EMBO/sd-character-level-ner huggingface dataset:
Figure 3(A) The overall structure of OTULINWT (3ZNV) is depicted with its catalytic triad (C129, H339, N341; pale cyan) and positions of variants identified in this study (M86, W167). (B) Close-up of M86 (wheat) and M86I (pale green) with the interaction network around OTULIN's catalytic center. (C) Close-up of OTULINC129A and its interaction with the proximal ubiquitin (Ub). M86, Y91, E95, and R122 of OTULIN and their interaction with each other and with the F4 patch of the proximal Ub are depicted. Interactions (dotted red lines) and water molecules (light blue) < 5 Å are shown. Residues of the catalytic triad (A129, H339, N341) in pale cyan. (D) OTULINW167S in complex with di-Ub. Contact sites of W167 (L237, I241, L273 and R274) in pale yellow. Positions of known homozygous OTULIN substitutions (Y244, L272, G281) in pale green. I44 patch of distal Ub in magenta and its contact sites L259, A262 and R263 in pale yellow. Close-up of W167 and S167: loss of size and gain of polarity from W to S.
Figure 6A. Western blot analysis of phosphorylation levels of tau and expression levels of indicated proteins in the cortical and hippocampal regions of the brain in APP/PS1mice after administration of Nec-1 (6.25 mg/kg) or vehicle (2.5% DMSO in PBS). The membranes analysed in (A) are identical to those in Fig 5F. B. Immunohistochemical analysis of cortical and hippocampal regions of the brains in wild-type (Wt) and APP/PS1mice after administration of Nec-1 (6.25 mg/kg) or vehicle (2.5% DMSO in PBS) for phosphorylated tau expression levels. Anti-tau (phospho Ser199) antibody (red colour) was applied for staining tau phosphorylation on Serine-199 and Hoechst 33342 (blue colour) for nuclear counterstaining in the brain sections. Scale bars = 200 μM. C. Western blot analyses of phosphorylated RIPK3 expressions in cortex and hippocampus of APP/PS1mice injected with Nec-1 (upper). The membranes analysed in (C) are identical to those in Fig 1G and 5F. C. Western blot analyses of phosphorylated RIPK3 expressions in primary cultured astrocytes treated with Nec-1 (lower). The membranes analysed in (C) are identical to those in Fig 1G and 5F. D, E. ThT assays for the inhibition of tau aggregation (D) and for disaggregation of tau (E). Fluorescence intensity was normalized to tauaggregates (100%, day 5). F. Surface plasmon resonance sensorgrams of Nec-1 targeting tau (left), and the corresponding dissociation fitting curve from the saturated region (right) under various concentrations of Nec-1.
Figure 3A. The coloning-formation rates of the trisomic and wild-type ES cells. Error bars, ±S.D. n=3. * P < 0.05, ** P < 0.01. B. Alkaline phosphatase (AP) staining of trisomic and wild-type ES cells after LIF withdrawal (LIF 0 U/ml) for 5 days. Scale bar, 500 μm.C. Evaluation of AP positive and negative colonies after LIF withdrawal. Black bars, undifferentiated colonies; orange bars, partially differentiated colonies; green bars, totally differentiated colonies. Error bars, ±S.D. n=3. **, P < 0.01. D. Boxplots of EB diameter at the indicated days of EB formation. Each boxplot was plotted for more than 20 EBs. * P < 0.05, ** P < 0.01. E. Time course analysis of marker genes expression during EB formation by RT-qPCR. Error bars, ±S.D. n=3. F. Quantification of Oct4, Nanog and Rex1 expression in trisomic and wild-type ES cell derived EBs at day 8 of differentiation. Error bars, ±S.D. n=3. ** P < 0.01.
