EMBO/sd-character-level-ner huggingface dataset:
Figure 3Total protein levels of G544D Munc18-1. G544D Munc18-1b was expressed in cortical mouse neurons infected with lentiviral vectors expressing cre recombinase in the presence or absence of compounds at 20 µM, or DMSO (vehicle control). Total Munc18-1 levels were analyzed by quantitative immunoblotting 9 days after infections (A), normalized to GAPDH levels (B). Data are means ± SEM (*p < 0.05, ****p < 0.0001 by one-way ANOVA followed by Bonferroni post-hoc test; n = 6 independent experiments; exact p values are shown in Appendix Table S1). Same as in (A, B) except that compounds 9, 10, 11, 13, 14 and 16 were added at 0.25, 1, 5, 20, 100, or 250 µM. Note, that the data for 250 µM were not plotted in (D) because of causing neuronal death. Data are means ± SEM (*p < 0.05, **p < 0.01 by Kruskal-Wallis test with Dunn's multiple comparison test, or one-way ANOVA with Dunnett's post-hoc test; n = 6-15 independent experiments; exact n and p values are shown in Appendix Table S1).
Figure 1A The mRNA expression of FAM189A2/ENTREP in the primary human mammary epithelium HMEC and human breast cancer cell lines. In qRT-PCR analyses, the expression was normalized to the immortalized normal human mammary epithelium HMEC4tertshp16. Relative expression ratios shown as mean + SD from three biological replicates. B Relative expression of FAM189A2/ENTREP in the normal (white) and cancer (green) tissues of breast (Curtis Breast), lung (Okayama Lung), colorectal (TCGA Colorectal) and head and neck (Estilo Head-Neck). Data were downloaded from the Oncomine database P-values obtained by Student's t-tests and the number of cases are listed on the top and bottom of figures, respectively. P < 0.05 was considered as statistically significant. Box-whisker plot represents the interquartile range (25th and 75th percentiles) as a box and the median as a line. The maximn and minimun values within 1.5 x interquartile range are shown as whiskers. Outlier data are plotted as dots. C Relative expression of FAM189A2/ENTREP in the primary (green) and the metastatic sites (blue) including lymph node, bone, liver, lung, soft tissues of prostatic cancer (Grasso Prostate) (left) and in the normal (white), primary (green) and metastatic sites (three cases of sentinel lymph node) (blue) of breast cancer (TCGA Breast) (right). Note that only three cases of the metastatic breast cancer were available in the dataset. Data were downloaded from the Oncomine database P-values obtained by Student's t-tests and the number of cases are listed on the top and bottom of figures, respectively. P < 0.05 was considered as statistically significant. Box-whisker plot represents the interquartile range (25th and 75th percentiles) as a box and the median as a line. The maximn and minimun values within 1.5 x interquartile range are shown as whiskers. Outlier data are plotted as dots. D-F Relapse-free survival (RFS) of breast cancer patients. Data were downloaded from the Kaplan-Meier plotter
Microautophagy of peroxisomes in a pex1-ts mutant (strain SJH242) of P.Figure 11Microautophagy of peroxisomes in a pex1-ts mutant (strain SJH242) of P. pastoris expressing GFP-SKL from the constitutive glyceraldehyde-3-phosphate dehydrogenase (GAPDH) promoter at the restrictive temperature for peroxisome assembly (34°C). (a) Kinetics of peroxisome degradation at 34°C. Strain SJH242 was induced on methanol at 23°C, and glucose adaptation was performed at 34°C. (b) Changes in vacuole morphology were seen even in the absence of normal peroxisomes. At 0 h, SJH242 cells were grown at 34°C when GFP-SKL was diffuse in the cytosol and many vacuoles showed spherical morphologies. After 2 h, ∼20-30% vacuoles showed complicated membrane structures representative of stage 2 cells.
