecedented control of cell membrane voltage
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In an impressive proof-of-concept demonstration, an interdisciplinary team of scientists has developed a bioelectronic system driven by a machine learning algorithm that can shift the membrane voltage in living cells and maintain it at a set point for 10 hours.
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Every living cell maintains a voltage across the cell membrane that results from differences in the concentrations of charged ions inside and outside the cell. Often called the membrane potential or resting potential, this voltage is regulated by ion channels in the cell membrane and plays important roles in cell physiology and functions such as proliferation and differentiation.Controlling cells with bioelectronics is difficult due to the complex ways cells respond to changes in their environment and the natural self-regulating feedback process known as homeostasis. Cells regulate ion movements to maintain a steady membrane voltage, so the researchers had to develop a system that could counteract this natural response."Biological feedback systems are fundamental to life, and their malfunctioning is often involved in diseases. This work demonstrates that we can tweak this feedback using a combination of bioelectronic devices actuated by machine learning, and potentially restore its functioning," said Marco Rolandi, professor and chair of electrical and computer engineering at the UC Santa Cruz Baskin School of Engineering.Rolandi is the senior corresponding author of the paper describing this work, published September 24 in the journal The researchers developed a system involving an array of bioelectronic proton pumps that add or remove hydrogen ions from solution in proximity to cultured human stem cells. The cells were genetically modified to express a fluorescent protein on the cell membrane that responds to changes in membrane voltage. The system is controlled by a machine learning algorithm that tracks how the membrane voltage responds to stimuli from the proton pumps."It is a closed-loop system, in that it records the behavior of the cells, determines what intervention to deliver using the proton pumps, sees how the cells react, then determines the next intervention needed to achieve and maintain the membrane voltage status we desire," Rolandi explained.Gomez, who developed the machine learning algorithm, said the algorithm is not trained on any data in advance and does not rely on a model of the system. Instead, the "learning" happens in real time as the neural network responds to input regarding the current state of the membrane voltage."The adaptive nature of biology -- that is, the ability of cells to change their response to external stimuli -- calls for an adaptive approach in controls, where static models and past information can become obsolete," Gomez said.Because the membrane voltage of stem cells is different from that of mature, differentiated cells, the researchers are interested in the possibility of using the system to induce and direct the differentiation of stem cells into specific cell types. They did not, however, explicitly look at cell differentiation in this proof-of-concept study.More broadly, the combination of bioelectronics and machine learning in a closed-loop biohybrid system has many potential applications in regenerative medicine and synthetic biology, Rolandi said. He noted that the results of this study will inform the team's work on a major effort to develop a "smart bandage" providing bioelectronic intelligent control of wound regeneration."This study is an important proof of concept for the use of bioelectronics and machine learning to control cell functions," he said.This research was funded by the Defense Advanced Research Projects Agency (DARPA).
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923164607.htm
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Berry good news: newly discovered compound from blueberries could treat inflammatory disorders
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Various plants and their products are known to contain "bioactive" ingredients that can alleviate human diseases. These "phytocompounds" often contain restorative biological properties such as anti-cancerous, antioxidant, and anti-inflammatory effects. Thus, understanding how they interact with the body can lead to potential treatment strategies against major immune disorders.
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A team of researchers at Tokyo University of Science, led by Prof Chiharu Nishiyama, has been working this direction for the past several years, to identify novel active components in functional foods and understand their effects on the body. Their efforts have now led to success: In their latest study, published in In patients with IBD, the gastrointestinal tract lining contains long-lasting ulcers caused by chronic inflammation due to an elevated immune response in the body. This involves the excessive production of immune system-related molecules called "cytokines." Moreover, two types of immune cells, "dendritic cells" (DCs) and "T cells," are also involved: at the onset of an immune response, DCs produce inflammatory cytokines and activate T cells to initiate a defense response. These processes together form a complex pathway that result in a "hyper" immune response. Thus, to find an effective compound that can suppress the immune system, it was crucial to test it on this population of immune cells.Thus, to begin with, the scientists studied the effects of a range of plant-derived compounds on DC-mediated T cell proliferation. Their initial research led them to PSB, which showed stronger immunosuppressive activity than the other candidates. When they dug deeper, they found that PSB treatment prevents T cells from differentiating into Th1 and Th17 (subtypes of T cells that elevate the immune response) while increasing their differentiation into regulatory T cells (another subtype known to inhibit inflammation). They also revealed that PSB treatment inhibits inflammatory cytokine production from DCs by attenuating the DNA-binding activity of a crucial transcription factor PU.1. When they further tested PSB in mice with IBD, they found that oral intake of PSB improved symptoms of IBD. Thus, the study confirmed that PSB is an extremely promising anti-inflammatory agent to fight IBD. Not just this -- it is easily absorbed by the body, making it an ideal drug candidate!Through these findings, the scientists have ushered in new possibilities for the treatment of not just IBD but also other inflammatory disorders. Dr Yashiro concludes, "For disease prevention, it is important to identify the beneficial components in foods and to understand the underlying mechanism by which immune responses and homeostasis are modulated in body. Our findings showed that PSB possesses a strong immunosuppressive property, paving the way for a new, natural treatment for IBD."
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923124732.htm
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Likely molecular mechanisms of SARS-CoV-2 pathogenesis are revealed by network biology
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Viral and bacterial pathogens wield pathogenic or virulent proteins that interact with high-value targets inside human cells, attacking what is known as the host interactome. The host interactome is the network map of all the protein-protein interactions inside cells.
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Such networks have been studied in organisms as diverse as plants, humans and roundworms, and they show a similarity to social networks like Facebook or airline route maps. In Facebook, a few people will have a huge number of friend connections, some will have many, and a vast majority will have much fewer. Similarly, airlines have a few hubs that many passengers pass through on the way to their destinations.Host interactomes show a limited number of high-powered hubs -- where a protein has a large number of connections -- and a limited number of important bottlenecks, which are sites with a large number of short paths to a node. These are key targets for pathogens as they seek to seize control of the infected cell, so it can rewire the cell's flow of information and cause disease.University of Alabama at Birmingham researchers, led by Shahid Mukhtar, Ph.D., associate professor of biology in the UAB College of Arts and Sciences, have now built an interactome that includes the lung-epithelial cell host interactome integrated with a SARS-CoV-2 interactome. Applying network biology analysis tools to this human/SARS-CoV-2 interactome has revealed potential molecular mechanisms of pathogenesis for SARS-CoV-2, the virus responsible for the COVID-19 pandemic. The UAB research, published in the journal So far in 2019, the SARS-CoV-2 virus has killed nearly 1 million people worldwide and 200,000 in the United States.The UAB researchers took many steps to generate the Calu-3-specific human-SARS-CoV-2 interactome, or CSI, that was the starting point for their network biology analyses.They began from a comprehensive human interactome of experimentally validated protein-protein interactions, posted online in 2015, and then manually curated other protein-protein interactions from four subsequent interactome studies. The resulting human interactome contained 18,906 nodes and 444,633 "edges" -- the term for the links between protein nodes.From two 2020 studies, the researchers compiled an exhaustive list of 394 host proteins that interact with the novel human coronavirus; these host proteins were called SARS-CoV-2 interacting proteins, or SIPs. The SIPs included 332 human proteins associated with the peptides of SARS-CoV-2 and 62 host proteins interacting with the viral factors of other human coronaviruses, including SARSCoV and MERS-CoV, the causes of SARS and MERS, which could also aid understanding the molecular pathogenesis of SARS-CoV-2By querying these 394 SIPs in the human interactome, they generated a subnetwork of 12,852 nodes and 84,100 edges that covered first and second neighbors of the 373 SIPs.Finally, they filtered these interactions in the context of temporal changes during COVID-19 infection, using a high-resolution temporal transcriptome derived from cultured human airway epithelial cells, or Calu-3, treated with SARSCoV and SARS-CoV-2 over time. Integrating this Calu-3 expression data with the SIPs-derived protein-protein interaction subnetwork resulted in a Calu-3-specific human-SARS-CoV-2 interactome, or CSI, that contained 214 SIPs interacting with their first and second neighbors, and forming a network of 4,176 nodes and 18,630 edges.The CSI had a power law degree distribution with a few nodes harboring increased connectivity compared to a random network, and thus exhibited properties of a scale-free network, similar to the other, previously generated human-viral interactomes.The robust, high-quality CSI was then further utilized for network-aided architectural and functional pathway analyses.Topological clustering and pathway enrichment analysis showed that the SARS-CoV-2 virus attacks central nodes of the host-viral network that participate in core functional pathways. Network centrality analyses discovered 33 high-value SARS-CoV-2 targets for possible drug therapy; these targets are possibly involved in viral entry, proliferation and survival to establish infection and facilitate disease progression. A probabilistic modeling framework elucidated critical regulatory circuitry and molecular events pertinent to COVID-19, particularly the host modifying responses and cytokine storm."In summary," Mukhtar said, "our integrative network topology analyses led us to elucidate the underlying molecular mechanisms and pathways of SARS-CoV-2 pathogenesis." Mukhtar's lab continues to work on network medicine and artificial intelligence to battle COVID-19 and other human inflammatory diseases.Co-first authors of the study, "Integrative network biology framework elucidates molecular mechanisms of SARS-CoV-2 pathogenesis," are graduate students Nilesh Kumar and Bharat Mishra, UAB Department of Biology.Other co-authors, along with Mukhtar, are Adeel Mehmood, UAB departments of Biology and Computer Science; and Mohammad Athar, Department of Dermatology, UAB School of Medicine.Support came from National Science Foundation grant IOS-1557796 and National Institutes of Health grant ES030246.
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923124718.htm
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Customizable synthetic antibiotic outmaneuvers resistant bacteria
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Antibiotic resistance is one of the world's most urgent public health threats. In the United States alone, tens of thousands of deaths result each year from drug-resistant strains of common bacteria such as Staphylococcus aureus and Enterococcus faecium, which can cause virtually untreatable hospital-acquired infections. Perilously few new classes of antibiotics are being developed to fight infections that have become resistant to traditional treatments, and bringing any new drugs to market could take decades.
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Researchers at UC San Francisco are tackling antibiotic resistance using a different approach: redesigning existing antibiotic molecules to evade a bacterium's resistance mechanisms. By devising a set of molecular LEGO pieces that can be altered and joined together to form larger molecules, the researchers have created what they hope is the first of many "rebuilds" of drugs that had been shelved due to antibiotic resistance. The research was published September 23, 2020, in "The aim is to revive classes of drugs that haven't been able to achieve their full potential, especially those already shown to be safe in humans," said Ian Seiple, PhD, an assistant professor in the UCSF School of Pharmacy's Department of Pharmaceutical Chemistry and the Cardiovascular Research Institute (CVRI), and lead author on the paper. "If we can do that, it eliminates the need to continually come up with new classes of drugs that can outdo resistant bacteria. Redesigning existing drugs could be a vital tool in this effort."In work descibed in the new Streptogramins disable bacteria by gumming up the works in the bacterial ribosome, making it impossible for the bacteria to make proteins. But bacteria resistant to streptogramins produce proteins called virginiamycin acetyltransferases (Vats), which recognize these antibiotics when they enter the bacterial cell. The Vats grab the drug and chemically deactivate it before it can bind to the ribosome, rendering it useless.Streptogramins, like most other antibiotics, are derived from naturally occurring antibiotic compounds produced by other organisms (usually bacteria) that are then tweaked to optimize their performance in the human body. Seiple figured that there must also be a way to make further changes to the drug molecule that would allow it to evade capture by the Vat proteins.Seiple set out to build new streptogramins from the ground up, rather than modifying existing structures. To make the building process easier, Qi Li, PhD, a postdoctoral fellow in the Seiple lab and co-first author on the paper, created seven molecular modules that can be tweaked as needed to build a set of variations on the streptogramin molecule."This system allows us to manipulate the building blocks in ways that wouldn't be possible in nature," said Seiple. "It gives us an efficient route to re-engineering these molecules from scratch, and we have a lot more latitude to be creative with how we modify the structures."Once Seiple and Li had their building blocks, the next step was to get a molecular-level view of the chemistry involved in order to better understand how to modify and piece together those molecular LEGOs.For that, Seiple teamed up with Fraser, who specializes in creating visual models of biological molecules."My lab's contribution was to say, 'Now that you've got the seven pieces, which one of them should we modify and in what way?'" said Fraser, whose work on the project was supported by the inaugural Sanghvi-Agarwal Innovation Award.To get answers to that question, Jenna Pellegrino, a graduate student in the Fraser Group and co-first author on the paper, used two complementary techniques, cryo-electron microscopy and x-ray crystallography, to create three-dimensional pictures of the drug at near-atomic resolution, as well as its target the bacterial ribosome, and its nemesis, the Vat protein.Using the models, Li, Pellegrino, Seiple, and Fraser could see which parts of the streptogramin molecule are essential to the antibiotic's function. Then Li was free to fiddle with the drug's non-essential regions to find modifications that prevented Vats from interacting with the drug while still allowing it to bind to its ribosomal targets and disable the bacterium.The team found that two of the seven building blocks seemed to offer potentially interesting sites for modification. They made variations of the drug that contained tweaks in those regions and found that these variations had activity in dozens of strains of pathogenic bacteria. The researchers also tested their most promising candidate against streptogramin-resistant S. aureus in infected mice, and found it was over 10 times more effective than other streptogramin antibiotics.Seiple points out that the knowledge gained through these collaborative experiments can be applied to modifying many other antibiotics."We learned about mechanisms that other classes of antibiotics use to bind to the same target," he said. "In addition, we established a workflow for using chemistry to overcome resistance to antibiotics that haven't reached their potential."Seiple will continue to refine these synthetic streptogramins and then hopes to move the work to the private sector where the reengineered antibiotics could be further developed and tested in human trials. He and Fraser plan to continue working together on reviving other antibiotics that have been shelved because of microbial resistance, refining a set of tools that can help researchers stay one step ahead of bacterial evolution."It's a never-ending arms race with bacteria," said Fraser. "But by studying the structures involved -- before resistance arises -- we can get an idea of what the potential resistance mechanisms will be. That insight will be a guide to making antibiotics that bacteria can't resist."
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923124653.htm
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Unraveling the genome in 3D-space
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The cells that make up our body are tiny, each of them measuring only micrometers in diameter. The ensemble of chromosomal DNA molecules that encode the genome, on the other hand, measures almost 2 meters. In order to fit into cells, chromosomal DNA is folded many times. But the DNA is not merely squeezed into the nucleus in a random manor but folded in a specific and highly regulated structure. The spatial organization of chromosomal DNA enables regulated topological interactions between distant parts, thereby supporting proper expression, maintenance, and transport of the genome across cell generations.
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Breaks in our DNA, which can occur spontaneously or result from irradiation or chemical insults, can lead to severe problems since they foster mutations and can ultimately lead to cancer. But not every DNA break has disastrous consequences, since our cells have ingenious ways of repairing the damage. One of the main DNA repair pathways involves copying the missing information on the damaged DNA from the replicated sister chromatid. For this to occur, the two DNA molecules of sister chromatids need to come close together at the exact same genomic position. How the two DNA molecules are organized relative to each other to support this important repair pathway, however, has remained unclear.The team around Daniel Gerlich developed a method that solves this problem. "Current methods to map the folding of DNA have a serious blind spot: They are not able to distinguish identical copies of DNA molecules. Our approach to solve this was to label DNA copies in a way such that we can discriminate them by DNA sequencing" explains Michael Mitter, doctoral student in Dr. Gerlich's lab and first author of the current publication in Nature. Using this approach, the researchers were able to create the first high resolution map of contact points between replicated chromosomes."With this new method, we can now study the molecular machinery regulating the conformation of sister chromatids, which will provide insights into the mechanics underlying the repair of DNA breaks and the formation of rod-shaped chromosomes in dividing cells, which is required for proper transport the genome to cell progeny," says Daniel Gerlich about the project, which is financed by the Vienna Science and Technology Fund (WWTF) and was a fruitful collaboration of several research groups at the Vienna BioCenter, including the Ameres and Goloborodko labs at IMBA, and the Peters lab at the neighboring Institute of Molecular Pathology (IMP).
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923124621.htm
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Structural biology: Ribosomes and Russian dolls
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Maturation of the ribosome is a complex operation. Work by an Ludwig-Maximilians-Universitaet (LMU) in Munich team now shows that the 90S precursor of the small 40S subunit undergoes a 'molting' process, during which it progressively discards its outermost components.
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Protein synthesis, programmed by the genetic information encoded in the DNA, is perhaps the most crucial process that takes place in biological cells. Proteins are indispensable for all organisms, because they are responsible for performing a vast range of biological functions. Indeed, the molecular machines that put proteins together -- which are known as ribosomes -- are themselves partly made up of specific proteins. The second vital ingredient of every ribosome is a small set of specific RNAs, which serve as scaffolds to which ribosomal proteins can be specifically attached. The synthesis of ribosomes is therefore an extremely complex, multistep process, which includes both assembly and maturation stages. This complexity explains why many of the details of the whole operation are still incompletely understood. Now a group of researchers led by Professor Roland Beckmann at LMU's Gene Center has obtained new insights into the maturation phase that gives rise to the small subunit of the functional ribosome in brewer's yeast. The study, which was carried out in collaboration with colleagues based in Heidelberg, appears in the leading journal In the cells of higher organisms, mature ribosomes are composed of two distinct subunits, each of which contains a long ribosomal RNA (rRNA) molecule (called 18S in the small and 25S in the large subunit in yeast). The subunits interact with one another and with the messenger RNAs that program the synthesis of each protein. In yeast, the smaller 40S subunit is derived from a much larger precursor complex called the 90S pre-ribosome. The 90S precursor particle contains a single (35S) RNA molecule. The RNAs ultimately associated with each mature subunit are produced by the removal of specific internal and end-fragments. However, one of the segments the RNA found in the 90S precursor plays an important role in ensuring that the mature 18S rRNA in the 40S subunit folds into its correct three-dimensional form.How the processing of the 35S rRNA is achieved has so far been unclear. The general idea was that, as the 40S subunit matures, the processing steps that give rise to the 18S rRNA take place, and the mature 40S particle eventually 'emerges' from the 90S precursor. The new study adds new details, which reveal that the process is rather more complicated than that. For a start, a specific enzyme (Dhr1) is required to ensure that the initial cleavage of the 35S rRNA precursor occurs at the right position. Dhr1 first exposes the cleavage site, enabling it to interact with the enzyme Utp24, which cuts the correct fragment off one end of the 35S rRNA.In addition, the 'emergence' of the 40S subunit entails an ordered series of reactions in which the outer shell of the 90S particle is progressively dissociated from the 40S. "It doesn't just go plop," Beckmann remarks. The process is actually reminiscent of the molting of an insect -- shedding of the integument takes place layer by layer. "It's rather like those Russian dolls. When you open one, you find a smaller one nestled inside," says Beckmann. -- And with the aid of cryo-electron microscopy, the specialists in Munich were able to discriminate between the different three-dimensional complexes characteristic of each step in the process. Earlier biochemical experiments performed by a team at the Center for Biochemistry at Heidelberg University (BZH), led by Professor Ed Hurt, had already cast doubt on the previous en bloc model by providing evidence for the idea that shedding of the outer layers of the 90S particle took place stepwise.The elucidation of such mechanisms is not only of interest from the point of view of basic research. As Beckmann points out, more and more disorders have been shown to be related to a lack of intact ribosomes. When errors occur in the assembly and maturation of these delicate and intricate molecular machines, they may ultimately lead to a relative dearth of ribosomes, which then perturbs the delicate equilibrium between protein synthesis and degradation. Among the resulting syndromes are diverse forms of muscle atrophy, growth anomalies, anemias and certain cancers.
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Biotechnology
| 2,020 |
September 23, 2020
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https://www.sciencedaily.com/releases/2020/09/200923124619.htm
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New mouse model of tau propagation
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Accumulation of assembled tau protein in the central nervous system is characteristic of Alzheimer's disease and several other neurodegenerative diseases, called tauopathies. Recent studies have revealed that propagation of assembled tau is key to understanding the pathological mechanisms of these diseases. Mouse models of tau propagation are established by injecting human-derived tau seeds intracerebrally; nevertheless, these have a limitation in terms of regulation of availability of human samples. To date, no study has shown that synthetic assembled tau induce tau propagation in non-transgenic mice.
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A study by researchers from TMIMS confirms that dextran sulphate, a sulphated glycosaminoglycan, induces the assembly of recombinant tau protein into filaments in vitro. As compared to tau filaments induced by heparin, those induced by dextran sulphate were shorter in length, and showed higher thioflavin T fluorescence and lower resistance to guanidine hydrochloride, which suggest that the two types of filaments have distinct conformational features. Unlike other synthetic filament seeds, intracerebral injection of dextran sulphate-induced assemblies of recombinant tau caused aggregation of endogenous murine tau in wild-type mice. Tau accumulation stained with a antibody against phosphorylated tau (AT8) was present at the injection site one month after injection, from where it spread to anatomically connected regions. Induced tau pathologies were also stained by anti-tau antibodies AT100, AT180, 12E8, PHF1, anti-pS396 and anti-pS422. They were thioflavin- and Gallyas-Braak silver-positive, indicative of amyloid. In biochemical analyses, accumulated sarkosyl-insoluble and hyperphosphorylated tau was observed in the injected mice.In conclusion, this study revealed that intracerebral injection of synthetic full-length wild-type tau seeds prepared in the presence of dextran sulphate caused tau propagation in non-transgenic mice. These findings establish that propagation of tau assemblies does not require tau to be either mutant and/or overexpressed.
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Biotechnology
| 2,020 |
September 22, 2020
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https://www.sciencedaily.com/releases/2020/09/200922135732.htm
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'Cheater mitochondria' may profit from cellular stress coping mechanisms
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Cheating mitochondria may take advantage of cellular mechanisms for coping with food scarcity in a simple worm to persist, even though this can reduce the worm's wellbeing.
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These findings, published today in Mitochondria are energy-producing units within cells that likely evolved from bacteria. They have their own DNA, take in resources from cells, and in exchange provide the cell with energy. But some so-called 'cheater mitochondria' have harmful DNA mutations that may reduce their energy output and harm the organism. Why these cheater mitochondria persist despite their harm to the larger organism is not currently clear."Cooperation and cheating are widespread evolutionary strategies," says lead author Bryan Gitschlag, a PhD student at the Department of Biological Sciences, Vanderbilt University, Nashville, Tennessee. "While cheating confers an advantage to individual entities within a group, competition between groups favours cooperation."Gitschlag and his colleagues studied the roundworm Caenorhabiditis elegans to see how competing evolutionary pressures within its cells and in its environment might enable the cheater mitochondria to persist.They measured the levels of cheater and typical mitochondria in the worm's cells. They found that, within the cells, a protein called DAF-16, which helps cells to survive stress, is necessary for cheater mitochondria to multiply. When the worms face food shortages, cheater mitochondria become more harmful to their hosts, but only in those lacking DAF-16. "This shows that food scarcity can strengthen evolutionary selection against worms carrying cheater mitochondria, but DAF-16 protects them from it," Gitschlag explains.The results suggest that competing selection pressures within an organism and in its environment may shed light on why selfishness and cooperation often exist side-by-side among populations."The ability to cope with scarcity can promote group-level tolerance to cheating, inadvertently prolonging cheater persistence," says senior author Maulik Patel, Assistant Professor of Biological Sciences at Vanderbilt University."As selfish mitochondrial genomes are implicated in numerous disorders, and cheating is a widespread evolutionary strategy, it will be interesting to apply our methods to study a broader collection of cheating variants and host species. This could allow us to better understand the development of mitochondrial disorders or the evolutionary principles underlying cooperation and cheating," Patel concludes.
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Biotechnology
| 2,020 |
September 22, 2020
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https://www.sciencedaily.com/releases/2020/09/200922112251.htm
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Mechanism that causes cell nuclei to grow
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By far the most important process in cell development is how cells divide and then enlarge in order to multiply. A research team headed by Freiburg medical scientist Prof. Dr. Robert Grosse has now discovered that bundled fibers of actin within a cell nucleus play an important part in how they enlarge after division. Fibers of the structural protein actin stabilize the outer form of the cell and transport substances into a cell. The mechanisms that influence the growth of the cell nucleus after division were less well known by scientists. The researchers have published their results in the journal
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After dividing, cell nuclei have to grow in order to reorganize and unpack the genetic information in chromatin, the basic genetic material, and so process and read it. With this work the scientists show that bundled fibers of actin -- which are normally responsible for exercising force -- work within the cell to expand the nucleus. Using a video microscope the researchers have measured in living cells how cell nuclei enlarge immediately after division. In order to observe the fibers of actin and skeletal structures in the cell nucleus, they also used a high-resolution super-resolution microscope.In the future Grosse and his team want to clarify whether mechanical forces work within the cell nucleus to re-organize them to sort the genetic information. If so, this process could for example be disrupted or changed in tumor cells or play a part in stem cells.
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Biotechnology
| 2,020 |
September 22, 2020
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https://www.sciencedaily.com/releases/2020/09/200922112234.htm
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Chemists make cellular forces visible at the molecular scale
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Scientists have developed a new technique using tools made of luminescent DNA, lit up like fireflies, to visualize the mechanical forces of cells at the molecular level.
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"Normally, an optical microscope cannot produce images that resolve objects smaller than the length of a light wave, which is about 500 nanometers," says Khalid Salaita, Emory professor of chemistry and senior author of the study. "We found a way to leverage recent advances in optical imaging along with our molecular DNA sensors to capture forces at 25 nanometers. That resolution is akin to being on the moon and seeing the ripples caused by raindrops hitting the surface of a lake on the Earth."Almost every biological process involves a mechanical component, from cell division to blood clotting to mounting an immune response. "Understanding how cells apply forces and sense forces may help in the development of new therapies for many different disorders," says Salaita, whose lab is a leader in devising ways to image and map bio-mechanical forces.The first authors of the paper, Joshua Brockman and Hanquan Su, did the work as Emory graduate students in the Salaita lab. Both recently received their PhDs.The researchers turned strands of synthetic DNA into molecular tension probes that contain hidden pockets. The probes are attached to receptors on a cell's surface. Free-floating pieces of DNA tagged with fluorescence serve as imagers. As the unanchored pieces of DNA whizz about they create streaks of light in microscopy videos.When the cell applies force at a particular receptor site, the attached probes stretch out causing their hidden pockets to open and release tendrils of DNA that are stored inside. The free-floating pieces of DNA are engineered to dock onto these DNA tendrils. When the florescent DNA pieces dock, they are briefly demobilized, showing up as still points of light in the microscopy videos.Hours of microscopy video are taken of the process, then speeded up to show how the points of light change over time, providing the molecular-level view of the mechanical forces of the cell.The researchers use a firefly analogy to describe the process."Imagine you're in a field on a moonless night and there is a tree that you can't see because it's pitch black out," says Brockman, who graduated from the Wallace H. Coulter Department of Biomedical Engineering, a joint program of Georgia Tech and Emory, and is now a post-doctoral fellow at Harvard. "For some reason, fireflies really like that tree. As they land on all the branches and along the trunk of the tree, you could slowly build up an image of the outline of the tree. And if you were really patient, you could even detect the branches of the tree waving in the wind by recording how the fireflies change their landing spots over time.""It's extremely challenging to image the forces of a living cell at a high resolution," says Su, who graduated from Emory's Department of Chemistry and is now a post-doctoral fellow in the Salaita lab. "A big advantage of our technique is that it doesn't interfere with the normal behavior or health of a cell."Another advantage, he adds, is that DNA bases of A, G, T and C, which naturally bind to one another in particular ways, can be engineered within the probe-and-imaging system to control specificity and map multiple forces at one time within a cell."Ultimately, we may be able to link various mechanical activities of a cell to specific proteins or to other parts of cellular machinery," Brockman says. "That may allow us to determine how to alter the cell to change and control its forces."By using the technique to image and map the mechanical forces of platelets, the cells that control blood clotting at the site of a wound, the researchers discovered that platelets have a concentrated core of mechanical tension and a thin rim that continuously contracts. "We couldn't see this pattern before but now we have a crisp image of it," Salaita says. "How do these mechanical forces control thrombosis and coagulation? We'd like to study them more to see if they could serve as a way to predict a clotting disorder."Just as increasingly high-powered telescopes allow us to discover planets, stars and the forces of the universe, higher-powered microscopy allows us to make discoveries about our own biology."I hope this new technique leads to better ways to visualize not just the activity of single cells in a laboratory dish, but to learn about cell-to-cell interactions in actual physiological conditions," Su says. "It's like opening a new door onto a largely unexplored realm -- the forces inside of us."Co-authors of the study include researchers from Children's Healthcare of Atlanta, Ludwig Maximilian University in Munich, the Max Planck Institute and the University of Alabama at Birmingham. The work was funded by grants from the National Institutes of Health, the National Science Foundation, the Naito Foundation and the Uehara Memorial Foundation.
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Biotechnology
| 2,020 |
September 22, 2020
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https://www.sciencedaily.com/releases/2020/09/200922083912.htm
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New drug candidate found for hand, foot and mouth disease
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A study appearing next week in the journal
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The compound of interest is a small molecule that binds to RNA, the virus's genetic material, and changes its 3-D shape in a way that stops the virus from multiplying without harming its human host.There are currently no FDA-approved drugs or vaccines for enterovirus 71, which affects hundreds of thousands of children each year, particularly in Southeast Asia. While most people get better within 7 to 10 days after suffering little more than a fever and rash, severe cases can cause brain inflammation, paralysis and even death.The work could pave the way for new treatments for other viral infections as well, says a team of scientists at Duke University, Case Western Reserve University and Rutgers University.Traditionally, most drugs are designed to bind to proteins to block or disrupt their role in causing disease. But much of the genome in humans and their microbial pathogens doesn't code for proteins, which means that only a fraction of their genetic material is targeted by existing drugs."For diseases that don't have good treatments, maybe the problem is we've been targeting the wrong thing," said co-author Amanda Hargrove, associate professor of chemistry at Duke.Instead of targeting proteins, Hargrove and others are looking for small molecules that target RNA, which most drug discovery programs have overlooked.When a virus like enterovirus 71 (or SARS-CoV-2, the virus that causes COVID-19) infects a human cell, it injects its RNA into the cell, hijacking the internal machinery to make copies of itself that eventually burst out to infect neighboring cells.Previous work on enterovirus 71 singled out one part of its RNA structure that helps the virus co-opt the host machinery it needs to replicate. This RNA region folds over on itself to form a hairpin, with a bulge in the middle where unpaired nucleotides balloon out to one side.If a drug can be developed to inhibit this region, the researchers say, we might be able to block the virus before it has a chance to spread.For the current study, Hargrove and colleagues screened a library of some 30 small molecules, looking for ones that bind tightly to the bulge and not other sites in the virus's RNA.RNA is a wiggly molecule; when it binds to other molecules such as host proteins or small molecule drugs it takes on different 3-D shapes.The researchers identified one molecule, dubbed DMA-135, that enters infected human cells and attaches itself to the surface of the bulge, creating a kink in this region.This shape change, in turn, opens access to another molecule -- a human repressor protein that blocks the "reading out" of the virus's genetic instructions, stopping viral growth in its tracks.In an experiment, the researchers were able to use the molecule to stop the virus from building up inside human cell cultures in the lab, with bigger effects at higher doses.Hargrove says it would take at least five years to move any new drug for hand, foot and mouth disease from the lab to medicine cabinets. Before their small molecule could reach patients, the next step is to make sure it's safe and effective in mice.In the meantime, the researchers are building on their success with enterovirus 71 and looking at whether RNA-targeting small molecules could be used to tackle other RNA viruses too, including SARS-CoV-2.This research was supported by grants from the National Institutes of Health (U54 AI150470, R35 GM124785, R01 GM126833).
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Biotechnology
| 2,020 |
September 21, 2020
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https://www.sciencedaily.com/releases/2020/09/200921152617.htm
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CRISPR-based malaria testing on-the-fly
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To achieve the goal of eradicating malaria set by the World Health Organization (WHO)'s Global Malaria Control Programme, it is critical that all local transmission of malaria parasites in defined geographic areas is eliminated. One important cornerstone on this path is the development of rapid, sensitive and species-specific diagnostic capabilities that are useful in the low-resource settings (LRSs) of many areas with endemic malaria.
