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D SPR binding isotherms of a representative experiment as in C. Calculated KD values of 3.2 and 10 µM for gephyrin and G375D, respectively, are indicated by vertical lines. The experiment was repeated three times with two independently purified protein batches and two sensor chips.
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(F) Results of RT-qPCR analysis for WT primary mouse myotubes subjected to inhibition of both miR-499 and miR-208b (Anti-miRs) (n = 4 independent experiments). *p = 0.037 (Ppargc1a), *p = 0.0019 (Fnip1).
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A: PAI-1 staining (left panel) and quantification (right panel) in glomeruli of kidney biopsies from young (< 40 years of age, n = 8) or old (> 80 years of age, n = 10) kidney donors at time of transplantation (M0). Original magnification X400. Scale bar = 20 μm. B: p16 staining (left panel) and quantification (right panel) in glomeruli of kidney biopsies from young (< 40 years of age, n = 8) or old (> 80 years of age, n = 10) kidney donors at time of transplantation (M0). The arrows point to the staining. Original magnification X400. Scale bar = 20 μm. Data information: Data are means ± SEM. Statistical analysis: t-student test
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G Taufibrils are endocytosed and induce lysosomal damage. Taufibrils were generated in vitro, fluorescently labelled by Dylight488 and fragmented by sonication (488-labelled tau). HeLa cells expressing mCherry-Gal3 were incubated with 300 nM taufibrils for 24 h. Arrows indicate compartments that contain 488-labelled tau and are positive for mCherry-Gal3 indicating lysosome rupture. Percentage of cells with Gal3-positive vesicles was quantified from three independent experiments (mean ± SD).
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Puf3 is phosphorylated in WT cells and is hyper-phosphorylated in ρ0 cells. Puf3-FLAG was immunoprecipitated from the indicated strains and treated with/without λ-phosphatase, and analyzed by regular and Phos-tag SDS-PAGE.
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HFFs were stimulated with for 24 h with 10U/mL IFNγ or left unstimulated and subsequently infected with RH, Pru or PruΔgra15 parasites for 3 h. The percentage of vacuoles that stained positive for (A) Total ubiquitin (n=3 for RH, n=5 for Pru and n=3 for PruΔgra15), is shown in the left bar diagram. On the right-hand side, a representative fluorescent image is shown for the Toxoplasma Pru strain, which expresses GFP. DNA was stained with Hoechst 33258. Scale bar is 10 µm. The yellow box inside each representative image is shown as inset pictures with magnification.
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E 35S−labelled in vitro−translated GFP or GFP−Bchs (aa 3326-3341) were incubated with GST or GST−dAtg8a and binding evaluated by ARG. 5% of the in vitro−translated protein used was loaded. Ponceau staining shows equal amounts of GST proteins used. Data are representative of three independent experiments.
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(E) GO term analysis of genes downregulated 72 hours after the knockdown. Negative log10 p-values of overrepresented GO terms, hypergeometric test.
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C. Histological spleen sections of Myc-Bcl2+LSK-, GMP- or DN2-transplanted mice displaying myeloperoxidasestaining. Leukemic cell engraftment in spleens ranged from 75.1%-95.5%.
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(E) Alignment of amino acid sequence of Munc18-1 in different species. ERK consensus site (P-X-S/T-P) (yellow) around phosphorylation site S241 (red) is highly conserved.
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E. KEGG pathway coverage in this dataset, with major metabolic pathways highlighted.
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The DOCK5 and Raptor binding sites were identified using mutant DOCK5 (DHR-2V1559A) and Raptor fragments (445 - 887 aa) based on a Co-IP assay. VA, mutant DOCK5 DHR-2; His, His-DHR-2.
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Proximity ligation assay between Ripk1 and Casp-8 performed in BN175 cells upon treatment with the indicated agents for 8 hrs (n = 3 biological replicates, shown are representative images). Scale bar represents 10 μm. DMSO (Unt), zVAD (10 µM), TNF (10 ng/ml), SM (100 ng/ml) and Mel (3.3 µM).
