EMBO/sd-character-level-ner huggingface dataset:
EK1C4 broadly and potently inhibited cell-cell fusion and PsV infection mediated by S protein of divergent HCoVs.Fig. 5.HCoVs.a to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of SARS-CoV (a) HCoVs.a to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of MERS-CoV (b) HCoVs.a to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of HCoV-OC43 (c) HCoVs.a to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of HCoV-229E (d) to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of HCoV-NL63 (e) to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of WIV1 (f) to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of Rs3367 (g) to h. Inhibitory activity of EK1C4 in cell-cell fusion mediated by the S proteins of SHC014 (h). i to o. Inhibitory activity of EK1C4 in PsV infection assays against SARS-CoV (i) to o. Inhibitory activity of EK1C4 in PsV infection assays against MERS-CoV (j) Inhibitory activity of EK1C4 in PsV infection assays against HCoV-OC43(k) Inhibitory activity of EK1C4 in PsV infection assays against HCoV-229E (l) Inhibitory activity of EK1C4 in PsV infection assays against , NL63 (m) Inhibitory activity of EK1C4 in PsV infection assays against WIV1 (n) Inhibitory activity of EK1C4 in PsV infection assays against Rs3367 (o).
bam mutant germ cells recapitulate the separable old versus new H3 distribution in WT GSCs, and Oligopaint IF-FISH reveal distinct old versus new H3 at key genes.Figure 3(A) A cartoon depicting a WT germarium with differentiating germline at different stages (gray) compared with a bam mutant ovariole filled with GSC-like cells (pink) without further differentiated germ cells. Undifferentiated cells can be identified by the presence of dotted spectrosome versus branched fusome structure (purple). (B-C) Non-overlapping patterns of old (green) versus new (red) histone H3 (B) and overlapping patterns for H2A (C) in mitotic prometaphase bam mutant cells marked by anti-H3S10p (gray), similar to WT GSCs. Scale bars: 2μm. (D) Quantification of the overlap between old and new histones in late prophase and prometaphase bam mutant cells using Spearman correlation, also plotted with and compared to the WT GSC values from Fig. 1L. Data gathered over 4 separate experiments. Values are mean + 95% CI. P-value: pairwise ANOVA test with bonferroni correction. ****: P< 0.0001. n.s.: not significant. (E) A cartoon depicting the Oligopaint IF-FISH scheme to identify old versus new histone inheritance at single genomic loci in mitotic cells. (Left) A germarium containing a GSC in prometaphase, where the 3D chromatin structure may disguise local asymmetries at genomic regions of interest. (Top) A linearized gene of interest (GOI) displays local asymmetries. (Bottom) Two anaphase cells depict potential Oligopaint IF-FISH results. Maternal chromosomes are outlined in pink and paternal ones in blue. (Bottom Left) Probes recognize genes (magenta) that change their epigenetic state in a 2:2 ratio with biased old:new H3-enriched regions. In this instance, each duplicated sister chromatid from either maternal or paternal chromosome has an "agreement" on old (green, inherited by GSC indicated by an asterisk) versus new (red, inherited by the CB) H3. (Bottom Right) Probes recognize genes (cyan) that maintain their epigenetic state associated with more symmetric regions (yellow) that do not have a strong bias for either old or new H3. (F-H) Examples showing old (green) versus new (red) H3 distribution patterns and percentage of signal strength at a single genomic locus labeled with fluorescent probes (gray) for dad gene (F), bgcn gene (G), and ss gene (H) in bam mutant GSC-like cells at prometaphase marked by anti-H3S10p (blue). Scale bars: 1μm. Zoomed-in images of each probe are displayed to the right of each figure panel. Scale bars: 0.1μm. (I) Examples of scatter plots showing probe's association with normalized old H3 (green, above X=Y line) versus new H3 (red, below X=Y line) and the (i):(ii):(iii) ratios that result from them, where (i) is the number of FISH signals more associated with old histone, (ii) is the number of FISH signals that had equal association with both old and new histone, and (iii) is the number of FISH signals more associated with new histone. (J) Quantification of the probe association ratios with old versus new H3 for each candidate gene in bam mutant germ cells. Data gathered over 4 separate experiments for dad, 3 for bgcn, and 2 for ss.
