EMBO/sd-character-level-ner huggingface dataset:
Figure 6A. Survival of Swiss-Webster mice following intravenous administration of purified recombinant toxins: LukED (10 μg per subunit), HlgAB (5 μg per subunit), HlgCB (35 μg per subunit) and LukSF-PV/PVL (35 μg per subunit). Buffer: n = 2 mice; LukED: n = 3 mice; HlgAB: n = 3 mice; HlgCB: n = 2 mice; PVL: n = 3. A representative of two independent experiments is shown. B. Survival of Swiss-Webster mice following prophylactic intravenous administration of dominant-negative mutants, followed by intravenous challenge with a lethal dose of WT LukED. Emut1Dmut = LukEmut1LukDmut, Amut1Bmut = HlgAmut1HlgBmut, Cmut1Bmut = HlgCmut1HlgBmut, Smut1Fmut = LukS-PVmut1LukF-PVmut. n = 6 mice per treatment. ***p = 0.0005 for Buffer vs Emut1Dmut, Buffer vs Amut1Bmut, and Buffer vs Cmut1Bmut. p = 0.1050 (not significant, ns) for Buffer vs Smut1Fmut. Statistics performed using Log-rank (Mantel-Cox) test. A representative of two independent experiments is shown. C. Survival of Swiss-Webster mice following prophylactic intravenous administration of dominant-negative mutants, followed by intravenous challenge with a lethal dose of WT HlgAB. Emut1Dmut = LukEmut1LukDmut, Amut1Bmut = HlgAmut1HlgBmut, Cmut1Bmut = HlgCmut1HlgBmut, Smut1Fmut = LukS-PVmut1LukF-PVmut. n = 6 mice per treatment. ***p = 0.0004 for Buffer vs Emut1Dmut and Buffer vs Amut1Bmut. **p = 0.0015 for Buffer vs Cmut1Bmut. p = 0.7955 (not significant, ns) for Buffer vs Smut1Fmut. Statistics performed using Log-rank (Mantel-Cox) test. A representative of two independent experiments is shown. D. Bacterial burden 96 hours post infection in the liver of mice infected systemically with ~1.5 x 107 CFU of S. aureus strain Newman and intravenously treated with buffer or LukEmut1LukDmut (Emut1Dmut, 10 μg per subunit) at t = 0, 24, and 48 h.p.i. Bars indicate Mean ± SEM, with n = 20 mice (Buffer) and n = 19 mice (Emut1Dmut2) combined from two independent experiments. *p = 0.0303 by unpaired, parametric, two-tailed Student's t-test.
Clearance of senescent cells prevents glomerulosclerosis, podocyte loss and PAI-1 overexpression during aging.Figure 4A: Morphology (PAS staining, upper panels) and quantification of glomerular lesions (lower panel) of kidneys from 28-month-old p16 INK-ATTAC mice treated with either vehicle (AP-) or AP20187 (AP+). Original magnification X400. Scale bar = 20 μm. n = 5. B: WT1 immunostaining (upper panels) and quantification of WT1-positive glomerular cells (lower panel) in 28-month-old p16 INK-ATTAC mice treated with either vehicle (AP-) or AP20187 (AP+). Original magnification X400. Scale bar = 20 μm. n = 5. C: PAI-1 immunostaining (upper panels) and quantification of PAI-1-positive glomeruli (lower panel) in 28-month-old p16 INK-ATTAC mice treated with either vehicle (AP-) or AP20187 (AP+). Original magnification X400. Scale bar = 20 μm. n = 5. Data information: Data are means ± SEM. Statistical analysis: t-student test: AP+ versus AP- mice.
Three-dimensional structure of 2019-nCoV Mpro, in two different views.Fig. 1: Three-dimensional structure of 2019-nCoV Mpro, in two different views. One protomer of the dimer is shown in light blue, the other one in orange. Amino-acid residues of the catalytic site are indicated as yellow and blue spheres, for Cys145 and His41, respectively. Black spheres indicate the positions of Ala285 of each of the two domains III (see text). Chain termini are labeled N and C for molecule A (light blue) and N* and C* for molecule B (orange).
