EMBO/sd-character-level-ner huggingface dataset:
Figure 5A Experimental design for the early-term treatment: 3 weeks after injection, mice were assigned randomly to vehicle (n=4) or BAPN (n=4) treatment (Top). Radiance-based tumor growth curves (mean ± SE) for the MM134 (left) and SUM44 xenografts (right) treated early (bottom). For statistical analysis, mixed-effects linear models with spline regression (when applicable) were used on log10(radiance) curves Likelihood-ratio tests from the ANOVA function in R were used for model comparison, i.e. to check whether the treatment/group covariate is significant. *** p < 0.001 (likelihood ratio statistic associated p-value). B Experimental design for treatment in metastatic settings: 5 months after injection, mice were randomized to vehicle (n=4) or BAPN (n=4) treatment (top). Radiance-based tumor growth curves (mean ± SE) for MM134 (left) and SUM44 (right) xenografts treated in the metastatic setting (bottom). For statistical analysis, mixed-effects linear models with spline regression (when applicable) were used on log10(radiance) curves Likelihood-ratio tests from the ANOVA function in R were used for model comparison, i.e. to check whether the treatment/group covariate is significant. *** p < 0.001 (likelihood ratio statistic associated p-value). C,D Bar graphs showing ex vivo radiance ± SEM in different organs harvested from females bearing MM134 or SUM44 intraductal xenografts for the mice described in B. Statistical significance determined by Student's unpaired t-test, two-tailed. * p < 0.05, ** p < 0.01, n.s not significant. E Radiance-based tumor growth curves (mean ± SE) for T47D treated with vehicle (n=8) or BAPN (n=12) 8 days after intraductal injections. n.s not significant. F Radiance-based tumor growth curves for (mean ± SE) ILC ER+ PDXs T125 and T137 treated with vehicle (n=6, 7) or BAPN (n=7, 8) 66 days after engraftment. ** p < 0.01. G Radiance-based tumor growth curves (mean ± SE) for non-ILC ER+ PDXs T99 and T157 treated with vehicle (n=7, 7) or BAPN (n=8, 12), 8 days after intraductal injection 3 weeks after intraductal injections. n.s not significant. H Representative micrographs showing picro-sirius red staining of sections from MM134 xenografts treated with PBS (left) or BAPN (right), n=4.Scale bars, 100 μm. I Representative Second-Harmonic Generation (SHG) image on SUM44 xenografts treated with vehicle (left) or BAPN (right), n=3. Collagen fibers in red and DAPI nuclear stain (blue). Scale bar, 50 μm. J Barplot showing Reactome pathway enrichment analysis of differentially down regulated genes K GSEA plots showing decreased early and late estrogen response and MYC targets. P values are computed by non-parametric permutation tests.
i.Figure 2CD68 immunoreactivity in white adipose tissue (scale bar = 25 μm, images representative of one animal each from a total of four animals per experimental group). Arrow points to a region stained with CD68 antibody. *P = 0.0109, Student's t-test. Epididymal fat mass was analyzed in mice (n = 6 animals/group) 7 days after i.c.v. injection of vehicle or AβOs. Data are representative of three independent experiments with similar results. *P = 0.0255. Relative expression of leptin (C), TNF-α (D) and IL-6 (E), respectively, in white adipose tissue of mice (n = 7 Veh; 9 AβOs) 7 days after i.c.v. injection of vehicle or AβOs. In (C), *P = 0.0394; in (D), **P = 0.0038; in (E), *P = 0.0305; Student's t-test. p-JNK (F; n = 5 animals/group) and IRS-1pSer312 (G; n = 6 animals/group) levels (normalized by total JNK and total IRS-1, respectively) in skeletal muscle of mice 7 days after i.c.v. injection of vehicle or AβOs. In (F), *P = 0.0464; in (G), *P = 0.0081; Student's t-test. Representative images of GLUT-4 immunofluorescence in insulin-stimulated skeletal muscle from mice that were i.c.v.-injected with vehicle (Veh) or 10 pmol AβOs. Bar graphs show quantification of GLUT-4 surface immunoreactivity in skeletal muscle of mice that received intraperitoneal injections of PBS or insulin (1 IU/kg body weight) 7 days after i.c.v. injection of vehicle or AβOs, as indicated (n = 5 animals/group). Scale bar = 25 μm. *P = 0.0144, one-way ANOVA followed by Bonferroni post hoc test. GLUT-4 mRNA (n = 4 animals/group) and total protein levels (normalized to actin levels; n = 5 Veh; 6 AβOs) were unchanged in skeletal muscle of Swiss mice injected with vehicle (Veh) or 10 pmol AβOs. Plasma levels of insulin (K; n = 12 animals/group), leptin (L; n = 11 Veh; 12 AβOs), cholesterol (M; n = 8 Veh; 6 AβOs), triglycerides (N; n = 8 Veh; 6 AβOs) or noradrenaline (O; n = 7 Veh; 8 AβOs) measured 7 days after i.c.v. injection of vehicle (Veh) or 10 pmol AβOs. In (O), *P = 0.0361, Student's t-test.