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Quantification of the light-mediated dissociation of OptoCAR in HEK293T cells by measuring the ratio of membrane-bound intracellular part of OptoCAR over time. Individual cells were illuminated after 25 seconds of imaging for 25 seconds to calculate dissociation rate (green) before returning to dark (signaling competent) state to calculate re-association rate (red line). A unitary value was assigned to the mean membrane fluorescence prior to illumination and zero to the minimum value. Raw plots are presented in Appendix Fig S1. Bounding area around datapoints shows mean ± SEM (n=12 cells).
|
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C) TOP: Aggregation kinetics of full length (fl) recombinant human TDP-43 (rec-hTDP-43) measured as an increase in absorbance. The addition of TEV protease induces the cleavage of the MBP tag and subsequent aggregation of rec-hTDP-43. Rec-hTDP-43 aggregates used for seeding were collected 18 hours after the addition of TEV protease. BOTTOM LEFT: rec-hTDP-43 aggregates stained by Thioflavin T (ThT). BOTTOM RIGHT: Transmission Electron Microscopy (TEM) image of the rec-hTDP-43 aggregates. All images are presented at the same scale of 1 μm.
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(E) Surface IgM or IgD MFI relative to WT in naïve splenic follicular B cells. N = 3 mice. Data show mean ± SEM.
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C The FIC index assay for synergy between AMSIN and HNP1&2 or hLZc to SSAL and SSAN. The dashed line represents the cutoff of the FIC index (0.5) for synergy.
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J Western blot analysis of ubiquitination of cellular proteins (Pan-Ubi) in HT1080 cells treated with NMS873 (10 μM) and additional NaCl (+34 mM) for 12 hrs. Data information: Data are representative of at least two biological replicates.
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Wild-type (WT) and YF-eNOS (YF) mice were treated with vehicle (Veh) or angiotensin II (AII, 1.44 mg/kg/day) for up to 28 days. PKM2 S-nitrosation in lungs from WT and YF mice administered AII for 28 days, n=7-8 mice per group (Unpaired Student's t test). DTT treatment was included to demonstrate specificity of the SNO signal. The blots show non adjacent lanes from the same membranes.
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The Il17a/Il17f locus, including qPCR primer sites covering SIEs (purple triangles) or AGG elements (green triangles).
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G: Relative mRNA expression of PAI-1 in whole kidneys from irradiated (RT+) and non-irradiated (RT-) control mice 12 months after irradiation. n = 5 for RT+ and RT- mice, respectively. Data information: Data are means ± SEM. Statistical analysis: t-student test: irradiated versus non-irradiated mice.
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Primary astrocytes from wild type mice were stimulated with IFN-γ (10 ng/ml) for indicated times. Proteins were immunoprecipitated from whole cell lysates with anti-OTUB1 antibody and analyzed by WB with antibodies against JAK1, JAK2, SOCS1, and OTUB1.
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(A) Liver with the extrahepatic biliary ducts (EHBDs) and gallbladder (Liver incl. Gb), liver lobes, EHBDs or gallbladder tissues were minced and cultured in Matrigel in the presence of R-spondin 1 and noggin. Phase contrast microscopy images were acquired one week later. Scale bar: 200 m.
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Immunocytochemistry of microglia isolated from Csf-1rflx/wt;Cx3cr1-Cre- and Csf-1rflx/wt;Cx3cr1-Cre+ P0 mouse pup brains. Microglia were stimulated with LPS and exposed to fluorescent opsonised latex beads for 1 hour before fixation and quantification of bead+ cells via microscopy. Cells were stained using MitoTracker(red). Arrowheads (white) indicate bead+ cells. Quantification of mouse microglial phagocytic activity expressed as percentage bead+ cells (n = 4 assays, error bars indicate SEM).
|
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(D) At 48h after triple-transfection of H7, HAT or TMPRSS2, wtSPINK6 or mutSPINK6, BHK21 cells were lysed for the detection of HA cleavage.