Figure 6(A, B) Binding activity of the full-length trimeric Spike coupled to an AFM cantilever tip to the surface of Vero E6 cells in the presence of (A) hCLEC4G and hCD209 or (B) mouse Clec4g or murine CD209c, probed at the indicated concentrations in SMFS experiments. A definite decrease in binding was observed at concentrations of 37 - 75 nM for Clec4g and 33 - 71 nM for hCLEC4G. (C) Infectivity of mouse Clec4g, CD209c, Asgr1 and hCLEC4G pre-treated SARS-CoV-2 virus in Vero E6 cells. Data represent two to three independent experiments for Clec4g, CD209c and hCLEC4G lectins (N=5-9) and, one experiment for Asgr1 (N=2-4). (D) Infectivity of mouse Clec4g, CD209c, Asgr1 and hCLEC4G pre-treated SARS-CoV-2 virus in human airway epithelial Calu3 cells measured in triplicates and compared to non-treated SARS-CoV-2 infected Calu3 cells measured in quadruplicates (technical replicates). : (C) and (D) Viral RNA was measured with qRT-PCR 15 hours after infection with 0.02 MOI (103 PFUs) of SARS-CoV-2. Data are presented as fold changes of viral loads over mock (only SARS-CoV-2 was added) treated controls (mean log10 values ± SD). All no virus controls were negative for SARS-CoV-2.
Figure 3Heatmap showing clustering analysis of 1,247 single cells, featuring 100 most variable genes (FDR<0.05). Single-cell RNA-seq results are obtained from 2 mice per group (1 female and 1 male each). 2D-tSNE representation of all single cells included in the study (n = 1,247) depicting the separation of microglia isolated from LPS-injected mice (770 cells in red) and steady state (477 cells in blue) in 2 main clusters. Expression of specific homeostatic (Tmem119, P2ry12, Siglech) and inflammatory (Ccl2, Gpr84) genes overlaid on the 2D-tSNE space. Bars represent log2 (Count+1). Representative multicolour flow cytometry analysis of 2 independent experiments showing TMEM119 and P2RY12 expression levels in CD11b+CD45int microglia of saline or LPS-injected mouse brains. For the unconjugated TMEM119 antibody, negative stands for primary antibody without secondary antibody. For P2RY12 antibody, negative represents isotype PE control.
Disruption of LRRC8A, but not of LRRC8D, abolishes ICl,vol and blocks volume regulation.Figure 3A, BVRAC currents (ICl,vol) of the HAP1 (A) and KBM7 (B) haploid cell lines. Left panels, example current traces of ICl,vol fully activated by hypotonic cell swelling measured with the voltage‐clamp protocol shown in (A). Dashed lines indicate zero current. Right panels, averaged current/voltage relationships of maximally activated ICl,vol. Consistent with VRAC currents, they needed hypotonic swelling for activation, displayed an I− > Cl− permeability sequence, and were blocked by DCPIB (Appendix Fig S2A-H). The difference in current inactivation between HAP1 and KBM7 cells can be explained by the fact that KBM7 cells hardly express LRRC8E (Fig EV1) which accelerates VRAC inactivation (Voss et al, 2014). At potentials > +100 mV, also KBM7 currents inactivated (Appendix Fig S2I). Data are presented as mean ± SEM; n = 5-10.C CDependence of regulatory volume decrease (RVD) of HEK cells on LRRC8 genes. Cells were exposed to hypotonic medium starting at t = 0, and intracellular calcein fluorescence was followed over ˜1 h as semiquantitative measure of cell volume. Data are presented as mean values ± SEM from sixteen wells.
Figure 7Northern blotting of mascRNA and 5S rRNA in the lysates of HEK cells before and after FBS starvation. Northern blotting of mascRNA and 5S rRNA in the lysates of human embryonic stem cells on different days of differentiation. Soft agar colony formation assay of HEK, HeLa or MCF-7 cells overexpressing mascRNA or expressing the the scrambled RNA.   Quantification of stained colones in Panel C.   Soft agar colony formation assay of HEK cells overexpressing mascRNA, or expressing the mutants (Anti-Mut1 and Mut1) or the the scrambled RNA (Scramble). Soft agar colony formation assay of QARS (shQARS) or GARS (shGARS) knockdown cells and the control cells stably overexpressing mascRNA (mascRNA) or expressing the the scrambled RNA (Scramble). Soft agar colony formation assay of MALAT1 knockdown cells (shMALAT1) and the control cells (Control) with stable QARS (QARS) or GARS (GARS) overexpression or without (PQ).