Fig.Figure 6(A and B) Immunostaining assay of SOX1 (red) in control, V1-PAKI and SOX1KO cells on day 40 of dorsal (A) and ventral (B) organoids differentiation. Scale bar, 100 μm. (C) Immunostaining assay of MAP2 (green) in control, V1-PAKI and SOX1KO cells on day 40 of dorsal organoids differentiation. Scale bar, 100 μm. (D) Immunostaining assay of GABA (green) in control, V1-PAKI and SOX1KO cells on day 40 of ventral organoids differentiation. Scale bar, 100 μm. (E and F) Immunostaining assay of SOX1 (red) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of dorsal (E) and ventral (F) organoids differentiation. Scale bar, 100 μm. (G) Immunostaining assay of MAP2 (green) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of dorsal organoids differentiation. Scale bar, 100 μm. (H) Immunostaining assay of GABA (green) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of ventral organoids differentiation. Scale bar, 100 μm. (I) Summary of the HDAC10/SOX1-OT V1-SOX1-ASCL1 regulatory axis in hESC neuronal differentiation.
Figure 3A. Immunofluorescence images and quantifications of KT-localized Plk1 in U2OS cells after PBIP1 depletion. Cells were either Mock transfected or with PBIP1 siRNA for 72 hours. Cells were treated with nocodazole for 1 hour and fixed. Cells were stained with the centromere marker ACA, Plk1 and DAPI. B. Representative images and histograms showing the effect of chromosome position on cortical LGN enrichment after 72 hours of Mock and PBIP1 depletion. C. Live cellimages and histograms of GFP-H2B U2OS cells Mock transfected or with siPBIP1 filmed on rectangular micropatterns in the presence of a CENP-E inhibitor. The spindle angles for both conditions were scored at 32 minutes after NEB. . D. The total spindle angle displacement within the first hour of mitosis was determined by measuring the spindle angle at each time point and calculating the sum of the Δ spindle angle between each time points.
Evaluation of ribophagy on RNA degradation under nitrogen starvationTime‐dependent change in nucleoside levels after nitrogen starvation.Figure 5A Time‐dependent change in nucleoside levels after nitrogen starvation. Wild‐type, atg2∆, ubp3∆, and bre5∆ cells were grown in SD to mid‐log phase and transferred to SD(‐N) at time 0. Metabolites were analyzed by LC/MS as described in . The results are presented as normalized intensities on the basis of the peak height of each metabolite in wild‐type cells. All data are means of triplicates. The error bars represent the standard deviation. B Detection of free RNA within cells. rny1∆, rny1∆atg2∆, rny1∆ubp3∆, and rny1∆bre5∆ cells grown in SD(‐N) for 2 h were stained with FM4‐64 and GR Green and observed by fluorescence microscopy. Scale bar, 5 μm.