Figure 3A Schematic of assay measuring CD8+ T cell mediated cytotoxicity. Cancer cells (grey) with or without genetic modifications were mixed with BiTE or bi-specific antibodies with or without IFN-γ and cytotoxic T lymphocytes (CTLs). Altered tumor cell numbers were determined after allowing for T cell engagement. B Relative cell numbers (± SD; n=3; biological replicates) of U2OS mock (black) or polyclonal MLLT6 knockout (grey) cells treated with varying amounts of EpCAM-CD3 BiTE and CTLs (Student's t-test; **p < 0.01; degrees of freedom (df)=4). C Kaplan Meier survival plot (left) showing changes in survival (percent) of mock (grey) and MLLT6 knockout (red) cells treated with CTLs and EpCAM-CD3 BiTE over a course of 9.5 hours. Representative images (right) of time-lapse fluorescence microscopy following U2OS mock (eGFP tagged, green) and MLLT6 knockout (mCherry tagged, red) cells immediately (top left and bottom left) after adding CTLs and EpCAM-CD3 BiTE and 9.5 hours later (top right and bottom right) (scale bars = 100 μm). D Relative cell numbers (left) (± SD; n=3; biological replicates) of mock (black), PD-L1 (dark grey) or MLLT6 (light grey) knockout cells treated with BiTE or bi-specific antibodies and CTLs as indicated (Student's t-test; **p < 0.01; df=4). Representative images (right) from bright field microscopy of polyclonal U2OS mock (top), PD-L1 (middle) or MLLT6 (bottom) knockout cells treated without or with EpCAM-CD3 BiTE or bispecific antibodies. E Relative normalized cell numbers (± SD; n=3; biological replicates) of U2OS mock (left), MLLT6 (middle) or PD-L1 (right) knockout cells treated with EpCAM-CD3 BiTE and CTLs in the presence (grey) or absence (black) of IFN-γ (Student's t-test; **p < 0.01; df=4 ).
Colorectal adenocarcinomas explants undergo collective invasion A) Representative confocal images of non-invasive or invasive human colorectal cancer explants collected from 8 patients with NOS adenocarcinoma (EV2A and B).Figure 2A) Representative confocal images of non-invasive or invasive human colorectal cancer explants collected from 8 patients with NOS adenocarcinoma The explants were fixed 4 days after recovery and stained for the lumen (Ezrin), F-actin (Phalloidin) and nuclei (DAPI). Boxed regions i, ii, iii and iv are displayed at high magnification. Arrowheads point to non-protruding cells, arrows point to protruding cells and white stars show off-centered nuclei. Scale bars: 20 μm. Graphs representing the percentage of adenocarcinoma explants displaying lumens (B) from 8 patients. Evaluation performed from DIC observation of live tissues, at least 50 explants were analysed per patients. The error bar represents the Standard Error of the Mean (Means ± SEM). P values were calculated using unpaired t-test (ns: non significant). Graphs representing the percentage of adenocarcinoma explants displaying protrusions (C) from 8 patients. Evaluation performed from DIC observation of live tissues, at least 50 explants were analysed per patients. The error bar represents the Standard Error of the Mean (Means ± SEM). P values were calculated using unpaired t-test (ns: non significant). D) Time-lapse sequences of an adenocarcinoma explant undergoing collective invasion into collagen-I gel monitored by DIC microscopy over 2 days Arrowheads point to non-protruding cells, arrows point to protruding cells and white stars point to lumens. Scale bars: 50 μm. E) Images of an invading gland of a colorectal NOS adenocarcinoma, at the light microscopic level (i, semithin section, 100x objective), and as seen by electron microscopy (ii, 1000x magnification; and iii, 4000x magnification). Note that the bulk of the invading neoplastic gland is in a state of glandular organization (regions denoted by arrowheads in the left panel): epithelia (E, middle panel) are polarized to form a lumen (L), and are attached to a basal membrane (asterisk) by hemidesmosomes; the gland is surrounded by fibroblasts (F). However, this organization is focally disrupted at the tubular invasion pole (arrows in the left panel) where neoplastic epithelial cells have lost polarization and directly contact the surrounding stroma as the basal membrane is missing (right panel).