Figure 6CHD8 subnetwork analysis in relationship to brain size and downstream CHD8-knockdown effects in vitro. A) CHD8 subnetwork analysis included all CHD8-targets from the Hc network. Network legend is the same of Figure 5: grey (downstream CHD8 target), cyan (upstream regulatory gene of brain-size relevant genes altered in ASD), red (differentially expressed in blood), green (cyan and red), diamond shape (brain size relevant gene altered in ASD), black circle (differentially expressed in ASD cortex from Voineagu et al, 2011). . B) Association analysis of gene expression levels with TBV measures in ASD and control toddlers. See permutation analysis in Appendix Table S6. ** p<0.001 C) Pathway analysis in Metacore. FDR<0.05.
PELP1, TEX10, WDR18 and MDN1 are involved in ribosome biogenesis.Figure 4(A) Diagram summarizing the main steps of rRNA processing. (B) HeLa cells were transfected with siRNA duplexes targeting PELP1, TEX10, WDR18, MDN1 or a control siRNA as indicated. Downregulation of the respective proteins was analysed by western blotting as indicated (WB, three lower panels). At 72 h after transfection, cells were pulse labelled with 32P‐orthophosphate for 1 h and chased for 2.5 h. RNA was separated on a denaturing agarose gel and detected by ethidium bromide (EtBr) staining and autoradiography after drying of the gel. (C) The signal intensities of the 28S and 32S rRNA forms were quantified by phosphoimager analysis and the ratio of 28S:32S rRNA was calculated. Values represent the average of four (two for siMDN1) independent experiments. Error bars indicate s.d. (D) PELP1 is associated with ITS2‐containing RNA species. Endogenous PELP1 was immunoprecipitated from HeLa cells. In total, 10% of the immunoprecipitated material was analysed by western blotting (lower panel). The remaining material was used for RNA isolation and cDNA synthesis. The cDNA was used as template for PCR amplification using primers within the ITS2/28S region or GAPDH as a control. Input corresponds to 10% of the cell lysate. Vertical line in the upper panel indicates removal of irrelevant adjacent lanes. (E) Pre‐ribosomal particles were separated by sucrose gradient centrifugation of nuclear extracts. Proteins were recovered from the fractions by TCA precipitation and further analysed by SDS-PAGE and western blotting using antibodies as indicated. Vertical lines solely indicate loading onto two separate gels. (F) Nucleolar export of pre‐60S particles is strongly compromised in cells depleted of PELP1, TEX10, WDR18, MDN1 or SENP3. HeLa cells expressing YFP‐RpL27 from a tetracycline‐inducible promoter were transfected with the indicated siRNAs. After 24 h, expression of YFP‐RpL27 was induced and 72 h later the localization of RpL27 was monitored. Pictures were taken using identical exposure times.
mediated acetylation of H4K14 mediates the repression of a set of genes with BAZ2A-bound promoter A.Figure 2A. Scatter plot showing the expression of EP300 and BAZ2A in two large cohorts of metastatic and primary PCa. B. EP300 acetylates H3K14 in PC3 cells. Western blot showing H3K14ac levels in PC3 cells treated for three days without and with the selective catalytic EP300 inhibitor A-485 (10 μM). Histone H3 serves as protein loading control. C. Genomic annotations of BAZ2A-bound regions in PC3 cells. D. Volcano plot showing fold change (log2 values) in transcript levels of PC3 cells upon BAZ2A knockdown. Gene expression values of two replicates were averaged and selected for 1.5 fold changes and P <0.05. E. Table showing expression changes of genes with BAZ2A-bound promoter upregulated upon BAZ2A-KD in PC3 cells in metastatic PCa vs. normal prostate tissue and in PC3 cells depleted of EP300 via siRNA. F. qRT-PCR showing increased expression levels of genes with BAZ2A-bound promoter and downregulated in metastatic tumors (AOX1, MANB2, DMPK, and DDB1 in PC3 cells treated with siRNA-EP300. Data were from three independent experiments. Values were normalized to GAPDH mRNA. Statistical significance (P-values) was calculated using two-tailed t-test (*<0.05, ****<0.0001). Error bars represent SD.