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B SDS-PAGE of E. coli expressed and purified 6His-gephyrins and CD spectra. Mean residue ellipticity of gephyrin and G375D show profiles with minima at 208 nm and 222 nm.
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(F) Confocal images of synaptic immunostaining in CA1 striatum radiatum at P10. (G, H) Quantification of synapse density in CA1 striatum radiatum in iCKO versus control at P10 (G), and CKO versus control at P21 (H). At P10, N = 5, p = 0.83. At P21, N = 5, p = 0.89.
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Predicted stress (tension normalized to the area) profiles within the cytoskeletal architecture as the synaptic contact progresses in time, corresponding to the three cases shown in (D).
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(F-G) Western blot for autophagy protein LC3-II, p62 and quantification normalized with GAPDH, mean ± SEM, n=3 biological replicates, *p < 0.05, **p < 0.001 by Student's t test.
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Distribution of each cluster across indicated time points after tamoxifen treatment.
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D The kinase is oriented with the ATP binding site facing the membrane. Key interactions between the kinase and the membrane are made by K621 and K627. The side chains of K621, K627 and K216 (the latter is within the KAKTLRK motif of a neighboring FERM domain) as well as AMP-PNP bound to the ATP binding site are shown in stick representation.
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(F) SIRT1 H363Y blocks CSAG2 regulation of p53 ac-K382 levels. HCT116 cells were transfected with the indicated constructs and blotted for the indicated proteins.
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D. Representative 2D class averages of the Elp123 subcomplex prepared through a high salt dissociation procedure.
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(B, C) Velocities (B) and frequencies (C) of anterograde and retrograde IFT of D1bLIC-YFP. n represents the number of cilia assayed from four independent experiments. Values show the mean ± S.D. Unpaired two-tailed Student's t test analysis, n.s.: no significance; ***, p<0.0001.
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D. Seurat UMAP plots with clusters 0-3 of aged (left panel) and young (right panel) samples from scRNAseq data.
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(D) INTERFEROME database classification of differentially expressed (DE) genes retained in dormant cells into predicted type I and/or II IFN regulated genes (IRGs).
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(C) Na+/K+ selectivities of the K+ sites in EAAT1 and GltPh (OFC) calculated from alchemical free-energy simulations. Mean ± SD of 1000 bootstrap samples.
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(B) HCT116 cells expressing XIAP‐specific or control siRNAs, HCT116 XIAP WT and XIAP KO cells were individually transfected with Flag‐XIAP or control vector. Cell lysates were analysed by western blotting. The data are representative of three biological replicates. The ratio of LCII/LC3I to actin is presented in Supplementary Figure S2D.
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E, Ribo-seq data over the FUS gene from the Gwips portal. Initiating ribosome reads are indicated by blue bars, and elongating ribosomes footprints are indicated by the blue curve. The graph captures the beginning of the FUS gene with FUS and altFUS methionines indicated by blue arrows. The genomic positions are indicated relative to the start of exon 1.
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A Scatter plot of RNA-seq expression analysis. 508 genes were upregulated and 391 genes were downregulated. Human MSCs were transfected with USP34 or control siRNAs and cultured in osteogenic medium for 2 days. Two biological replicates per group
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(H) Activities of major cell types in muscle regeneration. The colors of the heatmap show the log2FC to day 0 (uninjured) of TR and TSS levels. Representative genes with significant changes in TR are shown.
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B. ELISAs were used to analyze the inhibitory effect of OSMI-1 (10 µM, final) on cytokine and chemokine production from THP-1 cells exposed to Stx2a (10 ng/mL) for 9 h (n = 3 biological replicates). Data information: For graphs error bar represents mean ± S.E.M. Statistical analysis was performed using two-tailed Student's t-test. *P < 0.05; **P < 0.01; and ***P < 0.001. The effects of OSMI-1 were compared with those of the vehicle (DMSO) control at each time-point.