Infection with Mtb-Δeis induces cell death through JNK-dependent signaling.Figure 5(A and B) BMDMs were infected with Mtb-WT, Mtb-Δeis, or Mtb-c-eis (MOI = 1) for the indicated periods of time, and then subjected to Western blot analysis using Abs raised to p-ERK1/2, p-p38, p-JNK, and β-actin. Data shown are representative of three independent experiments that all yielded similar results (A). Expression of phospho-MAPK/β-actin protein in cytoplasmic extracts of BMDMs was quantified densitometrically (B). (C and D) BMDMs were pretreated with U0126 (5, 10, 20 µM), SB203580 (SB; 1, 5, 10 µM), or SP600125 (SP; 5, 10, 20 µM) for 45 min, and then infected with Mtb-Δeis for 30 min (C) or 36 h (D). (C) Cells were then incubated with DHE for 15 min, washed rapidly and thoroughly, and analyzed immediately for superoxide levels by flow cytometry. Quantitative DHE fluorescence data represent the mean±SD of four experiments. (D) Cell death after 36 h was assessed by PI staining and then examined by fluorescence microscopy. ( (E) Raw264.7 cells were transfected with siRNA specific for JNK1 (siJNK) or a non-specific control siRNA (siNS). At 24 h after transfection, cells were infected with Mtb-Δeis for 36 h. Cell death was then assessed by PI staining, and then examined by flow cytometry. Transfection efficiency was assessed by RT-PCR (inset). Data represent the mean±SD of five random fields and are representative of three independent experiments (D and E). *p<0.05, **p<0.01, ***p<0.001, vs. Mtb-WT-infected condition (B); SC (C and D); siNS (E). UI, uninfected; SC, solvent control (0.1% DMSO).
a) Fluorescence microscopy of Hoechst-counterstained HeLa cells transiently transfected with LC3–GFP subjected to 100 μM spermidine for 6 h.figf7(a) Fluorescence microscopy of Hoechst-counterstained HeLa cells transiently transfected with LC3-GFP subjected to 100 μM spermidine for 6 h. Representative pictures are shown. (b) Percentage of adherent cells exhibiting a clear LC3-GFP relocalization into cytoplasmic vacuoles. Numbers were determined using micrographs of Hoechst-counterstained HeLa cells as representatively shown in a. Data represent means ± s.d. (n = 3). (c) LysoTracker Red staining of vacuoles indicative of autophagy in oesophagus tissue from flies fed with 1 mM spermidine for two days, compared with controls (without spermidine). Nuclei were visualized by Hoechst staining. Scale bars, 10 μm. (d) Quantification of autophagic vesicles per nucleus in LysoTracker Red stained muscle tissue of female flies fed with supplementation of 1 mM spermidine or with 10% glucose (starved) for 48 h, compared with normal food (control). Data represent means ± s.e.m. of at least 20 flies for each group (*P 0.01). (e, f) Survival of Drosophila during ageing without (control) and with supplementation of food at various concentrations of spermidine (as indicated). Autophagy-deficient flies (f) homozygous mutant for Atg7 (Δatg7) were compared with flies capable of autophagy (e) and heterozygous for Atg7 (control). For details of strains and additional wild-type controls, see Supplementary Information Methods and Fig. S6h, i. (g) Fluorescence microscopy of C. elegans transgenic embryos expressing a full-length plgg-1DsRED::LGG-1 fusion protein indicative of autophagic activity. Shown are two representative pictures of embryos untreated (control) or treated with spermidine (0.2 mM) supplementation of food. (h) Quantification of autophagic activity through measurement of DsRED::LGG-1 pixel intensity from images of wild-type animals shown in g, and bec-1 RNAi knockdown animals (bec-1 RNAi). Data represent means ± s.e.m. (n = 3) with at least 25 images processed for each trial. (i) Survival of C. elegans during ageing with and without (control) supplementation of food (UV-killed E. coli) with spermidine (0.2 mM). Wild-type (N2) animals were compared with bec-1 RNAi animals deficient in autophagy induction. For mean life spans see Supplementary Information, Table S3.