a, b, Ciliated Atg5−/− MEFs in serum+ (*P = 0.f5a, b, Ciliated Atg5−/− MEFs in serum+ (*P = 0.038, 25 cells each per experiment, n = 3) (a) or serum− (*P = 0.006, nonlinear fit regression, 25 cells each per experiment, n = 3) (b). c, Scanning electron microscopy (SEM) of primary cilia. Arrows, cilia-associated vesicles; arrowheads, ciliary pocket. . d, Cilia length quantification from SEM (*P = 0.027 n = 28; **P = 0.001, ††P = 0.0001, n = 23). e, f, GLI1 (*P = 0.04, n = 3) (e) and GLI2 (***P = 0.0005, **P = 0.0046, †P = 0.0132, n = 3) (f) messenger RNA expression in Atg5−/− MEFs. g, h, IFT20 immunoblot in Atg5−/− (g) and WT MEFs with lysosomal inhibitors (NL) (h). i, IFT20 immunoblot in cytosol (Cyt), homogenate (Hom) and autophagosomes (APG). j, IFT20 protein levels in Atg5−/− MEFs (*P = 0.038, n = 6). k, IFT20 immunoblot in Atg5−/− MEFs. n.s., statistically non-significant. Mean ± s.d. in d and j and mean ± s.e.m. in other panels.
Figure 8(A) qRT-PCR was performed to analyze UHMK1 expression in BGC823 cells infected with or without H. pylori CagA+ strains 7.13 and J166. MOI, multiplicity of infection. Data were presented as mean ± standard deviation from three replicates. Data information: Unpaired, two-tailed statistical significance was assessed by Student's t‐test. Data represent mean ± SD. *P< 0.05, **P< 0.01,. (B) Immunoblotting of UHMK1, NCOA3 phosphorylated at S1062/T1067, NCOA3, and ATF4 in the samples (C) PMSS1, a H. pylori strain, was used to challenge mice orogastrically, with Brucella broth as the control. qRT-PCR was performed to measure UHMK1 (Ci) and PMSS1(Cii) expression in gastric tissues. Data were presented as mean ± standard deviation from three replicates. Data information: Unpaired, two-tailed statistical significance was assessed by Student's t‐test. Data represent mean ± SD. *P< 0.05, **P< 0.01,. (D) Immunoblotting of UHMK1, NCOA3 phosphorylated at S1062/T1067, NCOA3, and ATF4 in the samples (E) An ATF4 luciferase reporter assay was carried out in BGC823 cells with or without H. pylori infection. Data were presented as mean ± standard deviation from three replicates. (F) (Fi) The domain architecture of UHMK1 and the positions of human GC-associated mutations. (Fii and Fiii) IP kinase assay. Briefly, UHMK1 from HEK293 cells expressing WT-UHMK1 or its mutants was immunoprecipitated with anti-HA antibodies. The immunoprecipitated proteins were mixed with a synthetic peptide of the CATS protein (a known substrate of UHMK1) and [32P] ATP. A phosphocellulose paper assay was used to measure kinase activity. The results were normalized to a value of 1.0 for WT-UHMK1(three biological replicates). (G) The effects of human GC-associated UHMK1 mutations on the proliferation and colony formation of GC cells (three biological replicates). (H) The effects of human GC-associated UHMK1 mutations on GC cell invasion and migration (three biological replicates). Data information: Unpaired, two-tailed statistical significance was assessed by Student's t‐test. Data represent mean ± SD. *P< 0.05, **P< 0.01,.
Figure 3A The MAPKKK5-GFP protein was detected in the membrane fraction. The total (T), soluble (S), and microsomal membrane (M) fractions purified from Nb leaves expressing MAPKKK5-GFP were used for immunoblots with antibodies against GFP, PM-localized aquaporin PIP1 and cytosolic ascorbate peroxidase EAP1. B Subcellular localization of MAPKKK5-GFP and GFP in Nb leaves. The results were photographed at 38 hpi. Scale bar=50µm. C Detection of MAPKKK5-GFP and PBL27-HA transiently expressed in Nb leaves. Total proteins were purified from Nb leaves at 38 hpi. D Localization of MAPKKK5K375M-GFP was analyzed by using total (T), soluble (S), and microsomal membrane (M) fractions purified from Nb leaves expressing MAPKKK5 K375M -GFP E Co-immunoprecipitation assays show that MAPKKK5K375M-GFP forms the complex with PBL27-HA in Nb leaves. The microsomal fractions purified from Nb leaves co-expressing MAPKKK5K375M-GFP and PBL27-HA at 48 hpi were used for co-immunoprecipitation with α-GFP. F Visualization of the interaction between PBL27 and MAPKKK5K375M by bimolecular fluorescence complementation (BiFC) analysis in Nb leaves. The results were photographed at 38 hpi. Venus fluorescence indicates the interaction between PBL27 and MAPKKK5K375M. Scale bar=50µm.