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K. Heat map of PtdIns, PtdInsP and PtdInsP2 levels from 4 different murine brain regions. The colors from green to red indicate the relative contents of lipid isoforms in different brain regions from NPC1I1061T mice relative to wild-type brains; n=3 for both WT and NPC1I1061T animals.
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Separase is required for proper HDR but dispensable for NHEJ. U2OS DR-GFP (HDR reporter) and U2OS EJ5-GFP (NHEJ reporter) cells were separase-depleted by RNAi (SEP-1) or control-treated with GL2 siRNA, transfected to express HA-tagged ER-I-SceI, and then supplemented with OHT in G2-phase to induce nuclear accumulation of the homing endonuclease. Ethanol-supplemented samples served as negative controls. Two days later, cells were subjected t flow cytometry to quantify the percentage of GFP positive cells (B The GFP quantification in (B) displays averages (bars) of 3 independent experiments (dots)
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K. Basal R-GECO1 fluorescence (F) normalized to the maximum R-GECO1 fluorescence (Fmax) after ionomycin treatment at presynaptic boutons of hippocampal neurons (DIV18) co-expressing GFP, GFP-SCRN1 or GFP-SCRN1-F402A. N=3, n=70-89. L. Average basal R-GECO1 fluorescence (F0) normalized to the maximum R-GECO1 fluorescence intensity (Fmax) after ionomycin treatment at presynaptic boutons of hippocampal neurons (DIV18) co-expressing GFP, GFP-SCRN1 or GFP-SCRN1-F402A. N=3, n=70-89. Data information: Data represent Mean ± SEM; NS: not significant; *P < 0.05; **P < 0.01; ***P < 0.001, by Mann-Whitney U test.
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VEGFA (A) and VEGFR2 (B) mRNA levels quantified by qRT-PCR in control or CEP41-knockdown HUVECs under static or shear stress conditions. The expression of GAPDH was quantified for the normalization of the qRT-PCR results. Data are shown as mean ± SD of three independent experiments. Statistical significance was determined with the Brown-Forsythe ANOVA followed by Dunnett's T3 post hoc test (**P < 0.01, ***P < 0.001, ns: non-significant).
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D) U2OS cells were synchronized in G2 and damaged with 6 Gy of IR. Samples were taken at the indicated times after induction of damage with or without Tlk2-depletion. Samples were analyzed by western blot for the indicated proteins. Band signal intensity for Asf1A and Asf1B was measured and corrected for Ponceau S staining.
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(H) Primary hepatocyte expressing Dyn2-GFP, showing an absence of colocalization with the LD surface (stained with Oil Red O). Bar, 20 µM. Inset shows magnification of boxed region (bar, 2 µM).
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HEK293 cells: (A) Schematic outline of the experiment shown in (B). HA-Nrxn1α or HA-Nrxn1α (S>A), a mutant of the modified serine were expressed alone or in combination with V5-tagged CA10 in HEK293 cells. Neurexins were immunoprecipitated for HA and treated with heparinases ('Heps').
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D. HeLa cells were transfected with siCtrl or siRNA against Bclaf1 or p50, treated with TNF (10 ng/ml) for 6 hours followed by western blotting analysis.
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(G) Expression of Flag-EI24 in EI24 knockout cells inhibits IRE1 activity. Flag-Metap2 served as the control. Cells transfected with the indicated cDNAs were treated with or without 4 μM TM for 2 hr. Protein expression was analyzed by immunoblotting.
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(G) Survival rate of MEFs treated with tunicamycin (5 μg/ml), thapsigargin (1 μM), or hydrogen peroxide (100 μM). Error bars represent ±SD from four independent experiments. Data information: For graph the P values was determined by a Mann-Whitney U test. ns = not significant, *P < 0.05, **P < 0.01.
