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(B) 35S labeling of de novo mitochondrial protein synthesis of HeLa cells transfected as in (A). Polypeptide assignments flank the gel images. Coomassie stained gels are used as loading controls and immunoblots indicate the efficiency of LETM1 knockdown | [
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Gli1 protein levels were measured by Western blot in untreated (UT) and 100 nM SAG-treated WT and Mecp2 null MEF cells (n=5/6). Representative bands of Gli1, Mecp2 and α-tubulin are depicted above the histogram, which shows the mean ± S.E of the percentage of Gli1 expression levels, compared to the untreated WT samples (**p<0.01, ***p<0.001, by two-way ANOVA, followed by Bonferroni post hoc test). Two-way ANOVA reported a significant interaction (F (1, 18) = 6.652; p<0.05), a significant genotype effect (F (1,18) = 15.95; p<0.001) and a significant treatment (F (1, 18) = 20.15; p<0.001). | [
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E. Six evenly-sized densities are arranged in one Elongator lobe. A projection of the Elp456 hexamer X-ray structure (PDB code: 4A8J) is shown for comparison. | [
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(A) At 48 hours post transfection of the indicated protease plasmids in triplicate, BHK21 cells were incubated with a fluorogenic substrate and wtSPINK6 or loss-of-function mutSPINK6 or PBS for 2 hours and then applied to fluorescence assay. Data represent mean and SD of triplicated wells in a representative experiment performed 3 times. **, P<0.01. Student's t test. | [
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(F-I) In vitro proteasome activity assays were performed as described in Materials and Methods. Bars represent means +S.D. from three independent experiments. For statistical analysis, student t-test (two-sided, one type) were used. * P value: (F), 0.0039. | [
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Volcano plot of proteomic analysis of cell culture supernatant from 55d old CTRL and ECE2 KO COs shows two significantly downregulated proteins (red) in the secretome (n=3 batches of CTRL and ECE2 KO COs; FDR 0.05; s0 1; dashed lines indicating the cut-off). Non-significantly changed proteins of interest are highlighted in green below the cut-off curves. | [
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D,E HeLa cells expressing the indicated constructs. Flag-ITSN-s was visualized with anti-Flag and AlexaFluor561 or AlexaFluor488 in D and E respectively. Scale bar: 5 µm. Boxed regions are magnified on the bottom. White arrows and red arrows point to colocalizing and non-colocalizing structures, respectively. | [
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F. HeLa cells transfected with indicated siRNAs were pulse-labeled or not with 5-azadC for 30 min in late S phase according to the experimental setup in (A). Following 5-azadC withdrawal cells were incubated with nocodazole, collected at the indicated times and immunoblotted with indicated antibodies. | [
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(f,g) Immunofluorescence for endogenous Cx43 (green) in control NRK cells or those knocked down for Atg14 (f) maintained in the presence or absence of serum. | [
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(B) Thousands of genes were differentially expressed in Dex-induced GR-TFs compared to GR-TFs under mock conditions. | [
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ICC analyses of WT- or MT-HCT116 cells treated with 20 µM of KYA1797K for 24 hr. Scale bars, 10 µm | [
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(L) Transmission electron microscopy of hNSCs infected by ZIKV MR766 and Paraiba strains. Mitochondria (MT), endoplasmic reticulum (ER), Golgi bodies (GB), ribosomes (RS), phagophores (PH), lipid droplet (LD). Scale bars, first row, 1 μm (left), 500 nm (control right); 200 nm (Right). | [
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(A) The positions of the β6-L10-β7 and neck-coil peptides are shown in brown and green. The positions of the neck-linker peripheral peptides (α1-β3-P loop, α4-L12 and L12-α5) are shown in light blue. (B-D) Quantitation of site-specific intramolecular interactions. Representative MS/MS spectra of the indicated peptides, including the mutated amino acid (β6-L10-β7) (B and C) and neck-coil (D). In the inserted graphs, the intensities in the ADP-BeFx and ADP-AlFx states were measured with respect to the nucleotide-free state as 1.0. Grey bar graphs, WT; black bar graphs, E239K; (-), nucleotide-free. | [
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Molecular structure of the β-IgI3 & CC1 N-terminal domain co-complex. Residues that form the β-IgI3-CC1 interface are highlighted. | [