NPC2 inhibits IMC secretion of CCL6.Figure 8Immunoblot (left) and ELISA (right, n = 3) analyses of CCL6 in IMC-CM from primary Npc2+/+ IMCs cultured with or without 50 μg/ml bNPC2 for 48 h (immunoblots) or 72 h (ELISA). The data in the bar graph (ELISA) represent mean + SD. Quantification of secreted CCL6 into IMC-CM during the initial 24 h (0-24 h) and subsequent 48 h (24-72 h) of IMC culture by ELISA (n = 3, mean + SD). Immunoblot and qRT-PCR analyses of CCL6 expression in Npc2+/+ IMCs cultured with or without 50 μg/ml bNPC2 for 48 h. The data in the bar graph (qRT-PCR) represent mean + SD; n = 3. CLSM imaging of CCL6 (green)/LAMP1 (red) immunofluorescence of Npc2+/+ IMCs cultured with or without 50 μg/ml bNPC2 for 48 h. Scale bars, 10 μm. The right bottom image highlights the boxed area of the merged image of bNPC2-treated IMCs showing partial co-localisation of LAMP1 and CCL6. Immunoblot analysis of intracellular and secreted (CM) CCL6 in bafilomycin-treated Npc2+/+ IMC culture. IMCs without (left) or with (right) 50 μg/ml bNPC2 pre-loading for 30 min were chased in serum-free DMEM for 3 h and then treated with 200 nM bafilomycin A1 for 24 h.
Generation of Nrl.Figure 7a, Schematic representation of Nrl.Cre+/-/TdTomato+/- floxed chimera generation via morula aggregation, which results in chimeric mosaicism in the neural retina. Recombination and expression of TdTomato is only possible if Nrl.Cre+/- photoreceptors exchange material (Cre) with TdTomatofloxed cells via intercellular communication; b, Representative tile scan of un-injected Nrl.Cre +/-/TdTomato+/- chimera showing endogenous recombination (white); ROIs, as indicated with dashed or dot boxes; Scale bars = 50 µm; c & d, ROI from un-injected Nrl.Cre +/-/TdTomato+/- chimera showing spontaneous recombination presenting in mosaic stripe patterning; Scale bar = 50 µm in c and 10 µm in d; e & f, Representative images of Nrl.Cre+/-/TdTomato+/- chimera after receiving a subretinal injection of AAV-Nrl.Cre. The injected eyes show widespread recombination, which also shows the anticipated mosaic stripe patterning; Scale bar = 50 µm in (e) and 10 µm in (f); g, Quantification of number of recombined cells in un-injected (29,825 cells ± 2031) and AAV-Nrl.Cre-injected (131,562 ± 8478) Nrl-Cre+/-/TdTomato+/- chimeras, compared with un-injected Nrl-Cre-/-(wt)/TdTomato+/- chimera (0 ± 0). Graph shows mean ± SD
USP9X stabilizes XIAP to antagonize mitotic cell death independent of MCL1.Figure 2A Immunoblot analysis of WT, Xiap-/-, Mcl1-/- or cIap2-/- mouse embryonic fibroblasts that were lentivirally infected with shRNA constructs directed against a non-relevant mRNA (Ctrl) or against Usp9X mRNA and treated with taxol as specified. B Two-dimensional cell-cycle analysis (BrdU/PI) of cells described in (A). Sub G1 and G2/M fractions of cells were quantified and averaged with two additional, independent experiments (n=3, ± SD; WT: **, P = 0.0016; ***, P = 0.0004; Mcl1-/-: ***, P = 0.0003; **, P = 0.0017; cIap2-/-: **, P = 0.0043, Student's t test). Black bars exemplify shRNA Ctrl, white bars shRNA Usp9X samples. C Immunoblot analysis of HeLa cells transfected with a FLAG-tagged USP9X expression construct or empty vector (EV) and treated with taxol for the indicated times. D Immunoblot analysis of HeLa cells that were transfected with a FLAG-tagged XIAP expression construct or empty vector and treated as in (C). E Two dimensional cell-cycle analysis (BrdU/PI) of cells described in (C) and (D). Apoptotic indices represent ratios of sub G1 to G1/S cells and are shown for analyses at the indicated timepoints. **, P = 0.0020; ***, P = 0.0004; Student's t test. F Immunoblot analysis of HeLa cells that were transfected with siRNA oligonucleotides directed against a non-relevant mRNA (Ctrl) or against Xiap mRNA and treated with taxol as specified.