Comparison of ion binding in sodium and potassium channels.Figure 4Left: Sodium ions (cyan) in the selectivity filter of the NavMs sodium channel. Only two monomers (in white and pink) are shown for clarity. Coordinates are from the AB tetramer of PDBID 5BZB.Right: Potassium ions (blue) in the selectivity filter of the KcsA potassium channel. As for the NavMs structure, only two monomers (in grey and maroon) are shown for clarity. Coordinates are from PDBID 1K4C (Zhou et al., 2004).The two crystal structures were first superimposed using SSM (Krissinel and Henrick, 2004), and are drawn using the same scale, and with the correct ratio between the diameters of the ions used, so that the differences in the internal dimensions of their selectivity filters can be seen, thus making it clear how much more space there is available in the NavMs SF for intervening water molecules. In the KcsA channel, the ions are directly coordinated to the polypeptide backbone atoms, whereas in the NavMs SF, the closest atoms (side chains) are obviously at longer distances than direct bonding to the ions, enabling interactions via water molecules.
Arsenic directly binds to DNMT3A.Figure 2A LC-ESI-MS/MS spectrum of Dnmt3a pulled down by Biotin-As in NIH3T3 cells. NIH3T3 cells were treated with 10 μg/mL Biotin-As for 2 hours, followed by pull-down assay and determination using LC-ESI-MS/MS. B-D Validation of the binding between DNMT3A and Biotin-As at 5 μg/mL in NIH3T3 (B), MEFs (C) and CCRF-CEM (D) cells in the Biotin-As pull-down assay. E Schematic diagram of the structure of human DNMT3A. Upper panel: All cysteines in DNMT3A are shown. Lower panel: Structure of DNMT3A-ADD (PDB: 3A1A) and the four interested cysteine triads. F Measurement of DNMT3A-ADD thermodynamic stability upon ATO treatment. Purified DNMT3A-ADD was mixed with ATO at the indicated ratios in 20 mM HEPES (pH 7.5) for 1 hour. Melting curves were measured by DSF (left panel), and the calculated ΔTm values are shown (right panel). G LC-MS spectra of purified DNMT3A-ADD before and after ATO incubation at 1:5 molar ratio for 16 hours. H, I Biotin-As pull-down assays for the transfected DNMTs (H) and DNMT3A-C520A/C524A mutation (I) in 293T cells treated with 5 μg/mL Biotin-As.
Meth Inhibits Endosomal Antigen Processing and MHC II Restricted Antigen Presentation.Figure 2
Mesenchymal PDAC cells reduce proliferation and alter the morphology of PSCs.Figure 2(A) Brightfield images of PS-1 cells which were exposed for 72 hrs to CM from control (CM of PS-1 cells ), epithelial (blue) or mesenchymal (red) cell lines. Scale bar represents 100 µm. (B) Quantification of bead-calibrated cell count of PS-1 cells after treatment using flowcytometry. n=6 per group, 3 biological replicates for each cell line. (C) EdU assay performed after treatment showing percentage of PS-1 cells in G0/G1 phase, G2/M phase and S phase. n=6 per group, 3 biological replicates for each cell line..
Figure 6A. Schematic diagram showing the in vivo experiment. Two to three-month-old hemizygous M83 mice were injected into the striatum with sterile PBS or 10 μg of recombinant α-syn fibrils. The mice were treated with saracatinib at 10 mg/kg/day by oral gavage for 4 weeks. B. Fixed brain tissues containing the frontal cortex and the striatum were lysed. Western blot was performed with indicated antibodies. Values were derived from three individual mice (n=3). Two to three-month-old hemizygous M83 mice were injected into the striatum with sterile PBS or 10 μg of recombinant α-syn fibrils. The mice were treated with saracatinib at 10 mg/kg/day by oral gavage for 4 weeks. C. Immunohistochemistry of pSer129 α-syn was performed. Arrow indicates pSer129 α-syn positive inclusion. Scale bar indicates 50 μm. D. Scoring of pSer129 immunoreactivity in the cortex and the striatum of both regions. Values were derived from eight individual mice (n=8). *** P < 0.001, ** P < 0.01, * P < 0.05 compared to PBS group, unpaired t-test. Data are expressed as mean ± SEM. E. Schematic representation of the scoring of α-syn inclusion pathology detected by pSer129 staining.
