EMBO/sd-character-level-ner huggingface dataset:
Figure 6a) RNA-sequencing data showing expression of genes associated with neutrophil recruitment during liver aging; b, c) Quantification of b) neutrophil infiltrates (NIMP positive) and c) mean number of TAF positive hepatocytes in liver of young (3 months) and old (24 months) mice. Data are mean±S.E.M. of 3-5 mice per group ; d) Quantification of mean number of TAF in hepatocytes as a function of the distance from a Ly6G labelled neutrophil. Data are from 5 mice aged 29 months of age, line indicates linear relationship between mean TAF and distance (linear regression and F test: R2=0.35; P<0.0001); e) Representative micrograph showing hepatocyte from aged mouse containing a TAF located in close proximity to a neutrophil (labelled with Ly6G); f) schematic depicting experimental design; g) % of neutrophil migration towards conditioned media (CM) obtained from young (early passage) and replicatively senescent human fibroblasts. Media alone and fMLP were used as negative and positive controls respectively. Data are mean±S.E.M. of 3 independent experiments; h) Schematic depicting experimental design for panels i-k; i) Comparison between p16Ink4a mRNA levels of young (3 months old) and old INK-ATTAC mice (28-29 month old) treated with vehicle or AP20187. Data are mean ± SEM of n = 5-8 mice per age group; j) % of TAF positive hepatocytes in old INK-ATTAC mice (28-29 months old) treated with vehicle or AP20187. Data are mean ± SEM of n = 5-6 mice per group; k) Liver neutrophils (Ly6G) in old INK-ATTAC mice (28-29 months old) treated with vehicle or AP20187. Data are mean ± SEM of n = 5-7 mice per group; l) Intrahepatic leukocyte profiling by mass CyTOF. CyTOF was performed on young (6 months) and old (24 months) INK-ATTAC mice (n=5 mice per group); 23 unique clusters of intrahepatic leukocytes were defined by a 24-cell surface marker panel using the Rphenograph clustering algorithm and were visualized on a tSNE plot (t-distributed stochastic neighbor embedding plot).; m) (left) tSNE plots comparing the distribution of young and old intrahepatic leukocytes. Red indicates high frequency categorization of cells to a cluster while blue indicates low frequency; (right) tSNE plots comparing p16Ink4a expression in young and old intrahepatic leukocytes. Red indicates high p16Ink4a expression while blue indicates low expression. Data are from n=5 mice per group; n) % of p16Ink4a positive cells (detected by CyTOF) in different intrahepatic leukocyte populations isolated from young and old mice. Data are mean±s.e.m of n=5 mice per group; o) Schematic depicting experimental design for panels p and q. 3 month old wild-type mice were injected with murine Cxcl1 for 7 days; p) Quantification of liver neutrophils (Ly6G) in mice injected with vehicle or Cxcl1. Data are mean ± SEM of n = 7-10 mice per group ; q) Quantification of the % of TAF positive hepatocytes in mice injected with vehicle or Cxcl1. Data are mean ± SEM of n = 6-7 mice per group; r) Correlation between TAF positive hepatocytes and neutrophils (linear regression and F test: r2=0.70; p=0.0001).
Figure 3Principal component analysis (PCA) of Ronin fl/del, Ronin del/del (day 4) and Ronin del/del (day 8) transcriptomes; n = 3 independent RNA-seq datasets for each condition. Volcano plot of gene expression in Ronin del/del day 4 vs. Ronin fl/del (left panel) and Ronin del/del day 8 vs. Ronin fl/del ESC (right panel) with annotated naïve and primed pluripotency markers. Expression and binding of Ronin to core, naïve and primed pluripotency markers. Gene set enrichment analysis (GSEA) plot of Il6/Jak/Stat3 signalling, day 8 Ronin ko ESC. Expression and binding of Ronin to Il6/Lif signalling-related genes. K-means clustering of 5 major groups of genes in Ronin fl/del, Ronin del/del (day 4) and Ronin del/del (day 8) transcriptomes. Gene ontology (GO) enrichment analysis of the predefined gene clusters - molecular function (left panel) and cellular component (right panel).