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Currently, the presence of the four major malaria-causing Plasmodium species P. falciparum, P. vivax, P. ovale, and P. malariae is determined by microscopic analysis of blood samples in which parasites can be detected in red blood cells, or with so-called rapid diagnostic tests for specific Plasmodium proteins (antigens)."Unfortunately, available rapid diagnostic approaches cannot distinguish all four Plasmodium species from one other, which can be important to initiate the definitive course of treatment, and, most importantly, they are ineffective for detecting low numbers of Plasmodium parasites in asymptomatic individuals," said Nira Pollock, M.D., Ph.D., Associate Medical Director of Boston Children's Hospital's Infectious Diseases Diagnostic Laboratory and Associate Professor of Pathology and Medicine at Harvard Medical School. "These "asymptomatic carriers" are silent reservoirs for ongoing transmission by malaria-spreading mosquitoes and extremely important for ongoing global efforts to eradicate malaria," added Jeffrey Dvorin, M.D., Ph.D., Associate Professor of Pediatrics at Harvard Medical School and Senior Associate Physician in Infectious Diseases at Boston Children's Hospital.Now, a multi-disciplinary research collaboration which was led by Wyss Core Faculty member James Collins, Ph.D. at Harvard's Wyss Institute for Biologically Inspired Engineering and the Massachusetts Institute of Technology (MIT), and assembled by clinical fellow Rose Lee, M.D., MSPH, which also included Pollock and Dvorin, created a field-applicable, ultrasensitive diagnostic assay that specifically detects DNA sequences from all Plasmodium species in symptomatic and asymptomatic malaria. The new malaria diagnostic method combines an optimized 10-minute rapid sample preparation protocol with the CRISPR-based SHERLOCK system to enable highly specific and sensitive Plasmodium detection in another 60 minutes in simple reporter devices. It is published in PNAS."This field-ready SHERLOCK diagnostic malaria assay surpasses the sensitivity and specificity requirements set by the WHO for a desired test that can be used to detect low parasite density in asymptomatic carriers of all major Plasmodium species," said Wyss Founding Core Faculty member James Collins, Ph.D. "Its highly streamlined design could provide a viable solution to the present diagnostic bottleneck on the path to eliminate malaria, and more generally enabling malaria surveillance in low-resource settings." Collins is a lead of the Institute's Living Cellular Devices Focus Area, and also the Termeer Professor of Medical Engineering & Science at MIT.The research team demonstrated their engineered SHERLOCK (short for Specific High-sensitivity Enzymatic Reporter unLOCKing) assay to be capable of detecting less than two parasites per microliter of blood, the WHO's suggested "limit of detection" (LOD) for a test with broad utility in endemic areas. Showing the assay's clinical potential by analyzing clinical samples containing P. falciparum and P. vivax species, they called out them out with 100% sensitivity, by correctly identifying true positive samples, and 100% specificity, by also correctly identifying samples lacking a certain Plasmodium species in true negative samples. Near 100% sensitivity and specificity are key attributes of diagnostic assays to be used in real-world testing. Moreover, the assay is designed so that it can also determine the presence of frequently mutated P. falciparum strains that have lost their HRP2 antigen and thus escape the detection by common rapid diagnostic tests.Collins' group at the Wyss Institute and MIT co-developed the SHERLOCK technology with Feng Zhang's group at the Broad Institute. It was licensed to Sherlock Biosciences, a startup that used it to create a rapid molecular diagnostic for other disease applications, and recently received an emergency use authorization from the FDA for its COVID-19 rapid diagnostic.Other methods have been developed that, like the new SHERLOCK assay, amplify and detect the DNA (or RNA) nucleic acid material of Plasmodium species. However, to date, these methods remain limited by their need for expensive laboratory equipment, as in the case of polymerase chain reaction (PCR)-based methods, complicated sample preparation techniques, and trained personnel or, as in the case of simpler "isothermal" amplification methods performed at a single temperature, they have not shown the desired sensitivities in the field.The SHERLOCK malaria assay takes advantage of the CRISPR-Cas12a enzyme which can be programmed to become active with a so-called guide RNA that binds to a specific target nucleic acid target sequence, in this case a sequence from one of the four Plasmodium species. Activated Cas12a then non-specifically cleaves any single-stranded DNA strand in its vicinity with an extremely high turn-over rate of about 1,250 collateral cleavage reactions per second. The researchers leveraged this amplifying activity in their assay by integrating it with an optimized sample preparation, which does not require a specific nucleic extraction step like some other nucleic acid amplification tests (NAATs), and isothermal amplification of specific Plasmodium DNA and RNA sequences at the front end. Guide RNAs that recognize species-specific motifs in the amplified Plasmodium sequences then unleash Cas12a activity, which collaterally attacks single stranded DNA reporter sequences whose cleavage products help signal the presence of the pathogen-specific nucleic acids. At the backend of the assay, the signal is engineered to either cause a change in fluorescence in a hand-held device, or specific band on a lateral flow strip commonly used in clinical point-of-care devices."Importantly, the assay is compatible with different sample types, such as whole blood, plasma, serum, and dried blood, and all components required for amplification, Cas12a activation and signal generation can be lyophilized in a single test tube to work together in a one-pot-reaction after they are reconstituted and mixed with patient sample," said first-author Rose Lee, a clinical fellow in Collins' group and Boston Children's Hospital with a strong interest in infectious disease diagnostics and was instrumental in assembling the multi-disciplinary team together with Collins. "This avoids having to depend on a functional cold-chain and allows testing to be performed in low-resource settings with minimal expertise.""The collaboration's molecular assay for malaria diagnosis points the way in which Wyss Institute capabilities in the synthetic biology field, when paired with expertise in infectious disease biology and epidemiology, could change the course of truly debilitating diseases that paralyze large populations around the globe," said the Wyss Institute's Founding Director Don Ingber, M.D., Ph.D., who is also the Judah Folkman Professor of Vascular Biology at Harvard Medical School and Boston Children's Hospital, and Professor of Bioengineering at the Harvard John A. Paulson School of Engineering and Applied Sciences.
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Biotechnology
| 2,020 |
September 21, 2020
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https://www.sciencedaily.com/releases/2020/09/200921111704.htm
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Technique permits convenient, precise optical imaging of individual proteins
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Often considered the workhorses of the body, proteins are among the most important biomolecules critical to life processes. They provide structural foundation for cells and tissues and perform a dizzying array of tasks, from metabolizing energy and helping cells communicate with one another to defending the body from pathogens and guiding cell division and growth.
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Because protein dysfunction is implicated in so many serious diseases, proteins are the primary targets for most therapeutic drugs.In a new study, Shaopeng Wang and his colleagues describe a method for examining proteins in keen detail. To do this, his group makes clever use of a phenomenon known as surface plasmon resonance (SPR), incorporating it into an innovative type of microscope.While SPR has been a powerful technique for investigating the world of the very small, including the interactions of bacteria and viruses, the study marks the first occasion when SPR has successfully been used to image single molecules, in this case, proteins. The new method is known as plasmon scattering microscopy.According to Wang: "The race to develop this technology actually started 20 years ago." Along with lead author NJ Tao* the group calculated that a modified form of SPR should have the sensitivity to resolve single proteins, though much preparatory work was required to make this a reality.Wang is a researcher in the Biodesign Center for Bioelectronics and Biosensors. The new research appears in the advanced online addition of the journal Using SPR allows researchers to investigate the dynamics of cell surface proteins -- primary targets for drug design -- which are particularly challenging to observe using X-ray crystallography or NMR spectroscopy, the two conventional techniques usually brought to bear to characterize proteins.But what is a surface plasmon? "One property of metal is that you have a lot of free electrons," Wang says, referring to electrons not bound to atoms. "When the condition of incident light on these electrons is just right, the energy in the light causes these electrons to resonate. These oscillating electrons produce a wave across the metal surface. This is surface plasmon resonance."In order to detect the binding of an analyte molecule (like a protein) to a receptor molecule using SPR, the receptor molecule is usually immobilized on the sensor surface and the analyte molecule is added to an aqueous solution. Polarized light is typically directed under surface of a thin gold film, where surface plasmons are generated at a particular angle of the incident light. The surface confinement of light by the surface plasmon is seen as a decrease in intensity of reflected light.When protein molecules bind to immobilized receptor molecules, the refractive index at the gold surface changes, altering the surface plasmon resonance condition and producing an increase in signal intensity.To refine and calibrate the system, the researchers first observed binding events using polystyrene nanoparticles, whose size can be precisely controlled. The nanoparticles also have the advantage of producing higher contrast, aiding their detection by SPR. Using smaller and smaller nanoparticles allowed the group to reach the tiny dimensions of a biological protein.To achieve such impressive resolution, the researchers used a variant of the SPR technique, detecting light on the protein binding events from above, rather than below, which dramatically eliminates background noise, producing a crisp image. Because bound proteins scatter the SPR light in all directions, detection from the top avoids the reflected light, greatly improving image quality.Wang likens the effect to seeing stars against the background curtain of darkness, whereas stars are invisible to the eye against the noisy background of daylight. Detection of single proteins can be realized without a very powerful light source, since SPR produces strong enhancement to the light field near the sensor surface, clarifying the protein signal.By homing in on protein binding affinity, one of the key parameters critical for the design of safer, more effective drugs, the new SPR technique should have a bright future in the biomedical arena as well as shedding new light on foundational issues at the molecular scale.*NJ Tao, directed the Biodesign Center for Bioelectronics and Biosensors, prior to his unexpected death in March of this year. He was a leading figure in the advancement of techniques for nanoscale measurements in areas including molecular electronics, optical imaging and biosensing.
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Biotechnology
| 2,020 |
September 18, 2020
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https://www.sciencedaily.com/releases/2020/09/200918113347.htm
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Biologists create new genetic systems to neutralize gene drives
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In the past decade, researchers have engineered an array of new tools that control the balance of genetic inheritance. Based on CRISPR technology, such gene drives are poised to move from the laboratory into the wild where they are being engineered to suppress devastating diseases such as mosquito-borne malaria, dengue, Zika, chikungunya, yellow fever and West Nile. Gene drives carry the power to immunize mosquitoes against malarial parasites, or act as genetic insecticides that reduce mosquito populations.
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Although the newest gene drives have been proven to spread efficiently as designed in laboratory settings, concerns have been raised regarding the safety of releasing such systems into wild populations. Questions have emerged about the predictability and controllability of gene drives and whether, once let loose, they can be recalled in the field if they spread beyond their intended application region.Now, scientists at the University of California San Diego and their colleagues have developed two new active genetic systems that address such risks by halting or eliminating gene drives in the wild. On Sept.18, 2020 in the journal "One way to mitigate the perceived risks of gene drives is to develop approaches to halt their spread or to delete them if necessary," said Distinguished Professor Ethan Bier, the paper's senior author and science director for the Tata Institute for Genetics and Society. "There's been a lot of concern that there are so many unknowns associated with gene drives. Now we have saturated the possibilities, both at the genetic and molecular levels, and developed mitigating elements."The first neutralizing system, called e-CHACR (erasing Constructs Hitchhiking on the Autocatalytic Chain Reaction) is designed to halt the spread of a gene drive by "shooting it with its own gun." e-CHACRs use the CRISPR enzyme Cas9 carried on a gene drive to copy itself, while simultaneously mutating and inactivating the Cas9 gene. Xu says an e-CHACR can be placed anywhere in the genome."Without a source of Cas9, it is inherited like any other normal gene," said Xu. "However, once an e-CHACR confronts a gene drive, it inactivates the gene drive in its tracks and continues to spread across several generations 'chasing down' the drive element until its function is lost from the population."The second neutralizing system, called ERACR (Element Reversing the Autocatalytic Chain Reaction), is designed to eliminate the gene drive altogether. ERACRs are designed to be inserted at the site of the gene drive, where they use the Cas9 from the gene drive to attack either side of the Cas9, cutting it out. Once the gene drive is deleted, the ERACR copies itself and replaces the gene-drive."If the ERACR is also given an edge by carrying a functional copy of a gene that is disrupted by the gene drive, then it races across the finish line, completely eliminating the gene drive with unflinching resolve," said Bier.The researchers rigorously tested and analyzed e-CHACRs and ERACRs, as well as the resulting DNA sequences, in meticulous detail at the molecular level. Bier estimates that the research team, which includes mathematical modelers from UC Berkeley, spent an estimated combined 15 years of effort to comprehensively develop and analyze the new systems. Still, he cautions there are unforeseen scenarios that could emerge, and the neutralizing systems should not be used with a false sense of security for field-implemented gene drives."Such braking elements should just be developed and kept in reserve in case they are needed since it is not known whether some of the rare exceptional interactions between these elements and the gene drives they are designed to corral might have unintended activities," he said.According to Bulger, gene drives have enormous potential to alleviate suffering, but responsibly deploying them depends on having control mechanisms in place should unforeseen consequences arise. ERACRs and eCHACRs offer ways to stop the gene drive from spreading and, in the case of the ERACR, can potentially revert an engineered DNA sequence to a state much closer to the naturally-occurring sequence."Because ERACRs and e-CHACRs do not possess their own source of Cas9, they will only spread as far as the gene drive itself and will not edit the wild type population," said Bulger. "These technologies are not perfect, but we now have a much more comprehensive understanding of why and how unintended outcomes influence their function and we believe they have the potential to be powerful gene drive control mechanisms should the need arise."
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917181259.htm
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A scientific first: How psychedelics bind to key brain cell receptor
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Psychedelic drugs such as LSD, psilocybin, and mescaline cause severe and often long-lasting hallucinations, but they show great potential in treating serious psychiatric conditions, such as major depressive disorder. To fully investigate this potential, scientists need to know how these drugs interact with brain cells at the molecular level to cause their dramatic biological effects. Scientists at UNC-Chapel Hill and Stanford have just taken a big step in that direction.
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For the first time, scientists in the UNC lab of Bryan L. Roth, MD, PhD, and the Stanford lab of Georgios Skiniotis, PhD, solved the high-resolution structure of these compounds when they are actively bound to the 5-HT2A serotonin receptor (HTR2A) on the surface of brain cells.This discovery, published in "Millions of people have taken these drugs recreationally, and now they are emerging as therapeutic agents," said co-senior author Bryan L. Roth, MD, PhD, the Michael Hooker Distinguished Professor of Pharmacology at the University of North Carolina School of Medicine. "Gaining this first glimpse of how they act at the molecular level is really important, a key to understanding how they work. Given the remarkable efficacy of psilocybin for depression (in Phase II trials), we are confident our findings will accelerate the discovery of fast-acting antidepressants and potentially new drugs to treat other conditions, such as severe anxiety and substance use disorder."Scientists believe that activation of HTR2A, which is expressed at very high levels in the human cerebral cortex, is key to the effects of hallucinogenic drugs. "When activated, the receptors cause neurons to fire in an asynchronous and disorganized fashion, putting noise into the brain's system," said Roth, who holds a joint faculty appointment at the UNC Eshelman School of Pharmacy. "We think this is the reason these drugs cause a psychedelic experience. But it isn't at all clear how these drugs exert their therapeutic actions."In the current study, Roth's lab collaborated with Skiniotis, a structural biologist at the Stanford University School of Medicine. "A combination of several different advances allowed us to do this research," Skiniotis said. "One of these is better, more homogeneous preparations of the receptor proteins. Another is the evolution of cryo-electron microscopy technology, which allows us to view very large complexes without having to crystalize them."Roth credits co-first author Kuglae Kim, PhD, a postdoctoral fellow in his lab, for steadfastly exploring various experimental methods to purify and stabilize the very delicate serotonin receptors."Kuglae was amazing," Roth said. "I'm not exaggerating when I say what he accomplished is among the most difficult things to do. Over three years in a deliberate, iterative, creative process, he was able to modify the serotonin protein slightly so that we could get sufficient quantities of a stable protein to study."The research team used Kim's work to reveal the first X-ray crystallography structure of LSD bound to HTR2A. Importantly, Stanford investigators then used cryo-EM to uncover images of a prototypical hallucinogen, called 25-CN-NBOH, bound together with the entire receptor complex, including the effector protein Gαq. In the brain, this complex controls the release of neurotransmitters and influences many biological and neurological processes.The cryo-EM image is like a map of the complex, which Kim used to illustrate the exact structure of HTR2A at the level of amino acids -- the basic building blocks of proteins such as serotonin receptors.Roth, a psychiatrist and biochemist, leads the Psychoactive Drug Screening Program, funded by the National Institute of Mental Health. This gives his lab access to hallucinogenic drugs for research purposes. Normally, these compounds are difficult to study in the lab because they are regulated by the Drug Enforcement Agency as Schedule 1 drugs.Roth and colleagues are now applying their findings to structure-based drug discovery for new therapeutics. One of the goals is to discover potential candidates that may be able offer therapeutic benefit without the psychedelic effects."The more we understand about how these drugs bind to the receptors, the better we'll understand their signaling properties," Skiniotis says. "This work doesn't give us the whole picture yet, but it's a fairly large piece of the puzzle."
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917181243.htm
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Humans develop more slowly than mice because our chemistry is different
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Scientists from the RIKEN Center for Biosystems Dynamics Research, European Molecular Biology Laboratory (EMBL) Barcelona, Universitat Pompeu Fabra, and Kyoto University have found that the "segmentation clock" -- a genetic network that governs the body pattern formation of embryos -- progresses more slowly in humans than in mice because the biochemical reactions are slower in human cells. The differences in the speeds of biochemical reactions may underlie differences between species in the tempo of development.
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In the early phase of the development of vertebrates, the embryo develops into a series of "segments" that eventually differentiate into different types of tissues, such as muscles or the ribs. This process is known to be governed by an oscillating biochemical process, known as the segmentation clock, which varies between species. For example, it is about two hours in mice, and about five hours in humans. Why the length of this cycle varies between species has remained a mystery, however.To solve this mystery, the group began experiments using embryonic stem cells for mice and induced pluripotent stem (iPS) cells which they transformed into presomitic mesoderm (PSM) cells, the cells that take part in the segmentation clock.They began by examining whether something different was happening in the network of cells or whether there was a difference in the process within cells. They found, using experiments that either blocked important signals or put cells in isolation, that the latter is true.With the understanding that processes within cells were key, they suspected that the difference might be within the master gene -- HES7 -- which controls the process by repressing its own promoter, and did a number of complex experiments where they swapped the genes between the human and mouse cells, but this did not change the cycle.According to corresponding author Miki Ebisuya, who performed the work both at RIKEN BDR and EMBL Barcelona, "Failing to show a difference in the genes left us with the possibility that the difference was driven by different biochemical reactions within the cells." They looked at whether there were differences in factors such as the degradation rate of the HES7 protein, an important factor in the cycle. They looked at a number of processes including how quickly mouse and human proteins were degraded and found, confirming the hypothesis, that both proteins were degraded more slowly in human cells than in mouse cells. There were also differences in the time it took to transcribe and translate HES7 into proteins, and the time it took for HES7 introns to be spliced. "We could thus show," says Ebisuya, "that it was indeed the cellular environment in human and mouse cells that is the key to the differential biochemical reaction speeds and thus differential time scales."She continues, "Through this we have come up with a concept that we call developmental allochrony, and the present study will help us to understand the complicated process through which vertebrates develop. One of the key remaining mysteries is exactly what is difference between the human and mouse cells that drives the difference in reaction times, and we plan to do further studies to shed light on this."
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917180403.htm
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'Cellular compass' guides stem cell division in plants
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The stem cells tasked with creating and maintaining biological tissues have a difficult job. They have to precisely divide to form new specialized cells, which are destined to different fates even though they contain identical DNA. An obvious question then is: How do the cells divide in all the right ways to produce a healthy tissue? This was the grand motivating question for Andrew Muroyama, a postdoctoral scholar in the lab of Stanford University biologist Dominique Bergmann, as he monitored days of leaf development in the flowering plant Arabidopsis thaliana. There, amongst a thousand cells under his microscope, he noticed that the nucleus -- the DNA-containing control center in the cell -- moved in unexpected and strangely purposeful ways as stem cells divided.
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Previous research from the Bergmann lab identified a set of proteins that shuffle to one side of the stem cell before division. These proteins seemed to regulate how the stem cell divided, but the actual control mechanisms were unknown. These moving nuclei turned out to be a key to this mystery.In a paper published Sept. 17 in "I think our research highlights that the ability to watch the behaviors of cellular machines within living organisms can reveal unexpectedly elegant ways that individual cells cooperate to build tissues," said Muroyama, who is lead author of the paper. "You might think that something as fundamental as cell division would be completely solved by now but there is still so much to learn."The Bergmann lab makes Arabidopsis into fluorescent art under the microscope. Bright green nuclei wiggle within purple cell membranes. Watch closely, like Muroyama did, and you would see the usual process for asymmetric cell division: when an Arabidopsis stem cell first divides, the nucleus moves to one side. That way, the resulting daughter cells will be different sizes and will face different neighbors. Eventually, these two cells are destined to play different roles in the intricate final pattern of the leaf.But continue watching and the nucleus of one daughter cell moves again, hurrying to the opposite side of the cell where it will undergo a second asymmetric split."When Andrew showed me the videos of the cells, it was so bizarre," said Bergmann, who is a professor of biology in the School of Humanities and Sciences and senior author of the paper. "I thought, 'Why on Earth would a nucleus behave that way?' The first move makes sense but the second, in the complete opposite direction, was weird."In order to understand what they were seeing, the researchers conducted several experiments to tease apart the different factors that influence the cells during division.The researchers already knew about the cellular compass but were unsure what it was guiding or how it worked. By repelling the nucleus before the first division, the compass creates the first set of asymmetric daughters. But by attracting the nucleus immediately afterward, the compass can create a new set of asymmetric daughters on the other side."A critical step to understanding the function of the second migration was thinking about the longer history of the stem cells," said Muroyama. "The plant doesn't want to generate new stem cells right next to the ones that were just created. It wants to space them out, so moving the nucleus right after division sets it up for success when creating a second set of daughters."The researchers also discovered a protein that assists nuclear movement -- think a motor that powers the nucleus in the right direction. Disabling that motor prevented the second migration of the nucleus and the resulting leaves had fewer stomata than usual, which could impair the plant's ability to regulate water content and take in carbon dioxide.It was also known that cells in the leaf surface communicate with each other to regulate stem cell divisions. Curious about whether the cellular compass or cell-to-cell communication was the dominant cue to control how stem cells divide, the researchers modified cells so that they could not receive signals from neighboring cells and watched the bouncing nuclei. Without this communication, the compass appears in the wrong place within the cell, but could still move the nucleus around in predictable ways. This showed that, when it comes to leaf stem cells, the nucleus will follow the instructions from the cellular compass, even if it steers it wrong.As a next step, one graduate student in the Bergmann lab is already digging deeper into the purpose of the cellular compass, with particular interest into the different ways this compass can control cell divisions and fate.More broadly, these findings point to a different way of studying stem cells that focuses less exclusively on the journey of one cell. In some systems, the individual divisions that seem to define a cell's life may actually only be meaningful given what happens next and nearby."Looking back 10 years at what we thought was important for a stem cell, we've pretty much proven ourselves wrong," said Bergmann. "We were so focused on the details of what one stem cell did at a specific time and place. Now we understand that history and community matter. We have to look at the stem cell and its mother and grandmother and its neighbors."Graduate student Yan Gong is co-author of the paper. Bergmann is also a member of Stanford Bio-X and the Stanford Cancer Institute and an investigator of the Howard Hughes Medical Institute. This research was funded by the National Institutes of Health, Stanford University and the Howard Hughes Medical Institute.
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917105409.htm
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Engineered bacteria churn out cancer biomarkers
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Pity the glycan.
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These complex sugar molecules are attached to 80% of the proteins in the human body, making them an essential ingredient of life. But this process, known as glycosylation, has been somewhat overshadowed by flashier biomolecular processes such as transcription and translation."Glycosylation is absolutely essential for life on this planet. And yet, we still know relatively little about it," said Matthew DeLisa, the William L. Lewis Professor of Engineering in the Smith School of Chemical and Biomolecular Engineering. "While much attention has been given to understanding the genome and the proteome, the glycome -- which represents the entire complement of sugars, either free or present in more complex molecules such as glycoproteins, of an organism -- has been relatively understudied. We need new tools to advance the field forward."DeLisa's lab has created these very tools by commandeering simple, single-celled microorganisms -- namely The group's paper, "Engineering Orthogonal Human O-linked Glycoprotein Biosynthesis in Bacteria," published July 27 in Previously, DeLisa's team used a similar cell glyco-engineering approach to produce one of the most common types of glycoproteins -- those with glycan structures linked to the amino acid asparagine, or N-linked. Now the researchers have turned their attention to another abundant glycoprotein, namely O-linked, in which glycans are attached to the oxygen atom of serine or threonine amino acids of a protein.The O-linked glycans are more structurally diverse than their N-linked cousins, and they have important implications in the development of new therapeutic treatments for diseases such as breast cancer."Our cell-engineering efforts were quite complicated as we not only needed to equip When a cell turns cancerous, it expresses certain biomarkers, including abnormally glycosylated surface proteins, that indicate the presence of cancer. DeLisa's group equipped "The glycosylated version of MUC1 is one of the highest-priority target antigens for cancer therapy. It's been very challenging to develop therapies against this target," said DeLisa, the paper's senior author. "But by having a biosynthetic tool like the one we've created that is capable of replicating the MUC1 structure, we're hopeful that this could provide glycoprotein reagents that could be leveraged to discover antibodies or employed directly as immunotherapies, all of which could help in the fight against certain types of cancer."Both O-linked and N-linked glycans have also been discovered in one of the surface proteins of the SARS-CoV-2 virus, which causes COVID-19. DeLisa is hopeful his group's method of bacterial cell glyco-engineering will open the door for creating glycosylated versions of this S-protein that could lead to therapeutic antibodies against the coronavirus, or the development of a subunit vaccine.Because of their earlier work replicating N-linked glycans, the researchers were able to get the O-linked system up and running quickly. Now DeLisa's lab is primed to make proteins that carry both types of glycosylation, which is significant because many glycoproteins, such as the S-protein in SARS-CoV-2, carry both N- and O-linked glycan structures.The researchers are also exploring ways to increase the spectrum of glycoproteins that their engineered "We think of
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917105322.htm
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Discovery of microbes with mixed membranes sheds new light on early evolution of life
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Current research suggests that more complex life-forms, including humans, evolved from a symbiosis event between bacteria and another single-celled organism known as archaea. However, evidence of a transition period in which the two organisms mixed where nowhere to be found. That is, until now. In the deep waters of the Black Sea, a team of scientists found microbes that can make membrane lipids of unexpected origin.
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Cells are surrounded by a layer of membrane lipids that protect them from changes in their environment such as temperature, much in the same way that our skin changes when we are cold or exposed to the sun. Lead author and NIOZ senior scientist Laura Villanueva explains why they make such interesting biomarkers. 'When a cell dies, these lipids preserve like fossils and hold ancient-old information on Earths' early environmental conditions.' Our tree of life includes small and simple cells (Bacteria and Archaea) and more complex cells (Eukaryotes), including animals and humans. Bacteria and Eukaryotes share a similar lipid membrane. Looking at Archaea, their 'skin' or membrane looks very different and is primarily designed to help these microorganisms to survive in extreme environments. Villanueva: 'This "lipid divide," or difference in membranes between Bacteria and Eukaryotes on the one hand and Archaea on the other, is believed to have happened after the emergence of Bacteria and Archaea from the last universal cellular ancestor (LUCA).'The leading theory is that Eukaryotes evolved from a symbiosis event between archaeal and bacterial cells in which the archaeal cell was the host. But how does this work when their 'skins' are so different and share no sign of common ancestry? Villanueva: 'To explain the creation of more complex life-forms, the archaeal membrane must have made a switch to a bacterial type membrane. Such a switch likely needed a transition period in which the two membrane types were mixed.' However, mixed lipid membranes had never been found in microbes until the team of Villanueva made an unexpected discovery in de deep waters of the Black Sea.Villanueva: 'We found a possible missing piece of this puzzle in the Black Sea. Here, an abundant group of bacteria thrive in the deep-sea, absent of oxygen and with high sulfide concentration. We discovered that the genetic material of this group did not only carry pathway genes for bacterial lipids but archaeal ones as well.' The peculiarity was also found in the genetic material of other, closely related Bacteria and supports the idea that this ability to create 'mixed' membranes is more widespread than previously thought. This discovery sheds new light on the evolution of all cellular life forms and may have important consequences for the interpretation of archaeal lipid fossils in the geological record and paleoclimate reconstructions.
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Biotechnology
| 2,020 |
September 17, 2020
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https://www.sciencedaily.com/releases/2020/09/200917084555.htm
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Researchers discover how worms pass down knowledge through the generations
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When humans see their children about to eat something they oughtn't, we can simply tell them, "Don't eat that. It'll make you sick." Those who listen to this advice are spared the painful experience of learning that lesson for themselves. While other animals can't sit their offspring down for a good talking-to, that doesn't mean they are unable to instruct their descendants about potential harms.
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For instance, the microscopic roundworm "We wondered how the worms can know the identity of the bacteria they are eating," said Coleen Murphy, a professor of molecular biology and the Lewis-Sigler Institute for Integrative Genomics (LSI) and the senior author of a paper detailing the team's findings in the journal To investigate this question, two Murphy lab researchers -- associate research scholar Rachel Kaletsky and graduate student Rebecca Moore -- fed worms with a harmless bacterium that had been spiked with different materials isolated from pathogenic "Instead of a signal from the metabolites that the bacteria produce, as we had originally expected, we found that the worms 'read' the small RNAs that bacteria make, particularly small RNAs that correlate with the pathogenic state of the bacteria," said Murphy. In fact, the researchers discovered that inherited avoidance behavior requires one specific bacterial small RNA, called P11."The bacterial work that Geoff Vrla in Zemer Gitai's lab did was critical for proving that the key small RNA was P11," said Murphy."The P11 small RNA itself doesn't even make the worms sick -- just detecting the presence of P11 is enough to make the worms avoid the bacteria, and to pass it on to four generations of progeny," she adds.Kaletsky, Moore and their colleagues found that once a worm has eaten the bacterium, P11 is absorbed and processed by the worm intestine, then it finds its way into the worm's eggs and sperm. ("As far as we know, this is the first example found of an animal host 'reading' the small RNA of a pathogen and evolving a response that helps it stay healthier, a kind of nascent adaptive immune system response," said Murphy."There are only a few examples of such inter-species molecular communication via small RNAs, and even fewer examples of adaptive transgenerational behavioral changes in response to small RNAs," said Julie Claycomb, the Canada Research Chair in Small RNA Biology at the University of Toronto, who was not involved in the work."This study sets a high standard for understanding the molecular mechanisms governing such phenomena, and opens a novel area of investigation going forward," added Claycomb.
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Biotechnology
| 2,020 |
September 16, 2020
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https://www.sciencedaily.com/releases/2020/09/200916113426.htm
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Replicating a genome starts with a twist, a pinch, and a bit of a dance
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The most basic activity of a living thing is to turn one copy of its genome into two copies, crafting one cell into two. That replication event begins with a set of proteins -- the Origin of Replication Complex (ORC). And, with some cancers and developmental diseases linked to ORC proteins, structural biologists need to see how the complex works so they can understand how it might go wrong. Cold Spring Harbor Laboratory (CSHL) Professor & HHMI Investigator Leemor Joshua-Tor and colleagues published images of the human ORC in exquisite detail in
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The scientists think the first piece of the complex -- ORC1 -- finds the stretch of DNA where replication is supposed to begin and assembles the rest of the ORC (subunits 2-5) at that spot. Though, in yeast, a single sequence of DNA peppered throughout the genome spells out "start," there are no such simple signposts for the 30,000 start sites in humans. Our start signals are mysterious. Joshua-Tor says:"When the cell has to duplicate, the first thing that has to happen is that the genome has to duplicate. And so the positioning of ORC on these so-called "start" sites is really the first event that has to happen in order to start the duplication of the genome. You know in bacteria, there's usually one start site because it's a small genome, but in larger organisms like humans, in order to be able to replicate such a large genome, what the cell does is uses many, many start sites. And the interesting thing in mammalian systems is that we actually don't understand what a start site really looks like."To complicate things further, earlier on, as researchers looked at different organisms, they found differently shaped ORCs. But Joshua-Tor and colleagues found an explanation for those varying shapes. Parts of the ORC twist and pinch in dramatic ways, depending on what they are doing at the moment. A yeast ORC freezes mostly into one stable shape and a fly ORC into another. According to Kin On, a CSHL staff scientist, "the yeast complex is so stable, it is rock solid. But the human ORC assembly is very dynamic." Using cryo-electron microscopy (cryo-EM), sample preparation, and computer analysis techniques, the group was able to catch the human enzyme complex in many different shapes, including one that looks like a fly ORC and another that looks like yeast ORC. They assembled a series of images into a movie showing a wide range of motions. They even caught the first snapshot of a human ORC straddling a DNA molecule, which is key to understanding how ORCs do their jobs. According to Matt Jaremko, a postdoctoral fellow in Joshua-Tor's lab, "ORC is flexible, which helps the protein interact with DNA."The ORC was discovered at CSHL in 1992 by CSHL President and CEO Bruce Stillman, a collaborator of Joshua-Tor's on this study.Though a better understanding of ORCs may point to better treatments for cancer and developmental syndromes, Joshua-Tor says there is another reason to want to learn what we can about these beautiful cellular machines:"How we duplicate our genome is the most basic process of life, right? Really that's what life is all about. So, regardless of how we understand cancer and this developmental syndrome, you know, understanding ourselves and understanding the most basic process, that is part of the human endeavor really to understand ourselves. So it's not all about the utility of it. It's really, y'know, one of the basic endeavors of, of humanity is trying to understand life and ourselves. I think it's a big part of why we're doing it. At least a big part of why I'm doing it."