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Skin tumor outcomes in WT mice treated with DMBA/TPA carcinogenesis protocol in combination with SB431542 (test, n=9) or acetone (carrier control, n=5). (C) skin tumor counts per mouse over time
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(F) Surface expression levels of CD16:7 chimeras. Procedures and data expression were as in Figure 2E. The figure shows that the functional differences observed between CD16:7 chimeras (C-E) are not caused by differential surface expression.
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D. Sperm counts from the cauda epididymis of wild type and Gemc1-/- mice revealed no sperm in mice lacking GEMC1 (n=3). Mean and standard deviation are indicated. E. Examples of sperm morphology from spermatocyte spreads from 2 month old mice. Scale bars = 10 µm.
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overall mortality of DLBCL patients stratified according to subtype and disease stage pSTAT3 positivity was associated with higher overall- as well as disease-specific mortality in both ABC- and GCB-DLBCL subtypes
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C Relative positions of the nine sgRNAs. A ray diagram exhibits the locations of the nine sgRNAs in relation to the upstream regulatory regions of SNCA and the H3K4me3 and H3K27me3 peaks. The locations of the sgRNAs are as follows with respect to the TSS: sgA, 1,117 bp; sgB, 836 bp; sgC, 747 bp; sgD, 700 bp; sgE, 153 bp; sgF, 1,454 bp; sgG, 1,537 bp; sgH, 132 bp; sgI, 1,320 bp. The two non-coding exons (1A and 1B) along with the first coding exon (exon 2) are also shown. GRCh37/hg19 contig was used to describe the location of the gene and associated distribution of histone PTMs. The histone peaks shown were from midbrain regions of two adult postmortem human samples.
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a, clinical signs in infected hamsters. The clinical score is based on a cumulative 0-4 scale: ruffled fur; slow movements; apathy; absence of exploration activity. Symbols indicate the median ± interquartile range. Data information: Horizontal lines indicate medians. The p value is indicated in bold when significant at a 0.05 threshold. Mann-Whitney test M: male hamsters; F: female hamsters. Data were obtained from two independent experiments for each sex.
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D) Quantification of the distinct UIS4‐ and IBIS1‐positive membranous structures in fixed infected hepatoma cells at three different time points: 6 h after infection (the period of sporozoite transformation into exo‐erythrocytic forms), 24 h after infection (onset of parasite replication and growth), and 48 h after infection (mature liver stages). Values are shown as mean (± SD) from three separate wells (n = 100 per well) of one experiment and are representative of two experimental repetitions.
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D) Heat map of DEGs within the combined STAT3 gene sets representing mean z scores calculated from normalized RNAseq count data. Tumor-free control (black checkered), GTx-024-treated tumor-free (blue checkered), tumor-bearing control (black), GTx-024-treated tumor-bearing (blue), AR-42-treated tumor-bearing (red) and Combination-treated tumor-bearing (green).
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B Stable knockdown of ER-alpha within CTC-ITB-01 cells was performed by using lentiviral transfer of non-targeted (scramble) or two different targeted shRNAs against ER-alpha. CTC-ITB-01 cells were seeded on 6-well plates. After 10 days cells were fixed and stained (left upper panel). Macroscopic photos are shown of the fixed cells (left lower panel), scale bar 200 µm. Western blots of lysates from knockdown of ER-alpha expression in CTC-ITB-01 cells transduced with the same amounts of lentiviral vectors were probed with the indicated antibodies. The extracts were obtained 72h after lentiviral transduction (right panel). Where indicated, western blots were visualized at long exposure (l.e.) and/or short exposure (s.e.).
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A U2OS cells were treated with 10nM actinomycin D and cell extracts were prepared at indicated time points. Proteins from extracts were analyzed by western blot using the indicated antibodies.
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A. GST-pull down assays of purified GST-tagged Elp3 SAM domain (aa72-389) and KAT domain (aa348-557) with untagged full length Elp1 and Elp451-2706. GSH resin and proteins were used as input controls. Lower panel shows 5% of the input and upper panel shows bound fractions. All samples were analyzed by SDS-PAGE and stained by Coomassie. Identities of respective proteins are indicated on the right.