Figure 4A) Immunofluorescence staining for OCT4, ZSCAN4 and 2C::tbGFP in control and acetate treated ESC cultures. Green arrows indicate 2-cell-like cells, and inlets highlight their DAPI structure. Scale bar, 20 μm. B) RT-qPCR of the indicated genes in ESC cultures treated with sodium acetate for 24h. Shown are the mean ± s.d. of 3 independent cell cultures, performed in 2 technical replicates. C) Percentage of 2-cell-like cells obtained upon transfection of control or Dux-targeting siRNAs in control conditions or in combination with sodium acetate treatment. Measurements were obtained from 2 independent cell cultures, performed across 3 independent biological replicates. Boxes indicate the range between the first and third quantile, the band depicts the median and the whiskers extend no further than 1.5-times the interquartile range. Individual dots indicate the measurements obtained in each technical replicate. D) Percentage of 2-cell-like cells obtained upon transfection siRNAs targeting the indicated chromatin factors in control conditions or in combination with sodium acetate treatment. Shown are the mean ± s.d. of 4 independent cell cultures, performed across 2 independent biological replicates. E) Experimental design. ESC cultures were FACS-sorted to remove Zscan4+ and 2-cell-like cells and plated in a glass bottom 96-well-plate. Cells were then imaged for 96 hours in the presence of sodium acetate (32 mM), sodium L-lactate (32 mM) or in control conditions. F) Proportion of Zscan4+ cells at various timepoints during the time-lapse experiment. Shown are the mean ± s.d. of 3 independent experiments.
Hypoxia rescues muscle degeneration after chronic or acute inactivation of Rb by inducing glycolysis via HIF-1α.Figure 4.(A) Immunostaining for MHC (green) in the indicated myoblasts induced to differentiate under normoxia or hypoxia treated or not with lonidamine. Note the shortened myotubes in the presence of lonidamine. (B) Representative immunoblot (left) and mean expression of myogenin and MCK normalized for MHC expression (right; n = 3). (C) Rbf/f myoblasts transduced with Ad.GFP or Ad.Cre and immunoblotted for pRb. Tubulin was used as a loading control. (D) Rbf/f myoblasts transduced with Ad.GFP or Ad.Cre and immunostained for pRb (red). (insets) DAPI staining for nuclei (blue). (E) Immunostaining for MHC in Rbf/f myoblasts transduced with Ad.Cre and induced to differentiate in normoxia (top and middle) or hypoxia (bottom). (F, top) Brightfield images of Rbf/f myoblasts transduced with Ad.EV or Ad.Cre and induced to differentiate under hypoxia. (bottom) Larger view of the boxed field. Arrowheads point to rescued Rb−/− myotubes. This field is shown in Video 6. (G) Quantification of myotube formation in Rbf/f myoblasts transduced with Ad.EV or Ad.Cre and induced to differentiate under hypoxia. Counts are mean of six representative fields (n = 3). *, P = 0.004. (H) Quantification of myotube formation in the indicated myoblasts induced to differentiate under the indicated conditions. Myotube counts are the mean of six fields (n = 4). (I) MHC staining (red) of the indicated myoblasts transduced with Ad.NF-κB/HIF-1α and induced to differentiate in normoxia. (J) Quantification of myotube formation in the indicated myoblasts transduced with Ad.EV or Ad.NF-κB/HIF-1α and induced to differentiate in normoxia. Myotube counts are the mean of six fields (n = 3). Error bars represent SD.