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(A) The Venn diagram depicts the identification of 2511 QKI-targeted genes overlapping between three RBP-binding sites databases (POSTAR, EuRBPDB and doRiNA). | [
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(C) UbcH5 mediated polyubiquitination of MITA by RNF26. The RNF26 and MITA proteins were obtained by in vitro transcription and translation, then incubated with biotin-Ub, E1 and the indicated E2s. Polyubiquitination of MITA was examined by immunoblot analysis with HRP-streptavidin (top panel). The inputs of RNF26 and MITA were analyzed by immunoblots with anti-MITA and anti-RNF26 (bottom panels). | [
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Fluorescence polarisation profiles from incorporation of FITC-M451‑90 into excess unlabeled M451‑90 (green), RIPK1497‑583 (red), RIPK3387‑518 (blue) or ZBP1170-355 (purple). Average polarisation profile from smoothed triplicate samples plotted, with s.d. indicated. Experiment performed twice and representative results shown here | [
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A Schematic representation showing where the compounds act along the AC/PKA signalling pathway. | [
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Top panel: Y-axis shows the CAGE signal intensity (tag-per-million normalized number of mapped CAGE tags on plus strand). X axis shows LTR and 5' of Tf2 ORF. Three biological replicates are shown for each strain: fft2Δfft3 and WT. Bottom: LTR and tf2 ORF are colored: U3 (green), R (red), U5 (blue), self primer and primer binding sequence (PBS; gray), beginning of ORF (gray striped). TSS of all Tf2 transcripts from (Rhind et al, 2011) are shown as gray dots above the sequence elements. The black square indicates the 5' end of the Tf2 mRNA in WT and the green square indicates the 5' end in fft2Δ fft3Δ detected by 5'RACE. | [
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Immunoblot analyses of TDG and TDG-SUMO in whole cell extracts of mESCs Chromatin fractionation from mESCs expressing wildtype (ESC-TDGwt) or SUMOylation-deficient TDG (ESC-TDGsnm). Sol, chromatin unbound proteins; Chr1, chromatin associated proteins; Chr2, chromatin bound fraction; Histone H2B, stably chromatin bound marker. | [
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Control and si-IRGM transfected HT-29 cells uninfected or infected with CHIKV and qRT-PCR analysis were performed with (F) HERC5 | [
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Male C57BL/6J mice were injected with LV-shScrambled or shSUNCR1 lentivirus specifically into the gastrocnemius at 6 weeks of age. After two weeks of recovery, mice were fed with chow diet supplemented with 0 or 1% SUC for 6 weeks. Immunoblots and quantification of MyHC-I, IIb, NFAT, and PGC-1α protein in gastrocnemius (n=3). | [
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C) The slope of the linear regression of NF135 and NF175 on day 3 (circle) and day 5 (square) p.i.. Each dot represents one biological replicate. | [
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(F) HepG2 cells were transfected with vector, JFK, or/and ING5, or treated with control siRNAs or siRNAs against JFK, AMPKα1, or/and ING5. Culture media was collected and the concentration of 3-hydroxybutyric acid and acetoacetic acid was measured by ketone body assay kit. Error bars represent mean ± SD for triplicate experiments (*p < 0.05, paired two-tailed Student's t-test). | [
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F. In vitro ubiquitylation experiment as in B using E.coli-derived MCAK, Kif2a or Kif2b as substrates and Cdc34 as E2. | [
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C: Relative mRNA expression of senescence markers (p16 and p21) and SASP molecules (PAI-1, IL6, IL8) in GEnC at early (p9) or late (p17) passage. n = 5 independent experiments. Data information: Data are means ± SEM. Statistical analysis: t-student test | [
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G Representative western blot analysis of phosphorylated p53, and Parkin in Parkin -/- mice compared to WT. Quantification analysis of phosphorylated p53 and Parkin (Parkin; **p=0.0001 vs. WT, pp53; *p=0.019 vs. WT, n=4). | [
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cTAZ repressed 5×ISRE-luciferase activity induced by IFN-α. HEK293A cells were transfected with 5×ISRE-luciferase reporter along with the indicated plasmids, treated with or without IFN-α (50 ng/ml) for 12 hours. The expression of ectopic genes was determined by IB. Error bars indicate SD, n =3. ***P<0.001; one-way ANOVA test was used for statistical analysis. | [