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I-J) Circular histograms showing the number of phosphopeptides at each rhythmic phase. The distributions of rhythmic phosphopeptides in CKO cells was significantly different to WT cells (***p<0.001, Watson's two-sample test).
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(A) IHC images for vGluT2 staining in the dLGN of microglial Gpr56 conditional knockout mice (Gpr56fl/fl; Cx3cr1cre/+) and controls (Gpr56+/+; Cx3cr1cre/+). Both mice were intravitreally injected with AAV2 carrying Cdc50a shRNA. The yellow squares indicated the AAV2 non-infected areas and the dotted outlines indicated the AAV2 infected areas. Scale bar, 200 µm.
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(J) Western blot of GluR1 with microdissected P30 dLGN tissue. (K) Quantification of GluR1 expression in CKO mice and controls.
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Representative histopathological analysis for lung sections of Mavs -/- (D) mice that were infected with M.tb and subsequent left untreated (Mock) or treated with EVs from uninfected BMMs (EVs_Control), EVs from M.tb-infected BMMs (EVs), moxifloxacin (MXF), or combination of EVs and MXF (EVs+MXF). Data shown is representative of two independent experiments. The results are the mean ± SD (n = 4 mice per group). Scale bar, 100 μM
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(C-G) L6 myotubes were transduced with lentiviral expression plasmids for Flag-ALDH1L1 and GFP. NADPH levels (C), H2O2 levels (D, E) , Mitochondrial respiration (F, G) were measured from these cells in the presence and absence of NADPH oxidase inhibitor apocynin (n = 3 per group, for (C-E) and n = 5 for (F, G), means ± SEM, * p < 0.05, two-tailed student's t-test and two-way ANOVA followed by Bonferroni post-hoc testing).
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F. A representative trace of whole-cell current activated by 3 mmol/L of 2APB in the presence or absence of 10 µmol/L SKF in mouse differentiated brown adipocyte. The left inset indicates a voltage ramp-pulse protocol. The right inset indicates the current-voltage curves of basal current (a, black), current in the presence of 2APB+SKF (b, blue) and current in the presence of 2APB alone (c, red) at the time points of a, b and c, respectively.G. Comparison of the mean densities of basal currents (Base, black), currents in the presence of 2APB alone (red) and currents in the presence of 2APB+SKF (blue) at -60 mV and +100 mV in mouse differentiated brown adipocytes. Data are presented as mean ± SEM, n = 10; ** P < 0.01 vs. Base; ##P < 0.01 vs. 2APB alone. One-way ANOVA followed by 2-tailed t-test with Bonferroni correction.
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(A) Scheme of the genetic strategy to delete YAP and TAZ and activate the oncogene SmoM2 in tumor initiating cell
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H The S phase length of the GBCs in the MOE of the NC and miR-200b/a KD mice (the S phase length was calculated using the following formula: 3 h×BrdU+/BrdU+EdU-; n=3 mice each group; data represent the mean±SEM; *p<0.05; Student's t-test).
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A Representative still images of Alexa488-labeled TDP-43 condensates by spinning disc timelapse confocal microscopy. Wt condensates do not fuse, 5D condensates fuse slowly and 12D condensates readily fuse upon contact and relax into spherical droplets. Bar, 5 µm.
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J: Transacylations were measured as in I in the presence of 0.9 g BR/ g lipid. The ratio of total linoleoyl transfer over total oleoyl transfer was calculated.
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E Left, CD81-CD19 docking model in the closed conformation. The HDX-MS identified binding regions are colored as in C. Right, Modeled open conformation of CD81 and docking with CD19 (in the same manner as Left). MD simulation (1 µs) of this open conformation is shown in Fig 4 and Movie EV1.
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E - Lysates of cells treated or not for 6 h with ionomycin were incubated at 37 °C for 45 min with fructose-6-P and glutamine. The production of glucosamine-6-P was measured using HPAEC-PAD. Results of three independent experiments are shown, with mean ± SD, and p-value of the Student's paired t-test is indicated (*P < 0.05).