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Biotechnology
| 2,020 |
September 15, 2020
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https://www.sciencedaily.com/releases/2020/09/200915133159.htm
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Key role of immune cells in brain infection
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A new study has detailed the damaging role played by the immune system in a severe brain condition most commonly caused by the cold sore virus.
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Researchers have identified the specific type of immune cell that induces brain inflammation in herpes simplex virus (HSV) encephalitis. Crucially, they have also determined the signalling protein that calls this immune cell into the brain from the bloodstream.The findings, published in HSV encephalitis takes hold quickly and, despite rapid anti-viral drug treatment, many patients die. Most survivors are left with brain injury due to the inflammation and damage caused by the virus and immune cells gaining access to the brain, breaking down the blood-brain barrier."Determining the roles of specific immune cells and the factors that allow them to cross the protective blood-brain barrier is critical to develop targeted immune-therapies," explains Dr Benedict Michael, a Senior Clinician Scientist Fellow at the University of Liverpool, who led the research.Using a mouse model, the researchers showed that neutrophils (a type of immune cell) made the blood-brain barrier more permeable and contributed to the brain damage associated with HSV encephalitis. They also found that these neutrophils were not needed to control the virus.Meanwhile, monocyte immune cells were found to play a protective role and were needed to control the virus and prevent brain damage.The researchers also identified the exact signalling protein, called CXCL1, that drove the migration of these damaging neutrophils into the brain during HSV infection. By blocking this CXCL1 protein, neutrophils were prevented from crossing the blood-brain barrier and causing inflammation which resulted in less severe disease.The findings make the CXCL1 protein an attractive target for new therapies that can stop the influx of damaging white blood cells without limiting the roles of protective ones.Dr Michael said: "There is currently no licenced treatment for the severe brain swelling which occurs despite antiviral therapy in HSV encephalitis. Sometimes steroids are given, but as these suppress the immune system in a very broad way, there is a risk of uncontrolled viral infection."There is an urgent need for targeted treatment that prevents damaging immune cells from entering the brain without limiting the immune cells needed to control the virus."Now Dr Michael and colleagues are planning to examine the impact of the CXCL1 signalling protein in patients who have already had steroids as part of a clinical trial led by Professor Tom Solomon at the University of Liverpool.
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Biotechnology
| 2,020 |
September 15, 2020
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https://www.sciencedaily.com/releases/2020/09/200915105957.htm
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The Wnt pathway gets even more complicated
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The Wnt signalling pathway has been studied for decades, still it holds surprises in store. Bon-Kyoung Koo, group leader at IMBA -- Institute of Molecular Biotechnology of the Austrian Academy of Sciences and Tadasuke Tsukiyama at the Hokkaido University have now uncovered a new and unexpected role for a key component of the Wnt pathway, Casein Kinase-1, in regulating the pathway at the plasma membrane. This is the result of a study published today in
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In the Wnt pathway, Casein Kinase-1 is well-known as a part of the destruction complex. In the steady-state, when no Wnt signal is present, this complex destines the downstream mediator b-catenin for constant degradation. When a Wnt signal reaches the cell, the Wnt receptor Frizzled inhibits the destruction complex. This allows b-catenin to enter the nucleus, where it sets downstream responses in motion.In the newly published study, Tadasuke Tsukiyama and Bon-Kyoung Koo find that Casein Kinase-1 also regulates Wnt signalling at the plasma membrane. At the plasma membrane, the ubiquitin ligase RNF43 marks the Wnt receptor Frizzled for degradation, effectively shutting off the Wnt signalling pathway. The researchers discovered that Casein Kinase-1 triggers the switch for RNF43: When Casein Kinase-1 phosphorylates RNF43, RNF43 is activated and marks Frizzled with ubiquitin for degradation. When Casein Kinase-1 does not phosphorylate RNF43, RNF43 is inactive and signalling via Frizzled can continue. "We find that Casein Kinase-1 has an essential function in activating RNF43. With our work, we are effectively reintroducing Casein Kinase-1 to the field, defining a new role for this well-known regulator," Bon-Kyoung Koo explains.This new understanding could lead to a novel approach for reining the Wnt pathway in cancer cells. Tsukiyama and Koo found that a mutation in RNF43's extracellular domain interrupts its function in negative feedback regulation, the tumour suppressor function of RNF43. This mutation changes RNF43 into an oncogenic form that abnormally enhances Wnt signalling. The researchers found that mimicking the phosphorylation, by adding negatively charged residues to the mutant RNF43, can revert it back to a functional tumour suppressor. With this mimicked phosphoswitch, the mutant RNF43 was again able to inhibit Frizzled. "Some patients carry a mutation in RNF43's extracellular domain. We hope that, once we know how to mimic phosphorylation in cells, this phosphorylation would revive the RNF43 tumour suppressor, enabling it to again control the Wnt pathway," Bon-Kyoung Koo adds.
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Biotechnology
| 2,020 |
September 14, 2020
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https://www.sciencedaily.com/releases/2020/09/200914095903.htm
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Bioactive nano-capsules to hijack cell behavior
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Many diseases are caused by defects in signaling pathways of body cells. In the future, bioactive nanocapsules could become a valuable tool for medicine to control these pathways. Researchers from the University of Basel have taken an important step in this direction: They succeed in having several different nanocapsules work in tandem to amplify a natural signaling cascade and influence cell behavior.
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Cells constantly communicate with each other and have ways to pick up signals and process them -- similar to humans who need ears to hear sounds and knowledge of language to process their meaning. Controlling the cell's own signaling pathways is of great interest for medicine in order to treat various diseases.A research team of the Department of Chemistry at the University of Basel and the NCCR Molecular Systems Engineering develops bioactive materials that could be suitable for this purpose. To achieve this, the researchers led by Professor Cornelia Palivan combine nanomaterials with natural molecules and cells.In the journal In order to protect the enzymes from degradation in a cellular environment the research team loaded them into polymeric small capsules. Molecules can enter the compartment through biological pores specifically inserted in its synthetic wall and react with the enzymes inside.The researchers conducted experiments with nano-capsules harboring different enzymes that worked in tandem: the product of the first enzymatic reaction entered a second capsule and started the second reaction inside. These nano-capsuled could stay operative for days and actively participated in natural reactions in mammalian cells.One of the many signals that cells receive and process is nitric oxide (NO). It is a well-studied cellular mechanism since defects in the NO signaling pathway are involved in the emergence of cardiovascular diseases, but also in muscular and retinal dystrophies. The pathway encompasses the production of NO by an enzyme family called nitric oxide synthases (NOS). The NO can then diffuse to other cells where it is sensed by another enzyme named soluble guanylate cyclase (sGC). The activation of sGC starts a cascade reaction, regulating a plethora of different processes such as smooth muscle relaxation and the processing of light by sensory cells, among others.The researchers lead by Palivan produced capsules harboring NOS and sGC, which are naturally present in cells, but at much lower concentrations: the NOS-capsules, producing NO, act similarly to loudspeakers, "shouting" their signal loud and clear; the sGC-capsules, act as "ears," sensing and processing the signal to amplify the response.Using the intracellular concentration of calcium, which depends on the action of sGC, as an indicator, the scientists showed that the combination of both NOS and sGC loaded capsules makes the cells much more reactive, with an 8-fold increase in the intracellular calcium level."It's a new strategy to stimulate such changes in cellular physiology by combining nanoscience with biomolecules," comments Dr. Andrea Belluati, the first author of the study. "We just had to incubate our enzyme-loaded capsules with the cells, and they were ready to act at a moment's notice.""This proof of concept is an important step in the field of enzyme replacement therapy for diseases where biochemical pathways malfunction, such as cardiovascular diseases or several dystrophies," adds Cornelia Palivan.
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Biotechnology
| 2,020 |
September 13, 2020
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https://www.sciencedaily.com/releases/2020/09/200913162925.htm
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When methane-eating microbes eat ammonia instead
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Some microorganisms, the so-called methanotrophs, make a living by oxidizing methane (CH
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Nitric oxide is a highly reactive and toxic molecule with fascinating and versatile roles in biology and atmospheric chemistry. It is a signaling molecule, the precursor of the potent greenhouse gas nitrous oxide (NFor their study, Kartal and his colleagues used a methanotrophic bacterium named Methylacidiphilum fumariolicum, which originates from a volcanic mud pot, characterized by high temperatures and low pH, in the vicinity of Mount Vesuvius in Italy. "From this microbe, we purified a hydroxylamine oxidoreductase (mHAO) enzyme," reports Kartal. "Previously it was believed that mHAO enzyme would oxidize hydroxylamine to nitrite in methanotrophs. We now showed that it actually rapidly produces NO." The mHAO enzyme is very similar to the one used by "actual" ammonia oxidizers, which is quite astonishing, as Kartal explains: "It is now clear that enzymatically there is not much difference between aerobic ammonia- and methane-oxidizing bacteria. Using essentially the same set of enzymes, methanotrophs can act as de facto ammonia oxidizers in the environment. Still, how these microbes oxidize NO further to nitrite remains unknown."The adaptation of the mHAO enzyme to the hot volcanic mud pots is also intriguing, Kartal believes: "At the amino acid level, the mHAO and its counterpart from ammonia oxidizers are very similar, but the protein we isolated from M. fumariolicum thrives at temperatures up to 80 °C, almost 30 °C above the temperature optimum of their "actual" ammonia-oxidizing relatives. Understanding how so similar enzymes have such different temperature optima and range will be very interesting to investigate."According to Kartal, production of NO from ammonia has further implications for methane-eating microbes: "Currently there are no known methanotrophs that can make a living out of ammonia oxidation to nitrite via NO, but there could be methanotrophs out there that found a way to connect ammonia conversion to cell growth."
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Biotechnology
| 2,020 |
September 11, 2020
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https://www.sciencedaily.com/releases/2020/09/200911110749.htm
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Gene that drives ovarian cancer identified
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High-grade serious ovarian carcinoma (HGSOC) is the fifth-leading cause of cancer-related deaths in women in the United States, yet little is known about the origins of this disease.
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Now, scientists at the College of Veterinary Medicine have collaborated on a study that pinpoints which specific genes drive -- or delay -- this deadly cancer."We've taken the enormous collection of genomic mutation data that's been mined on cancer genetics and tried to make functional sense of it," said John Schimenti, professor of genetics in the Department of Biomedical Sciences and senior author of the study, which published Sept. 1 in Schimenti teamed with biomedical sciences colleague Alexander Nikitin, professor of pathology and director of the Cornell Stem Cell Program, and members of their respective labs to gain a better understanding of HGSOC.Cancer researchers have known for a while that the disease is almost always caused by multiple genetic "hits." One mutation alone does not turn a cell cancerous; generally at least two or three are required, and often different combinations of genes can cause the same cancer.Adding complexity, Schimenti said, is the fact that once one key genome-destabilizing mutation arises, others will follow. Sequenced tumors yield a plethora of mutations -- some are the originators of the cancer itself, while many others are spinoffs."It's a longstanding issue in cancer research," he said. "What are the genetic drivers, and what are the passengers in the process?"To address these complexities, the researchers wanted to test combinations of possible genetic suspects, and then parse out which of the many associated mutations were sparking the cancer.To do so, they turned to the Cancer Genome Atlas, an international collaborative database that compiles the genetic information from patient tumor samples and the mutated genes associated with them. They took a list of 20 genes known to mutate in HGSOC and, using CRISPR gene-editing technology, created random combinations of these mutations in cultured cells from the ovary surface, including regular epithelial cells and epithelial stem cells, to see which cell type was more susceptible to the mutations.The researchers then noted which combination of mutations turned which group of cells cancerous -- pinpointing both the genes driving the process and which cell type the cancer originated in.The study revealed what the team had originally suspected -- that ovarian surface stem cells were more apt to become cancerous when hit with mutations. They also unexpectedly discovered genes that had the opposite effect."We found there were various genes that would help the process along, but interestingly, there were other genes that, when mutated, actually inhibited the cancer initiation process," Schimenti said.Knowing which are the cells of origin and which genes are necessary in initiating this highly aggressive form of ovarian cancer can be powerful information, both for ovarian and other types of cancers. "The cancer driver screening methodology we used should be applicable to answering the same kinds of questions for cells and cancers in other organs and tissues," Nikitin said.Schimenti said the findings could be particularly useful for ovarian cancer patients who have their tumors biopsied and sequenced for genetic data."In the past, you would know which genes were mutated but you wouldn't know what role they played," he said. "Now you know which ones are important. And eventually, you could develop drugs to target the mutated genes that you know are causing the problem."This work was supported by grants from the Ovarian Cancer Research Fund, the New York State Stem Cell Science Program, the National Institutes of Health and the National Cancer Institute.
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Biotechnology
| 2,020 |
September 10, 2020
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https://www.sciencedaily.com/releases/2020/09/200910150330.htm
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Analysis of Australian labradoodle genome reveals an emphasis on the 'oodle'
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The creator of the Australian labradoodle set out to mix poodles and Labrador retrievers to develop a hypoallergic service dog. But, according to a new study by Elaine Ostrander at the National Institutes of Health, published September 10th in
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There are about 350 recognized dog breeds in the world today, many resulting from intense breeding programs that unintentionally created dogs at high risk for certain health problems. These high rates of disease were one motivating factor behind crossing two purebred dogs to create so-called "designer breeds," coupled with the desire to combine positive traits from the parental breeds.The Australian labradoodle is one of the most popular designer breeds, and so researchers analyzed genetic variations at more than 150,000 locations along its genome to understand how the breed has developed over the past 31 years. The findings show that genetically, the Australian labradoodle is mostly poodle, with smaller genetic contributions from the Labrador retriever and certain types of spaniel. Breeders appear to have preferentially chosen dogs with a poodle-like coat, which is associated with what many people consider hypoallergenicity, and without strong preference for specific traits from Labrador retrievers.The new study demonstrates that changes in very few genes, over a small number of generations, can define a new dog breed. The results of this genetic study may also inform the development of genetic tests that can be incorporated into thoughtful breeding programs to avoid some of the health problems that commonly afflict Australian labradoodles. Currently, Australian labradoodles supporters are lobbying to have the breed officially recognized by an international registry.
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Biotechnology
| 2,020 |
September 9, 2020
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https://www.sciencedaily.com/releases/2020/09/200909140314.htm
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Designed antiviral proteins inhibit SARS-CoV-2 in the lab
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Computer-designed small proteins have now been shown to protect lab-grown human cells from SARS-CoV-2, the coronavirus that causes COVID-19.
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The findings are reported today, Sept. 9, in In the experiments, the lead antiviral candidate, named LCB1, rivaled the best-known SARS-CoV-2 neutralizing antibodies in its protective actions. LCB1 is currently being evaluated in rodents.Coronaviruses are studded with so-called Spike proteins. These latch onto human cells to enable the virus to break in and infect them. The development of drugs that interfere with this entry mechanism could lead to treatment of or even prevention of infection.Institute for Protein Design researchers at the University of Washington School of Medicine used computers to originate new proteins that bind tightly to SARS-CoV-2 Spike protein and obstruct it from infecting cells.Beginning in January, more than two million candidate Spike-binding proteins were designed on the computer. Over 118,000 were then produced and tested in the lab."Although extensive clinical testing is still needed, we believe the best of these computer-generated antivirals are quite promising," said lead author Longxing Cao, a postdoctoral scholar at the Institute for Protein Design."They appear to block SARS-CoV-2 infection at least as well as monoclonal antibodies, but are much easier to produce and far more stable, potentially eliminating the need for refrigeration," he added.The researchers created antiviral proteins through two approaches. First, a segment of the ACE2 receptor, which SARS-CoV-2 naturally binds to on the surface of human cells, was incorporated into a series of small protein scaffolds.Second, completely synthetic proteins were designed from scratch. The latter method produced the most potent antivirals, including LCB1, which is roughly six times more potent on a per mass basis than the most effective monoclonal antibodies reported thus far.Scientists from the University of Washington School of Medicine in Seattle and Washington University School of Medicine in St. Louis collaborated on this work."Our success in designing high-affinity antiviral proteins from scratch is further proof that computational protein design can be used to create promising drug candidates," said senior author and Howard Hughes Medical Institute Investigator David Baker, professor of biochemistry at the UW School of Medicine and head of the Institute for Protein Design. In 2019, Baker gave a TED talk on how protein design might be used to stop viruses.To confirm that the new antiviral proteins attached to the coronavirus Spike protein as intended, the team collected snapshots of the two molecules interacting by using cryo-electron microscopy. These experiments were performed by researchers in the laboratories of David Veesler, assistant professor of biochemistry at the UW School of Medicine, and Michael S. Diamond, the Herbert S. Gasser Professor in the Division of Infectious Diseases at Washington University School of Medicine in St. Louis."The hyperstable minibinders provide promising starting points for new SARS-CoV-2 therapeutics," the antiviral research team wrote in their study pre-print, "and illustrate the power of computational protein design for rapidly generating potential therapeutic candidates against pandemic threats."
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Biotechnology
| 2,020 |
September 9, 2020
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https://www.sciencedaily.com/releases/2020/09/200909114854.htm
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Artificial intelligence aids gene activation discovery
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Scientists have long known that human genes spring into action through instructions delivered by the precise order of our DNA, directed by the four different types of individual links, or "bases," coded A, C, G and T.
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Nearly 25% of our genes are widely known to be transcribed by sequences that resemble TATAAA, which is called the "TATA box." How the other three-quarters are turned on, or promoted, has remained a mystery due to the enormous number of DNA base sequence possibilities, which has kept the activation information shrouded.Now, with the help of artificial intelligence, researchers at the University of California San Diego have identified a DNA activation code that's used at least as frequently as the TATA box in humans. Their discovery, which they termed the downstream core promoter region (DPR), could eventually be used to control gene activation in biotechnology and biomedical applications. The details are described September 9 in the journal "The identification of the DPR reveals a key step in the activation of about a quarter to a third of our genes," said James T. Kadonaga, a distinguished professor in UC San Diego's Division of Biological Sciences and the paper's senior author. "The DPR has been an enigma -- it's been controversial whether or not it even exists in humans. Fortunately, we've been able to solve this puzzle by using machine learning."In 1996, Kadonaga and his colleagues working in fruit flies identified a novel gene activation sequence, termed the DPE (which corresponds to a portion of the DPR), that enables genes to be turned on in the absence of the TATA box. Then, in 1997, they found a single DPE-like sequence in humans. However, since that time, deciphering the details and prevalence of the human DPE has been elusive. Most strikingly, there have been only two or three active DPE-like sequences found in the tens of thousands of human genes. To crack this case after more than 20 years, Kadonaga worked with lead author and post-doctoral scholar Long Vo ngoc, Cassidy Yunjing Huang, Jack Cassidy, a retired computer scientist who helped the team leverage the powerful tools of artificial intelligence, and Claudia Medrano.In what Kadonaga describes as "fairly serious computation" brought to bear in a biological problem, the researchers made a pool of 500,000 random versions of DNA sequences and evaluated the DPR activity of each. From there, 200,000 versions were used to create a machine learning model that could accurately predict DPR activity in human DNA.The results, as Kadonaga describes them, were "absurdly good." So good, in fact, that they created a similar machine learning model as a new way to identify TATA box sequences. They evaluated the new models with thousands of test cases in which the TATA box and DPR results were already known and found that the predictive ability was "incredible," according to Kadonaga.These results clearly revealed the existence of the DPR motif in human genes. Moreover, the frequency of occurrence of the DPR appears to be comparable to that of the TATA box. In addition, they observed an intriguing duality between the DPR and TATA. Genes that are activated with TATA box sequences lack DPR sequences, and vice versa.Kadonaga says finding the six bases in the TATA box sequence was straightforward. At 19 bases, cracking the code for DPR was much more challenging."The DPR could not be found because it has no clearly apparent sequence pattern," said Kadonaga. "There is hidden information that is encrypted in the DNA sequence that makes it an active DPR element. The machine learning model can decipher that code, but we humans cannot."Going forward, the further use of artificial intelligence for analyzing DNA sequence patterns should increase researchers' ability to understand as well as to control gene activation in human cells. This knowledge will likely be useful in biotechnology and in the biomedical sciences, said Kadonaga."In the same manner that machine learning enabled us to identify the DPR, it is likely that related artificial intelligence approaches will be useful for studying other important DNA sequence motifs," said Kadonaga. "A lot of things that are unexplained could now be explainable."This study was supported by the National Institute of General Medical Sciences (NIGMS) at the National Institutes of Health.
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Biotechnology
| 2,020 |
September 9, 2020
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https://www.sciencedaily.com/releases/2020/09/200909114750.htm
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Prediction of protein disorder from amino acid sequence
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Structural disorder is vital for proteins' function in diverse biological processes. It is therefore highly desirable to be able to predict the degree of order and disorder from amino acid sequence. Researchers have developed a prediction tool by using machine learning together with experimental NMR data for hundreds of proteins, which is envisaged to be useful for structural studies and understanding the biological role and regulation of proteins with disordered regions.
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In the last century, Anfinsen showed beyond a doubt that a protein can find its way back to its 'native' three-dimensional structure after it has been placed under 'denaturing conditions' where the protein structure is unfolded. The profound conclusion of his experiments was that apparently the information that governs the search back to the native state is hidden in the amino acid sequence. Thermodynamic considerations then set forth a view where the folding process is like rolling energetically downhill to the lowest point -- to the unique native structure. These findings have often been intertwined with the central dogma of molecular biology. Thus, a gene codes for an amino acid sequence, and the sequence codes for a specific structure.The next breakthrough came with the advent of cheap and fast genome sequencing in the wake of the human genome project; once thousands of genomes of various organisms were sequenced, scientists made a staggering discovery -- there were lots and lots of genes that coded for proteins with low-complexity. In other words, these proteins did not contain the right amino acids to fold up and experiments confirmed that they remained 'intrinsically disordered'. Also, the human genome turned out to have more than a third of its genes coding for protein Since disordered proteins are very flexible, they are not amenable to crystallization and therefore no information can be obtained from X-ray diffraction on protein crystals -- the approach that has been so pivotal for folded proteins. Instead, these proteins must be studied in solution, and for this purpose NMR (Nuclear Magnetic Resonance) spectroscopy is the most suited tool. In this method, a quantum physical property called 'spin' is measured in a strong magnetic field for each atom in the molecule. The exact precession frequencies of the spins are a function of their environment, and it is exactly this frequency that allows researchers to quantitatively measure to which extent each amino acid is ordered or disordered in the protein.In their new paper, published on 8 Sept 2020, Dr. Rupashree Dass together with Associate Professor Frans Mulder and Assistant Professor Jakob Toudahl Nielsen have used machine learning together with experimental NMR data for hundreds of proteins to build a new bioinformatics tool that they have called ODiNPred. This bioinformatics program can help other researchers making the best possible predictions of which regions of their proteins are rigid and which are likely to be flexible. This information is useful for structural studies, as well as understanding the biological role and regulation of intrinsically disordered proteins.
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Biotechnology
| 2,020 |
September 9, 2020
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https://www.sciencedaily.com/releases/2020/09/200909114748.htm
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Mysterious cellular droplets come into focus
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The world inside the human cell grew a bit more interesting in recent years as the role of a new biological structure became clearer.
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It was long believed that most important operations in the cell occur within organelles. "They're there to do certain functions. For instance, mitochondria generate the energy that everything runs on," explained Aleksei Aksimentiev, a professor of physics at the University of Illinois at Urbana-Champaign. "What is common to all of them is that they're surrounded by a lipid membrane. What people recently discovered is there are organelles that don't have lipid bilayers. They assemble spontaneously in the form of droplets. And those organelles have particular functions."In recent years, with improved imaging capabilities, the roles, occurrence, and behavior of these membrane-less organelles have become clearer. In 2017 they were given a name: biological condensates. They are thought to play a role in DNA repair and aging, and researchers believe a number of neurological diseases are related to the condensate not working properly, including Amyotrophic lateral sclerosis, or ALS, where nerve cells break down, leading to loss of muscular function."Let's say you have DNA and it suddenly has a break. It's usually a really bad thing, because it cannot replicate, but there's a machinery that will come and repair it," he explained. "A bubble of condensate forms that miraculously attracts only the molecules that are required to repair the DNA. There are all kinds of different condensates and they all recruit the right molecules somehow."How do these membrane-less organelles spontaneously form? And how do they recruit other molecules to help them?The physics of this process appears similar to phase separation, like how oil and water spontaneously form droplets in the right conditions, but with some differences. In normal phase separation, temperature usually motivates the separation. In biology, it is a change in concentrations."We don't know exactly how it works," Aksimentiev said. "I'm specifically interested in how this recruitment happens, and how molecules recognize other molecules."Aksimentiev is using the Frontera supercomputer at the Texas Advanced Computing Center (TACC), one of the fastest in the world, to better understand this process. Over the last decade, he and others developed the tools and methods to explore the behavior of biological systems at the atomic level using molecular dynamics simulations.Aksimentiev is able to simulate biological systems with millions of interacting atoms in a realistic environment for microseconds or even milliseconds -- the timescales at which biological systems operate. Today's supercomputers allow larger, faster simulations, and permit scientists to ask and answer new questions.Even by the standards of the field, biological condensates are challenging to study computationally. Unlike other ordered systems like proteins with known rigid structures, or disordered systems like water, biological condensates are what's known as 'partially disordered' -- a particularly hard type of structure to simulate.Writing in the The researchers showed that a particle-based molecular dynamics model can reproduce known phase separation properties of a FUS condensate, including its critical concentration and susceptibility to mutations.They also showed that they could use chain collapse theory to determine the thermodynamic properties of the condensate and to link them to changes in the shape of individual condensate molecules.The behavior of a biological condensate, with all its complex inter- and intramolecular interactions, can be described by a polymer physics model, they found. This makes computer modeling a useful tool for uncovering the behavior of these still-mysterious cellular actors.Aksimentiev's research sets the stage for future studies that will elucidate the molecular mechanisms driving the formation of droplets in more complex biological condensates, like those that repair RNA. The work is one step on a long path to fully elucidate the mystery of biological condensates in cells -- another trick of nature slowly uncovered.
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Biotechnology
| 2,020 |
September 9, 2020
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https://www.sciencedaily.com/releases/2020/09/200909100308.htm
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In ancient giant viruses lies the truth behind evolution of nucleus in eukaryotic cells
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Perhaps as far back as the history of research and philosophy goes, people have attempted to unearth how life on earth came to be. In the recent decades, with exponential advancement in the fields of genomics, molecular biology, and virology, several scientists on this quest have taken to looking into the evolutionary twists and turns that have resulted in eukaryotic cells, the type of cell that makes up most life forms today.
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The most widely accepted theories that have emerged state that the eukaryotic cell is the evolutionary product of the intracellular evolution of proto-eukaryotic cells, which were the first complex cells, and symbiotic relationships between proto-eukaryotic cells and other unicellular and simpler organisms such as bacteria and archaea. But according to Professor Masaharu Takemura of the Tokyo University of Science, Japan, "These hypotheses account for and explain the driving force and evolutionary pressures. But they fail to portray the precise process underlying eukaryotic nucleus evolution."Prof Takemura cites this as his motivation behind his recent article published in In a way, Prof Takemura's hypothesis has its roots in 2001 when, along with PJ Bell, he made the revolutionary proposal that large DNA viruses, like the poxvirus, had something to do with the rise of the eukaryotic cell nucleus. Prof Takemura further explains the reasons for his inquiry into the nucleus of the eukaryotic cell as such: "Although the structure, function, and various biological functions of the cell nucleus have been intensively investigated, the evolutionary origin of the cell nucleus, a milestone of eukaryotic evolution, remains unclear."The origin of the eukaryotic nucleus must indeed be a milestone in the development of the cell itself, considering that it is the defining factor that sets eukaryotic cells apart from the other broad category of cells -- the prokaryotic cell. The eukaryotic cell is neatly compartmentalized into membrane-bound organelles that perform various functions. Among them, the nucleus houses the genetic material. The other organelles float in what is called the cytoplasm. Prokaryotic cells do not contain such compartmentalization. Bacteria and archaea are prokaryotic cells.The 2001 hypothesis by Prof Takemura and PJ Bell is based on striking similarities between the eukaryotic cell nucleus and poxviruses: in particular, the property of keeping the genome separate in a compartment. Further similarities were uncovered after the discovery and characterization of a type of large DNA virus called "giant virus," which can be up to 2.5 µm in diameter and contain DNA "encoding" information for the production of more than 400 proteins. Independent phylogenetic analyses suggested that genes had been transferred between these viruses and eukaryotic cells as they interacted at various points down the evolutionary road, in a process called "lateral gene transfer."Viruses are "packets" of DNA or RNA and cannot survive on their own. They must enter a "host" cell and use that cell's machinery to replicate its genetic material, and therefore multiply. As evolution progressed, it appears, viral genetic material became integrated with host genetic material and the properties of both altered.In 2019, Prof Takemura and his colleagues made another breakthrough discovery: the medusavirus. The medusavirus got its name because, like the mythical monster, it causes encystment in its host; that is, it gives its host cell a "hard" covering.Via experiments involving the infection of an amoeba, Prof Takemura and his colleagues found that the medusavirus harbors a full set of histones, which resemble histones in eukaryotes. Histones are proteins that keep DNA strands curled up and packed into the cell nucleus. It also holds a DNA polymerase gene and major capsid protein gene very similar to those of the amoeba. Further, unlike other viruses, it does not construct its own enclosed "viral factory" in the cytoplasm of the cell within which to replicate its DNA and contains none of the genes required to carry out the replication process. Instead, it occupies the entirety of the host nucleus and uses the host nuclear machinery to replicate.These features, Prof Takemura argues, indicate that the ancestral medusavirus and its corresponding host proto-eukaryotic cells were involved in lateral gene transfer; the virus acquired DNA synthesis (DNA polymerase) and condensation (histones) genes from its host and the host acquired structural protein (major capsid protein) genes from the virus. Based on additional research evidence, Prof Takemura extends this new hypothesis to several other giant viruses as well.Thus, Prof Takemura connects the dots between his findings in 2019 and his original hypothesis in 2001, linking them through his and others' work in the two decades that come in between. All of it taken together, it becomes clear how the medusavirus is prime evidence of the viral origin of the eukaryotic nucleus.He says: "This new updated hypothesis can profoundly impact the study of eukaryotic cell origins and provide a basis for further discussion on the involvement of viruses in the evolution of the eukaryotic nucleus." Indeed, his work may have unlocked several new possibilities for future research in the field.
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Biotechnology
| 2,020 |
September 8, 2020
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https://www.sciencedaily.com/releases/2020/09/200908122457.htm
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More than just genetic code
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In photosynthesis, solar energy is converted into chemical energy, which is then used in nature to produce organic molecules from carbon dioxide. In plants, algae and cyanobacteria, the key photosynthesis reactions take place in two complex structures known as photosystems. These are located in a special membrane system, the thylakoids. However, many details of their molecular structure and the way the proteins are incorporated into the membranes have yet to be explored.
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A team led by Professor Conrad Mullineaux from the Institute of Biology and Chemistry at Queen Mary University London, UK, Professor Annegret Wilde and Professor Wolfgang Hess from the Institute of Biology III at the University of Freiburg and Professor Satoru Watanabe from the Institute of Biosciences at the Agricultural University of Tokyo, Japan, has published a study in the current issue of The researchers used molecular genetic, bioinformatics and high-resolution microscopic approaches at the single cell level for their investigations. The results confirm that mRNA molecules encode much more than just the sequence of the protein. They also carry signals that appear to control the position and coordination of the photosystem structure. The team was able to identify two proteins likely to be involved in this process by interacting with these mRNAs. The researchers say this opens the way to a detailed understanding of the molecular mechanisms involved and provides new approaches to make these processes useful for photobiotechnology.
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Biotechnology
| 2,020 |
September 8, 2020
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https://www.sciencedaily.com/releases/2020/09/200908113316.htm
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Cashing in on marine byproducts
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As exploitation of wild fisheries and marine environments threaten food supplies, Flinders University scientists are finding sustainable new ways to convert biowaste, algal biomass and even beached seaweed into valuable dietary proteins and other products.