Figure 1A Cell viability as assessed by GFP expression in HEK293T cells transfected with the pRetroX TETOne3G-eGFP plasmid only (vector) or pRetroX TETOne3G-eGFP harbouring the N-terminal fragment of GSDMD. Cells were treated with the indicated concentrations of doxycycline 24 h post transfection and the percentage of GFP-positive cells was determined 16 h later by flow cytometry. B LDH release from HEK293T cells transfected with the pRetroX TETOne3G-eGFP plasmid only (vector) or pRetroX TETOne3G-eGFP harbouring the N-terminal fragment of GSDMD. At 24 h post transfection, cells were treated with the indicated concentrations of doxycycline for 8 h and the percentage of LDH release was determined. Graphs show mean and s.d. of quadruplicate wells. C-D. PI staining of and LDH release from HEK293T cells transfected pRetroX TETOne3G-eGFP harbouring the N-terminal fragment of GSDMD in the presence of osmoprotectants. At 24 h post transfection, PEGs of the indicated molecular weights were added to a final concentration of 30 mM, cells were treated with 250 ng/ml doxycycline for 8 h and the level of PI staining (C) or LDH release (D) was determined. E-F. PI staining of and LDH release from LPS-primed primary BMDMs infected with log-phase S. typhimurium for the indicated timepoints in the presence of PEGs of the indicated molecular weight (numbers on the X-axis, 30 mM final concentration).* p<0.05 as determined by student's t test. Graphs show mean and s.d. of quadruplicate wells. Data are representative of at least three independent expeirments.
Figure 6(A) Cisplatin dose response curves of (i) MB002, (ii) Daoy, and (iii) MIC in the absence (EV) or presence of SOX9 by Alamar blue. Cells were pre-conditioned with doxycycline to induce expression of SOX9 (or EV) prior to treatment with increasing concentrations of cisplatin. The IC50 were calculated following 5 (MB002 and MIC) or 3 days (Daoy) of treatment. Data are mean + standard deviation from 3 independent repeats, each containing 5 technical replicates. (B) Cisplatin dose response curves of SOX9-expressing (i) Daoy and (ii) MIC in the absence or presence of FBW7α. Experiments and data analysis were performed as described in (A) (C) Overall survival analysis of mice bearing Daoy or Daoy-expressing dox-inducible SOX9 treated with cisplatin. The dox-preconditioned cells (105 cells) were orthotopically xenografted to Nude-Foxn1nu mice and left for 1 week to prior to being treated with vehicle control or cisplatin (2mg/kg) intraperitoneally for every other day for a total of 6 doses. (D) Heat map of the row-wise z-scores of 11 genes associated with cisplatin resistance in MB002 expressing Sox9-WT or Sox9-T236/T240A. Heat map was generated using the GenePattern software. (E) Quantitative analysis of ATP7A, DUSP2, and TTK mRNAs in MB002 following expression of SOX9-WT or SOX9-T236/240A. Total RNA were collected 24 hours following doxycycline treatment, from which cDNA were generated for qPCR. Data are mean mRNA level (normalized to B2M transcript) + standard deviation from 3 independent experiments with statistical significance were determined by Multiple comparisons 2-way ANOVA with Bonferroni's post-test. (F) Time course western blotting of HA-SOX9, ATP7A, DUSP2, ERK1/2 pThr202/Tyr204 and total ERK1/2 in MB002 cells following doxycycline induction of either EV, SOX9-WT or SOX9-T236/240A. GAPDH was used as a loading control.
Figure 3Bar graphs showing the relative cell numbers in the indicated cell types following the indicated treatments. Cell numbers were assessed after a 48-hour period of growth in cultures seeded at equal densities. All values represent mean±SEM. Data points represent values from biological replicates. Bar graphs showing the relative cell numbers in the indicated cell types following the indicated treatments. Cell numbers were assessed after a 48-hour period of growth in cultures seeded at equal densities. All values represent mean±SEM. Data points represent values from biological replicates. Bar graphs showing the relative cell numbers in the indicated cell types following the indicated treatments. Cell numbers were assessed after a 48-hour period of growth in cultures seeded at equal densities. All values represent mean±SEM. Data points represent values from biological replicates. Representative images showing Ki67-labeling in xenografts generated from implantation of HN31 cells stably expressing control shRNA (left) or MCOLN1 shRNA (right). Pseudocolored panels show the representative distribution Ki67 positive or negative nuclei. Scale bars shown (0.5 mm) apply to all panels. Bar graphs showing fraction (left) and total number (right) of Ki67 labeled nuclei in the indicated cell types. All values represent mean±SEM. Values in the bar graph on the right were normalized to control mean. Data points represent values from independent eggs. Statistical test employed was Student's t-test. Bar graphs showing the relative cell numbers in UMSCC-22A-HRASWT cells following the indicated perturbations. All values represent mean±SEM. Data points represent values from biological replicates. Statistical test employed was Student's t-test. Kaplan-Meier curves showing the survival of patients stratified on the basis of MCOLN1 expression in BLCA and HNSC patients whose tumors were wild-type for HRAS (right) or carried oncogenic HRAS mutations (left). Statistical test employed was the Mantel-Cox log-rank test.