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D, E, Western blotting analyses of proteins co-precipitated with anti-FLAG antibody from extracts of HeLa cells transfected with FLAG-tagged NANOS2, NANOS2 (ΔN10), NANOS2 (ΔC10), NANOS2 (C61A, C96A) or NANOS3, and HA-tagged DND1 (D) or with FLAG-tagged NANOS2, NANOS2 (C61A), NANOS2 (C96A) or NANOS2 (C61A, C96A), and HA-tagged DND1 (E).
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D Numbers of T and B cells found in pooled mesenteric and peripheral LNs of WT and Arhgap45-/- mice. Data information: In the results for each mouse are shown as a dot and correspond to three to four experiments involving a total of 12-25 mice. * P ≤ 0.033, ** P ≤ 0.002, *** P ≤ 0.001, **** P ≤ 0.0001; unpaired Student's t test. SEM are also shown.
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A. Western blot and quantification of phosphorylated PKD (S916/S876) in small intestine of Pkd2flox/flox and Pkd2gutΔ/Δ male mice. n=3. B. Western blot and quantification of phosphorylated PKD (S744-748) in small intestine of Pkd2flox/flox and Pkd2gutΔ/Δ male mice. n=3. C
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GFP signals (Bub1) co-localizing with Tomato signals (kinetochores) were scored on the movies (D).
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(C) PR1 and PR5 induction in transgenic lines. Leaves of four-week-old plants were inoculated with Pst DC3000 (OD600=0.01), and collected at 0, and 24 hpi for qRT-PCR analysis. The data in B and C are shown as mean ± SD (n=3) from three independent repeats with one-way ANOVA analysis and Tukey test (p<0.05). Different letters indicate significant differences.
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(A) The LC3-II level is reduced after 3 or 24 hr exposure to SIV-infected microglia supernatant and this decrease effect is blocked when is pretreated with rapamycin (2 µM). The protein level of elongation complex Atg12-Atg5 is also reduced, and again the effect is blocked in the presence of rapamycin. The protein level of p62 is increased for similar conditions with SIV-infected microglia supernatant, and the increase is blocked in the presence of rapamycin. GAPDH protein was used in these experiments as the loading control. One representative experiment of n = 4 is shown.
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Relative luciferase activities of indicated constructs in differentiating C2C12 cells transfected with control or Myoparr shRNA (Myoparr KD).
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B COS‐1 cells were treated with ATP8A1 siRNAs for 72 h. Cell lysates were prepared and then immunoblotted with anti‐ATP8A1 antibody. α‐tubulin was used as a loading control.
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Recombinant ISG20 and mA3A (mutant E72A, C72A) were mixed with indicated oligomers and analyzed in a single point binding mobility shift assay.
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(J) The number of motor neurons in the spinal cord (SC) in Epg5+/− and Epg5−/−mice.
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(B-C) Representative confocal images showing the uptake of double-labelled EVs (Ribogreen; green, PKH26, red) by astrocytes stained with GFAP (white) (B) and microglia cells stained with IBA1 (white) (C) incubated on mixed cortical cultures. Boxed areas are shown as zoomed images on right. Cell nuclei are stained with Hoechst (blue). The in vivo and in vitro uptake experiments were performed 3 and 2 times (n=3), respectively. Scalebar 100 = µm.
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cDNA analysis by Sanger sequencing of leukocyte RNA from patient A and a control sample. Reads beyond the alternatively spliced breakpoint yield double reads in patient samples, corresponding to the predicted mis-splicing and wildtype sequences.
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a mCherry-tagged truncated BRD4 mutant proteins and GFP-tagged ISX were detected in anti-GFP immunoprecipitates by Western blot. Data information: Each experiment was repeated at least three times.
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A. Representative fluorescence images of HeLa cells expressing the PtdIns3P probe GFP-2xFYVEHrs and MavQ with or without wortmannin treatment. Scale bars, 5 μm.