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In one of several projects under way at the Flinders Centre for Marine Bioproducts Development, researchers are looking to extract value from crayfish shells and other marine waste via a 'green' fluidic processing machine developed at the University."As world populations grow, so will demand for dietary proteins and protein-derived products and this cannot be met using traditional protein sources," says Professor Kirsten Heimann, who says millions of tonnes of sea catches produce bycatch, shells, bones, heads and other parts wasted during the processing of marine and freshwater species.Seafood processing by-products (SPBs) and microalgae are promising resources that can fill the demand gap for proteins and protein derivatives, they say in a new publication."These biomaterials are a rich source of proteins with high nutritional quality while protein hydrolysates and biopeptides derived from these marine proteins possess several useful bioactivities for commercial applications in multiple industries," adds Flinders University co-author Trung Nguyen in the paper published in "Efficient utilisation of these marine biomaterials for protein recovery would not only supplement global demand and save natural bioresources but would also successfully address the financial and environmental burdens of biowaste, paving the way for greener production and a circular economy."Value-adding also looks promising with many of the bioactive protein-derived products gaining attention to promote human health including in drug discovery, nutraceutical and pharmaceutical developments. Estimates of the commercial value of these therapeutic protein-based products in 2015 was US$174.7 billion and is predicted to reach US$266.6 billion in 2021, leading to a two-fold increase in demand of protein-derived products.Globally, 32 million tonnes of SPBs are estimated to be produced annually which represents an inexpensive resource for protein recovery while technical advantages in microalgal biomass production would yield secure protein supplies with minimal competition for arable land and freshwater resources.This comprehensive review article analyses the potential of using SPBs and microalgae for protein recovery and production critically assessing the feasibility of current and emerging technologies used for the process development.The nutritional quality, functionalities, and bioactivities of the extracted proteins and derived products together with their potential applications for commercial product development are also systematically summarised and discussed in the free online paper.
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Biotechnology
| 2,020 |
September 8, 2020
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https://www.sciencedaily.com/releases/2020/09/200908101624.htm
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Through enzyme testing, researchers sharpen CRISPR gene-editing tool
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One of the biggest scientific advances of the last decade is getting better thanks to researchers at The University of Texas at Austin; the University of California, Berkeley; and Korea University. The team has developed a new tool to help scientists choose the best available gene-editing option for a given job, making the technology called CRISPR safer, cheaper and more efficient. The tool is outlined in a paper out today in
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The CRISPR gene-editing technique holds tremendous potential to improve human health, agriculture and the future of people on the planet, but the challenge lies in the delicate nature of gene editing -- there is almost no room for error.To edit genes, scientists use dozens of different enzymes from a naturally occurring system called CRISPR. Researchers locate a problematic DNA sequence and use these specialized enzymes to snip it as if using a pair of scissors, allowing genetic material to be added, removed or altered. But these scissors are not perfect. Accuracy and effectiveness vary by the CRISPR enzyme and the project. The new tool guides users, so they can pick the best CRISPR enzyme for their high-stakes gene edit."We designed a new method that tests the specificity of these different CRISPR enzymes -- how precise they are -- robustly against any changes to the DNA sequence that could misdirect them, and in a cleaner way than has ever been done before," said Steve Jones, a UT research scientist who co-wrote the paper with Ilya Finkelstein, an associate professor of molecular biosciences.Problems can occur when a CRISPR enzyme targets the wrong sections of DNA. Each CRISPR enzyme has strengths and weaknesses in editing different sequences, so the researchers set out to create a tool to help scientists compare the different enzymes and find the best one for a given job."CRISPR wasn't designed in a lab. It wasn't made by humans for humans. It was made by bacteria to defend against viruses," said John Hawkins, a Ph.D. alumnus who was recently with UT's Oden Institute for Computational Engineering and Sciences. "There is incredible potential for its use in medicine, but the first rule of medicine is 'do no harm.' Our work is trying to make CRISPR safer."The team of researchers developed a library of DNA sequences and measured how accurate each CRISPR enzyme was, how long it took the enzyme to edit the sequences and how precisely they edited the sequence. For some tasks the commonly used enzyme CRISPR-Cas9 worked best; in others, different enzymes performed much better."It's like a standardized test," Hawkins said. "Every student gets the same test, and now you have a benchmark to compare them."The tool allows scientists to choose the best enzyme for editing on the first try, so the process becomes more efficient and cheaper. Additionally, it gives scientists information about where mistakes are most likely to occur for each enzyme, saving time."This technique gives us a new way to reduce risk," Jones said. "It allows gene edits to be more predictable."
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Biotechnology
| 2,020 |
September 8, 2020
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https://www.sciencedaily.com/releases/2020/09/200908091522.htm
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How mutations in DNA packaging machines cause cancer
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Like wrenches made of Legos, SWI/SNF chromatin remodeling complexes tighten or loosen DNA in our cells to control how genes are turned on and made into proteins. When assembled correctly, these complexes play a crucial role in the development of normal tissues, and when broken, they can lead to the development of cancer. These complexes are commonly disrupted by mutations in the genes that encode them -- but how this leads to cancer is poorly understood.
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New research from the Children's Medical Center Research Institute at UT Southwestern (CRI) determined how mutations in two key SWI/SNF proteins, ARID1A and ARID1B, can drive cancer development by disrupting the assembly of SWI/SNF complexes. The study, published in "While it is abundantly clear that SWI/SNF components are defective in almost all cancer types, it is still fuzzy how mutations in components lead to broken SWI/SNF complexes, and how broken complexes cause disease," says study leader Hao Zhu, M.D., an associate professor at CRI. "In this study, we tried to cleanly break one important type of SWI/SNF complex to study how it falls apart, and how this leads to uncontrolled cancer growth."SWI/SNF protein complexes help to pack and unpack DNA in the genome and are composed of 10-15 interacting proteins that can be arranged into different configurations in different tissues. Three main types of SWI/SNF complexes have been identified: cBAF, pBAF and ncBAF. But the roles they play in tissue development and disease have been unclear. To understand the importance of these complexes in animals, researchers at CRI focused on the cBAF complex. This complex was chosen because it is the most abundant one, and a subunit unique to this complex, ARID1A, is one of the most mutated genes in human cancer.ARID1A is closely related to another protein known as ARID1B, which is also unique to cBAF. It has been shown that some cancer cells need at least one ARID1 protein to survive. To examine whether simultaneous loss of both ARID1A and ARID1B would be more likely to cause or kill cancer cells, researchers eliminated or knocked out both genes in mice. Strikingly, the loss of both ARID1A and ARID1B genes resulted in aggressive liver and skin cancer formation within weeks."In cancers where ARID1A is gone or mutated, one proposed strategy to stop cancer growth is to inhibit the replacement protein ARID1B. This method was predicted to kill cancer cells that might need cBAF function to survive," says Zhu. "However, our findings suggest that therapeutically targeting ARID1B could make matters worse by accelerating aggressive cancer development."Researchers discovered that loss of these proteins led to the disassembly of the cBAF complex into many nonfunctional pieces.They were able to uncover how ARID1A and ARID1B proteins maintain stabilizing connections between different components within cBAF complexes. This helped them pinpoint a number of important regions within these ARID1 proteins, that when mutated can make cBAF complexes fall apart. Interestingly, the importance of these regions also explains why mutations accumulate in these regions in human cancers. When cBAF falls apart, the leftover components interfere with the composition and function of other types of SWI/SNF complexes, which further contributes to cancer."We hope that the findings in our paper will change the way people think about the molecular consequences of SWI/SNF disruption and how mutations in this complex drive malignancy," says Zixi Wang, Ph.D., a postdoctoral researcher at CRI, assistant instructor of pediatrics at UTSW, and lead author of the paper.The National Institutes of Health (R03ES026397-01 and R01DK111588), CPRIT (RP170267 and RP150596), Stand Up To Cancer (SU2C-AACR-IRG 10-16), and donors to the Children's Medical Center Foundation supported this work.
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Biotechnology
| 2,020 |
September 7, 2020
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https://www.sciencedaily.com/releases/2020/09/200907112333.htm
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A new twist on DNA origami
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A team of scientists from ASU and Shanghai Jiao Tong University (SJTU) led by Hao Yan, ASU's Milton Glick Professor in the School of Molecular Sciences, and director of the ASU Biodesign Institute's Center for Molecular Design and Biomimetics, has just announced the creation of a new type of meta-DNA structures that will open up the fields of optoelectronics (including information storage and encryption) as well as synthetic biology.
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This research was published today in It is common knowledge that the predictable nature of Watson-Crick base-pairing and the structural features of DNA have allowed DNA to be used as a versatile building block to engineer sophisticated nanoscale structures and devices."A milestone in DNA technology was certainly the invention of DNA origami, where a long single-stranded DNA (ssDNA) is folded into designated shapes with the help of hundreds of short DNA staple strands," explained Yan. "However it has been challenging to assemble larger (micron to millimeter) sized DNA architectures which up until recently has limited the use of DNA origami." The new micron sized structures are on the order of the width of a human hair which is 1000 times larger than the original DNA nanostructures.Ever since gracing the cover of "In this current research we developed a versatile "meta-DNA" (M-DNA) strategy that allowed various sub-micrometer to micrometer sized DNA structures to self-assemble in a manner similar to how simple short DNA strands self-assemble at the nanoscale level," said Yan.The group demonstrated that a 6-helix bundle DNA origami nanostructure in the sub-micrometer scale (meta-DNA) could be used as a magnified analogue of single-stranded DNA (ssDNA), and that two meta-DNAs containing complementary "meta-base pairs" could form double helices with programmed handedness and helical pitches.Using meta-DNA building blocks they have constructed a series of sub-micrometer to micrometer scale DNA architectures, including meta-multi-arm junctions, 3D polyhedrons, and various 2D/3D lattices. They also demonstrated a hierarchical strand-displacement reaction on meta-DNA to transfer the dynamic features of DNA to the meta-DNA.With the help of assistant professor Petr Sulc (SMS) they used a coarse-grained computational model of the DNA to simulate the double-stranded M-DNA structure and to understand the different yields of left-handed and right-handed structures that were obtained.Further, by just changing the local flexibility of the individual M-DNA and their interactions, they were able to build a series of sub-micrometer or micron-scale DNA structures from 1D to 3D with a wide variety of geometric shapes, including meta-junctions, meta-double crossover tiles (M-DX), tetrahedrons, octahedrons, prisms, and six types of closely packed lattices.In the future, more complicated circuits, molecular motors, and nanodevices could be rationally designed using M-DNA and used in applications related to biosensing and molecular computation. This research will make the creation of dynamic micron-scale DNA structures, that are reconfigurable upon stimulation, significantly more feasible.The authors anticipate that the introduction of this M-DNA strategy will transform DNA nanotechnology from the nanometer to the microscopic scale. This will create a range of complex static and dynamic structures in the sub-micrometer and micron-scale that will enable many new applications.For example, these structures may be used as a scaffold for patterning complex functional components that are larger and more complex than previously thought possible. This discovery may also lead to more sophisticated and complex behaviors that mimic cell or cellular components with a combination of different M-DNA based hierarchical strand displacement reactions.
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Biotechnology
| 2,020 |
September 7, 2020
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https://www.sciencedaily.com/releases/2020/09/200907112134.htm
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Viruses play critical role in evolution and survival of the species
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As the world scrambles to control the growing COVID-19 coronavirus pandemic, new research in
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Scientists in the Cincinnati Children's Perinatal Institute and at Azabu University in Japan obtained their data by studying laboratory mice and human germline cells.In two separate papers appearing in the same edition of the journal, they reveal two distinct and fundamental processes underlying germline transcriptomes. They also show that species-specific transcriptomes are fine-tuned by endogenous retroviruses in the mammalian germlineGermline transcriptomes include all the messenger RNA in germline cells, which contain either the male or female half of chromosomes passed on as inherited genetic material to offspring when species mate. This means that germline transcriptomes define the unique character of sperm and egg to prepare for the next generation of life.Although the studies are separate they complement one another, according to Satoshi Namekawa, PhD, principal investigator on both papers and a scientist in the Division of Reproductive Science at Cincinnati Children's."One paper, Maezawa and Sakashita "The second study, Sakashita Together the studies have significant potential ramifications for clinical practice, according to study authors, who include a multi-disciplinary mix of developmental biologists, bioinformaticians and immuno-biologists. Dysregulation of gene expression in the formation of male sperm is closely associated with male infertility and birth defects.Viruses, especially endogenous retroviruses (ERVs) that are an inherent part of mammalian biology, can dramatically influence gene expression, investigators report. ERVs are molecular remnants of retroviruses that infect the body and over time incorporate into the genome."What we learn from our study is that, in general, viruses have major roles in driving evolution," Namekawa explained. "In the long-term, viruses have positive impacts to our genome and shape evolution."The study, Maezawa and Sakashita Those tests revealed that the the genome-wide reorganization of super-enhancers drives bursts of germline gene expression after germ cells enter meiosis, a specialized form of cell division that produces the haploid genome of germ cells.The study further demonstrates the molecular process through whichsuper-enhancer switching takes place in germ cells. Super-enhancers are regulated by two molecules that act as gene-burst control switches -- the transcription factor A-MYB and SCML2, a critical silencing protein in sperm formation.Endogenous retroviruses are a group of transposable elements (TEs), mobile genetic elements that account for approximately 40-50 percent of a given mammalian genome. Also referred to as "jumping genes," TEs have long been considered genetic threats because transposition can be harmful if, for example, the process disrupts protein-coding genes.Building on findings from the 1950s that TEs can function as genetic regulatory elements, Namekawa and his collaborators (Sakashita Funding support for the ERV study came in part from: the National Institutes of Health (R01 GM122776, DP2 GM119134); a Lalor Foundation Postdoctoral Fellowship; a grant from the Azabu University Research Services Division; Japan's Ministry of Education, Culture, Sports, Science and Technology (MEXT)-Supported Program for the Private University Research Branding Project (2016-2019); a Grant-in-Aid for Research Activity Start-up (19K21196); the Uehara Memorial Foundation Research Incentive Grant; an Albert J. Ryan Fellowship, and a March of Dimes Prematurity Research Centre Collaborative Grant (#22-FY14-470).Support for the super-enhancer study came in part from a research project grant by the Azabu University Research Services Division; a grant from Japan's Ministry of Education, Culture, Sports, Science and Technology and (MEXT)-Supported Program for the Private University Research Branding Project (2016-2019); a Grant-in-Aid for Research Activity Start-up (19K21196) from the Takeda Science Foundation (2019); the Uehara Memorial Foundation Research Incentive Grant (2018); a Lalor Foundation Postdoctoral Fellowship; an Albert J. Ryan Fellowship; and a Cincinnati Children's Endowed Scholar Award; CpG grant awards and the National Institute of Health (DP2 GM119134, R01GM122776 and GM098605)
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Biotechnology
| 2,020 |
September 7, 2020
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https://www.sciencedaily.com/releases/2020/09/200904100623.htm
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Repulsion mechanism between neurons governs fly brain structure
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Researchers at Kanazawa University report in Nature Communications the discovery that in the developing fly brain, neurons stemming from the same parent cell experience repulsion. This lineage-dependent repulsion is regulated by a protein known as Dscam1.
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The brain's structure has columnar features, which are hypothesized to arise from nerve cells (neurons) stemming from the same parent cell, initially forming radial units. How exactly this process unfolds at the molecular level remains unexplained, however. Now, an important insight comes from Makoto Sato and colleagues from Kanazawa University who show how, in the fly brain, a gene known as Dscam regulates how neurons from one lineage repel each other, and project their axons to different columns. (Axons or nerve fibers are long protrusions of nerve cells, the function of which is to conduct electrical signals.) This finding corroborates the 'radial unit hypothesis', with the mechanism at play being lineage-dependent repulsion between sister neurons.The researchers first looked at the evolution of neuron growth in the medulla, a part of the fly's visual system featuring a columnar structure. Its development is similar to that of the cerebral cortex in the brain of mammals; it involves neuroblasts (neural stem-like cells) that produce radially oriented and clonally related groups of neurons. Sato and colleagues recorded the distances between sister neurons (i.e., neurons stemming from the same neuroblast and forming a radial unit) and between axon pairs. From the obtained distance data, the scientists were able to conclude that the sister neurons often repel each other -- this observation is consistent with the formation of columns. Sato and colleagues call this process 'lineage-dependent repulsion'.The mechanism that enables lineage-dependent repulsion must lie in daughter neurons derived from the same neuroblast 'remembering' the identity of their common mother neuroblast. Sato and colleagues put forward the explanation that the protein Dscam1 is involved. Dscam1 can develop nearly 20,000 variants, but when two identical Dscam1 molecules bind, they lead to a repulsive signal known to control self-avoidance in certain dendritic processes -- dendrites are branch-like extensions of nerve cells. The reasoning then is that daughter neurons stemming from the same neuroblast produce the same Dscam1 variant, and so repel each other, whereas neurons of different lineages express different Dscam1 variants that don't repel each other and can project to the same column.The scientists were able to support their argumentation by a series of experiments confirming the relation between Dscam1 and lineage-dependent repulsion. Sato and colleagues note that "the mechanism that we propose ... is very simple," and add that it will be "interesting to determine whether similar mechanisms exist in other biological systems including column formation in mammalian brains."The radial unit hypothesis is a theory describing the development of the cerebral cortex. (The cerebral cortex is the outer layer of neural tissue of the largest part of the human brain. It plays an important role in brain functions like perception, thought, memory, language, and consciousness.) The hypothesis asserts that in the early stages of development, the cerebral cortex forms as an assembly of interacting 'columns', or 'radial units', with each unit originating from a stem cell layer containing neural stem cells. Makoto Sato and colleagues from Kanazawa University now present results showing that, in the fly brain, neurons from one lineage project their axons to different columns, and that this mechanism is regulated by the gene Dscam (Down syndrome cell adhesion molecule).The fly Drosophila Dscam1 gene encodes more than 19,000 variants ('isoforms'). Binding of Dscam1 with identical isoforms results in a repulsive signal, which is important for certain self-avoidance mechanisms within cells. Sato and colleagues showed that in fly larvae, Dscam1 is expressed in neuroblasts, and therefore inherited by neurons having the same lineage. This results in lineage-dependent repulsion between neurons in radial units, which then plays an important role in the development of the columnar structure of the fly's medulla.
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Biotechnology
| 2,020 |
September 4, 2020
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https://www.sciencedaily.com/releases/2020/09/200904121315.htm
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Blood breakdown product commandeers important enzyme
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The hemoglobin in the red blood cells ensures that our body cells receive sufficient oxygen. When the blood pigment is broken down, "heme" is produced, which in turn can influence the protein cocktail in the blood. Researchers at the University of Bonn have now discovered in complex detective work that the "activated protein C" (APC) can be commandeered by heme. At the same time, APC can also reduce the toxic effect of heme. Perspectively, the findings may provide the basis for better diagnostic and therapeutic approaches to blood diseases. The study has been published online in advance in the journal
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"Blood is a juice of very special kind," is what Johann Wolfgang von Goethe had his Mephistopheles say. The hemoglobin gives blood its red color and ensures that the erythrocytes (red blood cells) can bind oxygen for breathing. This is managed by the hemoglobin-bound molecule "heme," which is a complex composing of a central iron ion and a porphyrin molecule. "The breakdown of erythrocytes results in a pool of so-called labile, regulatory heme," explains Prof. Dr. Diana Imhof from the Pharmaceutical Institute of the University of Bonn. As it can exert toxic effects in high concentrations, the body tries to keep the amount of heme in check.It has been known for quite some time that this "labile, regulatory heme" affects the function of biomolecules. The team around Diana Imhof has now discovered in meticulous detective work which of the many proteins in the blood is particularly under the control of heme. "Over the last few years, our research group has established a large database of model peptides," reports Imhof. The peptides are individual protein "snippets" from which the sometimes huge and complex structures are built. Instead of studying the giant molecules, the proteins, in their entirety, the researchers first took a shortcut with the snippets.The pharmacists at the University of Bonn proceeded in a similar way to profilers in thrillers, who draw conclusions about the perpetrator's behavior from crime scene traces, circumstantial evidence and the type of crime. The researchers used an algorithm to systematically search the database for protein fragments that might potentially interact with heme. Using these data, they were able to conclude that the "activated protein C" (APC) is a particular candidate for heme binding. This enzyme is known for its anticoagulant and clot-dissolving effect, but can also take over cell-protective and anti-inflammatory tasks."So far, the impact of heme on the function of APC has been unknown," says Imhof's colleague and lead author of the study, Marie-Thérèse Hopp. The researchers investigated the association with pure compounds in the test tube and by using blood plasma samples provided by the Institute of Experimental Hematology and Transfusion Medicine at the University Hospital Bonn. There, Prof. Dr. Bernd Pötzsch and Dr. Nasim Shahidi Hamedani also supported the pharmacists with know-how, APC samples, test systems and access to specific devices. "We demonstrated that the enzymatic and anticoagulant activity of APC is reduced in the presence of heme," reports Hopp. For example, if there is too little APC or its activity is restricted, the risk of a clot forming in the bloodstream increases, thereby causing thrombosis, heart attack or stroke. Indeed, diseases with an increased incidence of labile heme (hemolytic diseases), such as sickle cell disease, are often associated with thrombotic complications."For this reason, the influence of heme on the enzyme APC is more significant than has probably been suspected so far," says Imhof. Furthermore, the team discovered that APC might protect the cells of the inner blood vessel wall like a bodyguard against the cytotoxic effect of heme. The researchers cultivated human endothelial cells and exposed them to heme. If APC was present at the same time, the toxic effect of heme on the cells was suppressed."We are convinced that this interaction between APC and heme is significant, because many other blood proteins we were looking for did not bind heme," says Imhof. It might be worthwhile to further investigate the impact of labile, regulatory heme on APC in order to also gain new diagnostic and therapeutically relevant insights regarding blood coagulation disorders that occur in hemolytic diseases. Imhof: "The terrain should be explored much more thoroughly than has been the case to date."
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Biotechnology
| 2,020 |
September 4, 2020
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https://www.sciencedaily.com/releases/2020/09/200904092932.htm
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Coaxing single stem cells into specialized cells
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Researchers at the University of Illinois Chicago have developed a unique method for precisely controlling the deposition of hydrogel, which is made of water-soluble polymers commonly used to support cells in experiments or for therapeutic purposes. Hydrogel mimics the extracellular matrix -- the natural environment of cells in the body.
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The researchers noticed that their technique -- which allows for the encapsulation of a single cell within a minute hydrogel droplet -- can be used to coax bone marrow stem cells into specialized cells.Their findings are reported in the journal The new technique is an improvement over existing approaches that often mix much larger amounts of hydrogel with cells in an uncontrolled manner, which can make interactions between cells and their surroundings difficult to study. The new hydrogel deposition technique may also be useful for therapeutic purposes, such as for supporting stem cells used to create new tissues."Most experiments use a very high amount of hydrogels to interface with cells, which may not reflect what is happening in the body," said UIC's Jae-Won Shin, assistant professor of pharmacology and regenerative medicine at the College of Medicine, and assistant professor of bioengineering at the College of Engineering, and corresponding author on the paper.According to Shin, the team's deposition technique brings the ratio between hydrogels and cells in-line with what is seen in the body, and importantly, precisely controls the ratio on a single cell basis.Shin and colleagues also observed that stem cells in thinner gel droplets expanded more rapidly than they did in bulk gels."We observed that stem cells expand several orders of magnitude faster in thin gel droplets, and so they experience more tension than they do in bulk gels made of the same material," said Sing Wan Wong, a postdoctoral fellow in Shin's lab and first author on the study. "We believe this tension encourages stem cells in thin gel coatings to more readily become bone cells, compared to stem cells in bulk gels."The team believes the thin hydrogel deposition technique may help in the production of bone tissue from stem cells to use as regenerative therapeutics.Stephen Lenzini, Raymond Bargi, Celine Macaraniag, James C. Lee and Zhangli Peng of UIC and Zhe Feng of the University of Notre Dame are co-authors on the paper.This research was supported by grants from the National Institutes of Health (R01HL141255, R00HL125884) and the National Science Foundation (1948347-CBET).
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Biotechnology
| 2,020 |
September 3, 2020
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https://www.sciencedaily.com/releases/2020/09/200903171443.htm
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Treatment for canine ocular condition using turmeric
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Researchers at Texas A&M University have produced a therapeutic derived from turmeric, a spice long-praised for its natural anti-inflammatory properties, that shows promise in decreasing ocular inflammation in dogs suffering from uveitis, an inflammation of the eye that leads to pain and reduced vision.
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Uveitis -- a common condition in dogs, humans, and other species -- can have many causes, often occurring secondary to infectious diseases cancer, and autoimmune diseases; it also is found in patients with longstanding cataracts and after operations correcting cataracts."Uncontrolled inflammation inside the eye, also known as uveitis, is a leading cause of complications after cataract surgery in dogs. The management of postoperative ocular inflammation is a major challenge observed in both human and veterinary ophthalmology," said Dr. Erin Scott, an assistant professor at the Texas A&M University College of Veterinary Medicine & Biomedical Sciences.In a recent paper published in Oral medications currently used to treat uveitis must be adequately absorbed into the blood stream for their medicinal effects to be effective. This requires the medication to successfully pass through the intestinal barrier -- the physical barrier between the gut and the rest of the body via the circulatory system -- which limits the absorption of many drugs.Drug delivery to the eye presents additional challenges because of the blood-ocular barrier -- the physical barrier between blood vessels and tissues of the eye -- which tightly controls what substances can pass into the eye.Therefore, researchers must find ways to bypass such barriers to improve drug availability within the body.Scott and her colleagues' research implemented a novel formulation of curcumin that improved transport of the substance across both intestinal and ocular barriers. By adding nanoparticle molecules that interact with receptors on a ubiquitous transmembrane carrier protein, known as the transferrin receptor, curcumin is able to hitch a ride across crucial barriers, improving absorption of the substance and reducing ocular inflammation.Curcumin is especially attractive as a candidate for management of uveitis because it has no known side effects."Current treatments include a combination of systemic and topical anti-inflammatory medications, either in the form of steroids or non-steroidal anti-inflammatory drugs (NSAIDs)," Scott said. "While both these medications are effective in the treatment of uveitis, they can cause unwanted side effects, such as vomiting, diarrhea, stomach ulcers, negatively impact kidney and liver function, and increase glucose levels in diabetic patients."Scott and her colleagues hope to start a clinical trial in the Texas A&M Veterinary Medical Teaching Hospital using this new medication in the near future and are optimistic that the utility of their findings may benefit populations beyond dogs."This medication may translate to the treatment of cataracts and uveitis in humans," she said. "By studying animal patients with naturally occurring eye diseases, our findings may accelerate the development of medications to benefit both animals and humans."
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Biotechnology
| 2,020 |
September 3, 2020
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https://www.sciencedaily.com/releases/2020/09/200903145025.htm
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Structure of mRNA initiation complex could give insight into cancer and other diseases
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Researchers at the University of California, Davis and the MRC Laboratory of Molecular Biology in Cambridge, U.K. have solved the the structure of the complex formed when mRNA is being scanned to find the starting point for translating RNA into a protein. The discovery, published Sept. 4 in Science, provides new understanding of this fundamental process.
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"This structure transforms what we know about translation initiation in human cells and there has been a tremendous excitement from people in the field," said Christopher Fraser, professor of molecular and cellular biology at UC Davis and corresponding author on the paper.Although nearly all our cells contain our entire genome, cells use different subsets of genes to make the proteins they need to perform their various functions. This requires precise control over the processes by which the DNA is first transcribed to produce mRNA and then mRNA is translated to make protein.ranslation begins when a ribosome attaches to a piece of mRNA and scans along it until it finds a start codon, three letters of RNA that say "start translating here." There are over a dozen different proteins known as initiation factors involved in this process. Many of these initiation factors have been found to be dysregulated in various cancers.However, just how the factors come together and scan mRNA has been poorly understood, due to the lack of understanding of the structures of the entire complex.To investigate this, Fraser and postdoctoral researcher Masaaki Sokabe at the UC Davis Department of Molecular and Cellular Biology collaborated with Venki Ramakrishnan, Jailson Brito Querido, Sebastian Kraatz and Yuliya Gordiyenko at the LMB to visualise the structure of the complex. Ramakrishnan shared the 2009 Nobel Prize for Chemistry for his work on the structure of the ribosome.The team used an mRNA that lacked a start codon so that it would be trapped in the act of scanning. While big for a biological machine, you could fit about 3000 of these complexes across the width of a human hair. The team therefore used cryoelectron microscopy at the LMB to obtain a structure of the complex including the trapped mRNA. Cryoelectron microscopy allows biologists to capture three-dimensional movies of biological molecules down to the scale of single atoms.Based on this structure, the researchers proposed a model of how the mRNA slots into a channel in the small ribosomal subunit, and a mechanism for how the mRNA might be pulled through the ribosome for scanning, like a strip of film through an old-style projector.They were able to predict that for most mRNAs, the start codon would need to be sufficiently far from the front end of the mRNA for it to be found in the scanning process, which was the confirmed biochemically by Sokabe and Fraser. Further conformation of the model was obtained by mass spectrometry carried out by Mark Skehel of the LMB.The UC Davis College of Biological Sciences recently opened its own cryoelectron microscopy facility, which will make this kind of work possible on campus, Fraser said.The work was funded by UKRI MRC, Federation of European Biochemical Societies, Wellcome, Louis-Jeantet Foundation, and the National Institutes of Health.
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Biotechnology
| 2,020 |
September 3, 2020
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https://www.sciencedaily.com/releases/2020/09/200903133019.htm
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Cell division: Cleaning the nucleus without detergents
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A team of researchers, spearheaded by the Gerlich lab at IMBA, has uncovered how cells remove unwanted components from the nucleus following mitosis. The results, published in the journal
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Organization of cells into specific compartments is critical for their function. For instance, by separating the nucleus from the cytoplasm, the nuclear envelope prevents premature translation of immature RNAs. During mitosis, however, the nuclear envelope disassembles, allowing large cytoplasmic components such as ribosomes to mix with nuclear material. When the nuclear envelope reassembles following mitosis, these cytoplasmic components must once again be removed. "The nuclear envelope can contribute to this by actively importing or exporting substrates up to a certain size, but it was not clear what happens with very large cytoplasmic components," says Mina Petrovic, PhD student in the Gerlich lab and joint first author of the study.The research team from IMBA and EMBL have now shown that large components such as ribosomes are in fact removed from the forming nucleus before the nuclear envelope is assembled again. This exclusion process requires the protein Ki-67, which was the focus of an earlier publication in This important work shows how a single protein can dynamically change the material properties of cellular components to regulate compartmentalization of key processes within the cell.
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Biotechnology
| 2,020 |
September 3, 2020
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https://www.sciencedaily.com/releases/2020/09/200903133027.htm
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Excitable cells: Tracking the evolution of electrical signaling in plants
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A study led by researchers from Tasmania, Chile and Germany has furthered our understanding of plant evolution by tracking the origins of electrical signalling components that plants developed to communicate and adapt to life on land.
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The research team, including University of Tasmania plant scientist Dr Frances Sussmilch, studied DNA sequences of diverse plant species to map the evolutionary origins of important adaptations."By looking closely at the sequences for potassium channels in these species, we were excited to see evidence of ancient evolutionary events that have led to the complex cell-to-cell signalling mechanisms that can be seen in modern-day plants," Dr Sussmilch said.This story began around 500 million years ago when the algal ancestors of land plants emerged from water and conquered dry land.Plants faced new challenges on land, including difficult and changeable environmental conditions such as drought, heat, UV and wind, and interactions with new biological threats."These ancient plant pioneers made the landscape more hospitable for the first terrestrial animals and gave rise to the incredible diversity of land plants we see today, including bryophytes, ferns, gymnosperms and flowering plants," Dr Sussmilch said."A feature that likely helped plants to thrive on land is the presence of molecular mechanisms for cell-to-cell signalling. This enables communication between cells in the plant body, facilitating coordinated responses to environmental changes."One means of rapid cell-to-cell communication that operates in plants is action-potential-based electrical signalling, which is comparable to the electrical impulses that control the synchronised beat of the human heart.In plants, action potentials can be used for rapid responses to signals including wounding or touch, such as a Venus flytrap snapping shut. For electrical communication, plants need voltage-gated ion channels which control the flow of ions in and out of a cell in response to stressors.Dr Sussmilch said channels that enable the movement of potassium ions across the cell membrane in plants play a critical role in electrical excitability.The researchers made use of gene sequence resources for species representing each of the major groups of land plants and green algae.They examined the sequences of potassium channels from these diverse plants for sequence 'fingerprints' indicative of different structural and functional characteristics.The study found sequences with the fingerprint characteristic of one group of related efflux channels, that enable potassium ions to exit cells, was preserved between most land plant lineages -- liverworts, hornworts, ferns, gymnosperms and flowering plants -- and their closest living algal relatives."This suggests that this channel type may have already been present in plants when they first emerged from the water and was likely retained as land plants continued to evolve due to its importance for cell-to-cell communication," Dr Sussmilch said.However, this channel type was absent in all moss species examined, indicating that it may have become unnecessary in the niches that early mosses occupied and been lost from the genome of a common ancestor of the mosses alive today.In contrast, they found that the fingerprints for four distinct types of related influx channels, that enable potassium ions to enter cells, have diversified more recently, allowing for an increased variety of channel functions in flowering plants.The research findings, together with a sequence analysis pipeline that can be used to examine the evolution of other gene families in the same way, have recently been published in the journal
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902182421.htm
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New treatment for drug-resistant bacterial infections
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A new antibacterial agent that has been engineered by researchers at Dartmouth to essentially hide from the human immune system may treat life-threatening MRSA infections. A new paper, published today in
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The Centers for Disease Control and Prevention (CDC) has prioritized finding effective treatment of Methicillin-resistant Staphylococcus aureus (MRSA), one of the most common bacterial pathogens and the single most deadly drug-resistant bacteria in the United States. Now, a new study led by Dartmouth Engineering faculty shows promise for an engineered lysin-based antibacterial agent that may enable safe, repeated dosing to treat life-threatening infections by MRSA and other types of S. aureus.In recent years, lysins -- enzymes naturally produced by microbes and associated viruses -- have shown potential to treat S. aureus, which can rapidly acquire resistance to other types of antibiotic drugs."Lysins are one of the most promising next-generation antibiotics. They kill drug-sensitive and drug-resistant bacteria with equal efficacy, they can potentially suppress new resistance phenotypes, and they also have this laser-like precision," said Karl Griswold, corresponding author and associate professor of engineering at Dartmouth.While there is promise in lysins, development has been slowed due to concerns that they prompt humans' immune systems to develop antidrug antibodies, which can have negative side effects including life-threatening hypersensitivity reactions.That's why the Dartmouth Engineering team -- which also included researchers in Dartmouth's computer science department, The Lundquist Institute at Harbor-UCLA Medical Center, Lyticon, and Stealth Biologics -- engineered and patented F12, a new lysin-based antibacterial agent. F12 is essentially able to hide from the human immune system (due to T cell epitope deletion), and therefore does not cause the same negative side effects as unmodified, natural lysins.F12 is the first lysin-based treatment with the potential to be used multiple times on a single patient, making it ideal to treat particularly persistent drug-resistant and drug-sensitive infections. Preclinical studies showed the efficacy of F12 does not diminish with repeated doses, while two other anti-MRSA lysin treatments currently in clinical trials are only designed to be used a single time."We have engineered this super potent, super effective anti-MRSA biotherapeutic, and we've done it in a way that renders it compatible with and largely invisible to the human immune system. By making it a safer drug, we've enabled the possibility of dosing multiple times in order to treat even the most highly refractory infections," said Griswold.The team's paper, "Globally deimmunized lysostaphin evades human immune surveillance and enables highly efficacious repeat dosing," was published earlier today by The paper details the treatment's positive results in rabbits, mice with partially-humanized immune systems, and studies with extracted human immune cells. Griswold believes the antibacterial agent could be ready for human clinical trials as soon as 2023."This is the first report of a translation-ready deimmunized lysin, and F12 has serious, bonafide clinical potential," said Griswold.Further studies of F12 will examine synergy with standard-of-care antibacterial chemotherapies; preliminary results suggest the combinations are extremely potent and suppress drug-resistance phenotypes.