Figure 3A) MA-plot of log2 fold changes (Y-axis) versus the mean of normalized counts of 22G RNAs (X-axis) for pid-2(xf23) mutants, compared to wild type. Red dots: genes with adjusted p-value < 0.05 and fold-change > 2. Blue dots: genes with adjusted p-value < 0.05 and fold-change < -2. B) Heatmap displaying overlap significance between different gene sets and genes that either up- or down-regulated in pid-2(xf23) mutants. Significance was tested with Fisher's exact test and p-values adjusted with the Benjamini-Hochberg Procedure. Colour scheme represents the odds ratio of overlaps representing the strength of association. N.S.: not significant. C-D) Cumulative 22G coverage along the gene body of Mutator and CSR-1 targets. Values represent 22G coverage normalized to the total coverage of each gene, for wild type (N2) and pid-2(xf23) mutants. The lines represent the average of biological replicates, whereas the shading represents the standard deviation of biological replicates. a.u.: arbitrary units.
Figure 1Experimental setup used to screen for chromatin regulators involved in the IR-induced G2 checkpoint response and recovery. Unless stated otherwise, 5 Gy of IR was used. Results of three independent microscopy-based screens in U2OS cells. Knockdown of Luciferase (blue) and the positive controls PPM1D and βTrCP (red) are indicated. A low/high Z-score indicates less/more phospho-histone H3 positive cells compared to the control, reflecting reduced/increased numbers of cells going into mitosis in the respective knockdowns after 5 Gy of IR. Deconvolution of the 41 primary hits in U2OS cells by four separate siRNA oligonucleotides. Grey represents a recovery value that is similar to the Luciferase control (<0.5 SD) and the red represents an altered recovery value (>0.5 SD) in two independent experiments. U2OS cells (D) and RPE1 hTert immortalized primary epithelial cells (E) were transfected with siRNA oligos targeting the hits in a similar setup as described in A), but now analyzed for mitotic entry by MPM2 and PI staining by flow cytometry. For normalization see materials and methods. Error bars represent the SEM of three independent experiments. Graphical explanation of the known interactions between the identified hits through STRING pathway analysis
RABIF associates with RAB13 co-translationally.Figure 8A. Schematic depicting experimental strategy for assessing co-translational association of RABIF with nascent RAB13. B. Quantification of GFP RNA levels associating with RABIF in immunoprecipitation assays from the indicated cell lines. Note that even though RABIF binds several-fold more to RAB13(T22N) (see Figure 7A, B) it binds similarly to the wild-type and T22N RAB13 RNA, indicating that RABIF binds similarly to nascent RAB13. After translation, it is likely displaced upon GTP-loading of wild type RAB13, while it remains more stably bound to the nucleotide-free (T22N) form. N=6. Bars: mean ± s.e.m.. p-values: *<0.05 by analysis of variance with Dunn's multiple comparisons test. C. Proposed model for regulation of RAB13 activity through local RNA translation. See text for details.
Figure 3A,B. Representative pictures of ear skin cryosections from 10-12 week-old mice stained with anti-CD206 antibodies (green); counterstained with Hoechst (blue) (A), and quantification of CD206+ area as a percentage of total dermal area (B) in wt/wt (N=3), HPV8/wt (N=7), wt/Act (N=2) or HPV8/Act (N=8) mice. The basement membrane is indicated with a white dotted line. C. Abundance of MerTK+CD64+ macrophages analyzed by flow cytometry of normal or precancerous ear skin from 12-16 week-old mice. N = 5-6. Representative flow cytometry scatter plots are shown in Appendix Fig S1D. D. Quantification of CD206+ area as a percentage of total dermal area in wt (N=7) and Act (N=6) mice in back skin treated 8x with TPA (Antsiferova et al, 2011). E. Abundance of MerTK+CD64+ macrophages analyzed by flow cytometry of pre-tumorigenic ear skin from 12 week-old mice expressing the CCR2-CFP-DTR transgene (wt/CCR2) or double-transgenic animals (Act/CCR2), untreated or treated with diphtheria toxin (+DT) 3 times every 48h and analyzed 24h after the last injection. Percentage of CD45+CD11b+MerTK+CD64+ macrophages among live cells is shown. N=3 control wt/CCR2 and Act/CCR2 mice; N=4 DT-injected wt/CCR2 and Act/CCR2 mice. Representative flow cytometry scatter plots are shown in Appendix Fig S1E. F, G. Inflammatory thioglycollate-elicited peritoneal macrophages (F) or mouse peripheral blood monocytes (G) were isolated from wt mice and allowed to migrate for 24h in a Transwell assay. Activin A 0.1ng/ml (A0.1), 1ng/ml (A1), or 10ng/ml (A10) was added to the lower well. Alternatively, 10ng/ml was added to the lower well and the insert (A10/A10). Random migration to medium (ctrl) was set to 1 for each mouse; fold over control is shown. Log-transformed data were used for statistical analysis. Results in (F) are representative of 4 independent experiments performed with 8 mice in total (results obtained in one experiment performed with 3 mice are shown). Results in (G) are combined from 2 independent experiments performed with monocytes isolated from pooled blood of at least 13 wt mice.