Fig.Figure 3A Immunoblot analysis of purified H1N1 viruses A/California/7/2009 (Cal/7/9), A/Brevig Mission/1/1918 (BM/1/1918), A/Puerto Rico/8/1934 (PR8) or A/Brisbane/59/2007 (Bris/59/7) separated under reducing and denaturing conditions. Uncleaved HA0 and cleavage products HA1 and HA2 were detected with pooled pre-challenge sera (AAV-HA, -cHA, -GFPB Significant peptides identified in the epitope screen with AAV-HA pooled pre-challenge serum ((AAV-HA, -cHA, -GFP, WIV n = 18, AAV-NP n = 11; data shown in Fig.EV2) were mapped onto HA1 (red) and HA2 (blue) of the trimeric HA (PDB 3UBE generated with PyMol) (n = 3, technical duplicates). The position of the RBS (yellow) and the C179 epitope (dashed line) is marked. C to F Binding of C179 (C), or antibodies present in AAV-HA (D), AAV-cHA (E) or WIV (F) pooled pre-challenge sera (n = 18 mice per group) to Cal/7/9 or PR8 virus after incubation of the virions at pH = 7.2, 5.8, 5.4, 5.0, 4.4 or 4.4+DTT to induce conformational changes in HA or remove the HA1 subdomain. Mean ± SD (n = 3, in technical triplicates). Statistical significance between pH = 7.2 and other conditions was determined using Kruskal-Wallis test with Dunn's multiple comparison testing (*p<0.05, **p<0.01).
Figure 2A. Chemical structure of TP-16. B. Dose-effect curve of TP-16 in GloSensor™ cAMP assay in EP4- expressing 293 cells (n=3). C. Dose-effect curves of TP-16 in PGE2-induced calcium flux assay (n=3). D. The Schild plot of PGE2 in the presence of varying concentrations of TP-16. TP-16 shifted the dose-response curve of PGE2-induced intracellular cAMP levels in a dose-dependent manner (n=3). E. The pA2 value and slope of the Schild plot. F. Docked pose of TP-16 with critical residues in the putative binding pocket of human EP4 protein. EP4 is shown as a color cartoon, the residues important for the interaction are depicted in magenta sticks, and TP-16 is shown as a cyan stick figure. G. The LIGPLOT diagram summarizes key interactions between TP-16 (purple lines) and residues that originate from EP4. T69, T76, T168 and R316 establishes hydrogen bonds with TP-16. Semicircles with radiating lines indicate non-polar interactions.