| Application of MOFA to a single-cell multi-omics study.Figure 5 (a) Study overview and data types. Data modalities are shown in different rows (D = number of features) and samples in columns, with missing samples shown using grey bars (b) Fraction of the total variance explained (R2) by individual factors for each data modalit (c) cumulative proportion of total variance explained (d) Absolute loadings of Factor 1 (bottom) and Factor 2 (top) in the mRNA data. Labeled genes in Factor 1 are known markers of pluripotency (Mohammed et al, 2017) and genes labeled in Factor 2 are known differentiation markers (Fuchs, 1988) (e) Scatterplot of Factors 1 and 2. Colors denote culture conditions. The grey arrow illustrates the differentiation trajectory from naive pluripotent cells via primed pluripotent cells to differentiated cells
Figure 9Four-day-old seedlings of 35S:ARR12:YFP, 35S:ARR12S323A:YFP, and 35S:ARR12S323D:YFP were transferred to 1/2 MS medium supplemented with or without 50 mM or 75 mM NaCl. After three days of salt treatment, plant phenotypes were determined. Four-day-old seedlings of 35S:ARR12:YFP, 35S:ARR12S323A:YFP, and 35S:ARR12S323D:YFP were transferred to 1/2 MS medium supplemented with or without 50 mM or 75 mM NaCl. After three days of salt treatment, relative primary root lengths (B) were determined. After ten days of salt treatment, changes in survival rates (C) and fresh weights (D) were examined. Scale Bar, 1 cm.
Figure 2Principle of 5-azadC-induced DNMT1 DPC formation. See main text for details. HeLa cells were transfected with non-targeting control (CTRL) or DNMT1 siRNAs, exposed or not to 5-azadC and collected 2 h later. Chromatin-enriched fractions were immunoblotted with antibodies to SUMO2/3, DNMT1 and MCM6 (loading control). Representative images of HeLa cells that were treated and co-immunostained with antibodies to DNMT1, SUMO2/3 and PCNA. Scale bar, 10 μm. Proliferative capacity of HeLa/GFP-DNMT1 cells transfected with indicated siRNAs and exposed to the indicated 5-azadC doses for 2 h was assayed by measuring cell proliferation with the SRB assay (mean±SEM; n=3 independent experiments; ***p<0.001, Student's t-test). Extracts of HeLa/GFP-DNMT1 cells treated with 5-azadC for the indicated times were subjected to GFP immunoprecipitation (IP) under denaturing conditions followed by immunoblotting with SUMO2/3 and GFP antibodies. HeLa cells were subjected to consecutive rounds of transfection with DNMT1 siRNA targeting the UTR and expression plasmid encoding WT or catalytically inactive (CI) GFP-DNMT1. Cells were then left untreated or incubated with 5-azadC for 2 h, and SUMOylation of GFP-DNMT1 was analyzed Asterisk denotes unmodified GFP-DNMT1 recognized by the SUMO2/3 antibody due to weak cross-reactivity. Experimental set-up for global proteomic analysis of 5-azadC-induced SUMOylation changes. Mass spectrometry analysis of 5-azadC-induced SUMOylation changes. His10-SUMO2 conjugates from HeLa/His10-SUMO2 cells subjected or not to 5-azadC were purified on Ni-NTA under stringent conditions and analyzed by mass spectrometry. Proteins displaying significantly altered SUMOylation in response to 5-azadC treatment relative to the control condition were visualized through two-sample t-testing using permutation-based FDR to achieve q-values of <0.05 SUMO target proteins displaying altered SUMOylation upon DNMT1 knockdown are shown. Both control (CTRL) and DNMT1 siRNA-transfected cells were treated with 5-azadC.