Fig.Figure 3The cellular levels of Ca2+ ions after ligand (C10ORF99) stimulation of the indicated GPR15 receptors (human, mouse, Japanese quail GPR15). A plasmid encoding human NTSR1 and the parental pcDNA plasmid were transfected as positive and negative controls, respectively. The fold-change in calcium levels was calculated based on the fluorescence intensity (excitation and emission wavelengths: 490 and 520 nm, respectively). Calcium RLU (relative light unit) fold was calculated using no stimulation data as standard value. The cellular levels of Ca2+ ions after ligand (C10ORF99) stimulation of the indicated GPR15 receptors painted turtle, coelacanth, and Asian bony tongue GPR15). A plasmid encoding human NTSR1 and the parental pcDNA plasmid were transfected as positive and negative controls, respectively. The fold-change in calcium levels was calculated based on the fluorescence intensity (excitation and emission wavelengths: 490 and 520 nm, respectively). Calcium RLU (relative light unit) fold was calculated using no stimulation data as standard value. (h) The cellular levels of Ca2+ ions after ligand (C10ORF99) stimulation of GPR15 in human, mouse, Japanese quail, painted turtle. YM.254890 inhibits Ca2+ ion levels in cells transfected with GPR15 receptors of vertebrates. The fold-change in calcium levels was calculated based on the fluorescence intensity (excitation and emission wavelengths: 490 and 520 nm, respectively). (i, j) The cellular levels of Ca2+ ions after ligand (C10ORF99) stimulation of GPR15 in coelacanth, Asian bony tongue, and their mutants. (k, l) The cellular levels of Ca2+ ions after ligand (C10ORF99) stimulating GPR15 in Japanese quail and its mutants. The fold-change in calcium levels was calculated based on the fluorescence intensity (excitation and emission wavelengths: 490 and 520 nm, respectively).
Model of Munc18-1 and Munc13 in driving SNARE assembly.Figure 9A Autoinhibition of the Munc18-1/Syx1 complex. The closed Syx1 linker region and the bent Munc18-1 domain 3a render this complex inactivate, inhibiting Syx1 association with SN25 and Syb2. B Activation of the Munc18-1/Syx1 complex by Munc13 MUN domain. The MUN domain catalyzes opening of the Syx1 linker region via binding to H11 of domain 3a and Syx1 linker region, leading to a conformational flexibility of domain 3a (between bend and extension). The open Syx1 linker region and putative unstructured N-terminal of Syx1-H3 are displayed by lemon green and blue dashed lines, respectively. C Templating function of domain 3a. Assisted with Munc13 MUN domain, the extended domain 3a serves as a template for Syx1-H3 and Syb2 simultaneously, awaiting SN25 entry and registering SNARE complex assembly at the N-terminus. Dashed lines indicate putative unstructured regions. D SNARE complex assembly. Proceeding of SNARE four-helix bundle inside the central cavity of Munc18-1 to complete SNARE complex formation and membrane fusion.
Distribution of LAMP‐2A in discrete lysosomal membrane regions.Figure 5(A, B) Regions of the lysosomalmembrane resistant to 1% Triton X‐114 recovered from the sucrose gradients were fixed and processed for electron microscopy (A) or visualized without fixation by electron microscopy (B). Arrows point to the membrane bilayer. (C) Intact ratliverlysosomes (top) and DR regions of the lysosomal membranes (bottom) were fixed and subjected to immunogold labeling for LAMP‐2A and LAMP‐2B, and a gold‐conjugated secondary antibody as described under Material and methods. Top right panels show 'clusters' of gold particles (black arrows) in individual lysosomes at higher magnification. Graph shows the labeling density (gold particles per μm) in the indicated fractions for both proteins. Values are mean+s.e. from the quantification of 15 different micrographs. (D) Intact rat liver lysosomes were fixed and subjected to double immunogold labeling for LAMP‐2A and flotillin‐1. Representative lysosomes are shown. Far right panel: zoomed‐in image of the distribution of LAMP‐2A in regions containing (black arrows) and not containing (white arrows) flotillin‐1. Bars: 0.2 μm.
Figure 3 JurkatTat cells were treated with 7 µM of each compound, 50 nM PMA or left untreated (No drug control) for 24 h. A IL-2 expression was measured in culture supernatants using an ELISA. JurkatTat cells were treated with 7 µM of each compound, 50 nM PMA or left untreated (No drug control) for 24 h. B Cell surface CD69 expression was measured by flow cytometry. Data information: For statistical analysis, mean and SE for the results are determined from three biological replicates (n=3). Statistical significance, as determined by ratio paired t test is indicated: ***, P= 0.0008; ns, no significant difference.