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, E Serum-induced killing (CFU/ml) of different Gram-negative strains depends on MAC components C5 and C8, but not on lysozyme (10% serum). Dotted line represents the detection limit of the assay. Data information: (D, E) The cfu/ml (Fig. 1E, 2C) and sytox measurements (Fig. 1D, 2E) of "Buffer", "Serum", "ΔC5", "ΔC8" "Δlysozym" and "C5b6MAC" were all generated from the same experiment. (C-E) Data represent mean ± SD of 3 independent experiments. (D, E) Statistical analysis was done using a ratio paired two-tailed t-test and displayed only when significant as * p≤0.05, ** p≤0.01, *** p≤0.001 or **** p≤0.0001. | [
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D Relative superoxide levels were determined by DHE staining assay and compared (n=4) between NE4C cells with and without MARS knockdown by shRNA. Cells were cultured in 20 μM Hcy-containing media. MARS2 knockdown efficiencies were confirmed by western blot. | [
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(J) Immunoblot showing the levels of the MAVS protein in siCtrl- or siNDP52-transfected cells infected with VSV (MOI = 1, 12 hrs). α-Tubulin was used as a loading control. Cell-based studies were performed independently at least three times with comparable results. | [
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(F) Comparative expression by qPCR of Dux, two downstream targets of DUX (Zscan4 and Tdpoz4), and two DUX-independent targets of DPPA2 and DPPA4 (Mael and Tdrd1) in mESCs depleted of endogenous DPPA2 and DPPA4 and overexpressing ectopically DUX or GFP as a control. Expression was normalized to Actb. Bars represent the average, error bars the SD; n = 3 | [
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(G) Venn diagram showing the overlap of the genes selected by RIP-ChIP (1,334) and TargetScan7.0 (1,224) (http://www.targetscan.org/vert_70/). | [
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(B) The protein level of ATG-5 was detected by western blot after treatment with siRNAs. β -actin was used as internal controls. | [
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A Western blot analysis of various structural subunits of complex I in fibroblasts from Subjects 1 and 3 (S1 and S3 respectively) with three age-matched controls (C1, C2, and C3). | [
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(e) Quantification of fluorescence signal in AGO2 immunoprecipitates from HeLa cell extracts incubated with duplexes of let-7a-let-7a* labelled on the 3′ end of either let-7 or let-7a* with FITC. Cells were treated 16 h before with DMSO (control), BAF (200 nm) or geldanamycin (10 μM). Representative results of one experiment are shown. | [
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Representative images of E11.5 lung explants after treatment for 48 h with 100 μM Scra or mo142-3p reveals reduced size and branching morphogenesis after miR142-3p knockdown. | [
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Quantitation of resection activity in NBS1∆N cells complemented with wild type (WT) or NBS1 R28A K160M mutant and either mock depleted (siCtrl) or depleted of CtIP (siCtIP). At least 300 cells were assessed per condition. The horizontal line represents a median, the box spans from 25th to 75th percentile and the whiskers from 10th to 90th percentile. Statistical significance was determined using ordinary one-way ANOVA with Tukey's multiple comparison test, **** (p<0.0001). | [
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B Bioenergetic profiles measured with Seahorse flux analyzer, showing the contribution of OCR and ECAR to cellular bioenergetics in RKO and IMT1-resistant RKO cells treated with and without IMT1. Mean of n=3 independent experiments ± SEM. | [
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D Correlations of somatic insertion sites of the three most represented TE families (rover, copia and I-element) with modENCODE tracks for adult fly tissues. | [
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In the probe trial, AAV-Venus injected cDKOs (red) spend significantly less time in the target zone compared to AAV-Venus (###p<0.001) injected LM control animals (white) and failed to prefer it significantly over control zones in adjacent quadrants. Note that expression of APPsα (green) largely rescues the deficits of cDKOs. N=22 (LM controls: AAV-Venus), N=18 (cDKOs: AAV-Venus), N=18 (cDKOs: AAV-APPsα). Data represent mean ± SEM. Dashed lines: chance level. Data were analyzed using a mixed ANOVA model with conditions (LM control: AAV-Venus, cDKO: AAV-Venus and cDKO: AAV-APPsα) as between subject factor. Within subject factors were added to explore the dependence of genotype effects on day and place. Significant interactions and main effects were further explored by pairwise FDR-corrected two-tailed Student's t-tests. # indicates significant differences between LM control and cDKO injected with AVV-Venus, * between cDKO injected with AAV-Venus or APPsα, ● between LM control injected with Venus and cDKO injected with AAV-APPsα nsp>0.05, */#/●p<0.05, **/##/●●p<0.01, ***/###/●●●p<0.001. | [