regulates the production of inflammasome related cytokines in AECs (A-D) ELISA of IL-1β, IL-18, IFN-β and IL-6 in supernatant of AEC cells, which were stimulated for 16 h with RNA virus reovirus (MOI=5) or DNA virus HSV-1 (MOI=5).Figure 2(A-D) ELISA of IL-1β, IL-18, IFN-β and IL-6 in supernatant of AEC cells, which were stimulated for 16 h with RNA virus reovirus (MOI=5) or DNA virus HSV-1 (MOI=5). n = 3 primary samples. Data information: Each symbol represents an independent experiment; small horizontal lines indicate the average of triplicates. Mock indicated unstimulated cells. **P <0.01 and ***P<0.001 (unpaired t test). NS means no statistical difference. Data represent three independent biological replicates. (E) Immunoblot analysis of IL-1β and caspase-1 in AECs cell lysate or cell supernatant, β-actin served as a loading control throughout. Data information: Each symbol represents an independent experiment; small horizontal lines indicate the average of triplicates. Mock indicated unstimulated cells. **P <0.01 and ***P<0.001 (unpaired t test). NS means no statistical difference. Data represent three independent biological replicates.
a) COS-7 cells transfected with EGFP-LC3 construct for 4 h were treated with DMSO (control), 0.figf2(a) COS-7 cells transfected with EGFP-LC3 construct for 4 h were treated with DMSO (control), 0.2 μM rapamycin (positive control), 47 μM SMER10, 43 μM SMER18 or 47 μM SMER28 for 16 h, and analyzed by fluorescence microscopy. The effects of treatment on the percentage of EGFP-positive cells with >5 EGFP-LC3 vesicles are shown. Error bars denote s.e.m. P 0.0001 (all SMERs). (b) HeLa cells stably expressing EGFP-LC3 were treated with DMSO (control), 47 μM SMER10, 43 μM SMER18 or 47 μM SMER28 for 24 h. Confocal sections show cells containing EGFP-positive autophagic vesicles. Nuclei are stained with DAPI. Bar, 20 μM. (c) HeLa cells stably expressing EGFP-LC3 were treated for 4 h with DMSO (control) or 200 nM bafilomycin A1 (Baf), or with 200 nM bafilomycin A1 and 47 μM SMER10, 43 μM SMER18 or 47 μM SMER28. Cells were left untreated or pretreated with SMERs for 24 h before adding bafilomycin A1. Levels of EGFP-LC3-II were determined by immunoblotting with antibody against EGFP (above) and densitometry analysis relative to actin (below). Error bars denote s.e.m. P = 0.0259 (baf), P 0.0001 (SMER10), P = 0.0003 (SMER18 and SMER28) versus control; P = 0.0025 (SMER10), P = 0.0218 (SMER18), P = 0.0195 (SMER28) versus bafilomycin A1. ***P 0.001; **P 0.01; *P 0.05.
Figure 3(A) Immunofluorescence staining of FTCD and GM130, a Golgi marker. HepG2 cells were fixed, permeabilized, stained with a monoclonal antibody to FTCD, a polyclonal antibody to GM130 and DAPI, and observed by confocal microscopy. Scale bar = 10 μm. (B) Depletion of FTCD by FTCD siRNA duplexes. HepG2 cells were either mock transfected with water or transfected with two distinct siRNA duplexes specific to FTCD. After incubation for 48 hrs, the cells were analyzed by Western blotting with antibodies to FTCD and α-tubulin. (C) The Golgi in cells which were depleted of either FTCD or p47. HepG2 cells, which were treated with either FTCD siRNAs, p47 siRNA or mock for 48 hrs, were fixed and stained with a monoclonal antibody to GM130. Scale bar = 10 μm. (D) The results of quantification of (C). The values are from five sets of independent experiments. Results are expressed as the mean±SD (n=5), with 100 cells counted in each group of each set. Asterisks indicate a significant difference at P<0.01 compared with the mock-treated cells (Bonferroni method). (E) Representative EM images of the Golgi in HepG2 cells treated with either mock or FTCD siRNAs for 42 hrs. Scale bar = 0.5 μm. (F) The results of quantification of (E). Golgi membranes in HepG2 cells were classified into cisternae, vesicles and tubules, and counted as previously described (Shorter & Warren, 1999). The results are shown as the mean±SD of five sets of independent experiments, with 15-20 cells counted in each group in each independent experiment. An asterisk indicates a significant difference (P<0.05) compared with the mock-treated cells in each category (Bonferroni method).