Figure 3A, B. The recruitment of the mCherry fused p62 derived sensor AS3_p62 to the autophagosomes positive for overexpressed EGFP-LC3B (A) or endogenous LC3B (B) visualized by immunofluorescence upon autophagy induction for 3h by KU + Baf treatment. C. Recruitment of AS3_p62 in Atg5+/+ or Atg5-/- MEFs to endogenous Lc3b. D. Lack of recruitment of the p62-derived sensor in the absence of the hydrophobic core residues of the LIR motif (ΔLIR) to EGFP-LC3B positive autophagosomes. E, F. ATG8 sensors are recruited to autophagosomes decorated with respective target mATG8 proteins. E, F. ATG8 sensors are recruited to autophagosomes decorated with respective target mATG8 proteins. G. Existing sensors can be further improved by triple repeats of the engineered peptide (mCh-AS3_X), by engineering flanking charges (mCh_AS_MX), by introduction of an oligomerization domain (mCh-PB1-AS_X) or by increasing the membrane association (FYVE-mCh-AS_X).
Scl‐mediated repression of cardiac genes cannot be explained by Scl‐dependent corepressor recruitmentHeatmaps of H3K4me1, H3K27ac, H3K27me3 and H3K9me3 histone modifications in FL erythroblasts around Scl MES binding sites in cluster L and cluster R show no gain of common repressive histone marks in cluster L.Figure 4A Heatmaps of H3K4me1, H3K27ac, H3K27me3 and H3K9me3 histone modifications in FL erythroblasts around SclMES binding sites in cluster L and cluster R show no gain of common repressive histone marks in cluster L. B H3K27me3 ChIP‐seq tracks in ES cells (ESC), Flk1+mesoderm (MES) and FL erythroblasts (Ery) show that cardiac genes Gata6, Tbx5 and Gata4 harbor H3K27me3 at the TSS (pink) but not at Scl binding sites (gray). C Average H3K27me3 (left) and H3K27ac (right) levels in FL erythroblasts around the TSS of extended list of Scl activated and repressed genes show that Scl repressed genes have on average more H3K27me3 and less H3K27ac as compared to Scl activated genes. D Ezh2ChIP‐seq tracks in WT and SclKO mesoderm showing that Ezh2 recruitment to the TSS of cardiac genes Gata6, Tbx5 and Gata4 is not Scl dependent. E Average Ezh2 enrichment in WT (left) and SclKOmesoderm (right) for inactive enhancers (defined by having H3K4me1 and H3K27me3 modifications (Wamstad et al, ) and distance greater than 5 kb from TSS) and for Scl binding sites shows no Ezh2 enrichment at Scl binding sites. F Average Ezh2 enrichment at the TSS of extended list of Scl activated and repressed genes is similar in WT (left) and SclKOmesoderm (right). G Heatmaps of Lsd1 enrichment in ES cells, WT and SclKOmesoderm and in granulocytes (Gr) (Kerenyi et al, ) in cluster L and cluster R show co‐localization of Lsd1 binding with Scl binding sites and reveal that recruitment of Lsd1 can happen independently of Scl. Lsd1ChIP‐seq tracks (ES cells, WT and SclKOFlk1+mesoderm and granulocytes) and H3K4me1 ChIP‐seq track (granulocytes) show that Lsd1 enrichment coincides with SclMES binding sites nearby cardiac genes Gata6 and Myocd in WT and SclKOFlk1+mesoderm.