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902182419.htm
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Newly identified gene grants tomatoes resistance to bacterial speck disease
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Bacterial speck disease, which reduces both fruit yield and quality, has been a growing problem in tomatoes over the last five years. Because the culpable bacterium, Pseudomonas syringae, prefers a cool and wet climate, crops in places such as New York State have been particularly susceptible.
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Recent research at the Boyce Thompson Institute headed by postgraduates Carolina Mazo-Molina and Samantha Mainiero and overseen by faculty member Greg Martin may change this. Published in the August issue of Another resistance gene, Pto, which provides resistance to race 0 strains of Pseudomonas syringae, has been used for over 25 years. However, crops remain vulnerable to the increasingly common race 1 strain, resulting in significant losses for growers.With the discovery of this new gene, which the researchers have dubbed Pseudomonas tomato race 1 (Ptr1), damage caused by bacterial speck disease may soon become a thing of the past."We are working with plant breeders now to introduce the Ptr1 gene into tomato varieties that already have Pto," explains Martin, who is also a professor at Cornell University's School of Integrative Plant Science. "If you do that, then you will have resistance to all known bacteria that cause speck disease."The project started in 2015, after a chance outbreak of bacterial speck disease at one of Cornell's research farms, where BTI faculty member Jim Giovannoni was researching tomato fruit quality. Giovannoni is also a USDA scientist and an Adjunct Professor in Cornell's School of Integrative Plant Science."Speck disease wiped out their entire trial except for two plants," explains Martin. "Both of those plants turned out to have the Ptr1 resistance gene. This was a remarkable coincidence of natural selection and serendipity."The two plants that survived both contained a gene derived from Solanum lycopersicoides, a wild relative of the cultivated tomato. By collecting seeds of the plants and studying their inheritance patterns, the team determined that a single region on one chromosome is responsible for conferring resistance, work that was published in Mazo-Molina described the thrill of identifying the gene. "When we found Ptr1, I would always say it 'might be the gene' or 'could be the gene,'" she explains. "But at some point, I was able to tell myself, 'This is the gene. You don't have to doubt it.'"Ptr1 codes for a protein that indirectly detects the presence of a pathogenic protein called AvrRpt2. Both apple and the popular model plant Arabidopsis have genes that encode proteins that also recognize the same bacterial protein."The three proteins are completely different," says Martin. "There's no similarity at all. It looks like an example of convergent evolution, independent solutions in different plants to the same problem.""Because detection of AvrRpt2 evolved multiple times across evolutionary history, the AvrRpt2 protein likely plays a key role in the pathogen's ability to infect plants," Martin says.Now that the gene has been identified, the team is focused on developing tomato varieties that carry the Ptr1 gene. "The wild species in which Ptr1 is naturally found is really difficult to cross with cultivated tomatoes," explains Mainiero. "We can't just use traditional breeding methods."Thankfully, there may be another way."A defective form of the gene is present in many tomato varieties already," says Martin, "with natural mutations having made it nonfunctional. There is a new type of gene editing technology called CRISPR Prime Editing that might allow us to go in and repair this defective gene."Mainiero plans to work on the CRISPR Prime Editing project, and Mazo-Molina will focus on understanding the molecular mechanism of action underlying Ptr1.Martin emphasizes that the collaboration between the Mainiero and Mazo-Molina was key to the project's success. "It is a great example of a collaboration within a lab between two different lab members," he says. "They worked seamlessly as a team."
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902152836.htm
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Experimental vaccine that boosts antigen production shows promise against COVID-19
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A bioengineering technique to boost production of specific proteins could be the basis of an effective vaccine against the novel coronavirus that causes COVID-19, new research suggests.
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Scientists manipulated a natural cellular process to ramp up levels of two proteins used by the virus to infect other cells, packaged the protein-boosting instructions in nanoparticles and injected them into mice. Within a month, the mice had developed antibodies against the SARS-CoV-2 virus.The technique involves altering specific sequences of messenger RNA, molecules that translate genetic information into functional proteins. While these sequences are not translated to proteins, the researchers changed their structures to promote higher-than-usual levels of proteins. The sequences are known as untranslated regions, or UTRs."We've been engineering messenger RNA for four years, and earlier this year we made some progress identifying a role for UTRs -- and then COVID-19 happened," said Yizhou Dong, senior author of the study and associate professor of pharmaceutics and pharmacology at The Ohio State University.Though Phase 3 clinical trials of fast-tracked COVID-19 vaccine candidates are in progress, Dong said his lab's platform offers a potential alternative."If the current vaccines work well, that's wonderful. In case the field needs this, then it's an option. It worked as a vaccine is expected to, and we can scale this up very fast," he said. "For now, it's a proof of concept -- we've demonstrated we can optimize a sequence of messenger RNA to improve protein production, produce antigens and induce antibodies against those specific antigens."The study is published today in the journal The crux of the method is typical to vaccine development: using snippets of a pathogen's structure to produce an antigen -- the foreign substance that triggers an appropriate immune response -- and finding a safe way to introduce it to the body.But the engineering technique takes antigen design to a new level by making use of messenger RNA UTRs, Dong said.His lab worked with the two UTRs that bookend the start and finish of protein assembly, functioning as regulators of that process and influencing how the resulting protein interacts with others. UTRs themselves are strings of nucleotides, the molecules that compose RNA and DNA."For our application we tried to optimize the UTRs to improve the protein production process. We wanted as much protein produced as possible -- so we can give a small dose of messenger RNA that produces enough antigen to induce antibodies against the virus," Dong said.The team experimented with two potential antigens that the novel coronavirus is known to use to cause infection: a spike protein on its surface and a receptor binding domain, a component of the spike protein, that the virus uses to make its way into host cells -- a necessary step to make copies of itself. Both are used in other SARS-CoV-2 vaccine candidates.After manipulating the messenger RNA for these two proteins, the team encased them in lipid nanoparticles developed previously in Dong's lab. They injected mice with the experimental vaccine and gave them a booster two weeks later. A month after the first injection, immune cells in the mice had taken up the antigens of the two proteins and developed antibodies against them."It takes some time for the immune system to process the antigens and have cells produce antibodies," Dong said. "In this study, we detected antibodies after 30 days."And even if this vaccine candidate is not needed for COVID-19, he is continuing to refine this latest method of engineering messenger RNA."UTR is a platform that we can apply to any type of messenger RNA. We are exploring other therapeutics," Dong said.This work was funded by a National Institutes of Health Maximizing Investigators' Research Award, the National Institute of General Medical Sciences and the Ohio State College of Pharmacy startup fund.Co-authors, all from Ohio State, include Chunxi Zeng, Xucheng Hou, Jingyue Yan, Chengxiang Zhang, Wenqing Li, Weiyu Zhao and Shi Du.
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902115932.htm
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An unprecedented discovery of cell fusion
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Like humans, bacteria live together in communities, sometimes lending a hand -- or in the case of bacteria, a metabolite or two -- to help their neighbors thrive. Understanding how bacteria interact is critical to solving growing problems such as antibiotic resistance, in which infectious bacteria form defenses to thwart the medicines used to fight them.
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Now, researchers at the University of Delaware have discovered that bacteria do more than just work together. Bacterial cells from different species can combine into unique hybrid cells by fusing their cell walls and membranes and sharing cellular contents, including proteins and ribonucleic acid (RNA), the molecules which regulate gene expression and control cell metabolism. In other words, the organisms exchange material and lose part of their own identity in the process.This unprecedented observation, which was reported on Tuesday, Sept. 1 in The research team, led by Eleftherios (Terry) Papoutsakis, Unidel Eugene Du Pont Chair of Chemical and Biomolecular Engineering, studied interactions between The team found that "They mix their machinery to survive or do metabolism, and that's kind of extraordinary, because we always assumed that each and every organism has its own independent identity and machinery," said Papoutsakis.Previously, researchers have observed that bacteria could exchange some material through nanotubes. The combination into hybrid cells was unexpected."This is the first time we've shown this in this bacteria, and it's also a new mechanism of how material is exchanged," said Kamil Charubin, a doctoral student in chemical and biomolecular Engineering and first author of the paper.Although this phenomenon of interspecies microbial fusion is now being reported for the first time, it is likely ubiquitous in nature among many bacterial pairs.So why do bacteria bother to fuse together? The simple answer is likely because this process allows the microbes to share machinery that will increase their odds of survival.For example, some pathogenic bacteria -- those that can cause disease -- may borrow proteins from other antibiotic-resistant bacteria in order to shore up their own resistance. Some bacteria might borrow machinery from others in order to evade detection by the immune system. This could also help to explain why some bacteria are difficult to culture, or grow for study or medical diagnostic purposes. These difficult-to-culture bacteria might combine with or work with and depend on other microorganisms for their existence instead of growing and multiplying on their own.The team's findings may influence understanding of the evolution of biology because once bacterial species share machinery, they can evolve together instead of only evolving on their own, said Papoutsakis."These findings will guide new thinking in not just the field of microbial evolution, but also toward biotechnological solutions that can benefit the soldier," said Dr. Robert Kokoska, program manager, Army Research Office (ARO), an element of the U.S. Army Combat Capabilities Development Command's Army Research Laboratory. "These include studies of how the human microbiome shapes soldier human health and cognition and how microbial communities can be better designed for a broad range of advances including strategies for reliable in-field biological sensing, waste remediation and novel means of biosynthesis."This work was supported by the Army Research Office (award no. W911NF-17-1-0343, and W911NF-19-1-0274) and the U.S. Department of Energy (DE-SC0019155).
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902095126.htm
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Viruses on glaciers highlight evolutionary mechanism to overcome host defenses
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Viruses are often thought of as a human problem, however they are the most abundant biological entities on the planet. There are millions of viruses in every teaspoon of river, lake or seawater, they are found everywhere there is life and probably infect all living organisms. Most are completely harmless to humans and infect microscopic animals, plants and bacteria, which they hijack and reprogram to produce new virus particles, most often destroying these cells in the process. Every day, viruses destroy huge number of microorganisms in the environment, which changes the flow of energy in food webs on global scales. "Understanding how viruses evolve and function allows us to predict their role in the environment and how they interact with their hosts," says Christopher Bellas from the Department of Ecology at the University of Innsbruck. Together with colleagues from the Universities of Bristol, Reading and Aberystwyth in the UK, the University of Minnesota, USA, and Aarhus University in Denmark, he has sequenced and compared genomes (their total DNA) of viruses which infect microbes found on the surface of glaciers. The study, now published in the journal
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It is known from laboratory studies that viruses evolve rapidly in order to keep up with their hosts, which are also simultaneously evolving defenses against virus infection, this evolutionary arms race means that they should remain in balance relative to each other. This is known as the 'Red Queen' hypothesis after the character from Alice in Wonderland who states: It takes all the running you can do, to keep in the same place. "This means that when we sequence the genomes of viruses from two, long-term, isolated places, we should never find exactly the same virus genomes twice," says Christopher Bellas. The viruses studied by the research team originate in very unusual habitats on the surface of glaciers and ice sheets, called cryoconite holes. These small pools of meltwater on glaciers are ideal places to test how viruses evolve because they are miniature, replicated communities of microbes which are found on widely separated glaciers around the world.When the researchers looked at the virus genomes from isolated cryoconite holes, thousands of kilometers apart, they expected to find that they would each contain different viruses only distantly related to one another. What they actually found was that most bacterial infecting viruses (bacteriophages) were nearly identical between the Arctic and the Alps. However, when they looked closer at their stable genomes, they saw that there were many small sections in each genome where DNA from other related viruses was repeatedly swapped in and out, in a known process called recombination. In each different location, the viruses shuffled the genes present in these swappable regions like a kind of genetic fruit machine. "This means that in the natural environment, gene swapping between viruses by recombination creates much diversity in the virus population, specifically in genes which are involved in recognising and attaching to different hosts, this probably gives viruses the potential to quickly adapt to different hosts in the environment," explains Christopher Bellas.
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902091110.htm
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Microbial genetics: A protean pathogen
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The bacterium
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It is estimated that about half of the world's population is infected with the bacterium The researchers were able to make use of a unique set of samples obtained in the course of a vaccine trial performed in collaboration with colleagues from the Max Planck Institute for Infection Biology in Berlin in which adult volunteers were infected with H. pylori. Ten weeks later, bacterial samples were isolated from two different regions of the stomach in each case, and the donors were then treated with antibiotics to eliminate the pathogen. Complete genome sequences of single-cell isolates obtained from these samples were determined using state-of-the-art single-molecule, real-time (SMRT) sequencing methods and compared with that of the H. pylori strain with which the volunteers had been infected nearly 3 months previously.The results revealed that the bacterial genomes had undergone a surprising degree of diversification within this relatively short time span. Many of the mutations detected were located in genes that are directly involved in molecular interactions between the pathogen and host cells. The set of proteins affected included polypeptides that are expressed on the cell wall of the bacterium, as well as transport proteins that are found in its outer membrane. These findings suggest that specific genes are being positively selected to enable the bacterium to exploit metabolites that are available in different cellular niches in the lining of the stomach. In addition to these genetic mutations (which alter gene products), the authors noted a variety of epigenetic changes (which alter gene regulation) in the bacterial DNA during the early phase of infection. H. pylori possesses many enzyme systems that attach methyl groups to specific DNA sequences. The authors identified 24 such systems, and at least some of these may serve to regulate the expression of sets of genes in the bacterium. These results further support the notion that alterations in gene products and patterns of gene activity during the weeks and months following the primary infection play an important role in enabling the bacterium to adapt to, and become established in different niches in the stomach.
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Biotechnology
| 2,020 |
September 2, 2020
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https://www.sciencedaily.com/releases/2020/09/200902182435.htm
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Effective cancer immunotherapy further linked to regulating a cell 'suicide' gene
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Johns Hopkins Medicine researchers have added to evidence that a gene responsible for turning off a cell's natural "suicide" signals may also be the culprit in making breast cancer and melanoma cells resistant to therapies that use the immune system to fight cancer. A summary of the research, conducted with mice and human cells, appeared Aug. 25 in
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When the gene, called BIRC2, is sent into overdrive, it makes too much, or an "overexpression," of protein levels. This occurs in about 40% of breast cancers, particularly the more lethal type called triple negative, and it is not known how often the gene is overexpressed in melanomas.If further studies affirm and refine the new findings, the researchers say, BIRC2 overexpression could be a key marker for immunotherapy resistance, further advancing precision medicine efforts in this area of cancer treatment. A marker of this kind could alert clinicians to the potential need for using drugs that block the gene's activity in combination with immunotherapy drugs to form a potent cocktail to kill cancer in some treatment-resistant patients."Cancer cells use many pathways to evade the immune system, so our goal is to find additional drugs in our toolbox to complement the immunotherapy drugs currently in use," says Gregg Semenza, M.D., Ph.D., the C. Michael Armstrong Professor of Genetic Medicine, Pediatrics, Oncology, Medicine, Radiation Oncology and Biological Chemistry at the Johns Hopkins University School of Medicine, and director of the Vascular Program at the Johns Hopkins Institute for Cell Engineering.Semenza shared the 2019 Nobel Prize in Physiology or Medicine for the discovery of the gene that guides how cells adapt to low oxygen levels, a condition called hypoxia.In 2018, Semenza's team showed that hypoxia essentially molds cancer cells into survival machines. Hypoxia prompts cancer cells to turn on three genes to help them evade the immune system by inactivating either the identification system or the "eat me" signal on immune cells. A cell surface protein called CD47 is the only "don't eat me" signal that blocks killing of cancer cells by immune cells called macrophages. Other cell surface proteins, PDL1 and CD73, block killing of cancer cells by immune cells called T lymphocytes.These super-survivor cancer cells could explain, in part, Semenza says, why only 20% to 30% of cancer patients respond to drugs that boost the immune system's ability to target cancer cells.For the current study, building on his basic science discoveries, Semenza and his team sorted through 325 human genes identified by researchers at the Dana Farber Cancer Institute in Boston whose protein products were overexpressed in melanoma cells and linked to processes that help cancer cells evade the immune system.Semenza's team found that 38 of the genes are influenced by the transcription factor HIF-1, which regulates how cells adapt to hypoxia; among the 38 was BIRC2 (baculoviral IAP repeat-containing 2), already known to prevent cell "suicide," or apoptosis, in essence a form of programmed cell death that is a brake on the kind of unchecked cell growth characteristic of cancer.BIRC2 also blocks cells from secreting proteins that attract immune cells, such as T-cells and natural killer cells.First, by studying the BIRC2 genome in human breast cancer cells, Semenza's team found that hypoxia proteins HIF1 and HIF2 bind directly to a portion of the BIRC2 gene under low oxygen conditions, identifying a direct mechanism for boosting the BIRC2 gene's protein production.Then, the research team examined how tumors developed in mice when they were injected with human breast cancer or melanoma cells genetically engineered to contain little or no BIRC2 gene expression. In mice injected with cancer cells lacking BIRC2 expression, tumors took longer to form, about three to four weeks, compared with the typical two weeks it takes to form tumors in mice.The tumors formed by BIRC2-free cancer cells also had up to five times the level of a protein called CXCL9, the substance that attracts immune system T-cells and natural killer cells to the tumor location. The longer the tumor took to form, the more T-cells and natural killer cells were found inside the tumor.Semenza notes that finding a plentiful number of immune cells within a tumor is a key indicator of immunotherapy success.Next, to determine whether the immune system was critical to the stalled tumor growth they saw, Semenza's team injected the BIRC2-free melanoma and breast cancer cells into mice bred to have no functioning immune system. They found that tumors grew at the same rate, in about two weeks, as typical tumors. "This suggests that the decreased tumor growth rate associated with loss of BIRC2 is dependent on recruiting T-cells and natural killer cells into the tumor," says Semenza.Finally, Semenza and his team analyzed mice implanted with human breast cancer or melanoma tumors that either produced BIRC2 or were engineered to lack BIRC2. They gave the mice with melanoma tumors two types of immunotherapy FDA-approved for human use, and treated mice with breast tumors with one of the immunotherapy drugs. In both tumor types, the immunotherapy drugs were effective only against the tumors that lacked BIRC2.Experimental drugs called SMAC mimetics that inactivate BIRC2 and other anti-cell suicide proteins are currently in clinical trials for certain types of cancers, but Semenza says that the drugs have not been very effective when used on their own."These drugs might be very useful to improve the response to immunotherapy drugs in people with tumors that have high BIRC2 levels," says Semenza.
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Biotechnology
| 2,020 |
September 1, 2020
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https://www.sciencedaily.com/releases/2020/09/200901142725.htm
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Study tracks human milk nutrients in infant microbiome
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A new study in mice helps explain why gut microbiomes of breastfed infants can differ greatly from those of formula-fed infants.
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The study, "Dietary Sphinganine Is Selectively Assimilated by Members of the Mammalian Gut Microbiome," was published in July in the Sphinganine from milk Johnson Lab/Provided A new technique allows researchers to track specific nutrients as they are taken up by gut microbes in a mouse's digestive tract. The image shows certain microbes (red) taking in a nutrient common in human milk called sphinganine; blue microbes have not taken it in.The paper describes an innovative technique developed at Cornell to track the fate of metabolites -- nutrients formed in or necessary for metabolism -- through a mouse's digestive tract and identify how they interact with specific gut microbes."We think the methods are expandable to many different microbiome systems," said senior author Elizabeth Johnson, assistant professor of nutritional sciences in the College of Agriculture and Life Sciences. She noted that researchers investigating effects of a high-fat vs. low-fat diet, or a keto diet, might use the technique to track metabolites.The methodology could reveal how specific metabolites promote specific bacteria. This could allow nutritionists to prescribe that patients eat foods containing specific metabolites to intentionally change the composition of their microbiomes, Johnson said.Human milk and many other foods contain a class of lipid metabolites called sphingolipids. Previous research suggested that these metabolites help shape an infant's microbiome, but it was not known if they actually interact with the microbiome.The study identified two types of gut microbes, Bacteroides and Bifidobacterium, that use sphingolipids for their own metabolism.While very little is known about the specific roles of gut microbes in human health, Bacteroides have been implicated in both beneficial and not-so-beneficial effects, depending on context. They are generally associated with microbiomes of healthy breastfed infants. Bifidobacterium, shown for the first time in this study to process dietary sphingolipids, are considered the quintessential beneficial bacteria, comprising up to 95% of breastfed infants microbiome.They're also a highly popular over-the-counter probiotic."Our lab is very interested in how the diet interacts with the microbiome in order to really understand how you can best modulate it to have positive effects on health," Johnson said. "In this study, we were able to see that yes, these dietary lipids that are a big part of [breastfed] infants diets, are interacting quite robustly with the gut microbiome."Sphingolipids originate from three main sources: diet; bacteria that can produce them; and most host tissues.Johnson, along with first author Min-Ting Lee, a doctoral student, and Henry Le, a postdoctoral researcher, both in Johnson's lab, created a technique to specifically track dietary sphingolipids as they passed through the mouse gut."We custom synthesized the sphingolipid we added to the diet," Johnson said. "It is almost identical to ones derived from breast milk but with a small chemical tag so we could trace the location of the sphingolipid once it was ingested by the mice."Lee then used a fluorescent label that attached to cells or microbes that absorbed the tagged lipid, such that any bacteria that had taken up sphingolipids lit up red. Microbes from the mice's microbiomes were then isolated and analyzed. Populations with red microbes were separated from the others, and these were then genetically sequenced to identify the species of bacteria.With further investigation, Le was able to identify the metabolites that Bacteroides and Bifidobacterium produce when exposed to dietary sphingolipids. Further investigations are underway to determine whether these microbially-produced metabolites are beneficial for infant health.Johnson recently received a five-year, $1.9 million Maximizing Investigators' Research Award from the National Institutes of Health (NIH) to expand on this work, to better understand how lipid-dependent host-microbe interactions affect human health..The study was supported by seed funds from the Genomics Facility of the Biotechnology Resource Center at Cornell's Institute of Biotechnology.
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Biotechnology
| 2,020 |
September 1, 2020
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https://www.sciencedaily.com/releases/2020/09/200901125909.htm
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Scientists shed new light on pollen tube growth in plants
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New insight on how an enzyme ensures the correct growth of pollen tubes in flowering plants has been published today in the open-access journal
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The study reveals an unexpected role of KATANIN in moderating the mechanical properties of the papilla cell wall in Arabidopsis thaliana (A. thaliana), thereby preventing disordered pollen tube growth and allowing the tube to find its correct path to the underlying female plant tissues. These findings suggest that KATANIN has likely played a major role in the success of flowering plants on earth more widely.Seeds are produced in flowering plants when male and female germ cells called gametes fuse together. Male gametes are contained in the pollen grain while female gametes are found in the ovules, which are embedded in a female reproductive organ called the pistil. For successful seed production to happen, pollen grains need to meet with the surface of the pistil, which is composed of a layer of elongated cells called papillae. When a pollen grain lands on a papilla, it rehydrates and then produces a tube that will carry the male gametes toward the ovules.Pollen tubes grow first within the papilla cell wall, exerting a physical pressure on the cell. After crossing the papilla layer, they then grow in the intercellular space of underlying tissues. The pistil then produces compounds that guide the pollen tube to the ovules where it reaches the female gametes. But how the tube orients itself when it emerges from the pollen at the papilla surface remains unknown."It is striking that, whatever the position of the flower and hence the pistil on the stem, the pollen tube grows to the base of the papilla in the direction of the ovules. We wanted to explore the mechanisms that allow for this proper orientation of pollen tubes on the papilla cells," says lead author Lucie Riglet, who was a PhD student in senior author Thierry Gaude's lab at the Laboratory of Plant Reproduction and Development, ENS Lyon, France, at the time the study was carried out, and is now a postdoctoral researcher at the Sainsbury Laboratory, University of Cambridge, UK.Mechanical forces are known to play a major role in plant cell shape by controlling the orientation of cortical microtubules, which in turn mediate the deposition of cellulose microfibrils. For their study, Riglet and her team combined imaging, genetic and chemical approaches to show that the enzyme KATANIN, which cuts microtubules, also acts on cellulose microfibril orientation and confers mechanical properties to the papilla cell wall that allow for correct pollen tube orientation."By forcing the pollen tubes to take the right direction from their early places in the papilla, KATANIN has likely played a major role in the success of flowering plants on earth by promoting fertilisation," explains senior author Thierry Gaude, Group Leader at the Laboratory of Plant Reproduction and Development, ENS Lyon. "As KATANIN is found in most organisms, including humans, it is possible that the enzyme plays a role in regulating mechanical properties in other processes -- but this is a fascinating question that remains to be explored."
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Biotechnology
| 2,020 |
September 1, 2020
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https://www.sciencedaily.com/releases/2020/09/200901120743.htm
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Gene targets to combat microorganisms binding to underwater surfaces
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A group of synthetic biologists at NYU Abu Dhabi (NYUAD) have identified new genetic targets that could lead to safe, biologically-based approaches to combat marine biofouling -- the process of sea-based microorganisms, plants, or algae binding to underwater surfaces. Biofouling continues to present significant challenges for aquaculture and sea-based commercial activities, with one of the most common examples being found on the bottom of cargo ships, where the presence of attached marine organisms can change the hydrodynamics of ships, causing damage, and increasing fuel consumption.
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In addition to its financial and operational impacts, biofouling has ecological consequences as it can introduce invasive species to new environments when the ships change locations. The current method for preventing biofouling is a chemical-based substance that is toxic to marine ecosystems.A new study, led by NYUAD Research Scientist Weiqi Fu and Associate Professor of Biology Kourosh Salehi-Ashtiani, has identified 61 key signaling genes, some encoding protein receptors, that are turned on during surface colonization of a dominant group of phytoplankton (microscopic marine algae). The NYUAD researchers show that by increasing the level of the discovered genes and protein receptors, the biofouling activities of these marine-based planktonic cells can be manipulated. This study paves the way for the creation of new environmentally-friendly antifouling methods.In the paper, titled GPCR Genes as Activators of Surface Colonization Pathways in a Model Marine Diatom, published in the interdisciplinary journal "As marine biofouling on immersed artificial structures such as ship hulls, aquaculture cage facilities, and seawater handling pipes has had serious economic implications, there is a great need to discover a safe antifouling method," said Salehi-Ashtiani. "The receptors and signaling pathways described in this study pave the way for the targeted development of new antifouling techniques that are less harmful to global marine ecosystems," adds Fu, the lead author of the paper.At the beginning of the 21st century, the International Maritime Organization banned the use of many widely used antifouling methods that were chemically-based due to their high toxicity towards marine organisms. Since then, there has been a surge in research to discover an environmentally-friendly antifouling technique. As the mechanics of biofouling on both the cellular and molecular levels were previously unknown, the signaling genes and protein receptors identified by this study provide key insight into targets for future ecologically safe antifouling methods.
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Biotechnology
| 2,020 |
August 31, 2020
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https://www.sciencedaily.com/releases/2020/08/200831154410.htm
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Dodder uses the flowering signal of its host plant to flower
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The plant genus Cuscuta consists of more than 200 species that can be found almost all over the world. The parasites, known as dodder, but also called wizard's net, devil's hair or strangleweed, feed on other plants by attaching themselves to their hosts via a special organ, the haustorium, and withdrawing nutrients from them. They have neither roots nor leaves. Without leaves, they are hardly able to photosynthesize. Without roots they cannot absorb nutrients and water from the soil. On the other hand, they are integrated into the internal communication network of their host plants and can even pass on warning signals from plant to plant.
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A team of scientists led by Jianqiang Wu, who has been the leader of a Max Planck Partner Group at the Kunming Institute of Botany, Chinese Academy of Sciences, now asked how the parasites manage to synchronize flowering with their hosts. They had observed that plants of the Australian dodder (Cuscuta australis) adjusted the time of their flowering to that of their respective host plant species."The flowering time is controlled by leaves, as leaves can sense environmental cues and synthesize the flowering signal, a protein named FLOWERING LOCUS T (FT), which travels through the plant vascular system. We therefore wondered how a leafless parasite such as Cuscuta australis controls the timing of its flowering," says lead investigator Jianqiang Wu. In 2018, his team had sequenced the genome of C. australis and found that many genes important for regulation of flowering time were lost in C. australis genome. Therefore, C. australis seems to be unable to activate its own flowering mechanism.Based on the fact that FT proteins are mobile signals, the researchers hypothesized that dodder eavesdrops on the flowering signals produced by the leaves of its host and uses them for producing its own flowers. To prove this eavesdropping scenario, they used genetically modified host plants in which the expression of FT genes had been altered, and this indeed affected the flowering time of the parasite. They also coupled the FT protein to a green fluorescent protein (GFP) as a tag and detected the host plant's flower promoting signal in the parasite: The tagged FT protein had migrated from host to parasite.For dodder, it is the best strategy to synchronize flowering with that of its host. If it flowers much later than its host does, it may not be able to produce seeds at all, as the nutrients in the host are rapidly drained by the host's reproductive tissues. The host may even rapidly die before the parasite can even starts to produce seeds. However, if dodder flowers too early, its growth is likely prematurely ended and it may not be able to produce as many seeds as the dodder plants whose flowering time is synchronized with that of their hosts.In the course of evolution, plant parasites have lost certain traits and "outsourced" physiological processes. As a result, various genes in their genomes may be lost. "This work establishes that for a plant parasite, losing control over flowering processes can be advantageous, as it allows the parasite to hijack its host's mobile flowering signals for its own use. It can thereby readily synchronize its physiology with that of its host," says co-author Ian Baldwin, director of the Department Molecular Ecology at the Max Planck Institute for Chemical Ecology. Because of the gene loss, dodder may be able to better adapt to the parasitic lifestyle and ultimately increase its fitness.