QKI regulates thermogenic mRNA stability, nuclear export and translation efficacy.Figure 6The mRNA stability in adipocytes was measured by incubating cells with actinomycin D (5 μg/ml). Actinomycin D was added to stop transcription, and RNAs were harvested at the indicated times (x-axis) after transcription inhibition. Real-time PCR was used to determine remaining UCP1, PGC1α and Actin mRNA levels compared with the starting time. Increasing PGC1α mRNA half-life by depletion of QKI (ShScramble T1/2 = 4.73 h, ShQKI T1/2 = 8.04 h). (n=4 biological replicates) The nuclear and cytoplasmic fractionation was valid by immunoblotting using cytoplasmic and nuclear protein markers (GAPDH and Lamin B1) to show the purity of fractions. Real-time PCR analysis of mRNA transcript levels of the QKI targets UCP1 and PGC1α in the nuclear and cytoplasmic fractions of differentiated MEF cells transfected with ShScramble or ShQKI. Values represent the cytoplasmic-to-nuclear ratio of each transcript. Actin served as a negative control. (n=4 biological replicates) RNAs from isolated nuclear and cytoplasmic fractionation were harvested after RNA immunoprecipitation (RIP) of RNA-protein complexes using either anti-QKI antibody or control IgG. And RIP-RT-PCR showing the PGC1α and UCP1 transcripts majorly interacted with endogenous QKI in the nucleus. (n=3 biological replicates) Control (ShScramble) and QKI knockdown (ShQKI) adipocyte lysates were subjected to polysome profiling, and a representative polysome profile is shown. Total RNA was extracted from each fraction, and the distribution of UCP1 and PGC1α mRNA was measured via real-time PCR (K and L). The results are shown as the percentage of total mRNA (% mRNA) in polysome fractions and represent 3 independent experiments.
Npc2 haploinsufficiency causes stroma IMC accumulation.Figure 6Shortened survival of BVE mice following reduction in Npc2 gene dosage. Genotypes of mice are as follows: +/+ = Braf+/LSL−V600E;CreER+/0;Npc2+/+, +/hypo = Braf+/LSL−V600E;CreER+/0,Npc2+/hypo, hypo/hypo = Braf+/LSL−V600E;CreER+/0;Npc2hypo/hypo. Median survival time (MST) for each genotype is indicated. Representative H&amp;amp;E staining of lung sections from 5-week-old BVE mice on the Npc2+/+ (+/+) or Npc2+/hypo (+/hypo) background is shown. Scale bars, 500 μm. Stroma IMC quantification per lung in Npc2+/+ (+/+) or Npc2+/hypo (+/hypo) BVE mice at different ages (39-109 days p.p.). Ki67/Mac2immunofluorescence analysis of Npc2+/+ (+/+) and Npc2+/hypo (+/hypo) BVElung sections at 5 weeks of age. Merged images for Ki67 (green), Mac2 (red) and DAPI (blue) are indicated. Arrows indicate dual positive cells for nuclearKi67 and Mac2. Scale bars, 50 μm. The bar chart indicates % Ki67 in Mac2+lung stroma cells (mean + SD), obtained from 4 independent Npc2+/+ and Npc2+/hypoBVE pairs. In vivo BrdU incorporation of CD11c+ IMCs from Npc2+/+ (+/+) and Npc2+/hypo (+/hypo) BVE mice are shown. Representative flow cytometry plots of three independent experiments are indicated. Ki67 (green)/Mac2 (red) dual immunofluorescencestaining of lung sections from 5-week-old Npc2+/hypoBVEmice with nuclearDAPI (blue) staining, focusing on intratumour Mac2+cell clusters. Scale bar, 50 μm. Enlarged images of the boxed areas 1 and 2 highlight Ki67+Mac2− cells associated with Mac2+ cells (arrows). Lung SPC+/CD11c+cell numbers of Braf+/LSL−V600E;Npc2+/+ (+/+, n = 3) and Braf+/LSL−V600E;Npc2+/hypo (+/hypo, n = 7) mice at 10 weeks post-AdCre infection.