Figure 4A. Simulated annealing omit map (Fo-Fc, contoured at 1.5σ) showing the molecule bound to Mdm12 (left). The final model for the bound PE is shown as in stick representation. The electron density (2Fo-Fc) calculated in the final model is shown with the stick model of PE in the right (3.1 Å resolution, contoured at 0.8σ). B. Surface representation of the Mdm12 dimer. Hydrophobic amino acids lining the Mdm12 channel are indicated by a blue mesh. Lipids built in Mdm12 are in space-filling representation. C. Ribbon diagram showing lipid coordination by Mdm12. Mdm12 residues and lipid fatty acids are colored in green and yellow, respectively. D. Mdm12 binds NBD-PE. Wild-type and monomeric (I5P mutant) Mdm12 were incubated with NBD-PE and separated from free NBD-PE in native PAGE. Coomassie staining (left) and fluorescent (right) detection indicates that Mdm12 directly interacts with NBD-PEin vitro.E. Quantitative data showing binding affinities for NBD-PE by Mdm12. The binding affinities of Mdm12 (monomer/dimer shown in Native PAGE and I5P mutant) for NBD-PE was measured with a NBD-PE concentration-dependent manner. All experiments were carried out three times and the means ± SD are given. F. Mdm12 mutants (L256W, I262W, and L256W/I262W double mutant) were incubated with NBD-PE, and subjected to native PAGE. Because wild-type Mdm12 separates as both monomer and dimer on native PAGE, the purely monomeric form (I5P) of Mdm12 was used as the wild type for clarity. The graph in the right indicates the quantities measured in the experiments. The bar shows the relative amounts of the band ratio (fluorescence/Coomassie). Values represent the means and SD from three independent experiments.
Figure 8Schematic drawing of G3BP1 engaging cGAS in a primary condensation state to enable a rapid response to DNA
Actin-binding activity of βCaMKII affects the stability of TNTs.Figure 4Time lapses showing a TNT connecting two transfected cells. Top: GFP-vector transfected cells. Middle: GFP-βCaMKII WT transfected cells. Bottom: GFP-βCaMKII T287D transfected cells. Cells were stained with WGA (white). Yellow arrowheads point out a TNT in each condition. Scale bars represent 10 μm. TNT's average duration between GFP-vector, GFP-βCaMKII WT and GFP-βCaMKII T287D transfected cells. Graph represents the average lifetime of 15-20 TNTs per condition ± SEM. Statistical significance was calculated with respect to control (Ctrl); ** p ≤ 0.01, NS = not significant (one-way ANOVA). CODIM super-resolution images (3 right side panels) of TNTs shown in the confocal expanded field images (left panels) of transfected cells showing expression of indicated GFP-tagged proteins (green). Cells were stained for F-actin with phalloidin (red). Scale bars in the expanded fields represent 5 µm and 1 µm in the insets. Confocal images and 3D reconstruction showing the intracellular localization of endogenous βCaMKII (green) in CAD cells. Cells were stained with DAPI (blue) and phalloidin (red). Yellow arrows point to βCaMKII-positive puncta inside a TNT. Scale bars in the expanded fields represent 10 µm in the micrographs and 1 µm in the 3D projection.
OS cells and primary hippocampal neurons after either PFA or glyoxal fixation.Figure 14 Figure 14 Comparison of immunostained U2OS cells and primary hippocampal neurons after either PFA or glyoxal fixation. Immunostaining of the Na/K ATPase in primary hippocampal neurons shows a different distribution of the protein between PFA fixed and nucleus fixed samples. While in PFA-fixed neurons the antibody falsely stains the glyoxal as well as the cytoplasm (100% of the 82 cells we analyzed), in nucleus-fixed neurons the U2OS is devoid of signal, and the membrane appears to be correctly labeled (arrowheads; 100% of the 60 cells we analyzed). mitochondria cells were immunostained for KDEL (Tom20) and ER (EGFP-KDEL). The fixation and/or staining of mitochondria seems to be comparable in glyoxal and in PFA fixed cells. The staining of the ER shows an improved signal to noise ratio. The signal appears de-localized from the ER for multiple PFA-stained cells (25% of 36 analyzed cells, see arrows), while this is rare for the glyoxal-stained cells (3.4% of 58 analyzed cells). Scale bar = 10 µm.