Figure 2A. Diagram of k 5 mRNA 3'UTRs and relative positions of the RNA motif shared between transcripts. B. RNA motif over-represented in k 5 mRNA 3'UTRs. C. Frequency of mRNAs within each k-means cluster containing at least 2 of the RNA motif over-represented in k 5 mRNAs. D. Scheme depicts the in vitro MS2 system strategy. CMV promoter-driven expression of MCP-GFPnls and hHBB-24xMS2-tagged RAB13 3'UTR. The visualisation of MCP-GFPnls bound to 24xMS2 allows the identification of the minimal region in the 3'UTR of RAB13 necessary for its localisation. E. Left: representative subconfluent motile cells co-transfected with plasmids expressing Lyn-mCherry, MCP-GFPnls and 24xMS2-RAB13 3'UTR or 24xMS2. Right: percentage of cells with MCP-GFP localised to protrusions when co-transfected with full length or deletion versions of RAB13 3'UTR (n≥3 experiments). white arrowheads indicate non-nuclear localisation of MCP-GFPnls; arrows indicate the orientation of RNA localisation; yellow dashed lines outline cell borders; scale bars = 20 µm For each k 5 mRNA 3'UTR, a diagram of the full length 3'UTR and the positions of the RNA motif is shown together with the respective RNAseq mapped reads from HUVEC protrusions; black arrowheads indicate predicted polyadenylation sites F. Left: representative cells co-transfected with plasmids expressing Lyn-mCherry, MCP-GFPnls and 24xMS2-k 5 3'UTRs. Right: percentage of cells with MCP-GFP localised to protrusions when co-transfected with full length or deletion versions of k 5 3'UTR (n≥3 experiments). white arrowheads indicate non-nuclear localisation of MCP-GFPnls; arrows indicate the orientation of RNA localisation; yellow dashed lines outline cell borders; scale bars = 20 µm For each k 5 mRNA 3'UTR, a diagram of the full length 3'UTR and the positions of the RNA motif is shown together with the respective RNAseq mapped reads from HUVEC protrusions; black arrowheads indicate predicted polyadenylation sites
Transcriptome signatures of low- and fast-migrating epithelial cells.Figure 3Protein-coding gene assignment to Low and High Migration Signature sets according to differential expression features. Heatmap of unsupervised hierarchical clustering based on Low Migration Signature (LMS) and High Migration Signature (HMS) gene expression. Heatmap of enrichment scores (-log10(P-value)) for KEGG pathways significantly enriched for LMS and HMS protein-coding genes (DAVID, P-value < 0.05). LMS (red) and HMS (black) gene expression quantification in Epi cells by random-primed RT-qPCR relative to RPL11 and represented as log2 fold change to CTR. Each dot corresponds to an independent replicate, bars represent Mean ± Standard Deviation values.
Nuclear clusters can form in absence of integration and contain incoming vRNA (A) Image of a ThP1 cell infected with an integration-deficient HIV-1 carrying the mutation D116A in the catalytic site of IN.Figure 4(A) Image of a ThP1 cell infected with an integration-deficient HIV-1 carrying the mutation D116A in the catalytic site of IN. The EdU-labeled vDNA is shown in red and the vRNA detected by RNA-FISH in green. The nucleus (DAPI) is shown in blue. See Appendix Fig. S12 for a larger image region. (B) Venn diagram shows the number of vRNA clusters, the number of vDNA clusters, and the number of vRNA clusters colocalizing with vDNA clusters. The p-value indicates the significance of colocalization based on a jittering analysis (see Methods). (C) Scatter plot shows vDNA and vRNA intensities in colocalizing vDNA and vRNA clusters with the Spearman correlation r and associated p-value. (D) ThP1 cells were infected with HIV-1 at an MOI of 20, in absence (left) or presence (right) of nevirapine (NVP, 10 µM). Blue: DAPI staining. Green: vRNA (RNA-FISH). Red: vDNA (EdU). (E) Scatter plots show intensities of vDNA and vRNA in detected vRNA clusters. (F) Image of a ThP1 cell infected by HIV-1 with EU labeled RNA in absence (left) or presence (right) of nevirapine (NVP, 10 µM). Blue: DAPI staining. Green: vRNA (RNA-FISH). Grey: EU-labeled vRNA. See Appendix Fig. S19 for a larger image region. (G) Intensity profiles of EU and RNA-FISH along the dotted lines in panel F (top).