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Susceptibility weighted imaging (SWI) of an ALSP patient with the ΔA781_N783 CSF-1R variant showing iron deposition. | [
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ATP measurements in: (H) age-1(hx546); gas-1(fc21) (a; g) and age-1(hx546); aak-2(ok524); gas-1(fc21) (a; aa; g) mutant nematodes; | [
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(F,G) BATs from mice exposed to normal conditions (30℃) or cold (4℃) were lysed and immunoprecipitated with antibodies against FUNDC1, LC3B or IgG, then immunoblotted with the indicated antibodies. The ratio between immunoprecipitated LC3B to FUNDC1 or FUNDC1 to LC3B were quantified and plotted (G). | [
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(g) U20S cells stably expressing HaloTag-p62 were labelled with HaloTag ligand for 15 min and washed out followed by treatment with purmorphamine. After 48 h, cells were lysed, run on a SDS-polyacrylamide gel electrophoresis and fluorescent HaloTag was visualized using a Typhoon 8600 variable mode imager. Fluorescence was normalized to total protein levels, detected on the same gels by Kryton Fluorescence protein stain. A representative gel and its quantitated normalized levels are shown. In all panels, graphs show mean values and error bars represent s.d. from a triplicate experiment representative of at least three independent experiments. Statistical analyses were performed by two-tail Student's t-test unless indicated: ***P0.001; **P0.01; *P0.05. SDS-PAGE, SDS-polyacrylamide gel electrophoresis. | [
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C57BL/6 mice were immunized with 200 µg MOG35-55 emulsified in CFA and 500 ng of purified Bordetella pertussis toxin. When EAE symptoms occurred, the mice were injected i.p. with 25 µg hTAPBPL-Ig or control Ig protein 3 times per week. (A) Mean clinical scores. Deletion of Tregs partly reverses the effect of TAPBPL on EAE. Groups of EAE mice as in (A) were injected i.p. with anti-CD25 antibody. Mean clinical scores are shown. | [
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Engraftment ratio of transplanted cells in mouse lungs on indicated days. n=3. Error bars, S.E.M. *P=0.0474. | [
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High-resolution CFP FLIM of HEK sensor cells expressing CFP/YFP-TauRDP301L and treated with Tau condensates. FRET between CFP and YFP induces CFP-lifetime reduction in densely packed CFP/YFP-TauRDP301L accumulations. Fit-free analysis of the lifetime distribution in phasor plots enables the assignment of distinct lifetime clusters (circles in phasor plot) to different Tau accumulation types. LT-2=NE Tau, LT-3=CYT and NUC Tau, LT-4=fully aggregated CYT Tau. LT-1 corresponds to soluble CFP-TauRD in the cytosol of cells without accumulations. Asterisks indicate position of individual cell nuclei. Scale bars=10 μm. | [
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(c) Rotarod performance; n = 11 for all three groups (*, P = 0.0055; **, P = 0.002; ***, P = 0.0455; ****, P = 0.0242). | [
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Primary astrocytes from OTUB1fl/fl and GFAP-Cre OTUB1fl/fl mice were treated with IFN-γ (10 ng/ml) for indicated times. Cytoplasmic and nuclear extracts were separated and analyzed by WB with indicated antibodies. | [
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GRdim/dim mice (n=4) were treated with PBS or 1.5 mg/kg LPS with or without 10 mg/kg DEX pretreatment. Ileum was isolated and gene expression was measured via qPCR. Fold changes are depicted on the graphs. | [
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(A) SCPO, STIPO and STAPO for the 51 genes validated by two out of four individual siRNA duplexes reproducing the GFP-LC3 spot phenotype, above a cutoff of greater or less than 20% of control, and passing additional criteria (Supplementary Table 3). Red boxes indicate an increase >1.2; pink boxes an increase between 1.1 and 1.2; white boxes are between 0.9 and 1.1; light blue boxes a decrease between 0.9 and 0.8; dark blue boxes a decrease to 0.8. Hits underlined were taken forward as three out of four4 hits, (§) not chosen because of restricted tissue expression, (*) are three out of four when alternate method of normalisation is used. | [