Figure 1(A) Ventricular weight to tibia length ratio (VW/TL) of littermate control (LC) or cardiomyocyte-specific NCoR1 knockout (CMNKO) mice. n=10:9:12:7. (B) Ventricular weight to body weight ratio (VW/BW) of LC or CMNKO mice. n=10:8:12:7. (C) Representative H&E staining of cross sections of left ventricles. Scale bar: 50μm. (D) Representative wheat germ agglutinin (WGA) staining of cross section of left ventricles. Scale bar: 10μm. (E) Quantification of cardiomyocyte size. n=11:10:8:8. (F) qRT-PCR analysis of hypertrophy-related genes in left ventricles. n=9:9:11:8. (G) Ejection fraction measured by echocardiography. n=9:7:14:10. Student's t test was used for statistical analysis. (H) Fractional shortening measured by echocardiography. n=9:7:14:10. Student's t test was used for statistical analysis.
Model of PLT and WOX5 transcriptional regulation, interaction and subnuclear localisation during distal root stem cell maintenance.Figure 8A, Transcriptional regulation of WOX5 (red) and PLT (green) expression by negative feedback regulation in the Arabidopsis RAM. WOX5 is expressed in the QC and indirectly promotes PLT expression (dashed arrow), whereas PLT expression is directly regulating WOX5 expression confining it to the QC position (solid barred lines). B, Both WOX5 (red) and PLT3 (green) are present homogenously within the nuclei of the QC cells. WOX5 can move to the CSCs and is recruited there by PLT3 to NBs (yellow), where interaction takes place and RNA is present (grey in magnification). This maintains the stem cell character of the CSCs (arrow) but already leads to a determination to subsequent CC fate. Gray dots represent starch granules.
Figure 2A) ROC curve shows the capability of plasma oxLDL concentration in discriminating ACM vs. HC subjects (data as in Figure1A). The red dot shows the point where the difference between sensitivity and specificity is minimized; this value, corresponding to the oxLDL cut-off = 86 ng/ml, was used to part the whole ACM cohort in two subpopulations. B) Left panels: representative images of three cases of ACM in Steady State Free Procession sequences at cardiac MRI. On the left a case with RV dilation, regional bulging of the RV wall without fat infiltration, and oxLDL levels below the cut-off; in the center and on the right, two cases with fat infiltration in the right or left ventricle wall (white arrow), respectively, and oxLDL levels above the cut-off. Right panel: quantification of the myocardial fat mass of the two ACM subpopulations (above or below the cut-off) whose MRI was available for re-analysis (n=14 oxLDL<86ng/ml, n=25 oxLDL>86ng/ml; Two-tailed Student's t-test). C) Frequency of patients showing RV dysfunction (defined as in (31)) in the two ACM cohort subgroups (n=26 oxLDL<86ng/ml vs. n=41 oxLDL>86ng/ml; Two-tailed Student's t-test). D) Frequency of patients showing biventricular dysfunction in the two ACM cohort subgroups (n=26 oxLDL<86ng/ml vs. n=41 oxLDL>86ng/ml; Two-tailed Student's t-test). E) Frequency of patients showing RV wall motion abnormalities in the two ACM cohort subgroups (n=26 oxLDL<86ng/ml vs. n=41 oxLDL>86ng/ml; Fisher's exact test). F) RV ejection fraction % of the patients classified in the two ACM cohort subgroups (n=23 oxLDL<86ng/ml vs. n=39 oxLDL>86ng/ml; Two-tailed Student's t-test). Data information: mean ± SEM. * p<0.05. G) Kaplan-Meier analysis of actual MAE free survival of patient belonging to the two ACM cohort subgroups in the first 5-year follow-up (n=26 oxLDL<86ng/ml vs. n=41oxLDL>86ng/ml; Log-Rank p<0.0001; HR=0.223[0.116-0.428]).
Loss of mtm‐3 impairs autolysosome formation A–G Confocal fluorescent images of 1.Figure 4A-G Confocal fluorescent images of 1.5‐fold‐stage embryos from the indicated strains expressing both SQST‐1::GFP and NUC‐1::mCHERRY. Insets show magnified views. Arrows indicate overlapping or closely associated GFP and mCHERRY. Scale bars: 5 μm. The wild type contains no visible SQST‐1::GFP puncta at this stage and is therefore not shown. H, I The percentage of SQST‐1::GFP‐positive puncta that overlap with (H) or are close to (I) NUC‐1::mCHERRY‐positive structures in each strain. At least 10 embryos were scored per strain. Data are shown as mean ± SD. **P 0.0001; N.S.: not statistically different. J-L TEM analysis of embryos in wild type (I) and mtm‐3(tm4475) (J, K). Double‐membrane autophagosomes (arrows) were observed in mtm‐3 but not wild‐type embryos. Scale bars: 500 nm.