phase.Figure 1A Protein expression E2F7 and E2F8 in RPE cells treated with 100 µg/mL cycloheximide (CHX) and 10 µM MG132. The drugs were added simultaneously. B Protein levels of E2F7 and E2F8 in RPE and U2OS cells after 16 hours of treatment with the CDK4/6 inhibitor PD0332991, or the CDK2 inhibitor NU6140. C Protein expression of E2F7 and E2F8 after 8 hours of PD0332991 treatment, in presence or absence of the proteasome inhibitor MG132 (10 µM) for two hours prior to harvesting. D Schematic overview of conserved KEN motifs in human/mouse E2F7 and E2F8 proteins. E FACS profile showing expression of cell cycle markers in RPE cells with stable expression of the FUCCI system. Encircled areas indicate the gates used to sort cell cycle-specific populations. F Immunoblots of FACS-sorted RPE cells with stable expression of the FUCCI system. Cells were sorted based on expression of truncated versions of and Azami Green-tagged Geminin (amino acids 1-130) and Kusabira Orange-tagged CDT1 (amino acids 30-120), respectively. Blots are representative examples of four independent replicates derived from two different stable RPE-FUCCI clones. G Transcript levels of atypical E2Fs in sorted RPE-FUCCI cells measured by qPCR. Bars represent average ± s.e.m. of fold change, normalized to G1 abundance.
Macrophage-derived VEGF depletion restores blood vessel architecture in glioma.Figure 7A. qPCR analysis of CCR7, IL12, CCL19, CXCL10, MRC1, TGFβ, CD209 and VEGF on RNA samples from MHCII or MRC1 tumor extracted macrophages (n=5 isolation per group).Statistical analysis: A. G. Mann-Whitney u-test. Error bars: A-D-E-F: meanSD B. sFlt1 binding assay on 5 weeks growth glioma section implanted in ROSAmT/mG::Pdgfb-iCre mouse. At late stage tumor growth, M2 macrophages surrounding blood vessels present very high amount of VEGF (binding sFlt1) to the neighboring endothelial cells (5µm depth stack). C. F4/80 immunohistochemistry and sFlt1 binding on sections of 4 weeks growth glioma in Vegfafl/fl::LysMCre+ or Vegfafl/fl ::LysMCre- mice (50µm depth stack). D. Quantification of F4/80 positive macrophages. VEGF depletion in myeloid cells does not alter macrophages recruitment (n=7 mice per group). Statistical analysis: D.E.F. t-test; Error bars: A-D-E-F: meanSD; E. Quantification of F4/80 positive macrophages binding sFlt1. VEGF depletion is effective in 61% of the macrophages invading the tumor (n=7 mice per group). Statistical analysis: D.E.F. t-test; Error bars: A-D-E-F: meanSD; F. Vessel diameter quantification. Myeloid cell derived VEGF depletion induces blood vessel "normalization" compared to control (n=7 mice per group). Statistical analysis: D.E.F. t-test; Error bars: A-D-E-F: meanSD; G. Tumor volume quantification in 4 weeks growth glioma in Vegfafl/fl::LysMCre+or Vegfafl/fl::LysMCre- mice (n=7 mice per group). Statistical analysis: G. Mann-Whitney u-test. Error bars: G: median interquartile. Scale bars: 100µm.