PMEPA1 is a target gene involved in the response to Activin signaling (A) Ranking of top genes whose expression is correlated with INHBB in Group 3 MB patient samples.Figure 5(A) Ranking of top genes whose expression is correlated with INHBB in Group 3 MB patient samples. Spearman's rank correlation coefficient ρ and p-value are indicated. The p-values were determined by Spearman rank correlation test *p < 0.05, **p < 0.01, ***p < 0.001. (B) Boxplots representing PMEPA1 expression levels in the different MB groups (WNT in blue, SHH in red, Group 3 in yellow and Group 4 in green) and in fetal and adult cerebellum (grey) in the dataset " Data ref: Cavalli et al, 2017". Only p-values corresponding to comparisons between Group 3 and the other groups are indicated. The p-values were determined by Wilcoxon rank-sum test *p < 0.05, **p < 0.01, ***p < 0.001. (C) Scatter plot of INHBB and PMEPA1 gene expression levels in all MB groups. Colored dots represent each patient samples and colors represent the MB groups (WNT in blue, SHH in red, Group 3 in yellow and Group 4 in green). The p-values were determined by Spearman rank correlation test *p < 0.05, **p < 0.01, ***p < 0.001. (D) Boxplot represents the quantification of PMEPA1 protein level normalized to β-actin levels across groups. The p-values were determined by unpaired t-test for D *p < 0.05, **p < 0.01, ***p < 0.001. (E) Scatter plot represents Log2 relative protein level of P-Smad2 (x axis) and PMEPA1 (Y axis) normalized to β-actin in each individual samples. The p-values were determined by Wilcoxon rank-sum test *p < 0.05, **p < 0.01, ***p < 0.001. (F) Immunoblots of phosphorylated Smad2 (P-Smad2), total Smad2, MYC, PMEPA1, OTX2 and β-actin were performed on extracts from 1603MED or D283 or D458 cells treated with either DMSO (vehicle), LY364947, SB431542, blocking antibody against ActivinB (Ab α-ActB), Follistatin, PBS or ActivinB (ActB) for 24 hours. Immunoblot analysis of PMEPA1 levels in 1603MED (G) cells 48 hours after transfection with either siCTRL (blue) or siPMEPA1 (red). Bar graphs on the right represent the quantification of the relative level of PMEPA1 protein level normalized to β-actin. The p-values were determined by unpaired t-test *p < 0.05, **p < 0.01, ***p < 0.001. Bars represent the mean ± SD. Number of replicates is n≥3. Growth curves of 1603MED (H) cells after transfection with either siCTRL (blue) or siPMEPA1 (red). The p-values were determined by two-way ANOVA *p < 0.05, **p < 0.01, ***p < 0.001. Immunoblot analysis of PMEPA1 levels in D283 (I) cells 48 hours after transfection with either siCTRL (blue) or siPMEPA1 (red). Bar graphs on the right represent the quantification of the relative level of PMEPA1 protein level normalized to β-actin. The p-values were determined by unpaired t-test *p < 0.05, **p < 0.01, ***p < 0.001. Bars represent the mean ± SD. Number of replicates is n≥3. Growth curves of D283 (J) cells after transfection with either siCTRL (blue) or siPMEPA1 (red). The p-values were determined by two-way ANOVA *p < 0.05, **p < 0.01, ***p < 0.001.
Proteomics and metabolomics show enhanced ribosome- and TCA/OXPHOS- activity in PtenStat3pc-/- -mice A.Figure 4A. Differentially expressed proteins in PtenStat3pc-/- vs. Ptenpc-/- proteomic samples. Colors indicate adj.p-value and log2-FC. (Black = Log2-FC ≤ 1 & adj. p-value ≥ 0.05, orange = Log2-FC > 1 & adj. p-value ≥ 0.05, red = Log2-FC > 1 & adj. p-value < 0.05). FC = fold change. B. STAT3 immunohistochemistry- staining of wild type, Ptenpc-/-, PtenStat3pc-/- mouse prostates. Scale bar = 100µm. C. Significantly enriched KEGG pathways in PtenStat3pc-/- versus Ptenpc-/- groups. Dotted line: adj. p-value = -log10(0.05). D. Significantly enriched metabolic KEGG pathways in PtenStat3pc-/- versus wild type (=WT) groups. Dotted line: adj. p-value = -log10(0.05). Red = up-regulated, blue = down-regulated. E. Metabolite concentrations in nmol/µg of 5 metabolites in WT, Ptenpc-/- and PtenStat3pc-/- prostates. Box-plot shows median, 1st and 3rd quartiles, and whiskers extend to ± 1.5 interquartile range. Jitter represents biological replicates. ANOVA tests and Tukey multiple comparisons were applied. Red = PtenStat3pc-/- (n = 3), blue = Ptenpc-/- (n = 5), grey = wild type (WT, n=5), q = adj. p.-value, n.s. = not significant.