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Biotechnology
| 2,020 |
August 31, 2020
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https://www.sciencedaily.com/releases/2020/08/200831154350.htm
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Novel Dual CAR T cell immunotherapy holds promise for targeting the HIV reservoir
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A recent study published in the journal
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"This study highlights how relatively straightforward alterations to the way T cells are engineered can lead to dramatic changes in their potency and durability," Riley said. "This finding has significant implications for using engineered T cells to fight both HIV and cancer."The global HIV epidemic impacts more than 35 million people around the world. Antiretroviral therapy (ART) is a daily treatment that can control, but not cure, HIV infection. However, access and lifelong adherence to a daily regimen is a significant barrier for many people living with HIV. A major hurdle to HIV cure is the viral reservoir, copies of HIV hidden away in the genome of infected cells. If ART treatment is stopped, the virus is able to rapidly make new copies of itself, ultimately leading to the development of AIDS.CAR T cells are a powerful immunotherapy, currently used in cancer treatments, in which a patient's own immune T cells are engineered to express Chimeric Antigen Receptors (CARs). These CARs re-program the T cells to recognize and eliminate specific diseased or infected cells, such as cancer cells or, potentially, HIV-infected cells.Allen's and Riley's research groups worked together to design a new HIV-specific CAR T cell. They needed to design a CAR T cell that would be able to target and quickly eliminate HIV-infected cells, survive and reproduce once in the body, and resist infection by HIV itself, since HIV's primary target is these very same T cells."By using a stepwise approach to solve each issue as it arose, we developed protected Dual CAR T cells, which provided a strong, long-lasting response against HIV-infection while being resistant to the virus itself," Allen said.This Dual CAR T cell, a new type of CAR T cell, was made by engineering two CARs into a single T cell. Each CAR had a CD4 protein that allowed it to target HIV-infected cells and a costimulatory domain, which signaled the CAR T cell to increase its immune functions. The first CAR contained the 4-1BB co-stimulatory domain, which stimulates cell proliferation and persistence, while the second has the CD28 co-stimulatory domain, which increases its ability to kill infected cells.Since HIV frequently infects T cells, they also added in a protein called C34-CXCR4, developed in the lab of James Hoxie, MD, a professor of Hematology-Oncology at Penn. C34-CXCR4 prevents HIV from attaching to and then infecting the cell. The final CAR T cell was long-lived, replicated in response to HIV infection, killed infected cells effectively, and was partially resistant to HIV infection.When the protected Dual CAR T cells were given to HIV-infected mice, the team saw slower HIV replication and fewer HIV infected cells than in untreated animals. They also saw reduced amounts of virus and preservation of CD4+ T cells, HIV's preferred target, in the blood of these animals. In addition, when they combined Dual CAR T cells with ART in HIV-infected mice, the virus was suppressed faster, which led to a smaller viral reservoir than in mice who were only treated with ART."The ability of these protected Dual CAR T cells to reduce the HIV burden in a variety of tissues and cell types, including long-lived memory CD4+ T cells, we believe supports the approach of using CAR T cell therapy as a new tool to target the HIV reservoir towards a functional cure for HIV," said Allen.
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Biotechnology
| 2,020 |
August 31, 2020
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https://www.sciencedaily.com/releases/2020/08/200831131640.htm
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How antibiotics interact
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It is usually difficult to predict how well drugs will work when they are combined. Sometimes, two antibiotics increase their effect and inhibit the growth of bacteria more efficiently than expected. In other cases, the combined effect is weaker. Since there are many different ways of combining drugs -- such as antibiotics -- it is important to be able to predict the effect of these drug combinations. A new study has found out that it is often possible to predict the outcomes of combining certain antibiotics by quantitatively characterizing how individual antibiotics work. That is the result of a joint study by Professor Tobias Bollenbach at the University of Cologne with Professor Gasper Tkacik and the doctoral researcher Bor Kavcic at the Institute of Science and Technology Austria. The paper 'Mechanisms of drug interactions between translation-inhibiting antibiotics' has been published in
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'We wanted to find out how antibiotics that inhibit protein synthesis in bacteria work when combined with each other, and predict these effects as far as possible, using mathematical models,' Bollenbach explained. As head of the research group 'Biological Physics and Systems Biology' at the University of Cologne, he explores how cells respond to drug combinations and other signals.Bacterial ribosomes can gradually translate the DNA sequence of genes into the amino acid sequence of proteins (translation). Many antibiotics target this process and inhibit translation. Different antibiotics specifically block different steps of the translation cycle. The scientists found out that the interactions between the antibiotics are often caused by bottlenecks in the translation cycle. For example, antibiotics that inhibit the beginning and middle of the translation cycle have much weaker effects when combined.In order to clarify the underlying mechanisms of drug interactions, the scientists created artificial translation bottlenecks that genetically mimic the effect of specific antibiotics. If such a bottleneck is located in the middle of the translation cycle, a traffic jam of ribosomes forms, which dissolves upon introducing another bottleneck at the beginning of the translation cycle. Using a combination of theoretical models from statistical physics and experiments, the scientists showed that this effect explains the drug interaction between antibiotics that block these translation steps.Tobias Bollenbach concluded: 'A quantitative understanding of the effect of individual antibiotics allows us to predict the effect of antibiotic combinations without having to test all possible combinations by trial and error. This finding is important because the same approach can be applied to other drugs, enabling the development of new, particularly effective drug combinations in the long term.'
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Biotechnology
| 2,020 |
August 31, 2020
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https://www.sciencedaily.com/releases/2020/08/200831131632.htm
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Study finds missing link in the evolutionary history of carbon-fixing protein Rubisco
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A team led by researchers at the University of California, Davis, has discovered a missing link in the evolution of photosynthesis and carbon fixation. Dating back more than 2.4 billion years, a newly discovered form of the plant enzyme rubisco could give new insight into plant evolution and breeding.
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Rubisco is the most abundant enzyme on the planet. Present in plants, cyanobacteria (also known as blue-green algae) and other photosynthetic organisms, it's central to the process of carbon fixation and is one of Earth's oldest carbon-fixing enzymes."It's the primary driver for producing food, so it can take CO2 from the atmosphere and fix that into sugar for plants and other photosynthetic organisms to use. It's the primary driving enzyme for feeding carbon into life that way," said Doug Banda, a postdoctoral scholar in the lab of Patrick Shih, assistant professor of plant biology in the UC Davis College of Biological Sciences.Form I rubisco evolved over 2.4 billion years ago before the Great Oxygenation Event, when cyanobacteria transformed the Earth's atmosphere by producing oxygen through photosynthesis. Rubisco's ties to this ancient event make it important to scientists studying the evolution of life.In a study appearing Aug. 31 in The new version, called form I-prime rubisco, was found through genome sequencing of environmental samples and synthesized in the lab. Form I-prime rubisco gives researchers new insights into the structural evolution of form I rubisco, potentially providing clues as to how this enzyme changed the planet.Form I rubisco is responsible for the vast majority of carbon fixation on Earth. But other forms of rubisco exist in bacteria and in the group of microorganisms called Archaea. These rubisco variants come in different shapes and sizes, and even lack small subunits. Yet they still function."Something intrinsic to understanding how form I rubisco evolved is knowing how the small subunit evolved," said Shih. "It's the only form of rubisco, that we know of, that makes this kind of octameric assembly of large subunits."Study co-author Professor Jill Banfield, of UC Berkeley's earth and planetary sciences department, uncovered the new rubisco variant after performing metagenomic analyses on groundwater samples. Metagenomic analyses allow researchers to examine genes and genetic sequences from the environment without culturing microorganisms."We know almost nothing about what sort of microbial life exists in the world around us, and so the vast majority of diversity has been invisible," said Banfield. "The sequences that we handed to Patrick's lab actually come from organisms that were not represented in any databases."Banda and Shih successfully expressed form I-prime rubisco in the lab using Form I rubisco is built from eight core large molecular subunits with eight small subunits perched on top and bottom. Each piece of the structure is important to photosynthesis and carbon fixation. Like form I rubisco, form I-prime rubisco is built from eight large subunits. However, it does not possess the small subunits previously thought essential."The discovery of an octameric rubisco that forms without small subunits allows us to ask evolutionary questions about what life would've looked like without the functionality imparted by small subunits," said Banda. "Specifically, we found that form I-prime enzymes had to evolve fortified interactions in the absence of small subunits, which enabled structural stability in a time when Earth's atmosphere was rapidly changing."According to the researchers, form I-prime rubisco represents a missing link in evolutionary history. Since form I rubisco converts inorganic carbon into plant biomass, further research on its structure and functionality could lead to innovations in agriculture production."Although there is significant interest in engineering a 'better' rubisco, there has been little success over decades of research," said Shih. "Thus, understanding how the enzyme has evolved over billions of years may provide key insight into future engineering efforts, which could ultimately improve photosynthetic productivity in crops."
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Biotechnology
| 2,020 |
August 31, 2020
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https://www.sciencedaily.com/releases/2020/08/200831094713.htm
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Cells can remain functional despite damage to mitochondria
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Mitochondria are the power plants of our cells and play an important role in providing energy for normal function of the tissues in our body. Nerve cells are particularly dependent on mitochondria for their activity and decreased mitochondrial function is seen in both inherited and more common age-associated forms of degenerative diseases. A long-standing view has been that neurons, in contrast to other cell types, cannot adjust their metabolism to compensate for mitochondrial dysfunction, and therefore irreversibly degenerate. In a new study, researchers from the Max Planck Institute for Biology of Aging in Cologne, Germany, and the Karolinska Institute in Stockholm, Sweden, challenge this dogma by showing that neurons have the potential to counteract degeneration and promote survival by adapting their metabolism.
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In our increasingly ageing society, neurodegenerative diseases are posing a significant burden. A growing body of evidence has linked mitochondrial dysfunction to some of the most devastating forms of neurodegeneration, such as Parkinson´s disease, different ataxias and several peripheral neuropathies. However, despite the urge to find strategies to prevent or arrest neurodegeneration, our understanding of the precise events underlying neuronal death caused by mitochondrial dysfunction is very limited. "We generally tend to consider neurons as terminally differentiated cells with very limited or no capacity to adapt their energy metabolism to challenging conditions," says Elisa Motori, a lead author of this study. "For some neurological diseases there is ample evidence that mitochondrial dysfunction can be tolerated for lengthy periods of time. We therefore asked the question whether degenerating neurons may activate a program of metabolic resilience."The researchers devised an innovative approach to purify degenerating neurons from the mouse brain and analyze the global protein content (proteome) of these neurons. "Unexpectedly, the proteomic data showed the existence of a precisely coordinated, neuron-specific metabolic program that becomes activated in response to mitochondrial dysfunction," continues Motori.In particular, the authors identified a form of metabolic rewiring (Krebs cycle anaplerosis) that makes neurons resistant to an otherwise very rapidly progressing degeneration. This type of metabolic adaptation was previously only thought to occur in peripheral tissues or supporting cells (glia cells) in the brain. "The finding that neurons can induce anaplerosis was not only intriguing, but we could further demonstrate that it had a protective role. When we blocked anaplerosis neurons died at a much faster pace and the disease became more severe." explains Elisa Motori.The identification of certain forms of metabolic rewiring in dysfunctional neurons provide new mechanistic insights into the processes leading to neurodegeneration. Based on these new findings, the authors hope that it may be possible to develop therapeutic approaches to prolong neuronal survival and improve function in patients with mitochondrial diseases and other types of neurodegeneration.
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Biotechnology
| 2,020 |
August 28, 2020
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https://www.sciencedaily.com/releases/2020/08/200828140314.htm
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Plant scientists study the interaction of heat stress responses in corn
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Environmental extremes driven by climate change create stresses in crops, and plant breeders are attempting to untangle the genetic factors that endow plants with tolerance to stress. A new study from Iowa State University scientists shows how two seemingly unrelated responses in corn plants interact to help the crop survive heat stress.
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The study, published on Tuesday in the academic journal "These two systems have been thought to operate independently," said Stephen Howell, Distinguished Professor of Genetics, Development and Cell Biology and senior author of the study. "We've been able to show these systems sometimes work together to mitigate damage caused by heat and to protect the plant from stress."Heat stress causes proteins to denature and misfold in the endoplasmic reticulum, an organelle inside cells. Misfolded proteins can be toxic, and their buildup sets off an alarm that activates the expression of genes that protects plants from heat stress. A similar response plays out in different locations of the cell, including the cytoplasm, where excessive heat activates the expression of a different set of genes encoding heat-shock proteins.The new study shows that, although the two responses take place in different parts of the cell, they actually work in concert during heat stress: a powerful transcription factor produced in the unfolded protein response activates the expression of a key factor helping to trigger the heat shock response.The scientists found that knocking out the unfolded protein response made corn plants more susceptible to heat stress and hindered the heat shock response. That raises the question if overexpressing the misfolded protein response could strengthen the ability of corn plants to withstand high heat, but Howell said doing so may have other undesirable consequences."There's a seesaw balance, if you will, between defense and growth," he said. "The more you contribute to defense, the more you sacrifice growth. It may be that you could provide somewhat greater defense to crops but you might do so at the expense of growth."In their study, the researchers drew on data gathered in the Enviratron, a state-of-the-art facility at the ISU Ag Engineering/Agronomy Research Farm that utilizes a robotic rover that travels through a series of specialized growth chambers that carefully control the environments in which the plants are raised. Development of the Enviratron was funded through a grant from the National Science Foundation. Zhaoxia Li, first author of the paper and a postdoctoral scientist in Howell's lab, said the facility allows researchers to control variables such as temperature, moisture, light and carbon dioxide concentrations to study their effect on plant development.Howell said previous scientific papers have described the design and construction of the Enviratron, but this is the first publication in a journal based on data generated in the facility."We hope that studies like this will emphasize the value of conducting such research under controlled environmental conditions offered by the Enviratron," he said.
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Biotechnology
| 2,020 |
August 28, 2020
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https://www.sciencedaily.com/releases/2020/08/200828140310.htm
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'Jumping' DNA regulates human neurons
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The human genome contains over 4.5 million sequences of DNA called "transposable elements," these virus-like entities that "jump" around and help regulate gene expression. They do this by binding transcription factors, which are proteins that regulate the rate of transcription of DNA to RNA, influencing gene expression in a broad range of biological events.
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Now, an international team of scientists led by Didier Trono at EPFL has discovered that transposable elements play a significant role in influencing the development of the human brain. The study is published in The scientists found that transposable elements regulate the brain's development by partnering up with two specialized proteins from the family of proteins known as "Krüppel-associated box-containing zinc finger proteins, or KZFPs. In 2019, another study led by Trono showed that KZFPs tamed the regulatory activity of transposable elements in the first few days of the fetus's life. However, they suspected that these regulatory sequences were subsequently re-ignited to orchestrate the development and function of adult organs.The researchers identified two KZFPs as specific only to primates, and found that they are expressed in specific regions of the human developing and adult brain. They further observed that these proteins kept controlling the activity of transposable elements -- at least in neurons and brain organoids cultured in the lab. As a result, these two KZFPs influenced the differentiation and neurotransmission profile of neurons, as well as guarded these cells against inflammatory responses that were otherwise triggered if their target transposable elements were left to be expressed."These results reveal how two proteins that appeared only recently in evolution have contributed to shape the human brain by facilitating the co-option of transposable elements, these virus-like entities that have been remodeling our ancestral genome since the dawn of times," says Didier Trono. "Our findings also suggest possible pathogenic mechanisms for diseases such as amyotrophic lateral sclerosis or other neurodegenerative or neurodevelopmental disorders, providing leads for the prevention or treatment of these problems."
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Biotechnology
| 2,020 |
August 28, 2020
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https://www.sciencedaily.com/releases/2020/08/200828115357.htm
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How bacteria adhere to fiber in the gut
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Researchers have revealed a new molecular mechanism by which bacteria adhere to cellulose fibers in the human gut. Thanks to two different binding modes, they can withstand the shear forces in the body. Scientists of the University of Basel and ETH Zurich published their results in the journal
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Cellulose is a major building block of plant cell walls, consisting of molecules linked together into solid fibers. For humans, cellulose is indigestible, and the majority of gut bacteria lack the enzymes required to break down cellulose.However, recently genetic material from the cellulose-degrading bacterium R. champanellensis was detected in human gut samples. Bacterial colonization of the intestine is essential for human physiology, and understanding how gut bacteria adhere to cellulose broadens our knowledge of the microbiome and its relationship to human health.The bacterium under investigation uses an intricate network of scaffold proteins and enzymes on the outer cell wall, referred to as a cellulosome network, to attach to and degrade cellulose fibers. These cellulosome networks are held together by families of interacting proteins.Of particular interest is the cohesin-dockerin interaction responsible for anchoring the cellulosome network to the cell wall. This interaction needs to withstand shear forces in the body to adhere to fiber. This vital feature motivated the researchers to investigate in more detail how the anchoring complex responds to mechanical forces.By using a combination of single-molecule atomic force microscopy, single-molecule fluorescence and molecular dynamics simulations, Professor Michael Nash from the University of Basel and ETH Zurich along with collaborators from LMU Munich and Auburn University studied how the complex resists external force.They were able to show that the complex exhibits a rare behavior called dual binding mode, where the proteins form a complex in two distinct ways. The researchers found that the two binding modes have very different mechanical properties, with one breaking at low forces of around 200 piconewtons and the other exhibiting a much higher stability breaking only at 600 piconewtons of force.Further analysis showed that the protein complex displays a behavior called a "catch bond," meaning that the protein interaction becomes stronger as force is ramped up. The dynamics of this interaction are believed to allow the bacteria to adhere to cellulose under shear stress and release the complex in response to new substrates or to explore new environments."We clearly observe the dual binding modes, but can only speculate on their biological significance. We think the bacteria might control the binding mode preference by modifying the proteins. This would allow switching from a low to high adhesion state depending on the environment," Professor Nash explains.By shedding light on this natural adhesion mechanism, these findings set the stage for the development of artificial molecular mechanisms that exhibit similar behavior but bind to disease targets. Such materials could have applications in bio-based medical superglues or shear-enhanced binding of therapeutic nanoparticles inside the body. "For now, we are excited to return to the laboratory and see what sticks," says Nash.
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Biotechnology
| 2,020 |
August 28, 2020
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https://www.sciencedaily.com/releases/2020/08/200828091957.htm
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Engineers uncover biomechanical effects of skin rubbing
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Understanding the skin damage caused by rubbing could lead to better topical skin treatments and help prevent the formation of new routes for viral and bacterial infection.
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Tohoku University biomechanical engineers have developed a better understanding of the damage that can be caused by something rubbing against the skin. The study was published in the The uppermost part of the skin, called the stratum corneum, is formed of layers of keratinocyte cells suspended in a lipid matrix. The stratum corneum plays an important protective role, forming a barrier against the invasion of viruses and bacteria, maintaining skin hydration, and managing skin recovery following damage.Kikuchi and colleagues at Tohoku University in Japan wanted to develop a more in depth understanding of the mechanics of skin damage caused by rubbing. This could help scientists develop more effective drugs that are applied through the skin and to understand how to better prevent viruses and bacteria from gaining access to the body through damaged skin.The researchers measured the mechanical effects of rubbing on pig skin, which is very similar to human skin. A gear rotated against skin samples at known rates and pressures. The scientists then measured the damage this caused by exposing the skin samples to a fluorescent dye. The more damaged the skin, the more the fluorescent dye was able to permeate it.The team found that the mechanical rubbing caused keratinocytes to shrink and wrinkle in the direction of the rubbing. Gaps also formed between the keratinocytes, degrading the skin's barrier function. They then developed a mathematical formula to describe the permeability of rubbed skin, which can be estimated from the amount of strain applied."We believe our findings could improve transdermal drug delivery and we plan to investigate the development of novel topical drugs that can be applied to the skin by rubbing," says Kikuchi.He adds that their findings could apply to the current pandemic if rubbing face masks were found by further research to cause skin damage, potentially allowing another avenue for COVID-19 infection. The team did not specifically investigate this topic and Kikuchi encourages people to continue to wear face coverings that fit comfortably over the nose and mouth.
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Biotechnology
| 2,020 |
August 28, 2020
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https://www.sciencedaily.com/releases/2020/08/200828091953.htm
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Giant nanomachine aids the immune system
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Cells that are infected by a virus or carry a carcinogenic mutation, for example, produce proteins foreign to the body. Antigenic peptides resulting from the degradation of these exogenous proteins inside the cell are loaded by the peptide-loading complex onto so-called major histocompatibility complex molecules (MHC for short) and presented on the cell surface. There, they are specifically identified by T-killer cells, which ultimately leads to the elimination of the infected cells. This is how our immune system defends us against pathogens.
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The peptide-loading complex ensures that the MHC molecules are correctly loaded with antigens. "The peptide-loading complex is a biological nanomachine that has to work with atomic precision in order to efficiently protect us against pathogens that cause disease," says Professor Lars Schäfer, Head of the Molecular Simulation research group at the Centre for Theoretical Chemistry at RUB.In previous studies, other teams successfully determined the structure of the peptide-loading complex using cryo-electron microscopy, but only with a resolution of about 0.6 to 1.0 nanometres, i.e. not in atomic detail. Based on these experimental data, Schäfer's research team in collaboration with Professor Gunnar Schröder from Forschungszentrum Jülich has now succeeded in creating an atomic structure of the peptide-loading complex."The experimental structure is impressive. But only with our computer-based methods were we able to extract the maximum information content contained in the experimental data," explains Schröder. The atomic model enabled the researchers to perform detailed molecular dynamics computer simulations of the peptide-loading complex and thus to study not only the structure but also the dynamics of the biological nanomachine.Since the simulated system is extremely large with its 1.6 million atoms, the computing time at the Leibnitz Supercomputing Centre in Munich aided this task considerably. "Using the high-performance computer, we were able to push into the microsecond time scale in our simulations. This revealed the role of sugar groups bound to the protein for the mechanism of peptide loading, which had previously only been incompletely understood," outlines Dr. Olivier Fisette, postdoc researcher at the Molecular Simulation research group.The atomic model of the peptide-loading complex now facilitates further studies. For example, some viruses try to cheat our immune system by selectively switching off certain elements of the peptide-loading complex. "One feasible objective we'd like to pursue is the targeted intervention in these processes," concludes Schäfer.
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827155006.htm
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Antiviral used to treat cat coronavirus also works against SARS-CoV-2
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Researchers at the University of Alberta are preparing to launch clinical trials of a drug used to cure a deadly disease caused by a coronavirus in cats that they expect will also be effective as a treatment for humans against COVID-19.
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"In just two months, our results have shown that the drug is effective at inhibiting viral replication in cells with SARS-CoV-2," said Joanne Lemieux, a professor of biochemistry in the Faculty of Medicine & Dentistry."This drug is very likely to work in humans, so we're encouraged that it will be an effective antiviral treatment for COVID-19 patients."The drug is a protease inhibitor that interferes with the virus's ability to replicate, thus ending an infection. Proteases are key to many body functions and are common targets for drugs to treat everything from high blood pressure to cancer and HIV.First studied by U of A chemist John Vederas and biochemist Michael James following the 2003 outbreak of severe acute respiratory syndrome (SARS), the protease inhibitor was further developed by veterinary researchers who showed it cures a disease that is fatal in cats.The work to test the drug against the coronavirus that causes COVID-19 was a co-operative effort between four U of A laboratories, run by Lemieux, Vederas, biochemistry professor Howard Young and the founding director of the Li Ka Shing Institute of Virology, Lorne Tyrrell. Some of the experiments were carried out by the Stanford Synchrotron Radiation Lightsource Structural Molecular Biology program.Their findings were published today in the peer-reviewed journal "There's a rule with COVID research that all results need to be made public immediately," Lemieux said, which is why they were posted before being peer-reviewed.She said interest in the work is high, with the paper being accessed thousands of times as soon as it was posted.Lemieux explained that Vederas synthesized the compounds, and Tyrrell tested them against the SARS-CoV-2 virus in test tubes and in human cell lines. The Young and Lemieux groups then revealed the crystal structure of the drug as it binds with the protein."We determined the three-dimensional shape of the protease with the drug in the active site pocket, showing the mechanism of inhibition," she said. "This will allow us to develop even more effective drugs."Lemieux said she will continue to test modifications of the inhibitor to make it an even better fit inside the virus.But she said the current drug shows enough antiviral action against SARS-CoV-2 to proceed immediately to clinical trials."Typically for a drug to go into clinical trials, it has to be confirmed in the lab and then tested in animal models," Lemieux said. "Because this drug has already been used to treat cats with coronavirus, and it's effective with little to no toxicity, it's already passed those stages and this allows us to move forward.""Because of the strong data that we and others have gathered we're pursuing clinical trials for this drug as an antiviral for COVID-19."The researchers have established a collaboration with Anivive Life Sciences, a veterinary medicine company that is developing the drug for cats, to produce the quality and quantity of drug needed for human clinical trials. Lemieux said it will likely be tested in Alberta in combination with other promising antivirals such as remdesivir, the first treatment approved for conditional use in some countries including the United States and Canada.The U of A researchers' work was funded by the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council of Canada, Alberta Innovates, Li Ka Shing Institute of Virology and the GSK Chair in Virology.
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827155005.htm
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Researchers develop a yeast-based platform to boost production of rare natural molecules
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Many modern medicines, including analgesics and opioids, are derived from rare molecules found in plants and bacteria. While they are effective against a host of ailments, a number of these molecules have proven to be difficult to produce in large quantities. Some are so labour intensive that it is uneconomical for pharmaceutical companies to produce them in sufficient amounts to bring them to market.
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In a new study published in One of the principal ingredients in this new technique developed by the biology professor and Concordia University Research Chair in Microbial Engineering and Synthetic Biology is simple baker's yeast.The single-cell organism has cellular processes that are similar to those of humans, giving biologists an effective substitute in drug development research. Using cutting-edge synthetic biology approaches, Martin and his colleagues in Berkeley, California were able to produce a large amount of benzylisoquinoline alkaloid (BIA) to synthesize an array of natural and new-to-nature chemical structures in a yeast-based platform.This, he says, can provide a blueprint for the large-scale production of thousands of products, including the opioid analgesics morphine and codeine. The same is true for opioid antagonists naloxone and naltrexone, used to treat overdose and dependence.Martin has been working toward this outcome for most of the past two decades. He began with researching the genetic code plants use to produce the molecules used as drugs by the pharmaceutical industry. Then came transplanting their genes and enzymes into yeast to see if production was possible outside a natural setting. The next step is industrial production."We showed in previous papers that we can get milligrams of these molecules fairly easily, but you're only going to be able to commercialize the process if you get grams of it," Martin explains. "In principle, we now have a technology platform where we can produce them on that scale."This, he says, can have huge implications for a country like Canada, which has to import most of the rare molecules used in drugs from overseas. That's especially relevant now, in the midst of a global pandemic, when fragile supply chains are at risk of being disrupted."To me, this really highlights the importance of finding alternative biotech-type processes that can be developed into a homemade, Canadian pharmaceutical industry," he adds. "Many of the ingredients we use today are not very difficult to make. But if we don't have a reliable supply process in Canada, we have a problem."Martin admits he is curious to see where the technology leads us. He believes researchers can and will use the new platform for the commercialization and discovery of new drugs."We demonstrate that by using this platform, we can start building what is called new-to-nature molecules," he says. "By experimenting with enzymes and genes and the way we grow things, we can begin making these into tools that can be used in the drug discovery process. We can access a whole new structural space."This study was financially supported by a Natural Sciences and Engineering Research Council of Canada (NSERC) Industrial Biocatalysis Grant, an NSERC Discovery Grant and by River Stone Biotech ApS.
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827141339.htm
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DNA repair: Locating and severing lethal links
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Chemical lesions in the genetic material DNA can have catastrophic consequences for cells, and even for the organism concerned. This explains why the efficient identification and rapid repair of DNA damage is vital for survival. DNA-protein crosslinks (DPCs), which are formed when proteins are adventitiously attached to DNA, are particularly harmful. DPCs are removed by the action of a dedicated enzyme -- the protease SPRTN -- which cleaves the bond between the protein and the DNA. Up to now, how SPRTN recognizes such crosslinks, which can differ significantly in structure, has remained unclear. Now a team led by Professor Julian Stingele (LMU Gene Center), in cooperation with Professor Michael Sattler (Helmholtz Zentrum München and Technical University of Munich), has shown that the enzyme utilizes a modular recognition mechanism to detect such sites, such that it is activated only under highly specific conditions. The new findings appear in the journal
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DPCs can be created by interactions with highly reactive products of normal metabolism or with synthetic chemotherapeutic agents. These lesions are extremely toxic because they block the replication of DNA -- and therefore inhibit cell division. Timely and effective repair of these crosslinks by SPRTN is crucial for cell viability and the suppression of tumorigenesis. In humans, mutations that reduce the activity of the enzyme are associated with a high incidence of liver cancer in early life and markedly accelerate the aging process. "SPRTN has a difficult job to do because, depending on the protein and the DNA subunit involved, the structure of the crosslink can vary widely. So the enzyme has to be able to identify many different structures as aberrant," explains Hannah Reinking, first author of the study. "We therefore asked ourselves what sorts of properties a DPC should have in order to be recognized and cleaved."To answer this question, Reinking and colleagues constructed model substrates consisting of proteins attached to defined positions within DNA strands, and examined whether the SPRTN protease could repair them in the test-tube. This approach revealed that SPRTN interacts with structures that are frequently found in the vicinity of DPCs. With the aid of nuclear magnetic resonance spectroscopy, they went on to show that SPRTN contains two recognition domains. One binds to double-stranded, and the other to single-stranded DNA. "So the protein uses a modular system for substrate recognition. Only when both domains are engaged is the enzyme active -- and DNA in which double-stranded and single-stranded regions occur in close proximity is often found in the vicinity of crosslinks," says Stingele.These results are also of clinical relevance. The action of many chemotherapeutic drugs depends on their ability to form crosslinks with DNA. Since tumor cells divide more frequently than non-malignant cells, they are particularly sensitive to this type of DNA damage. DNA repair enzymes like SPRTN are therefore of great interest as potential drug targets for use in the context of personalized cancer therapies, and agents that specifically inhibit the protease could eventually be employed to boost the efficacy of chemotherapy. "Our work now makes it possible to conceptualize such therapeutic strategies," says Stingele.
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827141309.htm
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Synthetic compound could serve as prototype for novel class of drugs to treat neurological damage
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Researchers from the German Center for Neurodegenerative Diseases (DZNE), UK and Japan have developed a neurologically acting protein and tested it in laboratory studies. In mice, the experimental compound ameliorated symptoms of certain neurological injuries and diseases, while on the microscopic level it was able to establish and repair connections between neurons. This proof-of-principle study suggests that biologics, which act on neuronal connectivity, could be of clinical use in the long term. The results are published in the journal
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The human brain's neuronal network undergoes life-long changes in order to be able to assimilate information and store it in a suitable manner. This applies in particular to the generation and recalling of memories. So-called synapses play a central role in the brain's ability to adapt. They are junctions through which nerve signals are passed from one cell to the next. A number of specific molecules -- known as "synaptic organizing proteins" -- make sure that synapses are formed and reconfigured whenever necessary.An international team of researchers has now combined various structural elements of such naturally occurring molecules into an artificial protein called CPTX and tested its effect in different disease models. To this end, the compound was administered to mice with neurological deficits that occur in similar fashion in humans. Specifically, the tests focused on Alzheimer's disease, spinal cord injury and cerebellar ataxia -- a disease that is characterized primarily by a failure of muscle coordination. All these conditions are associated with damage to the synapses or their loss. The study was a collaborative effort by experts from several research institutions, including the DZNE's Magdeburg site, MRC Laboratory of Molecular Biology in UK, Keio University School of Medicine in Tokyo, and, also in Japan, Aichi Medical University."In our lab we studied the effect of CPTX on mice that exhibited certain symptoms of Alzheimer's disease," said Prof. Alexander Dityatev, a senior researcher at the DZNE, who has been investigating synaptic proteins for many years. "We found that application of CPTX improved the mice's memory performance." The researchers also observed normalization of several important neuronal parameters that are compromised in Alzheimer's disease, as well as in the studied animal model. Namely, CPTX increased the ability of synapses to change, which is considered as a cellular process associated with memory formation. Furthermore, CPTX was shown to elevate what is called "excitatory transmission." This is to say that the protein acted specifically on synapses that promoted activity of the contacted cell. And finally, CPTX increased the density of so-called dendritic spines. These are tiny bulges in the cell's membrane that are essential for establishing excitatory synaptic connections.Further research by the study partners in the UK and Japan revealed that application of CPTX to mice with motor dysfunction -- caused either by spinal cord injury or pathological conditions similar to cerebellar ataxia -- improved the rodent's mobility. And at the cellular level, the drug was shown to repair and promote excitatory synaptic connections.CPTX combines functional domains present in natural synaptic organizing proteins in a unique way. The compound was designed to act as a universal bridge builder for excitatory connections between nerve cells. Where two neurons meet, either in adhesive contact or actually in synaptic connection, CPTX links to specific molecules on the surfaces of both involved cells, and thereby either triggers the formation of new synapses or strengthens already existing ones."At present, this drug is experimental and its synthesis, the credit for which goes to our UK partners, is quite demanding. We are far off from application in humans," Dityatev emphasized, who in addition to his research at the DZNE is also a member of the Medical Faculty of the University Magdeburg. "However, our study suggests that CPTX can even do better than some of its natural analogs in building and strengthening nerve connections. Thus, CPTX could be the prototype for a new class of drugs with clinical potential." Application would be in disorders that are associated with impaired neuronal connectivity. "Much of the current therapeutic effort against neurodegeneration focuses on stopping disease progression and offers little prospect of restoring lost cognitive abilities. Our approach could help to change this and possibly lead to treatments that actually regenerate neurological functions. Based on the principles we have used in designing CPTX, we thus intend to develop further compounds. In future studies, we want to refine their properties and explore possible therapeutic applications."