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J Deformation of the mandible walls under a series of stress levels with different Young's moduli. Data were obtained from multiple tests as in I. Gray horizontal lines indicate upper and lower limit values of mean mandible wall displacement (79.74 and 36.48 µm, respectively); dashed colored lines indicate results of the free outer surface; solid colored lines indicate results of the fixed outer surface (with consideration of good linearity of the simulation results, the corresponding result points of the simulation series were omitted and replaced by solid or dashed colored lines for clarity); the actual stress value should be between the two extreme boundary conditions. Results show that the probable stress level ranged from 3-20 kPa. n = 3. | [
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(G) Bar graph illustrating the mitotic index in cells treated with low concentrations of nocodazole for 12 h | [
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(A) NSCLC expressing single site mutants of EGFR, PC9 (3X106) or H3255 (8X106), were seeded in 10-cm dishes. On the next day, complete media were replaced with media containing serum (1%) and the cells were treated for 24 hours with different EGFR-specific TKIs (erlotinib, 50 nM; osimertinib, 50 nM, or afatinib, 10 nM), either alone or in combination with 2XmAbs (cetuximab and trastuzumab, 5 μg/ml each). Thereafter, cells were washed with cold saline and extracted. Proteins were separated using gel electrophoresis and transferred onto nitrocellulose membranes. After blocking, membranes were incubated overnight with the indicated primary antibodies, followed by incubation with horseradish peroxidase-conjugated secondary antibodies (60 minutes), and treatment with Clarity™ Western ECL Blotting Substrates (Bio-Rad). ECL signals were detected using the ChemiDoc™ Imaging System (Bio-Rad) and images were acquired using the ImageLab software. Signals (relative to Control) were quantified and normalized to the signals of GAPDH (numbers shown below each lane). | [
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D. Fluctuation analysis for Bu-1 loss in two independent timeless/ddx11 double-mutant clones (#1 and #2), ddx11 expressing DDX11KAK and fancj and fancj/ddx11 double-mutants. Fluctuation analyses for wild type, timeless #1 and ddx11 #1 are shown for comparison. | [
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(b) We transfected MEFs stably expressing mCherry-EGFP-LC3 with BFP-tagged AR25Q or AR125Q. Cells were imaged after 24 h, and yellow puncta (autophagosomes) and red puncta (autolysosomes) in AR-expressing (blue) cells were counted. Autophagosomes: n = 3 independent experiments, F = 20.65; autolysosomes: n = 3 independent experiments, F = 4.49. One-way ANOVA with post hoc Tukey test. *P 0.05, **P 0.01, ***P 0.001. n = 33 cells per genotype. | [
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The gross appearance of BAT, eWAT and ingWAT fat pads after administration of AAV-ShQKI or AAV-NC vectors in local adipose (ingWAT and BAT). | [
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A) Kaplan-Meier survival analysis (Log-Rank test with Holm-Sidak's correction for multiple comparisons). Mean age of death (± SEM) for G93A was 169.3 ± 2.5 days and for PKO/G93A was 185.6 ± 2.2 days, indicating that Parkin knockout prolongs lifespan of SOD1-G93A mice; n=19 for Non Tg (9 males and 10 females), n= 20 for PKO (10 males and 10 females), n= 18 for G93A (8 males and 10 females) and n= 19 for PKO/G93A (9 males and 10 females); p= 0.0000285. | [
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(E) Quantification of the mean (±SEM) number of DVGlut+ boutons per muscle 4 NMJ in each genotype. [WT n = 23, hiw n = 21 , hiw > Wnd-RNAi n= 16, hiw > dNmnat-RNAi n=19. 1-way ANOVA w/Tukey's multiple comparisons, DF=78, F=169.1, p < 0.0001. WT vs hiw p < 0.0001 (****), WT vs hiw > Wnd-RNAi p = 0.9988 (NS), WT vs hiw > dNmnat-RNAi p < 0.0001 (****), hiw vs hiw > dNmnat-RNAi p = 0.9695 (NS)]. | [
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Analysis of Rrm3 recruitment in WT and rpb1‐1. Rrm3 IP histogram bars in the y‐axis show the average signal log2 ratio of loci enriched in the immunoprecipitated fraction along the indicated regions in the x‐axis. When the ratio fulfills the P‐value criteria (P 0.01) it is shown in red. | [
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(G) Western blotting of SKOV3-Empty "E" and OPCML "O" cells transfected with non-targeting siRNA (NTrg) or siRNA smartpool for PTPRG, stimulated with Gas6 over a 24 hour time course. GAPDH was used as loading control | [
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Arp2/3 complex subunits are colored as follows: Arp3, orange; Arp2, red; ARPC1, green; ARPC2, cyan; ARPC3 (not shown for clarity); ARPC4, blue; ARPC5, yellow. Actin monomers recruited by WASP are not shown. Purple arrows indicate contact between actin filaments or SPIN90 with ARPC4 that may stimulate a structural change required for activation. Thick grey arrow in left panels indicates potential rigid body rotation of Arp2, ARPC1, ARPC5 and the globular portion of ARPC4 hypothesized to move Arp2 into the short pitch arrangemen | [