Figure 5A, B. HP1γ-dependent differential splicing junctions detected by MAJIQ between transcriptomes of HP1γ and KO cells. Top, Schemes of splice junctions showing annotated alternative splicing events (blue and green, panel A), or an annotated exon-exon junction and de novo cryptic junctions increased in KO (red and green, respectively, panel B). C. Box plot depicting the length of exon junctions found differentially regulated by MAJIQ between HP1γ (green) and KO (orange) in three categories as in Fig EV5A, all classified differential HP1γ/KO junctions, annotated alternative exon junctions, non-annotated de novo cryptic junctions detected in KO only; n, differential events. Box boundaries represent 25th and 75th percentiles; center line represents median; whiskers indicate ±1.5×IQR; points are values of outliers. Data information: All graphes represent mean and s.d. (** P < 0.005, ***P < 0.001; two-tailed Student's t-test). D. RNAchIP read density per kb on the indicated introns for input and IP in samples from both untreated and PMA stimulated cells, n=6 biological replicates, or 3-times randomized intervals, n=18 samples. Dots represent individual data points. HP1γ/KO introns are all 272 intronic intervals between exon junctions differentially detected in HP1γ and KO. Random introns are a matching library of random intronic intervals. E. RNAchIP read density per kb on the genes corresponding to the introns tested in (D), and analyzed as in (D), n=6 biological replicates, or 3-times randomized intervals, n=18 samples. Data information: All graphes represent mean and s.d. (** P < 0.005, ***P < 0.001; two-tailed Student's t-test). F. Reads from input and IP counted on the indicated features in combined unstimulated and PMA stimulated triplicate samples, n=6 biological replicates. Dots represent individual data points. Data information: All graphes represent mean and s.d. (** P < 0.005, ***P < 0.001; one-tailed Student's t-test). G. Average distribution profiles of sense SINE repeats on indicated exons and on ±0.7kb of surrounding introns.
Figure 5A. Hierarchical clustering of the 82 differentially expressed (DE) genes identified in Jurkat cell lines overexpressing miR-34c-5p (J34c-2 and J34c-5) vs control (JØ-1, 2 and 3 clones). Color scale represents mean centered log2 fold expression level across samples. Pie charts represent the relative proportion of predicted miR-34c-5p binding sites by the Miranda or PITA algorithms (or both) in down-regulated (top) and up-regulated (bottom) gene sets. B. qRT-PCR validation of selected genes. Plot represents the average percentage of target gene mRNA (normalized to GAPDH) in J34c vs JØ cells (all differences were found to be significant with a paired two tailed t test, P<0.05; n=5).
Figure 5A Pull-down assay of lysates from untransfected P1.HTR cells with unphosphorylated (YENM) and tyrosine-phosphorylated (pYENM) IDO1 peptides, analyzed by immunoblotting with anti-p85, -p110α, -p110β, -p110γ, and -p110δ antibodies. Whole cell lysates (WCL) were used as control of lysate amount. B Confocal immunofluorescence microscopy images of P1.HTR cells transfected with IDO1.WT incubated with class IA PI3K p110 isoform inhibitors (indicated) or medium alone (Ctrl) for 24 h and co-stained with antibodies recognizing IDO1 (green) and EEA1 (red). Nuclei were stained with DAPI (blue). The merge imagine is shown for all stainings or without DAPI (no DAPI). Scale bar, 10 µm. C Pearson's coefficient of IDO1 colocalization with EEA1 in P1.HTR cells transfected and treated as in (B), with each point representing colocalization within an image stack. Data represent mean ± SD (n > 20 cells from two independent experiments). D Tyrosine phosphatase activity in anti-IDO1 immunoprecipitates from P1.HTR cells transfected with IDO1.WT cells and treated with PI3K catalytic inhibitors (indicated) or medium alone (ctrl). Data represent the mean ± SD (n = 3 technical replicates).