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827101839.htm
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Gut microbes could unlock the secret to healthy aging
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Bacteria and other microorganisms in the digestive tract are linked with dozens of health conditions including high blood pressure, high blood lipids, and body mass index (BMI) according to research presented today at ESC Congress 2020.
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"Our study indicates that microbiota might have an important role in maintaining health and could help us develop novel treatments," said study author Dr. Hilde Groot of University Medical Centre Groningen, the Netherlands.The human gut microbiome is the totality of microorganisms (generally bacteria and single-celled organisms called archaea) and their collective genetic material present in the digestive tract. Small-scale studies have suggested a link between the gut microbiome and individual diseases.This study, for the first time, investigated multiple diseases and other traits in one cohort -- revealing the staggering extent to which the microbiome influences sickness and health. The analysis used genetic data as a proxy for microbiome composition.Dr. Groot explained: "Previous research has shown that the human gut microbiome composition could be partially explained by genetic variants. So, instead of directly measuring the make-up of the microbiome, we used genetic alterations to estimate its composition."The study included 422,417 unrelated individuals in the UK Biobank who had undergone genotyping to identify their genetic make-up. Information was also collected on a wide range of diseases and other characteristics including BMI and blood pressure. The average age of participants was 57 years and 54% were women.The researchers found that higher levels of eleven bacteria (estimated from genetic data) were associated with a total of 28 health and disease outcomes. These included chronic obstructive pulmonary disease (COPD), atopy (a genetic tendency to develop allergic diseases like asthma and eczema), frequency of alcohol intake, high blood pressure, high blood lipids, and BMI.To take one example, higher levels of the genus Ruminococcus were linked with increased risk of high blood pressure.Regarding alcohol consumption, Dr. Groot said: "What we eat and drink is connected to microbiome content, so we studied the links with meat, caffeine, and alcohol. We observed a relationship between raised levels of Methanobacterium and drinking alcohol more often. It is important to stress that this is an association, not a causal relation, and more research is needed."A real strength of the study was conducting a broad analysis in the same group of people. Dr. Groot said: "Considering that the results were observed in one cohort, this cautiously supports the notion that microbiota and the substances they produce (called metabolites) provide links between numerous diseases and conditions. The findings may help identify common pathways. Nevertheless, more research (for example in other cohorts) is needed to validate our findings."She concluded: "Follow-up studies are required to study causality before giving concrete advice to the public and health professionals. This study provides clues where to go."
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827101815.htm
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Rejuvenating old organs could increase donor pool
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Despite the limited supply of organs available for patients on waitlists for transplantation, organs from older, deceased donors are frequently discarded or not utilized. Available older organs have the potential to close the gap between demand and supply that is responsible for the very long wait-times that lead to many patients not surviving the time it takes for an organ to become available. Older organs can also often provoke a stronger immune response and may put patients at greater risk of adverse outcomes and transplant rejection. But, as the world population ages, organs from older, deceased donors represent an untapped and growing resource for patients in need. Investigators from Brigham and Women's Hospital are leading efforts to breathe new life into older organs by leveraging a new class of drugs known as senolytics, which target and eliminate old cells. Using clinical and experimental studies, the team presents evidence that senolytic drugs may help rejuvenate older organs, which could lead to better outcomes and a wider pool of organs eligible for donation. Results are published in
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"Older organs are available and have the potential to contribute to mitigating the current demand for organ transplantation," said corresponding author Stefan G. Tullius, MD, PhD, chief of the Division of Transplant Surgery at the Brigham. "If we can utilize older organs in a safe way with outcomes that are comparable, we will take a substantial step forward for helping patients."As organs age, senescent cells accumulate. These cells, which no longer divide, escape the body's usual means of destroying older, unneeded cells. Senescent cells release cell-free mitochondrial DNA (mt-DNA), which also accumulates in older organs. Recent studies have suggested that this rise in mt-DNA is tied to organ rejection.In their Nature Communications paper, Tullius and colleagues identified senescent cells as the key source of mt-DNA and present evidence that the accumulation of mt-DNA provokes an immune response leading to organ failure and rejection. Senolytic drugs force senescent cells back into the cell cycle, allowing the body to clear them. The researchers therefore examined whether senolytic drugs could be used to improve outcomes. In a mouse model, they treated organ donors with a combination of the senolytic drugs dasatinib and quercetin. The drugs reduced the number of senescent cells, reduced mt-DNA levels and decreased inflammation. Most relevantly, the survival of old organs treated with senolytics was as comparable to that of organs originating from young donors.Since the authors carried out their therapeutic experiments in a mouse model, further mechanistic studies are needed to evaluate whether senolytic drugs may have the same effects on human organs from older donors and the same degree of success in clearing senescent cells, as well as whether organs can be treated effectively with senolytic drugs after they are harvested. The authors have already started with first steps in humans and determined that augmented levels of mt-DNA circulate in older organ donors."We have not yet tested the effects clinically, but we are well prepared to take the next step toward clinical application by using a perfusion device to flow senolytic drugs over organs and measure whether or not there are improvements in levels of senescent cells," said Tullius. "Our data provide a rationale for considering clinical trials treating donors, organs, and/or recipients with senolytic drugs to optimize the use of organs from older donors. The goal is to help to close the gap between organ availability and the needs of the many patients currently on transplant waiting lists."
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200827101812.htm
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Japanese sake: the new pick-me-up? Yeast strain makes fatigue-fighting ornithine
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Fans of sake, the traditional Japanese alcoholic beverage, may have even more reason to enjoy it now: Japanese scientists have discovered that a mutant strain of sake yeast produces high levels of the amino acid ornithine.
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In a study published this month in Ornithine is a non-protein-making amino acid and a precursor to two amino acids -- arginine and proline. It has been found to perform several physiological functions, such as reducing fatigue and improving sleep quality."We wanted to obtain sake yeast strains with improved ethanol tolerance," says a first author of this article, Masataka Ohashi. "During sake fermentation, the yeast is exposed to high concentrations of ethanol, which impedes yeast cell growth, viability and fermentation. Increased ethanol tolerance in sake yeast strains could improve ethanol production and reduce fermentation time."To find ethanol-tolerant yeast strains, the researchers isolated mutants that accumulated proline, which can alleviate ethanol toxicity, using a conventional mutagenesis (i.e., one that doesn't involve genetic modification). They also conducted whole genome sequencing analysis, and performed brewing tests with sake yeast strains. Then they identified and analyzed a new mutation in a gene that encodes a variant of "We previously constructed self-cloning industrial yeast strains that accumulate proline to increase ethanol tolerance and productivity of yeast," explains Prof. Hiroshi Takagi, a corresponding author. "But those yeasts have not been yet acceptable to consumers because they're considered to be genetically modified, even though a self-cloning yeast has no foreign genes or DNA sequences -- they only have yeast DNA."The researchers successfully isolated non-genetically modified yeasts that produced 10 times the amount of ornithine compared with the parent strain, which is widely used in Japanese sake breweries, and the sake brewed with them contained 4-5 times more ornithine.The results of this study will contribute to the development of improved yeast strains for production of high levels of ornithine, and the strain obtained in this study could be readily applied to sake, wine, and beer brewing. Ornithine-accumulating yeast strains could also be used in the production of ornithine-rich dietary supplements made from these yeasts and their products.Prof. Takagi also describes "There are two major purposes for breeding of industrial yeast: improvement of fermentation ability with enhanced tolerance to environmental stresses during fermentation processes and diversity of product taste and flavor with modified metabolic pathways. In yeast, amino acid metabolisms vary under different growth environments and the metabolic styles form a complicated but robust network. The elucidation of metabolic regulatory mechanisms and physiological roles for amino acids is important fundamental research for understanding life phenomenon. The yeast is reliable and safe in food production, and thus the development of novel strains that overproduce 'functional amino acids' such as ornithine, proline and branched-amino acids, would greatly contribute to food-related industries."
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Biotechnology
| 2,020 |
August 27, 2020
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https://www.sciencedaily.com/releases/2020/08/200826141403.htm
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A protein with an unprecedented fold helps bacteria uptake thiosulfate as a sulfur source
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A new study led by researchers at Nara Institute of Science and Technology (NAIST), Nara, Japan, in
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Most people know that water is made of hydrogen and oxygen atoms, and carbon compounds are found in all life on earth. However, other elements like sulfur are also indispensable for life, and sulfur-based molecules like L-cysteine are essential for many of our proteins. L-cysteine is also commercially important, as it is heavily used by the food, cosmetic and pharmaceutical industries."In nature, L-cysteine is produced by microorganisms that collect inorganic sulfur in the soil. If the secretion and production efficiency of L-cysteine by microorganisms can be dramatically improved, this procedure will be superior to existing methods, such as in the production of glutamic acid by Bacteria can take up both sulfate and thiosulfate ions from the environment in order to synthesize L-cysteine. The efficiency of the synthesis from thiosulfate is higher because of fewer chemical reaction steps. To examine which proteins are crucial for thiosulfate uptake, the researchers conducted a series of genetic studies, finding YeeE.To understand how YeeE physically conforms for the transport, the researchers uncovered crystal structures of YeeE that revealed an unprecedented fold forming an hourglass shape."Both the inner and outer surfaces of YeeE are indented toward the center. We think this shape is crucial for initiating the uptake and conducts thiosulfate," explains Dr. Yoshiki Tanaka, the first author of the study.Molecular dynamics simulations implied that the uptake occurs by passing the thiosulfate ion through three key sites in the YeeE structure. In the model, the first site attracts a thiosulfate ion to the positively charged surface. Then using S-H-S type hydrogen bonds, YeeE passes the ion to the other two sites and on to the cytoplasm without itself undergoing any major conformational changes.This mechanism for uptake is quite unusual among membrane transporters according to Tanaka."There is a lot less movement compared with transporters that have inward- and outward-facing structures or use rocking bundle motions. YeeE is not structurally designed to use these other mechanisms," he says.Tsukazaki adds that the new mechanism expands our knowledge of nutrient transport into a cell, knowledge that can be exploited for industrial purposes."Comparatively little is known about the YeeE family of transport proteins. Through more study of the structure for the uptake and genetic modifications, we might be able to artificially design 'super' YeeE that enhance L-cysteine productivity via high thiosulfate uptake" he says.
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Biotechnology
| 2,020 |
August 26, 2020
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https://www.sciencedaily.com/releases/2020/08/200826141358.htm
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Key immune system protein discovered in plants
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Plants have a unique ability to safeguard themselves against pathogens by closing their pores -- but until now, no one knew quite how they did it. Scientists have known that a flood of calcium into the cells surrounding the pores triggers them to close, but how the calcium entered the cells was unclear.
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A new study by an international team including University of Maryland scientists reveals that a protein called OSCA1.3 forms a channel that leaks calcium into the cells surrounding a plant's pores, and they determined that a known immune system protein triggers the process.The findings are a major step toward understanding the defense mechanisms plants use to resist infection, which could eventually lead to healthier, more resistant and more productive crops. The research paper was published on August 26, 2020 in the journal "This is a major advance, because a substantial part of the world's food generated by agriculture is lost to pathogens, and we now know the molecular mechanism behind one of the first and most relevant signals for plant immune response to pathogens -- the calcium burst after infection," said José Feijó, a professor of cell biology and molecular genetics at UMD and co-author of the study. "Finding the mechanism associated with this calcium channel allows further research into its regulation, which will improve our understanding of the way in which the channel activity modulates and, eventually, boosts the immune reaction of plants to pathogens."Plant pores -- called stomata -- are encircled by two guard cells, which respond to calcium signals that tell the cells to expand or contract and trigger innate immune signals, initiating the plant's defense response. Because calcium cannot pass directly through the guard cell membranes, scientists knew a calcium channel had to be at work. But they didn't know which protein acted as the calcium channel.To find this protein, the study's lead author, Cyril Zipfel, a professor of molecular and cellular plant physiology at the University of Zurich and Senior Group Leader at The Sainsbury Laboratory in Norwich, searched for proteins that would be modified by another protein named BIK1, which genetic studies and bioassays identified as a necessary component of the immune calcium response in plants.When exposed to BIK1, one protein called OSCA1.3 transformed in a very specific way that suggested it could be a calcium channel for plants. OSCA1.3 is a member of a widespread family of proteins known to exist as ion channels in many organisms, including humans, and it seems to be specifically activated upon detection of pathogens.To determine if OSCA1.3 was, in fact, the calcium channel they were looking for, Zipfel's team reached out to Feijó, who is well known for identifying and characterizing novel ion channels and signaling mechanisms in plants. Erwan Michard, a visiting assistant research scientist in Feijó's lab and co-author of the paper, conducted experiments that revealed BIK1 triggers OSCA1.3 to open up a calcium channel into a cell and also explained the mechanism for how it happens.BIK1 only activates when a plant gets infected with a pathogen, which suggests that OSCA1.3 opens a calcium channel to close stomata as a defensive, immune system response to pathogens."This is a perfect example of how a collaborative effort between labs with different expertise can bring about important conclusions that would be difficult on solo efforts," Feijó, said. "This fundamental knowledge is badly needed to inform ecology and agriculture on how the biome will react to the climatic changes that our planet is going through."Feijó, will now incorporate this new knowledge of the OSCA1.3 calcium channel into other areas of research in his lab, which is working to understand how the mineral calcium was co-opted through evolution by all living organisms to serve as a signaling device for information about stressors from infection to climate change."Despite the physiological and ecological relevance of stomatal closure, the identity of some of the key components mediating this closure were still unknown," Zipfel said. "The identification of OSCA1.3 now fills one of these important gaps. In the context of plant immunity this work is particularly apt in 2020, the UN International Year of Plant Health."
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Biotechnology
| 2,020 |
August 26, 2020
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https://www.sciencedaily.com/releases/2020/08/200826123123.htm
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Novel PROTAC enhances its intracellular accumulation and protein knockdown
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Cancer therapies sometimes involve drugs that mediate the breakdown of specific intracellular proteins that participate in cancer formation and proliferation. Proteolysis-targeting chimeras or PROTACs are a promising type of protein-degrader molecules, but their effectiveness has been challenged by their limited ability to accumulate inside cells.
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In this study published in the journal "PROTACs are molecules made of two parts: one binds to the target protein and the other links to the enzyme ubiquitin ligase. The two parts are joined by a chemical linker," said corresponding author Dr. Jin Wang, associate professor of pharmacology and chemical biology and of molecular and cellular biology at Baylor.Once PROTAC binds to its target protein and the ubiquitin ligase, the ligase will attach ubiquitin groups to the surface of the target protein, which tags it for degradation by the proteasome inside the cell.Wang and his colleagues were interested in improving PROTAC's ability to degrade target proteins inside cells. Drawing from their years of expertise in the fields of organic chemistry and chemical biology, they experimented with different chemical binders to determine how they affected PROTAC's efficacy. Serendipitously, they discovered that a specific type of chemistry can enhance PROTAC's intracellular accumulation.The researchers worked with a well-known target of PROTAC, the enzyme Bruton's tyrosine kinase (BTK). They used different types of binders -- reversible noncovalent, reversible covalent, and irreversible covalent -- to construct PROTACs that targeted BTK."When we compared the different constructs in their ability to degrade BTK, we were excited to discover that the cyano-acrylamide-based reversible covalent chemical binder significantly enhanced the intracellular accumulation and target engagement of PROTACs, much better than the others," said co-first author Dr. Wen-Hao Guo, a postdoctoral associate in the Wang lab."Furthermore, we developed RC-1, a reversible covalent BTK PROTAC that was effective as both a BTK inhibitor and degrader," said co-first author Dr. Xiaoli Qi, assistant professor in pharmacology and chemical biology at Baylor. "This represents a novel mechanism of action for PROTACs. Our work suggests the possibility that this strategy to improve PROTACs can be applied to target other molecules."
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Biotechnology
| 2,020 |
August 26, 2020
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https://www.sciencedaily.com/releases/2020/08/200826110326.htm
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Unique HIV reservoirs in elite controllers
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Xu Yu, MD, Ragon group leader, recently published a study entitled "Distinct viral reservoirs in individuals with spontaneous control of HIV-1," in
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HIV affects more than 35 million people worldwide and can be effectively controlled, but not cured, with a daily regimen of ART. Upon infection, retroviruses like HIV place copies of their viral genetic material into cells' genomes, creating viral reservoirs, sanctuaries where HIV persists despite ART, throughout the body. When a complete copy of the virus, or intact viral genome, is incorporated into a cell's genome, it can be used to create new copies of HIV. For people living with HIV, this means that if they stop taking ART, the intact viral genomes previously integrated into the cells' genomes start making new copies of the virus, leading to rapid viral rebound and disease progression. The HIV viral reservoir has remained a major obstacle to an HIV cure.Elite controllers' immune systems use a T-cell mediated immune response to control the virus without medication, to the point that the virus is completely undetectable by standard assays. Understanding the interplay between their immune system and HIV may hold the key to helping the immune systems of people living with HIV to suppress the virus without daily treatment, achieving what is known as a functional cure.Yu's group studied the viral reservoir in elite controllers, using next-generation sequencing techniques to precisely map the locations of intact HIV genomes in the human genome. They found that in elite controllers, HIV was often found in locations of the genome that researchers call gene deserts. In these inactive parts of the human genome, human DNA is never turned on, and HIV cannot be effectively expressed but remains in a "blocked and locked" state. This means that HIV is locked in the cell's genome, and the viral genome is blocked from being used to create more viruses and is therefore incapable of causing disease."This positioning of viral genomes in elite controllers," Yu, says, "is highly atypical, as in the vast majority of people living with HIV-1, HIV is located in the active human genes where viruses can be readily produced."When the authors collected cells from elite controllers and infected them with HIV in the lab, they found the virus integrated into active sites in the cell genomes, not in the inactive gene deserts. This suggests that the elite controllers' unique viral reservoirs may be a result of their HIV-suppressing T cell response eliminating intact viral genomes from active sites.If researchers are able to identify which viral reservoirs can make new copies of the virus after treatment stops, it may help them to target a treatment against the active, or rebound-competent, reservoirs. This study suggests that if researchers can activate the kind of T cell immunity that is present in elite controllers, they may be able to eliminate rebound-competent viral reservoirs in people living with HIV, achieving a functional cure. The remaining viral DNA, located in non-active parts of the human genome, could be allowed to exist without causing disease.Yu's group had one more finding: one of their elite controller participants had no intact HIV found in over 1.5 billion cells analyzed. This raises the possibility that a "sterilizing cure" of HIV, in which the participant's immune system has removed all intact HIV genomes from the body, may be achieved naturally in extremely rare instances.This project was supported by the National Heart, Lung, and Blood Institute, the National Institute of Allergy and Infectious Diseases, the National Institute of Drug Abuse, the National Institute of Health, the Mark and Lisa Schwartz Family Foundation, the Ragon Institute of MGH, MIT and Harvard, the Bill & Melinda Gates Foundation, and the Foundation for AIDS Research (amfAR).
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Biotechnology
| 2,020 |
August 26, 2020
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https://www.sciencedaily.com/releases/2020/08/200826110324.htm
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Tag team gut bacteria worsen symptoms of multiple sclerosis
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Researchers at the RIKEN Center for Integrative Medical Sciences (IMS) have discovered that a particular combination of microorganisms in the gut can worsen symptoms in a mouse model of multiple sclerosis. The study published in the scientific journal
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Multiple sclerosis is an autoimmune disease in which the immune system attacks the myelin that covers the nerve cells of the brain and spinal cord. Demyelination affects how rapidly neurons communicate with each other and with muscles, causing a variety of symptoms including numbness, weak muscles, tremors, and the inability to walk. Gut microorganisms have been reported to affect symptoms of multiple sclerosis, but how bacteria in the intestines can affect myelin of the brain and spinal cord remained a mystery.Researchers led by Hiroshi Ohno at RIKEN IMS set out find this connection using a mouse model of the disease. These mice experience similar demyelination of the spinal cord that results from autoimmune attacks by T cells that produce the cytokine IL-17A. However, giving these mice the antibiotic ampicillin reduced demyelination. The treatment also prevented the activation of a particular type of T cell. As Ohno explains, "we found that treatment with ampicillin, and only ampicillin, selectively reduced activity of T cells that attack an important protein called myelin oligodendrocyte glycoprotein [MOG], which helps myelin stick to neurons."This was confirmed by taking immune cells from the small intestines and other regions and measuring their cytokine production in the presence of MOG. Production was only reduced by ampicillin and only when the T cells came from the small intestine. At this point, the team knew that microorganisms in the small intestine activate MOG-specific T cells, which can then go and attack myelin. The next step was to figure out which bacteria were responsible.Because only ampicillin reduced symptoms in the model mice, they looked for microbiota that were almost completely deleted only in ampicillin-treated mice. They found only one such bacteria, a new strain called OTU002. To test the hypothesis that OTU002 was the culprit, they examined mice that lacked all bacteria except OTU002. They found that symptoms in these mice were more severe than those in germ-free mice. At this point, the team knew that their newly discovered gut bacterium was responsible for the worsening symptoms."But, there was a problem," says first author Eiji Miyauchi. "Symptoms in the OTU002-only mice were not as bad as those in the regular model mice. This means that the original effect must involve more than one microorganism." The team hypothesized that a different bacterium was cross-reacting with MOG-specific T cells, mimicking the location on MOG that the T cells recognize. Shotgun genome sequencing revealed that a protein expressed by "Other studies have focused on fecal microbes, or a single microbe, in patients with multiple sclerosis or in model mice," says Miyauchi. "Our data emphasize the necessity of considering the synergistic effects of intestinal microbes on autoimmune diseases and give hope to people looking for effective treatments for multiple sclerosis.""But, because gut microbes and T cell binding locations on myelin differ between mouse and human, further studies using human microbes and autoreactive T cells are now needed."
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Biotechnology
| 2,020 |
August 25, 2020
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https://www.sciencedaily.com/releases/2020/08/200825165201.htm
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Using light's properties to indirectly see inside a cell membrane
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For those not involved in chemistry or biology, picturing a cell likely brings to mind several discrete, blob-shaped objects; maybe the nucleus, mitochondria, ribosomes and the like.
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There's one part that's often overlooked, save perhaps a squiggly line indicating the cell's border: the membrane. But its role as gatekeeper is an essential one, and a new imaging technique developed at the McKelvey School of Engineering at Washington University in St. Louis is providing a way to see into, as opposed to through, this transparent, fatty, protective casing.The new technique, developed in the lab of Matthew Lew, assistant professor in the Preston M. Green Department of Electrical and Systems Engineering, allows researchers to distinguish collections of lipid molecules of the same phase -- the collections are called nanodomains -- and to determine the chemical composition within those domains.The details of this technique -- single-molecule orientation localization microscopy, or SMOLM -- were published online Aug. 21 in Editors at the journal -- a leading one in general chemistry -- selected Lew's paper as a "Hot Paper" on the topic of nanoscale papers. Hot Papers are distinguished by their importance in a rapidly evolving field of high interest.Using traditional imaging technologies, it's difficult to tell what's "inside" versus "outside" a squishy, transparent object like a cell membrane, Lew said, particularly without destroying it."We wanted a way to see into the membrane without traditional methods" -- such as inserting a fluorescent tracer and watching it move through the membrane or using mass spectrometry -- "which would destroy it," Lew said.To probe the membrane without destroying it, Jin Lu, a postdoctoral researcher in Lew's lab, also employed a fluorescent probe. Instead of having to trace a path through the membrane, however, this new technique uses the light emitted by a fluorescent probe to directly "see" where the probe is and where it is "pointed" in the membrane. The probe's orientation reveals information about both the phase of the membrane and its chemical composition."In cell membranes, there are many different lipid molecules," Lu said. "Some form liquid, some form a more solid or gel phase."Molecules in a solid phase are rigid and their movement constrained. They are, in other words, ordered. When they are in a liquid phase, however, they have more freedom to rotate; they are in a disordered phase.Using a model lipid bilayer to mimic a cell membrane, Lu added a solution of fluorescent probes, such as Nile red, and used a microscope to watch the probes briefly attach to the membrane.A probe's movement while attached to the membrane is determined by its environment. If surrounding molecules are in a disordered phase, the probe has room to wiggle. If the surrounding molecules are in an ordered phase, the probe, like the nearby molecules, is fixed.When light is shined on the system, the probe releases photons. An imaging method previously developed in the Lew lab then analyzes that light to determine the orientation of the molecule and whether it's fixed or rotating."Our imaging system captures the emitted light from single fluorescent molecules and bends the light to produce special patterns on the camera," Lu said."Based on the image, we know the probe's orientation and we know whether it's rotating or fixed," and therefore, whether it's embedded in an ordered nanodomain or not.Repeating this process hundreds of thousands of times provides enough information to build a detailed map, showing the ordered nanodomains surrounded by the ocean of the disordered liquid regions of the membrane.The fluorescent probe Lu used, Nile red, is also able to distinguish between lipid derivatives within the same nanodomains. In this context, their chosen fluorescent probe can tell whether or not the lipid molecules are hydrolyzed when a certain enzyme was present."This lipid, named sphingomyelin, is one of the critical components involved in nanodomain formation in cell membrane. An enzyme can convert a sphingomyelin molecule to ceramide," Lu said. "We believe this conversion alters the way the probe molecule rotates in the membrane. Our imaging method can discriminate between the two, even if they stay in the same nanodomain."This resolution, a single molecule in model lipid bilayer, cannot be accomplished with conventional imaging techniques.This new SMOLM technique can resolve interactions between various lipid molecules, enzymes and fluorescent probes with detail that has never been achieved previously. This is important particularly in the realm of soft matter chemistry."At this scale, where molecules are constantly moving, everything is self-organized," Lew said. It's not like solid-state electronics where each component is connected in a specific and importantly static way."Every molecule feels forces from those surrounding it; that's what determines how a particular molecule will move and perform its functions."Individual molecules can organize into these nanodomains that, collectively, can inhibit or encourage certain things -- like allowing something to enter a cell or keeping it outside."These are processes that are notoriously difficult to observe directly," Lew said. "Now, all you need is a fluorescent molecule. Because it's embedded, its own movements tell us something about what's around it."
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Biotechnology
| 2,020 |
August 25, 2020
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https://www.sciencedaily.com/releases/2020/08/200825142335.htm
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Study shines new light on young tree seedlings
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The first few weeks of a tree seedling's life can be the most precarious.
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As it pushes thin new roots into the ground it's also reaching up with tiny new leaves. Water and energy are precious. Most seedlings never make it past their first month on the ground.But while much is known about the growing process, there remains a layer of mystery around the mechanisms within these small plants. Now, a new study by a University of Georgia researcher sheds some light on the microscopic tissues that help tree seedlings grow. The results could change how researchers and growers view the first weeks of a tree's life."I've been working on newly germinated seedlings for 20 years, and I feel this is one of the first breakthroughs for me about how different they are, even from a 20-week-old seedling," said Dan Johnson, an assistant professor of tree physiology and forest ecology at the UGA Warnell School of Forestry and Natural Resources. "It's these first few weeks of life that seem to be fundamentally different."Johnson and a team of researchers used a high-powered X-ray called a synchrotron to take extremely detailed cross-section images of ponderosa pine seedlings at various stages of hydration. Located at the University of California-Berkeley, the synchrotron accelerates electrons to nearly the speed of light, and while they will instantly kill a human cell, plants, it turns out, can withstand the intense power for a short period of time.So, Johnson and his collaborators X-rayed the intact stem of the pine seedlings over several days, taking images of what was going on inside the plant. The pictures show extremely detailed black-and-white images that detail pockets of hydrated cells in gray. As the images progress and the seedlings dry out, black pockets of air can be seen on the images, almost as if the stems are being eaten from the outside in.He and other researchers thought the plant's xylem -- a central nervous system of the plant, in a sense -- would quickly dry up if it went without water. Turns out, they were wrong -- and the resulting images offer never-before-seen insights into the first few weeks of a tree seedling's life."The way we thought these seedlings were going to fail, hydraulically, as they dried out, was not at all how they failed," he said. "We thought the vascular tissue -- the xylem -- was going to be filled with air. We call it embolism in humans. But what we found was, it wasn't the xylem that dried out, it was all the tissue surrounding it. Even in some of the seedlings that looked like they were ripped apart (for lack of water), the xylem is fully hydrated."All plants have xylem tissue; it transports water throughout the plant. And in older plants, the xylem often does dry out as a plant faces drought. But the images that Johnson captured show that seedlings' plumbing is completely different from their older cousins.The findings were published in the August issue of the "To me, this is the most vulnerable life stage. If a seedling is going to die, it's going to die in the first few weeks of life," said Johnson. "In the field, we see 99% of natural regeneration seedlings die -- you'll come back to the field one day and thousands have died. And they die in places where it just dries out too quickly."Johnson said his findings point to how sensitive tissues outside the xylem are to water loss in the first few weeks of a seedling's life. When a wild-sewn seedling survives, it's often because that particular site had more favorable conditions, such as more moisture or the seed landed in a depression where it was more protected from the elements.In addition to the detailed black-and-white images, the team also made corresponding color images of the seedling stems with a laser, using a process called confocal microscopy. Different cells reflect in different colors, creating a rainbow of circles that researchers can use to better identify parts of the stem.But while the yellows, reds and blues are striking on the laser-produced images, the real eye-opener for Johnson was the black-and-white reality of the decimated, dried out stems and their central core, which was the last to give up."I was completely shocked. It was not what any of us on the paper expected," said Johnson, pointing to one image of a withered stem that looks almost chewed up. "That's at a desiccation level that would kill that plant. So, to have that xylem so full when it's so dead is counter-intuitive."While the discovery may bring more questions than answers, Johnson notes that the survival of the xylem may change how plants' first few weeks are understood. It's almost as if, he said, the first leaves to emerge from a seedling are connected to a completely different set of tissues. "The xylem might not be the plumbing to the first few leaves of the plant, which is bizarre because that's what we learned in plant physiology," he added.
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Biotechnology
| 2,020 |
August 25, 2020
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https://www.sciencedaily.com/releases/2020/08/200825113619.htm
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Accumulating extra genome copies may protect fly brain cells during aging
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Scientists have discovered a novel anti-aging defence in the brain cells of adult fruit flies: producing extra copies of the genome, according to a new study published today in
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The findings could help explain how the brain, which rarely produces new cells, is able to cope with the accumulation of cell damage over time and prevent excess cell loss during aging. They may also help scientists answer questions about human age-related brain diseases.Most cells have two copies of each chromosome, one from each parent. This provides each cell with two complete copies of DNA, referred to as the genome. But some cells accumulate extra copies of chromosomes, although how and why they do this is not entirely clear. Some scientists have suggested that excess chromosomes might lead to age-related brain diseases."The brain of the fruit fly Drosophila melanogaster is an ideal model for studying age-related changes in the brain because the fly has a relatively short lifespan, its brain cells rarely multiply, and we have excellent tools for manipulating fly genetics," says lead author Shyama Nandakumar, a doctoral student in the department of Molecular, Cellular, and Developmental Biology at the University of Michigan, Ann Arbor, US.In the study, Nandakumar and colleagues examined adult fly brain cells and found that some accumulate extra copies of their genome, especially in parts of the brain responsible for vision -- the region of the brain which shows more DNA damage with age.Next, they subjected fly brain cells to oxidative stress or ultraviolet radiation, which damage DNA and can cause cell death. They found that this exposure increased the production of extra copies of chromosomes in the cells, and the cells were less likely to die as a result of the damage. "These data suggest that cells with extra copies of the genome are more resistant to cell death and may serve a beneficial or protective role in the aging brain," Nandakumar explains.Previous studies have found that patients with early stages of Alzheimer's disease have extra chromosomes in their brain cells than people of the same age who do not have the condition. This has led scientists to question whether accumulating extra chromosomes leads to brain cell death and brain disease."Our study suggests that the production of extra copies of chromosomes might actually be a normal response to the accumulation of age-related damage in flies and may even help protect against cell death," concludes senior author Laura Buttitta, Associate Professor of Molecular, Cellular and Developmental Biology at the University of Michigan. "Further studies are now needed to determine if this is also true in humans."