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F Usp26-deficient mice produced XY aneuploid round spermatids. FISH analysis of Chr X (green) and Chr Y (red) was performed in 2-month-old and 6-month-old Usp26+/Y and Usp26−/Y round spermatids. Nuclei were stained with DAPI (blue). The arrows indicate the Y chromosome, and the arrowheads indicate the X chromosome. G Quantification of different types of round spermatids in 2-month-old Usp26 +/Y, Usp26-/Y mice (n= 3 independent experiments) and 6-month-old Usp26+/Y (n= 3 independent experiments), Usp26-/Y mice (n= 4 independent experiments). P=0.0026 for XY spermatozoa in 6-month-old Usp26+/Y and Usp26−/Y mice. P=0.0216 for O spermatozoa in 6-month-old Usp26+/Y and Usp26−/Y mice. H Usp26-deficient mice produced XY aneuploid spermatozoa. FISH assay of Chr X (green) and Chr Y (red) was performed in 2-month-old and 6-month-old Usp26+/Y and Usp26−/Y spermatozoa. Nuclei were stained with DAPI (blue). The arrows indicate the Y chromosome, and the arrowheads indicate the X chromosome. I Quantification of different types of spermatozoa in 2-month-old Usp26+/Y, Usp26-/Y mice (n= 3 independent experiments) and 6-month-old Usp26+/Y, Usp26-/Y mice (n= 5 independent ex experiments). P=0.0195 for XY spermatozoa in 6-month-old Usp26+/Y and Usp26−/Y mice. Data are presented as means ± SD. | [
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I-L) Muscle force was evaluated in ex vivo soleus muscles from old control and Mfn2KO mice. Maximal tetanic force (I), twitch to tetanus ratio (J), and half relaxation (K) and contraction times (L) were measured (n=4 soleus per genotype). | [
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mRNA expression analysis (RTqPCR) of p53 target genes in thymocytes 6 hours after 6 Gy irradiation. Shown are expression values normalized to β-actin. | [
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(D) Distance distribution between enhancer midpoints and promoter TSSs for E-P pairs obtained with the CN (light grey) and CW (dark grey) approach. The histogram shows the 500 kbp distance range in 20 kbp bins. | [
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Groups of five mice were intranasally infected with IAV (1918), and recombinant DDX3 protein was administered intranasally. Two days post-infection, virus titers in lung homogenates were determined by plaque assay. A p value of <0.05 was considered significant (** p < 0.01). | [
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(D) Antibody TB24PB037 binds to MTB-LAM but not to LAM from M. smegmatis. ELISA performed as described in (C). | [
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C) Flow cytometry analysis of surface DC-SIGN in human MoDCs after infection with Pg381, Mfa1+Pg and FimA+Pg. The analysis of the intensity used Kruskal-Wallis test analysis of different groups and Dunn's test for multiple comparisons 3 different experiments (* p<0.01). | [
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(M) Principal component analysis by combining all the values related to motor and cognitive tasks from wt mice (blue dots), R6/2 ACSF mice (green dots), and R6/2 chol-high mice (red dots). | [
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G, H Immunoblotting using an anti-P-Akt, anti-P-S6, anti-4EB-P1 or anti-GAPDH (loading control) antibody of protein extracts from CAFs treated with the indicated molecules (PDGF receptor inhibitor, 5-10 μM; Jak1/2 inhibitor ruxolitinib, 5 or 10 μM; EGFR inhibitor, 150 or 300 nM and recombinant PDGF, 5 μg/ml) (representative of n = 3). | [
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(A) Schematic overview of cloning, expression and purification of 143 carbohydrate recognition domain (CRD) - mouse IgG2a-Fc fusion proteins, from 168 annotated murine CRD containing proteins. The constructs were expressed in HEK293F cells and secreted Fc-fusion proteins were purified using protein A columns. | [
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OVCAR-3 cell confluency by IncuCyte® during 72h of treatment with APR-246 +/- MK-571 (n = 3). | [
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B. Visualization of the relative gene expression for characteristic genes along the pseudotime of the three trajectories. The genes visualized are color-coded to represent their respective pseudotemporal expression. | [