ssRNA40- and CL097-induced autophagy in macrophages is mediated through CYP27B1 and the VDR.Figure 3A-B) Macrophages were treated for 24 h with CL097, ssRNA40 or ssRNA41. (A) Top, qRT-PCR for CYP27B1 after 6 h. Bottom, immunoblots of CYP27B1 using antibody to CYP27B1 or β-actin. (B) Top, qRT-PCR for VDR after 6 h. Bottom, immunoblots of VDR using antibody to VDR or β-actin. (C-D) Macrophages transduced with non-specific scrambled shRNA (shNS), CYP27B1 shRNA (shCYP27B1) or VDR shRNA (shVDR) were treated for 24 h with CL097, ssRNA40, ssRNA41 or rapamycin (Rapa). (C) Left, immunoblots performed with antibodies to CYP27B1, VDR and β-actin. Right, immunoblots of LC3B isoforms using antibody to LC3B or β-actin. (D) Cells were fixed and permeabilized then stained with antibody to LC3B. Bar charts represent the percentage of cells with LC3B puncta and are presented as the means ± s.e.m., n = 4. (E) Macrophages were treated for 24 h with CL097, ssRNA40, and rapamycin in the presence of 45 nmol/L or 100 nmol/L 25D3 and lysed. Immunoblots of LC3B isoforms and SQSTM1 using antibody to LC3B, SQSTM1 or β-actin. Bar charts are reported as mean ± s.e.m., n = 4. * p<0.05.
Highlighted TALEN-TBC1D15-L and TALEN-TBC1D15-R indicate the TALEN binding pair that targets exon 9 of the TBC1D15 gene.Figure 1--figure suppleme
Figure 3(A) CHA19, ENAP1, LBD16 and WRI3 vary in the pathways and the number of genes targeted based on Y1H. (B) Thousands of genes were differentially expressed in Dex-induced GR-TFs compared to GR-TFs under mock conditions. (C) Gene ontologies significantly enriched in the up-regulated DEGs included metabolic pathways in central carbon metabolism. (D) Gene ontologies significantly enriched in the down-regulated DEGs related to central carbon metabolism were found mainly in GR-LBD16. (E) WRI3 were enriched for targets in glycolysis/gluconeogenesis and the TCA cycle. The WRI3 network consists of interactions identified in Y1H and through expression profiling of GR-WRI3. Solid gray lines indicate WRI3-target interactions found in the Y1H network only. Dashed lines signify interactions detected by RNA-Seq only. Colored solid lines indicate interactions identified via Y1H and RNA-Seq. Lines are colored by whether GR-WRI3 up-regulates (blue) or down-regulates (red) target gene expressions.
Figure 7MDA-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. Cell lysates treated in A were analysed by western blot with indicated antibodies (n=3). MDA-MB-231 cells were treated with Olaparib (10 μM) combined with ADR (0.5 μM), CPT (0.5 μM) or VP16 (10 μM) for 72 h, and CCK8 activity of the cells was detected (n=3). Results represent mean ± SEM of three experiments and Error bars indicate SEM. N.S., not significant; **P < 0.01, Student's t test. MDA-MB-231 cells were treated with Olaparib (10 μM) combined with ADR (0.5 μM) for 72 h, and their lysates were subjected to IP using anti-BRD7 antibodies and analysed by Western blot with indicated antibodies (n=3). MDA-MB-231 cells were transfected twice with BRD7 siRNAs, and these cells were subjected to CCK8 assay after treatment with Olaparib (10 μM) combined with either ADR (0.5 μM) or VP16 (10 μM) for 72 h (n=3). Results represent mean ± SEM of three experiments and Error bars indicate SEM. N.S., not significant; **P < 0.01, Student's t test. Resistance of BRD7-depleted cells to Olaparib was reversed by the expression of wild-type BRD7 but not mutant BRD7. shRNA-resistant BRD7 wild-type and BRD7 mutant plasmids were transduced into BRD7-depleted MDA-MB-231 cells, and cell viability was determined as indicated above (n=3). Results represent mean ± SEM of three experiments and Error bars indicate SEM. N.S., not significant; **P < 0.01, Student's t test.