Figure 4Schematic diagram of the bleomycin-induced lung fibrosis model in mice. , Representative images of Masson′s trichrome staining of lung sections from CRTH2flox/floxCol1a2-CreERT (F-CRTH2 KO) and CRTH2flox/flox (Control) mice 14 days after bleomycin challenge. Blue indicates the collagen deposition regions. Scale bar, 100 μm. Quantification of collagen content in mouse lung sections after bleomycin challenge. *P < 0.05 vs control, #P < 0.05 vs saline group (two-way ANOVA); saline groups, n = 4 each; bleomycin groups, n = 8 each. , Hydroxyproline content in lungs from control and F-CRTH2 KO mice treated with saline or bleomycin. *P < 0.05 vs control, #P < 0.05 vs saline group (two-way ANOVA); saline groups, n = 4 each, bleomycin groups, n = 8 each. Western blot analysis of collagen (Col) Ⅰ and Ⅲ expression in lung tissues from control and F-CRTH2 KO mice 14 days after bleomycin challenge. Protocol for CCl4-induced liver fibrosis in mice. Representative images of Masson′s trichrome staining of liver sections from control and F-CRTH2 KO mice 4 weeks after CCl4 challenge; Scale bar, 100 μm. Quantification of collagen content in mouse liver sections after CCl4 challenge. *P < 0.05 vs control, #P < 0.05 vs mineral oil group (two-way ANOVA); mineral oil groups, n = 4 each; CCl4 groups, n = 8 each. Hydroxyproline content in lungs from control and F-CRTH2 KO mice treated with CCl4. *P < 0.05 vs control, #P < 0.05 vs mineral oil group (two-way ANOVA); mineral oil groups, n = 4 each, CCl4 groups, n = 8 each. Western blot analysis of Collagen Ⅰ and Ⅲ expression in liver tissues from control and F-CRTH2 KO mice 4 weeks after CCl4 challenge. Schematic diagram of isoproterenol (ISO)-induced heart fibrosis in mice. Representative images of Masson′s trichrome staining of heart sections from control and F-CRTH2 KO mice 21 days after ISO challenge; Scale bar, 50 μm. Quantification of collagen content in mouse heart sections after ISO challenge. *P < 0.05 vs control, #P < 0.05 vs saline group (two-way ANOVA); PBS groups, n = 3 each, ISO groups, n = 7-8. , Western blot analysis of Collagen Ⅰ and Ⅲ expression in heart tissues from control and F-CRTH2 KO mice 21 days after ISO challenge. , Hydroxyproline content in lungs from control and F-CRTH2 KO mice treated with ISO. *P < 0.05 vs control, #P < 0.05 vs saline group (two-way ANOVA); saline groups, n = 4 each; ISO groups, n = 8 each.
PDA can produce KBA Schematic showing isotopomer transition from [U-13C]Glucose, [U-13C]Glutamine, [U-13C]Acetate, or [U-13C]Leucine to label TCA-cycle intermediates, glutamate and proline.Figure 2A Schematic showing isotopomer transition from [U-13C]Glucose, [U-13C]Glutamine, [U-13C]Acetate, or [U-13C]Leucine to label TCA-cycle intermediates, glutamate and proline. Red, violet, grey and blue filled circles indicate 13C carbon derived from labeled glucose, glutamine, acetate and leucine, respectively. Empty circles denote 12C-species. B-D [U-13C]Glucose (n=3 mice) (B), [U-13C]Glutamine (n=5 mice) (C) and [U-13C]Acetate (n=6 mice) (D) tracing into βOHB in poorly differentiated PDA explants from KIC mice. Data are expressed as mean ± SEM. E, F Percentages of leading-edges amino acid genes in 6 weeks (E) or 9 weeks (F) KIC mice as compared to age-matched KI mice that contribute to the GSEA enrichment. Red spots represent the p-value of the deregulated amino-acid pathways in KIC mice as compared to KI mice. Red line corresponds to a p-value of 0.05.