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(C) HeLa cells were transfected with siRNA control, and siRNA for p38IP. After incubation for 2 h as in (A), samples were immunoblotted with anti‐LC3 and anti‐actin antibodies. Endogenous LC3II/LC3I levels were quantified and the ratio presented as arbitrary units. In (B), n=4, ***P=0.0001 and *P=0.0324, Students t‐test. In (C), data are representative of two experiments. | [
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C The p97-EQ mutant stabilizes K48-conjugates. Lysosome damage was induced as in (A) in cells transfected with GFP-tagged p97-wt or p97-EQ. Cells were chased after washout for 12 h and stained for K48 chains.D Quantification of K48-positive lysosomes 2 h and 12 h after damage in cells treated as in (A). Data represent mean ± SD of three independent experiments. | [
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G The fragment encompassing the PB1 and ZZ domains of p62 is sufficient for mediating its interaction with MOAP-1. p62 KO LO2 cells were transfected with plasmid encoding Myc-p62 or the indicated fragments and Flag-MOAP-1. 24 hours later, the transfected cells were subjected to co-IP assay with anti-Myc antibody. | [
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Multi-Dimensional-Scaling (MDS) plot based on the gene expression profiles (microarrays) of 3 healthy FT and 8 tumor tissue samples and their respective organoid cultures shows 4 clusters: Normal FT tissue, normal FT organoids, cancer organoids and cancer tissue. | [
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(B) Averaged fluorescence intensity profiles of GFP-dynein (green squares) and Alexa647-BicD2-N (magenta circles) at growing microtubule plus ends. Mean values from three separate experiments per condition (with a total of at least 75 kymographs) are shown; error bars are s. e. | [
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(I) Flow-cytometric analysis of lymph node cells from a control Pax5+/+ mouse and a 10-week-old Pax5Jak2/+ tumor mouse. (J) Flow-cytometric analysis of B220lowCD19+ B cells from the bone marrow of a 4-week-old Pax5Jak2/+ mouse. | [
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Differentiated lung bud tip organoids at air-liquid interface in co-culture with donor-specific human fetal lung fibroblasts. After 14 days of differentiation at air-liquid interface, cells express LPCAT1, green, markers in areas containing one cell layer. Dotted lines indicate the barrier between multilayered and single layered epithelium. Data information: Nuclei are stained with Hoechst Scale bars indicate 50 μm. | [
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MDA-MB-231 cells were transfected twice with indicated siRNAs, the absorbance values of indicated cell lines was measured at different time points by CCK8 assay (n=3). Data from three independent experiments are expressed as mean ± SEM. Error bars indicate SEM. | [
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Serum pancreatic lipase titers of mice immunized with cInsulin (n=5) or control immunization (n=4) measured by ELISA. Dots represent individual mice. Mean ± SD, statistical significance was calculated by using Mann-Whitney-U test, **P < 0.01. | [
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Transcriptome analysis of gene expression regulated by auxin, UV-B, and UVR8. (A) Heat map of UV-B-, UVR8-, and auxin-regulated genes. The parameter measured by color key shows the Log-fold change. (B) Auxin signaling genes are up-regulated by auxin treatment but down-regulated by UV-B in a UVR8-dependent manner. Three biological replicates were analyzed and final Log fold-change is shown. | [
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Venn diagram comparing the number of epistatic pairs detected in various phenotypes. | [
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Coimmunoprecipitation and immunoblot analysis of LIF-treated HEK293T cells cotransfected with the indicated constructs. | [
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D. Heatmaps highlighting the gene expression changes observed with each treatment. Left: the top 50 ISGs, ranked by IFN-induced expression (B), were compared with respect to the impact that dBRD9-A pre-treatment had on induction. Right: comparative fold change in induction for each ISG in the presence or absence of dBRD9-A. | [
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A. Representative Coomassie stained SDS-PAGE gels of UTRN-ABD and UTRN-V87D co-sedimentation with F-actin. Pellet (top) and supernatant (bottom) fractions of individual co-sedimentation reactions of increasing utrophin concentrations, uncropped gel images of all the co-sedimentation reactions are presented in Supplement Figure 6. | [