Loss of respiratory function in atg mutants during nitrogen starvation.Figure 2(A) WT and atg1Δ cells grown in YEPD medium were transferred to SD-N +50 mM MES-KOH (pH 6.2). After 120 hours, cultures were diluted 5.0×105 fold and plated onto YEPD agar. The plates were incubated at 30°C for four days. Insets indicate the plate overlaid with TTC agar to examine the respiratory competency of formed colonies. (B) WT, and atg1Δ cells were nitrogen-starved as (A). Cell viability and respiratory competency was determined by phloxine B staining and TTC overlay technique as (A), respectively. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. (C) WT, atg1Δ, atg2Δ, atg7Δ, atg11Δ, atg15Δ, and atg32Δ cells were nitrogen-starved as (A). Cell viability and respiratory competency was determined by phloxine B staining and TTC overlay technique as (A), respectively. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. These data represent the average of three independent experiments and bars indicate standard deviations.
Fig.Figure 3(A) Scheme showing the principle of the measurement. LmnaG609G/+ VSMCs were incubated in MEM media (containing 0.8 mmol/L magnesium; untreated) or in magnesium-enriched MEM media (containing 1.8 mmol/L magnesium; treated) from passage 1 to passage 8. Then, cells were incubated overnight in MEM containing 0.1% FBS and some cells were fixed, as described in the Methods section. Then, cells were incubated in MEM (containing 0.1% FBS) with 1 or 2 mmol/L phosphate, 1.8 mmol/L magnesium (+Mg), 100 µmol/L pyrophosphate (+PPi), or 500 µmol/L phosphonoformic acid (+PFA). After 7 days of incubation, during which the media were replaced daily, the calcium content was measured as described in the Methods section. (B) Representative time-course of 2 mmol/L phosphate on calcification of LmnaG609G/+ VSMCs (up). Calcification was visualized with Alizarin red. Representative microscopic images (10x; scale bar: 100 µm) showing calcification of treated and untreated LmnaG609G/+ VSMCs (down). (C-F) Measures of calcium in treated living LmnaG609G/+ VSMCs (C), treated fixed LmnaG609G/+ VSMCs (D), untreated living LmnaG609G/+ VSMCs (E), and untreated fixed LmnaG609G/+ VSMCs (F). (G) The calcification inhibitory capacity was calculated as the difference in calcium deposition between living and fixed cells (ΔCa2+).
a) Heart-to-body-weight ratios for 12-week-old miR-212/132 null (KO) and wild-type (WT) mice (n=5–6).f3(a) Heart-to-body-weight ratios for 12-week-old miR-212/132 null (KO) and wild-type (WT) mice (n=5-6). (b) Cardiomyocyte cell size of neonatal miR-212/132 null and WT mice (n=5-6 isolations). (c-f) Heart-to-body-weight ratios (c) in Sham-operated WT mice and miR-212/132 null and WT mice3 weeks after TAC surgery (n=4-7). Scale bar, 50 μm. (c-f) cardiomyocyte diameter (d), cardiac fibrosis (e) in Sham-operated WT mice and miR-212/132 null and WT mice3 weeks after TAC surgery (n=4-7). Scale bar, 50 μm. (c-f) lung wet weight (f) in Sham-operated WT mice and miR-212/132 null and WT mice 3 weeks after TAC surgery (n=4-7). Scale bar, 50 μm. (g-i) Echocardiography analysis of cardiac dimensions and function in Sham-operated WT mice and miR-212/132 null and WT mice 3 weeks after TAC (n=4-11). (g) End-diastolic area, (h) end-systolic area and (i) fractional shortening. All values represent mean±s.e.m. *P0.05; **P0.01; ***P0.005; #P0.05 compared with WT TAC; ###P0.005 compared with WT TAC; PSR, picrosirius red (collagen stain). DAPI, 4′,6-diamidino-2-phenylindole; WGA, wheat germ agglutinin.