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Biotechnology
| 2,020 |
August 25, 2020
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https://www.sciencedaily.com/releases/2020/08/200825110648.htm
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Study revealing structure of a protein complex may open doors to better disease research
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More than two decades ago scientists discovered the Arp2/3 complex, an actin (cellular protein) cytoskeketal nucleator which plays a crucial role in cell division, immune response, neurodevelopment other biological processes. But there has been no determined structure of the activated state of the complex until now, an achievement that may lay the foundation for uncovering its role in biology and in the development of disease. Researchers at Stony Brook University led by Saikat Chowdhury, PhD, determined the structure of Arp2/3 and describe it in a paper published in
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Chowdhury, and graduate student and first author Mohammed Shaaban determined the first near-atomic resolution structure of Arp2/3 in its active state by using cryo-electron microscopy. The structure shows the complex in its active form and bound to a signaling molecule. It also shows the nucleated actin filament, thus providing a structural snapshot of the global and local conformational changes in the Arp2/3 complex that help grow new actin filament in cells."Obtaining the macromolecular structure of activated Arp2/3 complex has been a long-standing goal for scientists," says Chowdhury, senior author and an Assistant Professor in the Department of Biochemistry and Cell Biology in the College of Arts and Sciences at Stony Brook University. "Our structure reveals a level of molecular details which show the individual components of the complex and how they are positioned relative to each other in the active state."Having a structure of Arp2/3 in its active state will help drive more detailed research of the complex. Chowdhury explains that this is extremely important because when Arp2/3 is deregulated in the biological state, it is associated with cancer metastasis, neurodegeneration, bacterial and viral infections and wound healing problems."So not only does this structure enable us to fill a knowledge gap in the actin cell biology field, it potentially helps to build our understanding of the underlying causes of a number of diseases with the ultimate goal of developing new therapeutics," emphasizes Chowdhury.Determining the Arp2/3 structure in its activated state required the researchers to use technologies available in Stony Brook University's Cryo-Electron Microscopy Facility, a center supported by the National Institutes of Health (NIH), and the High Performance Computing capabilities in the Division of Information Technology.Chowdhury is also affiliated with the Institute of Engineering Driven Medicine and Institute of Chemical Biology & Drug Discovery at Stony Brook University, as well as an affiliated scientist at Brookhaven National Laboratory. The research was done in collaboration with Brad Nolen at the University of Oregon and supported in part by the NIH and SUNY, Stony Brook University.
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Biotechnology
| 2,020 |
August 24, 2020
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https://www.sciencedaily.com/releases/2020/08/200824120048.htm
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Large molecules need more help to travel through a nuclear pore into the cell nucleus
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A new study in the field of biophysics has revealed how large molecules are able to enter the nucleus of a cell. A team led by Professor Edward Lemke of Johannes Gutenberg University Mainz (JGU) has thus provided important insights into how some viruses, for example, can penetrate into the nucleus of a cell, where they can continue to proliferate and infect others. They have also demonstrated that the efficiency of transport into a cell decreases as the size of the molecules increases and how corresponding signals on the surface can compensate for this.
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"We have been able to gain new understanding of the transport of large biostructures, which helped us develop a simple model that describes how this works," said Lemke, a specialist in the field of biophysical chemistry. He is Professor of Synthetic Biophysics at JGU and Adjunct Director of the Institute of Molecular Biology (IMB) in Mainz.A typical mammalian cell has about 2,000 nuclear pores, which act as passageways from the cell cytoplasm into the cell nucleus and vice versa. These pores in the nuclear envelope act as gatekeepers that control access and deny entry to larger molecules of around five nanometers in diameter and greater. Molecules that have certain nuclear localization sequences on their surface can bind to structures within nuclear pores, allowing them to enter into the nucleus rapidly. "Nuclear pores are remarkable in the diversity of cargoes they can transport. They import proteins and viruses into the nucleus and export ribonucleic acids and proteins into the cell cytoplasm," explained Lemke, describing the function of these pores. "Despite the fundamental biological relevance of the process, it has always been an enigma how large cargoes greater than 15 nanometers are efficiently transported, particularly in view of the dimensions and structures of nuclear pores themselves."With this is mind and as part of their project, the researchers designed a set of large model transport cargoes. These were based on capsids, i.e., protein "shells" in viruses that enclose the viral genome. The cargo models ranging from 17 to 36 nanometers in diameter were then fluorescently labeled, allowing them to be observed on their way through cells. Capsid models without nuclear localization signals on their surface remained in the cell cytoplasm and did not enter the cell nucleus. As the number of nuclear localization signals increased, the accumulation of the model capsid in the nucleus became more efficient. But even more interestingly, the researchers found that the larger the capsid, the greater was the number of nuclear localization signals needed to enable efficient transport into the nucleus.The research team looked at a range of capsids of various viruses including the hepatitis B capsid, the largest cargo used in this study. But even increasing the number of nuclear localization signals to 240 did not result in the transport of this capsid into the nucleus. This corresponds with the results of earlier studies of the hepatitis B virus that have indicated that only the mature infectious virus is capable of passage through a nuclear pore into the nucleus.In cooperation with Professor Anton Zilman of the University of Toronto in Canada, a mathematical model was developed to shed light on the transport mechanism and to establish the main factors determining the efficiency of transport. "Our simple two-parameter biophysical model has recreated the requirements for nuclear transport and revealed key molecular determinants of the transport of large biological cargoes on cells," concluded first author Giulia Paci, who carried out the study as part of her PhD thesis at the European Molecular Biology Laboratory (EMBL) in Heidelberg.
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Biotechnology
| 2,020 |
August 21, 2020
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https://www.sciencedaily.com/releases/2020/08/200821155747.htm
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Nasal vaccine against COVID-19 prevents infection in mice
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Scientists at Washington University School of Medicine in St. Louis have developed a vaccine that targets the SARS-CoV-2 virus, can be given in one dose via the nose and is effective in preventing infection in mice susceptible to the novel coronavirus. The investigators next plan to test the vaccine in nonhuman primates and humans to see if it is safe and effective in preventing COVID-19 infection.
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The study is available online in the journal Unlike other COVID-19 vaccines in development, this one is delivered via the nose, often the initial site of infection. In the new study, the researchers found that the nasal delivery route created a strong immune response throughout the body, but it was particularly effective in the nose and respiratory tract, preventing the infection from taking hold in the body."We were happily surprised to see a strong immune response in the cells of the inner lining of the nose and upper airway -- and a profound protection from infection with this virus," said senior author Michael S. Diamond, MD, PhD, the Herbert S. Gasser Professor of Medicine and a professor of molecular microbiology, and of pathology and immunology. "These mice were well protected from disease. And in some of the mice, we saw evidence of sterilizing immunity, where there is no sign of infection whatsoever after the mouse is challenged with the virus."To develop the vaccine, the researchers inserted the virus' spike protein, which coronavirus uses to invade cells, inside another virus -- called an adenovirus -- that causes the common cold. But the scientists tweaked the adenovirus, rendering it unable to cause illness. The harmless adenovirus carries the spike protein into the nose, enabling the body to mount an immune defense against the SARS-CoV-2 virus without becoming sick. In another innovation beyond nasal delivery, the new vaccine incorporates two mutations into the spike protein that stabilize it in a specific shape that is most conducive to forming antibodies against it."Adenoviruses are the basis for many investigational vaccines for COVID-19 and other infectious diseases, such as Ebola virus and tuberculosis, and they have good safety and efficacy records, but not much research has been done with nasal delivery of these vaccines," said co-senior author David T. Curiel, MD, PhD, the Distinguished Professor of Radiation Oncology. "All of the other adenovirus vaccines in development for COVID-19 are delivered by injection into the arm or thigh muscle. The nose is a novel route, so our results are surprising and promising. It's also important that a single dose produced such a robust immune response. Vaccines that require two doses for full protection are less effective because some people, for various reasons, never receive the second dose."Although there is an influenza vaccine called FluMist that is delivered through the nose, it uses a weakened form of the live influenza virus and can't be administered to certain groups, including those whose immune systems are compromised by illnesses such as cancer, HIV and diabetes. In contrast, the new COVID-19 intranasal vaccine in this study does not use a live virus capable of replication, presumably making it safer.The researchers compared this vaccine administered to the mice in two ways -- in the nose and through intramuscular injection. While the injection induced an immune response that prevented pneumonia, it did not prevent infection in the nose and lungs. Such a vaccine might reduce the severity of COVID-19, but it would not totally block infection or prevent infected individuals from spreading the virus. In contrast, the nasal delivery route prevented infection in both the upper and lower respiratory tract -- the nose and lungs -- suggesting that vaccinated individuals would not spread the virus or develop infections elsewhere in the body.The researchers said the study is promising but cautioned that the vaccine so far has only been studied in mice."We will soon begin a study to test this intranasal vaccine in nonhuman primates with a plan to move into human clinical trials as quickly as we can," Diamond said. "We're optimistic, but this needs to continue going through the proper evaluation pipelines. In these mouse models, the vaccine is highly protective. We're looking forward to beginning the next round of studies and ultimately testing it in people to see if we can induce the type of protective immunity that we think not only will prevent infection but also curb pandemic transmission of this virus."This work was supported by the National Institutes of Health (NIH), grant and contract numbers 75N93019C00062, R01 AI127828, R01 AI130591, R01 AI149644, R35 HL145242, HHSN272201400018C, HHSN272201200026C, F32 AI138392 and T32 AI007163; the Defense Advanced Research Project Agency, grant number HR001117S0019; a Helen Hay Whitney Foundation postdoctoral fellowship; and the Pulmonary Morphology Core at Washington University School of Medicine.Diamond is a consultant for Inbios, Vir Biotechnology, NGM Biopharmaceuticals, and on the scientific advisory board of Moderna. The Diamond laboratory has received unrelated funding support from Moderna, Vir Biotechnology, and Emergent BioSolutions. Diamond, Curiel, Ahmed Hassan and Igor Dmitriev have filed a disclosure with Washington University for possible development of ChAd-SARS-CoV-2. Michael Holtzman is a member of the DSMB for AstroZeneca and founder of NuPeak Therapeutics. The Baric laboratory has received unrelated funding support from Takeda, Pfizer and Eli Lily.
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Biotechnology
| 2,020 |
August 21, 2020
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https://www.sciencedaily.com/releases/2020/08/200821141154.htm
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Bioluminescent tag to detect DNA break repair
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A new bioluminescent reporter that tracks DNA double stranded break (DSB) repair in cells has been developed by researchers from Massachusetts General Hospital (MGH) and the Academia Sinica in Taiwan. The international team's novel bioluminescent repair reporter (BLRR)-based system can be used to monitor DNA repair pathways directly in animals as well as cell lines. No such system previously existed for in vivo studies. These pathways play a crucial role in multiple conditions, including cancer.
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"One of the main reasons cancer cells are resistant to treatment is that they can inherently repair the DNA damage caused by radiation and chemotherapy," explains Christian Elias Badr, PhD, investigator in the Department of Neurology at MGH and co-senior author of the paper. The study's other co-senior author is Charles Pin-Kuang Lai, PhD, at the Academia Sinica in Taiwan.Their study appeared this month as an online advance paper in DSB damage repair is key to maintaining genomic integrity and cell viability. It also plays a role in cancer treatment, which often includes chemoradiotherapy (radiation and chemotherapy), which disrupts DSB. A cell may recognize the damage and use its intrinsic DNA damage response (DDR) to reduce DSB-caused cell death. As a result, the cancer cell's own DNA repair mechanisms can promote drug resistance and recurrence in some malignancies. Researchers would like to know more about them.The BLRR approach builds on earlier work members of the team did on enzymes called luciferases. These produce bioluminescence, making them useful for tracking molecules in cells. BLRR uses secreted Gaussia and Vargula luciferases to detect homology-directed repair (HDR) and non-homologous end joining (NHEJ) -- the two major pathways to DSB repair. Using BLRR. Researchers can track HDR and NHEJ-related activities over time in cells. It also detects DSB repairs in xenografted tumors in vivo."You can study DNA damage in cells with next generation sequencing (NGS), but that's more costly and time consuming," Badr says. "And our system's accuracy is comparable to NGS."The researchers used their new tag to carry out multiple studies. In one, they found a significant difference in the efficiency of CRISPR/Cas9-mediated editing with guide RNAs 1-10bp apart. They also used BLRR analysis to detect altered dynamics for DSB repair induced by small molecule modulators. Lastly, they used the system to discover HDR-suppressing functions of anticancer cardiac glycosides in human glioblastomas and glioma cancer stem-like cells by inhibiting DNA repair protein RAD51 homolog 1.In their paper, the authors describe the BLRR system as: "A highly sensitive platform to simultaneously and longitudinally track HDR and NHEJ dynamics that is sufficiently versatile for elucidating the physiology and therapeutic development of DSB repair." The authors plan on using this reporter system in high throughput drug screening to identify novel therapeutics that sensitize cancer cells to radiation and chemotherapy.
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Biotechnology
| 2,020 |
August 20, 2020
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https://www.sciencedaily.com/releases/2020/08/200820102454.htm
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Studying viral outbreaks in single cells could reveal new ways to defeat them
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Many viruses, including HIV and influenza A, mutate so quickly that identifying effective vaccines or treatments is like trying to hit a moving target. A better understanding of viral propagation and evolution in single cells could help. Today, scientists report a new technique that can not only identify and quantify viral RNA in living cells, but also detect minor changes in RNA sequences that might give viruses an edge or make some people "superspreaders."
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The researchers will present their results at the American Chemical Society (ACS) Fall 2020 Virtual Meeting & Expo."For studying a new virus like SARS-CoV-2, it's important to understand not only how populations respond to the virus, but how individuals -- either people or cells -- interact with it," says Laura Fabris, Ph.D., the project's principal investigator. "So we've focused our efforts on studying viral replication in single cells, which in the past has been technically challenging."Analyzing individual cells instead of large populations could go a long way toward better understanding many facets of viral outbreaks, such as superspreaders. That's a phenomenon in which some cells or people carry unusually high amounts of virus and therefore can infect many others. If researchers could identify single cells with high viral loads in superspreaders and then study the viral sequences in those cells, they could perhaps learn how viruses evolve to become more infectious or to outwit therapies and vaccines. In addition, features of the host cell itself could aid various viral processes and thus become targets for therapies. On the other end of the spectrum, some cells produce mutated viruses that are no longer infectious. Understanding how this happens could also lead to new antiviral therapies and vaccines.But first, Fabris and colleagues at Rutgers University needed to develop an assay that was sensitive enough to detect viral RNA, and its mutations, in single living cells. The team based their technique on surface enhanced Raman spectroscopy (SERS), a sensitive method that detects interactions between molecules through changes in how they scatter light. The researchers decided to use the method to study influenza A. To detect the virus's RNA, they added to gold nanoparticles a "beacon DNA" specific to influenza A. In the presence of influenza A RNA, the beacon produced a strong SERS signal, whereas in the absence of this RNA, it did not. The beacon produced weaker SERS signals with increasing numbers of viral mutations, allowing the researchers to detect as few as two nucleotide changes. Importantly, the nanoparticles could enter human cells in a dish, and they produced a SERS signal only in those cells expressing influenza A RNA.Now, Fabris and colleagues are making a version of the assay that produces a fluorescent signal, instead of a SERS signal, when viral RNA is detected. "SERS is not a clinically approved technology. It's just now breaking into the clinic," Fabris notes. "So we wanted to provide clinicians and virologists with an approach they would be more familiar with and have the technology to use right now." In collaboration with virologists and mathematicians at other universities, the team is developing microfluidic devices, or "lab-on-a-chip" technologies, to read many fluorescent samples simultaneously.Because SERS is more sensitive, cheaper, faster and easier to perform than other assays based on fluorescence or the reverse transcriptase-polymerase chain reaction (known as RT-PCR), it could prove ideal for detecting and studying viruses in the future. Fabris is now collaborating with a company that makes a low-cost, portable Raman spectrometer, which would enable the SERS assay to be easily conducted in the field.Fabris and her team are also working on identifying regions of the SARS-CoV-2 genome to target with SERS probes. "We're in the process of obtaining funding to work on possible SARS-CoV-2 diagnostics with the SERS method we developed," Fabris says.A brand-new video about the research is available at
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Biotechnology
| 2,020 |
August 20, 2020
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https://www.sciencedaily.com/releases/2020/08/200820102420.htm
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Biochemistry: Move over Michaelis-Menten!
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Cells send signals through enzyme cascades, where one enzyme passes the signal to the next. In such cascades, it is crucial that the enzyme recognizes the right substrates to ensure that, for example, a hormone activates the right cellular activities. Protein kinases, the enzymes in such cascades, are usually not sufficiently specific on their own, and therefore they rely on other proteins to physically connect them to the right substrates.
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"Currently, we describe signalling enzymes with equations developed for metabolic enzymes," Magnus Kjærgaard explains. "Metabolic enzymes that make energy for our bodies, for example, need to process many substrates per minute. In contrast, signalling enzymes act as switches, and often only need to convert a single substrate to have an effect. Therefore, the equations developed to describe metabolic enzymes are less relevant for signalling enzymes."For more than a hundred years, biochemists have described the activity of enzymes using the Michaelis-Menten equation, which describes how activity increases with increased substrate equation. When the enzyme is connected to its substrate, it does not matter how much substrate is present. Instead, the speed of the reaction depends on how the enzyme is connected to the substrate and thus on the connector molecule. Until now, we have not had any description of how the structure of such molecules affected enzymatic reactions."Normally, the question you are trying to answer is what graph shape describes the enzyme activity. We had a much more fundamental problem," says first-author Mateusz Dyla. "What should we put on the X-axis instead of concentration?"The authors made a model system where they could change the connection between the enzyme and the substrate. They used this to measure how the length of a flexible connector affects the first round of catalysis by the enzyme, which took place in milliseconds. Finally, they ended up with an equation that describes how the speed of the enzyme depends on the connection between enzyme and substrate. This equation suggested that connector molecules play an overlooked role in controlling cellular signalling.The connection between enzyme and substrate also affects which substrates the enzyme prefers. Substrates that look similar can be very different when the enzyme only processes a single connected substrate."It is like the difference between how long it takes me to eat a single hotdog, and how many hotdogs I can eat over a whole week," Magnus explains. "Over the course of a week, I will be limited by how fast I can digest the hotdogs. This is irrelevant to the time it takes to eat the first hotdog. Therefore, the two types of measurements give different results. If you want to understand kinase switches, you have to focus on the first round of catalysis."In the long-term, this may have implications for the development of drugs targeting kinases in, for example, cancer. Mateusz explains: "We hope that one day it will be possible to make drugs that not only target the enzyme, but also target how it is connected to its substrate."
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Biotechnology
| 2,020 |
August 19, 2020
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https://www.sciencedaily.com/releases/2020/08/200819110925.htm
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Understanding the inner workings of the human heart
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Researchers have investigated the function of a complex mesh of muscle fibers that line the inner surface of the heart. The study, published in the journal
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In humans, the heart is the first functional organ to develop and starts beating spontaneously only four weeks after conception. Early in development, the heart grows an intricate network of muscle fibers -- called trabeculae -- that form geometric patterns on the heart's inner surface. These are thought to help oxygenate the developing heart, but their function in adults has remained an unsolved puzzle since the 16th century."Our work significantly advanced our understanding of the importance of myocardial trabeculae," explains Hannah Meyer, a Cold Spring Harbor Laboratory Fellow. "Perhaps even more importantly, we also showed the value of a truly multidisciplinary team of researchers. Only the combination of genetics, clinical research, and bioengineering led us to discover the unexpected role of myocardial trabeculae in the function of the adult heart."To understand the roles and development of trabeculae, an international team of researchers used artificial intelligence to analyse 25,000 magnetic resonance imaging (MRI) scans of the heart, along with associated heart morphology and genetic data. The study reveals how trabeculae work and develop, and how their shape can influence heart disease. UK Biobank has made the study data openly available.Leonardo da Vinci was the first to sketch trabeculae and their snowflake-like fractal patterns in the 16th century. He speculated that they warm the blood as it flows through the heart, but their true importance has not been recognized until now."Our findings answer very old questions in basic human biology. As large-scale genetic analyses and artificial intelligence progress, we're rebooting our understanding of physiology to an unprecedented scale," says Ewan Birney, deputy director general of EMBL.The research suggests that the rough surface of the heart ventricles allows blood to flow more efficiently during each heartbeat, just like the dimples on a golf ball reduce air resistance and help the ball travel further.The study also highlights six regions in human DNA that affect how the fractal patterns in these muscle fibers develop. Intriguingly, the researchers found that two of these regions also regulate branching of nerve cells, suggesting a similar mechanism may be at work in the developing brain.The researchers discovered that the shape of trabeculae affects the performance of the heart, suggesting a potential link to heart disease. To confirm this, they analyzed genetic data from 50,000 patients and found that different fractal patterns in these muscle fibers affected the risk of developing heart failure. Nearly five million Americans suffer from congestive heart failure.Further research on trabeculae may help scientists better understand how common heart diseases develop and explore new approaches to treatment."Leonardo da Vinci sketched these intricate muscles inside the heart 500 years ago, and it's only now that we're beginning to understand how important they are to human health. This work offers an exciting new direction for research into heart failure," says Declan O'Regan, clinical scientist and consultant radiologist at the MRC London Institute of Medical Sciences. This project included collaborators at Cold Spring Harbor Laboratory, EMBL's European Bioinformatics Institute (EMBL-EBI), the MRC London Institute of Medical Sciences, Heidelberg University, and the Politecnico di Milano.
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Biotechnology
| 2,020 |
August 18, 2020
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https://www.sciencedaily.com/releases/2020/08/200818175417.htm
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Cryo-EM study yields new clues to chicken pox infection
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Despite decades of study, exactly how herpesviruses invade our cells remains something of a mystery. Now researchers studying one herpesvirus, the varicella zoster virus (VZV) that causes chicken pox, may have found an important clue: A key protein the virus uses to initiate infection does not operate as previously thought, researchers at Stanford University and the Department of Energy's SLAC National Accelerator Laboratory report August 18 in
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The results were made possible by high-resolution cryo-electron microscopy (cryo-EM), which showed that the immune system can prevent infection by attacking a spot on the protein in an unexpected place, said Stefan Oliver, a senior research scientist in pediatrics at Stanford and the new study's first author.Herpesviruses including VZV -- along with HIV, coronaviruses, and a number of other virus families -- are enclosed in a protective membrane, and the first step in the process of invading a cell is for the viral envelope to fuse with the cell's membrane. In VZV's case, a protein called gB that sits on the outside of the viral envelope uses a set of molecular fingers to grab onto and fuse with cells.But it turns out that's only part of the story. To investigate what was happening in more detail, Oliver and colleagues used an antibody from a patient that prevented VZV fusion with cells in cryo-EM experiments to discover where the antibody attacks gB.To Oliver and colleagues' surprise, the antibody bound to a spot on gB far from the fusion fingers, indicating that it may not need to target the fingers to prevent fusion with a cell. This result suggests that there may be more involved in the process of fusion, which causes infection, than was realized.Figuring out exactly how the fusion process works will take further studies that could inform the design of treatments and vaccines for other herpesviruses, Oliver said, since they also rely on gB to infect cells. "Vaccines are currently not available for herpesviruses, with the exception of the one that prevents VZV, so the development of vaccines that target this newly identified region of gB has the potential to solve an important medical need."Oliver added, "It was only possible to uncover this mechanism by generating one of the highest resolution structures of a viral protein-antibody pair using cryo-EM. Without the cryo-EM capabilities at SLAC these fascinating insights into the molecular mechanisms of fusion function would not have been achievable."
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Biotechnology
| 2,020 |
August 18, 2020
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https://www.sciencedaily.com/releases/2020/08/200818142109.htm
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Heart attack damage reduced by shielded stem cells
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Bioengineers and surgeons from Rice University and Baylor College of Medicine (BCM) have shown that shielding stem cells with a novel biomaterial improves the cells' ability to heal heart injuries caused by heart attacks.
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In a study using rodents, a team led by Rice's Omid Veiseh and Baylor's Ravi Ghanta showed it could make capsules of wound-healing mesenchymal stem cells (MSCs) and implant them next to wounded hearts using minimally invasive techniques. Within four weeks, heart healing was 2.5 times greater in animals treated with shielded stem cells than those treated with nonshielded stem cells, the researchers found.The study is available online in the Royal Society of Chemistry journal Someone has a heart attack every 40 seconds in the United States. In each case, an artery that supplies blood to the heart becomes blocked and heart muscle tissue dies due to lack of blood. Hearts damaged by heart attacks pump less efficiently, and scar tissue from heart attack wounds can further reduce heart function."What we're trying to do is produce enough wound-healing chemicals called reparative factors at these sites so that damaged tissue is repaired and restored, as healthy tissue, and dead tissue scars don't form," said Veiseh, an assistant professor of bioengineering and CPRIT Scholar in Cancer Research at Rice.Ghanta, associate professor of surgery at Baylor, a cardiothoracic surgeon at Harris Health's Ben Taub Hospital and co-lead author of the study, said prior studies have shown that MSCs, a type of adult stem cell produced in blood marrow, can promote tissue repair after a heart attack. But in clinical trials of MSCs, "cell viability has been a consistent challenge," Ghanta said."Many of the cells die after transplantation," he said. "Initially, researchers had hoped that stem cells would become heart cells, but that has not appeared to be the case. Rather, the cells release healing factors that enable repair and reduce the extent of the injury. By utilizing this shielded therapy approach, we aimed to improve this benefit by keeping them alive longer and in greater numbers."A few MSC lines have been approved for human use, but Veiseh said transplant rejection has contributed to their lack of viability in trials."They're allogenic, meaning that they're not from the same recipient," he said. "The immune system perceives them as foreign. And so very rapidly, the immune system starts chewing at them and clearing them out."Veiseh has spent years developing encapsulation technologies that are specifically designed not to activate the body's immune system. He co-founded Sigilon Therapeutics, a Cambridge, Massachusetts-based biotech company that is developing encapsulated cell therapeutics for chronic diseases. Trials of Sigilon's treatment for hemophilia A are expected to enter the clinic later this year."The immune system doesn't recognize our hydrogels as foreign, and doesn't initiate a reaction against the hydrogel," Veiseh said. "So we can load MSCs within these hydrogels, and the MSCs live well in the hydrogels. They also secrete the same reparative factors that they normally do, and because the hydrogels are porous, the wound-healing factors just diffuse out."In previous studies, Veiseh and colleagues have shown that similar capsules can keep insulin-producing islet cells alive and thriving in rodents for more than six months. In the heart study, study co-lead author Samira Aghlara-Fotovat, a Rice bioengineering graduate student in Veiseh's lab, created 1.5-millimeter capsules that each contained about 30,000 MSCs. Several of the capsules were placed alongside wounded sections of heart muscle in animals that had experienced a heart attack. The study compared rates of heart healing in animals treated with shielded and unshielded stem cells, as well as an untreated control group."We can deliver the capsules through a catheter port system, and that's how we imagine they would be administered in a human patient," Veiseh said. "You could insert a catheter to the area outside of the heart and inject through the catheter using minimally invasive, image-guided techniques."Veiseh said capsules in the study were held in place by the pericardium, a membrane that sheaths the heart. Tests at two weeks showed that MSCs were alive and thriving inside the implanted spheres.More than 800,000 Americans have hearts attacks each year, and Ghanta is hopeful that encapsulated MSCs can one day be used to treat some of them."With further development, this combination of biomaterials and stem cells could be useful in delivering reparative therapy to heart attack patients," he said.Veiseh said the pathway to regulatory approval could be streamlined as well."Clinical grade, allogenic MSCs are commercially available and are actively being used in patients for a range of applications," he said.Veiseh credited Aghlara-Fotovat with doing much of the work on the project."She basically executed the vision," he said. "She developed the hydrogel formulation, the concept of how to package the MSCs within the hydrogel, and she did all the in vitro validation work to show that MSCs remained viable in the capsules."Aghlara-Fotovat is co-mentored by Ghanta and worked in his lab at Baylor alongside research assistant Aarthi Pugazenthi, including assisting in rodent surgeries and experiments."What attracted me to the project was the unmet clinical need in (heart attack) recovery," Aghlara-Fotovat said. "Using hydrogels to deliver therapeutics was an exciting approach that aimed to overcome many challenges in the field of drug delivery. I also saw a clear path to translation into the clinic, which is the ultimate goal of my Ph.D.""I think one of the things that attracts students to my lab in particular is the opportunity to do translational work," Veiseh said. "We work closely with physicians like Dr. Ghanta to address relevant problems to human health."
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Biotechnology
| 2,020 |
August 18, 2020
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https://www.sciencedaily.com/releases/2020/08/200818103818.htm
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RNA as a future cure for hereditary diseases
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Short RNA molecules can be used as medication. Their effectiveness is based on the genetic information they carry: therapeutic RNA can bind to the body's own RNA and thus influence how it functions. However, only a handful of such drugs are available so far.
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"That's mainly because it's tricky to get the RNA molecules to precisely the organ in the body where they need to take effect. Currently, that's the biggest hurdle in the development of RNA drugs," says Jonathan Hall, Professor of Pharmaceutical Chemistry at ETH Zurich. Together with Daniel Schümperli, Emeritus Professor from the University of Berne, and colleagues from ETH, University Hospital Zurich and Triemli Hospital Zurich, he has now succeeded in developing an RNA molecule that can compensate for the effect of gene mutations in bone marrow cells.This therapeutic approach could one day be applied to a rare hereditary disease called erythropoietic protoporphyria (EPP), which affects people whose mother and father both have a genetic predisposition to the disease. Those who suffer from EPP experience a painful sensitivity to sunlight.Gene mutations cause the body of these patients to produce less of a certain enzyme, ferrochelatase. Ferrochelatase is central to the production of haemoglobin, the protein that transports oxygen in the blood and makes it appear red. This ferrochelatase deficiency causes a metabolic molecule, protoporphyrin, to accumulate in the red blood cells. Protoporphyrin reacts to rays of visible light, forming molecules that attack tissue and can cause painful inflammation when the patient is exposed to sunlight or a strong artificial light.Hall and his colleagues developed several short RNA molecules, which bind to the RNA copy of the ferrochelatase gene in the body's cells. In cell culture experiments, they identified certain molecules that were able to restore a sufficient production of the enzyme and thus compensate for the negative effects of the known EPP gene mutations.However, developing the RNA molecule was only the first part of the task. "This molecule must also be able to reach the right organ in the body and from there penetrate the interior of the cells," Hall says. In the case of EPP, these are the blood stem cells in the bone marrow. To this end, the researchers fused one of the RNA molecules with various chemically active compounds, which they tested in a mouse model of EPP. They identified one fusion molecule -- the RNA molecule fused with cholesterol -- that was able to compensate for the gene mutation in this animal model.Hall stresses that it is too early to label the molecule he has identified as an RNA drug. In demonstrating that such molecules can be used to increase the amount of functional ferrochelatase in mice, the researchers are at only an early stage of their work. "This is the first step and it shows that our approach holds promise," Hall says. Next, the researchers need to optimise the fusion molecule or identify other fusion molecules that are even more effective, he explains, adding that they also require additional, more refined mouse models for the EPP disease. Further research is essential to find an optimum drug candidate whose effect can then be investigated in humans.
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Biotechnology
| 2,020 |
August 18, 2020
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https://www.sciencedaily.com/releases/2020/08/200818103807.htm
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Researchers discover novel molecular mechanism that enables conifers to adapt to winter
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In boreal forest during late winter, freezing temperatures are typical but at the same time the sun can already shine very brightly. This combination is especially dangerous to evergreen plants, such as conifers. The chlorophyll pigment-proteins in their needles absorb light, but the enzyme activity, stopped by the cold, prevents the plants from using the light for photosynthesis. This exposes the cells to damage.
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Dissipating the excess light energy as heat, the so-called non-photochemical quenching, is a common, fast, and dynamic but intermittent regulation mechanism in all plants and algae, and it is employed to protect the plant from damage caused by high light intensity. However, the combination of freezing temperatures and high light intensity results in a particular form of quenching in conifers: sustained non-photochemical quenching.Researchers from the University of Turku, Finland, discovered an essential part of the mechanism associated to sustained non-photochemical quenching in conifers. The discovery is significant as the mechanism in question is still poorly understood in science."We collected needle samples from nature for four years and studied spruce branches in simulated conditions mimicking late winter. On the basis of biophysical and molecular biology analyses, we could show that the triply phosphorylated LHCB1 isoform and phospho-PSBS protein in chloroplast appear to be prerequisites for the development of sustained non-photochemical quenching that safely dissipates absorbed light energy as heat," say Doctoral Candidate Steffen Grebe and Postdoctoral Researcher Andrea Trotta from the Molecular Plant Biology unit of the Department of Biochemistry at the University of Turku.In the phosphorylation of a protein, a phosphoryl group is added to certain amino acids, which is a common mechanism for protein regulation in cells. The phosphorylation of the proteins discovered in spruce has not been described in science before.The researchers believe that together with the limited photoinhibition of photosystem II, the phosphorylations lead to structural changes in pigment-proteins so that the needles can effectively dissipate the excess light energy.The regulation mechanisms of photosynthesis have been previously studied on a molecular level mainly on fast-growing species regularly used in plant biology, such as thale cress (Arabidopsis thaliana) and the alga Chlamydomonas reinhardtii. However, it is not possible to study the winter acclimatisation with these plants and easily transfer the knowledge to conifer species. The molecular biology research of conifers became possible after the spruce genome sequencing was published in 2013."The spruce genome is approximately ten times larger than that of humans. The genome sequencing of spruce led by our long-time partner, Professor Stefan Jansson from the Umeå University, enabled the molecular photosynthesis study we have now conducted in Turku, says Principal Investigator," Academician Eva-Mari Aro.The new information on spruces' adaptation to their environment can be used in assessing the impact of climate change on photosynthesis of conifers and their carbon sink capacity as photosynthesis in conifer forests is one of the most important carbon sinks on a global scale.
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Biotechnology
| 2,020 |
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