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Nissl staining of the Purkinje cell layer of the brains from various knockout mice shown in (C) confirms the absence of Purkinje cells in Ccp1-/-, and the presence of a wild-type-like Purkinje-cell density in Ccp1-/- Ttll1flox/flox L7-cre mice. Scale bar: 20 µm. Purkinje cell layer (PC) and granule cell layer (GC) are indicated.
|
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293T cells were co-transfected with NS3 expression plasmid (or empty plasmid as control) and siRNA for Beclin-1 (or siRNA control or no RNA as controls). Lysates were obtained from FACS-sorted transfected cells. Immunoblots of sorted cells were probed for STAT1; STAT2; Beclin-1 FLAG-tagged NS3; LC3 and GAPDH.
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VEGFR2 expression is regulated by TFEB through a miR-dependent mechanism. VEGFR2 in human scr-shRNA and sh-TFEB ECs and in control treated with a specific miR-control, miR-15a-5p and miR-16-5p mimics Representative western blot of VEGFR2 expression under the same experimental conditions previously reported. The bar graph shows the densitometric analysis expressed as the ratio between VEGFR2 and α-tubulin
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] |
(B) Cytoplasmic TDP-43 immunostaining is elevated not only in poly-GA transduced neurons, but also in the non-transduced receiver cells. White and red arrows indicate cells with cytoplasmic TDP-43 in GFP positive and negative cells, respectively.
|
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D. GFP-TgAtg8-expressing parasites were grown in host cells in the presence of 3H-ethanolamine and then starved in HBSS for 8 hours, still in the presence of the radioactive PE precursor. Tachyzoites were then lysed and GFP-TgAtg8 was immunoprecipitated using anti-TgAtg8 antibody and analysed by urea SDS-PAGE. The immunoprecipitated forms of GFP-TgAtg8 were detected by Western blot using anti-GFP antibody and radioactive ethanolamine was found to be incorporated into immunoprecipitated GFP-TgAtg8, as detected by fluorography (3H-etn).
|
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F. Representative hematoxylin and eosin (H&E)-stained images of mouse brain sections with GSC1123 xenografts expressing shC, shL115-1, or shL115-2. Images represent results of five mice per group of two independent experiments. Scale bars: 1 mm. G. Quantification of tumor volume in F.
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C. HA-MBP-control, HA-AGO1 or HA-AGO2 was co-expressed with GFP-SIM1 and the cell lysates were subjected to IP with HA antibodies and WB analysis with indicated antibodies to detect the presence of proteins in the immunoprecipitates.
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Photograph showing the custom pressure controller used for programmatic pressure control during automated microinjection. Scale bar is 8 mm.
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(B) Immunoblot analysis was performed on MDA-MB-157 and SUM159PT cells treated with single or in combination with AZD6244 and BI2536 inhibitors after 96 hours and cleaved PARP and Caspase-3 were determined along with CEP55, MYC and both phosphorylated and total ERK1/2. COX-IV as a loading control (left panels). Percentage of sub-G1 population identified using propidium iodide staining and quantified by FACS following single and combination treatment with AZD6244 and BI2536 inhibitors after 96h. Graph represents the mean±SEM of two independent experiments (middle panels). Representative images of colony forming capacity at 14 days determined using crystal violet staining in cells treated with single and combination inhibitors (middle panels). Combination index (CI) values calculated through CompuSyn software (right panels) where 1 indicative of additive (no interaction), >1 indicative of antagonistic, and <1 indicative of synergistic.
|
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Images of mESCs transduced with Rptor siRNA on different days in 2iL. siNC, transduced with a negative control; day 0 was the day of siRNA transfection, also hereafter in similar experiments. Scale bar, 400 µm.
|
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A. Mice were co-colonized with E1 and either N1 WT (n=4) or N1 ΔtssC (n=3). Five days post-inoculation, fecal RNA was extracted and tested for BFT expression via qRT-PCR.
|
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