Figure 5A-F. Endocytic activity of R522 and P522 M-MOP and microglia was assessed using FITC soluble Aβ1-42 oligomers and pHrodo Red Dexran (10,000 MW). Endocytosis arbitrary units (A.U) describes the amount of bioparticle cellular fluorescence emission at each time point. Aβ1-42 oligomer uptake in M-MOP (A), primary microglia (B), and hIPSC-derived microglia (C). Dextran uptake in M-MOP (D), primary microglia (E) and hIPSC-derived microglia (F). For hIPSC-derived microglia, 3 isogenic P522 and 3 isogenic R522 clones were examined with at least 6 wells in 3 independent experiments. All microglia and M-MOP data shows the mean±SD of 3 independent experiments were analysed by two-way ANOVA using Sidak's multiple comparison test. *=p<0.05, **=p<0.01, ***=p<0.001. (blue: P522; red: R522)
Fig.Figure 7Number of Us11-GFP expressing neurons at 3 days post-LY294002-induced reactivation in P6 SCG neuronal cultures transduced with shRNA targeting PML at 1 day post-infection with HSV-1 in the presence or absence of IFNα (150 IU/ml). n=9 biological replicates. Statistical comparisons were made using a Mann-Whitney test (ns not significant, * P<0.05). Quantification of colocalization of vDNA foci detected by click chemistry to PML in primary SCG neurons transduced with shRNA targeting PML at 1 day post-infection with HSV-1EdC in the presence or absence of IFNα (150 IU/ml). Each point represents the percentage of 20 vDNA foci that colocalized to PML from 3 biological replicates. Number of Us11-GFP expressing neurons at 3 days post- reactivation in P6 SCG neuronal cultures transduced with shRNA targeting PML at 1 day post-infection with HSV-1 in the presence of IFNα (150 IU/ml). Reactivation was induced by LY294002 in the presence of the DLK inhibitor GNE-3511 (4μM). n=9 biological replicates.
Fig.Figure 1(A) Detection of VgpA in supernatants from indicated strains by immunoblotting (IB) with anti-VgpA antisera. (B) T3SS2 translocation of VgpA into Caco-2 cells using CyaA-based assay; VopV is a known T3SS2 effector. Error bars represent mean ± standard deviation (n=3 biologically independent experiments). (C) Localization of translocated VgpA (upper panel) and VgpAL10A (lower panel) during infection.
| Multi-Omics Factor Analysis: model overview and downstream analyses.Figure 1 (a) Model overview: MOFA takes an arbitrary number of M data matrices as input (Y1,..., YM), one or more from each data modality, with co-occurrent samples but features that are in general unrelated and that differ in numbers. MOFA decomposes these matrices into a matrix of factors, Z, for each sample and M weight matrices, one for each data modality (loadings W1,.., WM). White cells in the weight matrices correspond to zeros, i.e. inactive features, whereas the cross symbol in the data matrices denote missing values (b) The fitted MOFA model can be queried for different downstream analyses, including (i) variance decomposition, assessing the proportion of variance explained by each factor in each data modality, (ii) semi-automated factor annotation based on the inspection of loadings and gene set enrichment analysis, (iii) visualization of the samples in the factor space and (iv) imputation of missing values, including missing assays
Figure 3(A Dissociation constants for the K+ sites in EAAT1 (A) from unguided MD simulations. Values are calculated by dividing off rates by second-order rate constants of ion binding. Mean ± SD of 1000 bootstrap samples.. B) Dissociation constants for the K+ sites in GltPh (B) from unguided MD simulations. Values are calculated by dividing off rates by second-order rate constants of ion binding. Mean ± SD of 1000 bootstrap samples.. (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. (D) Mean current responses to rapid switching from 140 mM CholGluc to 140 mM KGluc of HEK293T cells expressing WT EAAT1 (n=3) or WT EAAT2 (n=3). Cells were intracellularly dialyzed with a 115mM KGluc-based solution and held at 0 mV. (E) Simulated charge displacements associated with K+ binding to outward, transporter translocation, and K+ dissociation from inward-facing GltPh in units of e0; SDs determined by bootstrap sampling range from 7.10-3- 8·10-3 e0.