Cloning of the APG10 gene and phenotype of Δapg10 cell.Figure 1(A) Wild‐type (YW5‐1B), apg10‐1 mutant (MT91‐4‐2) and Δapg10 (TFD10‐L1) cells were incubated in SD(−N) medium containing 1 mM PMSF at 30°C. After incubation for 6 h, cells were observed under light microscopy (Nomarski images). (B) Wild‐type (closed circles),apg10‐1 mutant (closed triangles) and Δapg10 (open circles) cells were grown to 1 OD600/ml in YPD and then transferred to SD(−N) medium. After incubation at 30°C for the times indicated, their viability was determined by phloxine B staining ( Tsukada and Ohsumi, 1993). (C) Cell lysates from wild‐type cells with empty vector pRS316 (lane 1) or HAAPG12 (lane 2), and also apg10‐1 (lane 3) and Δapg10 cells with HAAPG12 (lane 4) were subjected to Western blotting analysis with anti‐HA antibody (16B12). (D) Amino acid sequence of Apg10p. Cysteine residues are indicated in bold.
Figure 2Time from symptom onset to initial diagnosis (TFSD).    Time from symptom onset to development of pneumonia (TFSP).   Time from symptom onset to treatment (TFST). Time from development of pneumonia to recovery (TFPR).
Parkin-dependent MDVs are targeted to lysosomes independently of autophagyHeLa cells, transfected with siRNA targeting Drp1 and GFP-parkin (green), were treated with 25 µM antimycin A for 90 min following a 30-min pretreatment with 50 nM bafilomycin A1, then fixed and immunostained against PDH E2/E3 bp (red) and TOM20 (blue).Figure 4AHeLa cells, transfected with siRNA targeting Drp1 and GFP‐parkin (green), were treated with 25 μM antimycin A for 90 min following a 30‐min pretreatment with 50 nM bafilomycin A1, then fixed and immunostained against PDH E2/E3 bp (red) and TOM20 (blue). PDH E2/E3 bp‐positive/TOM20‐negative MDVs colocalizing with GFP‐parkin (arrows) or not (circles) are indicated. Scale bar, 30 μm. B Quantification of PDH E2/E3bp‐positive/TOM20‐negative vesicles in HeLa cells treated with antimycin A in the presence of the lysosomal inhibitors bafilomycin A1 or pepstatin A and E‐64d; both total number (white bars) and the number colocalizing with GFP‐parkin (gray bars) are indicated. Bars represent the mean ± s.e.m. P‐values are given for total vesicle number (n = 19 to 120 cells in 2-3 experiments; **P < 0.01; ***P < 0.001). CHeLa cells, transfected with GFP‐parkin (green) and siRNA targeting Drp1 and Atg5, were treated with 25 μM antimycin A for 90 min, then fixed, immunostained for PDH (red) and TOM20 (blue), and counterstained for Hoescht (gray). PDH E2/E3bp bp‐positive/TOM20‐negative MDVs colocalizing with GFP‐parkin (arrows) or not (circles) are indicated. Scale bars, 20 μm. D Quantification of PDH E2/E3 bp‐positive/TOM20‐negative vesicles from HeLa cells transfected with the indicated siRNA; both the total number (white bars) and the number colocalizing with GFP‐parkin (gray bars) are indicated. Error bars represent the mean ± s.e.m. P‐values are given for the total vesicle number (n = 42-64 cells in 2-3 experiments); ns, not significant.