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B. The various CTRL#3 or BRD9-KO#3 cells described in (A) were stimulated with 1000 IU/mL of IFN-α2 for 16h prior to infection with VSV-GFP at an MOI of 0.6 PFU/cell. Total Integrated Green Fluorescent Intensities were determined using the Incucyte live-cell analysis system at 48h post-infection. Data information: data represent means and standard deviations from n=3 biological experiments (individual data points shown). Numbers above IFN-α2-treated bars indicate their approximate difference to the respective untreated conditions. Dotted lines are a visual guide for minimum virus replication in control cells in the presence of IFN-α2. Statistical significance was determined by 1-way ANOVA on log-transformed intensity values (***p-value < 0.001; ****p-value < 0.0001; n.s. not significant). | [
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Flow cytometry of CFSE-labeled CD45.1+CD4+ T cells stimulated 3 d alone or together with CD45.2+GFP(Foxp3)+NGFR+-sorted CD4+ naïve T cells from Foxp3EGFP mice transduced with control retrovirus (Min) or retrovirus encoding C/EBPβ and cultured for 3 d in the presence of TGF-β, anti-IFN-γ and anti-IL-4 Abs. Histograms are gated for CD45.1+. The ratios shown are responder to suppressor. Data are representative of two independent experiments with consistent results | [
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MPSIIIA GAG was 2-O-desulphated and the resulting oligosaccharides applied to a WT mixed glial culture for 24 hours. The intracellular production of IL-1β was measured (n=3 independent experiments each with 3 inter-experimental replicates). Data are expressed as mean ± STDEV and were tested by one-way ANOVA with Tukey's post-test; MPSIIIA GAG vs. 2ODS MPSIIIA GAG P<0.0001. 2ODS, 2-O-desulphated. | [
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(C) HEK cells transfected for 8 h with SODG85R‐GFP along with FLAG‐BAG3 and HA‐Hsp70, as indicated, were treated for 24 h with BafA1 (0.1 μM). Indicated proteins were analysed by immunoblotting. Diagram shows the ratio of SODG85R‐GFP levels in BafA1‐treated cells to those in untreated cells. | [
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(C) Validation of gene candidates affecting soluble antigen uptake in primary mouse B cells. Follicular B cells isolated from CRISPR-targeted bone marrow chimera mice internalize anti-mouse IgM F(ab')2. Data show mean and SEM of N = 3 mice. P, statistics from 2-way ANOVA; at 5 min all 3 genotypes show p<0.0005 versus control; at 15 min p values are indicated in corresponding color. | [
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In metastatic melanoma, BACE2 activity assists the secretion of protein aggregates into the extracellular space. The presence of these aggregates might be sensed by Agrin, known to activate the YAP signalling cascade, and is able to induce YAP mediated CTGF transcription. In turn, melanoma cells treated with BACE inhibitors produce fewer protein aggregates and show a lower YAP transcriptional activity. | [
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A. HeLa cells were co-transfected with FLAG-DRP1 and FUNDC1-MYC for 24 h. Cells were lysed and immunoprecipitated using anti-FLAG antibody. FLAG-DRP1 and FUNDC1-MYC were analyzed with the corresponding antibodies. | [
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5D) HeLa cells were transfected with scrambled siRNA or PINK1 siRNA and GFP or GFP-Parkin or PINK1-GFP or PINK1G309D-GFP. After transfection cells were starved for additional 24 h with HBSS and afterwards the amount of adherent cells was determined. The number of adherent cells transfected with scrambled siRNA and the indicated plasmid was set as 1. Starvation-mediated increased cell loss after PINK1 knockdown could be rescued by PINK1 or PINK1G309D-GFP but not by GFP or Parkin; GFP: n = 5, p<0.005; Parkin: n = 3, p<0.05; PINK1: n = 5, ns; PINK1G309D-GFP: n = 3, ns. | [
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(D) Representative immunofluorescent images used for the quantification in C, scale bar 20 μm, collagen-I was assessed with a specific primary antibody staining | [
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Co-staining of CD31 (red) and phage (green) in brain sections comprising tumor and healthy brain. Scale bar, 100μm. A high-magnification view from the low-magnification insert of the tumor section is shown. Scale bar, 25 μm. | [
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J, K) Hippocampal neurons from Nrg3-/- (J) and Nrg3-/-;ErbB4-/- (K) mice were transduced with the SypCFP/Nrg3 expressing virus and immunostained for Nrg3, ErbB4/parvalbumin and SypCFP. Nrg3 and SypCFP were strongly enriched in synapses on dendrites of PV neurons in Nrg3-/- (J) but not Nrg3-/-;ErbB4-/- cultures (K) | [
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