Figure 1AIB1 and YAP co-activate and corepress a subset of TEAD target genes. Venn diagrams showing differentially regulated genes in MCF10A cell lines that overexpress YAP or YAP S94A in the presence of control shGFP or shAIB1 as indicated. RNA-sequencing performed in triplicate. RNA-sequencing was performed in triplicate and statistics in Panel (A-B) were calculated by EdgeR with Benjamini-Hochberg multiple test correction in R. Heatmaps of AIB1-YAP co-activated, YAP activated, AIB1-YAP corepressed, and YAP repressed genes shown in panel (A). Red and blue indicate high and low levels, respectively. RNA-sequencing was performed in triplicate and statistics in Panel (A-B) were calculated by EdgeR with Benjamini-Hochberg multiple test correction in R. Select genes are regulated by YAP and AIB1, others are YAP regulated independently of AIB1. Endogenous gene expression changes assessed by RT-qPCR. Significant genomic co-occupancy between AIB1 and TEAD in normal breast and breast cancer cells. Overlap of AIB1 and TEAD4 ChIP-seq peaks in MCF10A and MCF7 cell lines. Select peaks from MCF10A ChIP-seq shown. ChIP-sequencing performed in triplicate. AIB1 and TEAD are localized at promoters and enhancers globally. Feature distribution based on overlapping AIB1-TEAD4 peaks with respective MCF10A and MCF7 histone ChIP-seq peaks. TEAD binding motifs are enriched within AIB1 occupied sites across the genome. HOMER de novo motif analysis of AIB1 ChIP-seq peaks in MCF10A and MCF7 cells. AIB1 modulates TEAD transcriptional activity in a YAP dependent manner. HEK293T cells were transfected with CTGF-promoter reporter and expression vectors of indicated proteins. AIB1 is engaged with TEAD on co-activated target promoters in normal cells. Sequential ChIP-reChIP performed with indicated antibodies in MCF10A cells. Levels of precipitated chromatin assessed by qPCR. AIB1 is engaged with TEAD on co-activated target promoters in early stage breast cancer cells. Sequential ChIP-reChIP performed with indicated antibodies in MCFDCIS cells. Levels of precipitated chromatin assessed by qPCR.
TEFM interacts with RNA processing factors.Figure 6Volcano plot showing TEFM-BioID. TEFM-BirA*-HA was stably expressed in HEK 293 cells, followed by streptavidin affinity purification and mass spectrometry analysis to detect the associated proteins. Mitochondrial targeted BirA* (MTS-BirA*) was used as a control. The significantly enriched proteins are presented above the horizontal dashed line with 5% false discovery rate (FDR) significance. The blue dots highlight proteins involved in mtDNA transcription and the green dots represent proteins involved in RNA metabolism. All other proteins are in grey.
Figure 2A Colony formation for cb9Δmyc cells infected with retroviral vectors expressing the indicated Myc proteins; all cells were expanded with doxycycline prior to the final plating step, upon which the compound was either maintained (+dox), or removed (-dox) to switch off the tet-Myc transgene. One representative experiment out of 3 is shown. B Cumulative cell counts for cb9Δmyc cells upon serial passaging with or without dox (removed at day 2). The data are presented as mean ± SD; n = 3 (biological replicates) C C Percentage of BrdU positive cb9Δmyc cells, in the presence or absence (24h after removal) of doxycycline. Data are presented as mean ± SD; n = 3. Two-tailed Student t-test was used to compare between two groups and expressed as p-values. D Immunoblot analysis of c-myc-/- HO15.19 rat fibroblasts infected with retroviral vectors expressing the indicated Myc proteins. Parental TGR1 cells serve as control for endogenous levels of the MycWT protein. E Colony formation for the same cells as in (D). One representative experiment out of 3 is shown. F Cumulative cell counts for the same cells as in (D). Data are presented as mean ± SD; n = 3 (biological replicates). Two-tailed Student t-test was used to compare between two groups and expressed as p-values.
Figure 2(A) Line plots depicting fluctuations in gene expression levels from vsn normalized probe intensities across human T cell development for both αβ- and γδ-lineages (n=2 biological replicates and error bars indicate SEM). (B) Dendrogram classifying the T cell developmental subsets and replicates, based on the normalized mRNA expression. (C) Diffusion map, displaying diffusion component (DC) 1, 2 and 4, generated from the mRNA expression profiles reconstructing the developmental trajectories of the αβ and γδ developmental lineages across human thymocyte subsets. The line connecting the different subsets is reflecting the developmental stages of human T cell development. (D-E) Scatterplot displaying the up (red) or down (blue) regulation of probes (log2 fold change >= 2) identified by sequential pairwise comparisons across either αβ (D) or γδ (E) development.
Disaggregation of aggregated proteins during recovery from heat shock.Figure 3A Line graphs showing solubility after heat shock (HS) and during different time points of recovery. Each line corresponds to one protein. Orange lines show the mean solubility. B Quantification of disaggregation rates. Disaggregation rate for each protein is estimated as a slope from linear fits of data used in A. Histograms of slopes (binwidth = 0.02 Solubility/h) are shown for aggregators and soluble proteins. C Different disaggregation profiles for aggregating proteins. Solubility line graphs as in A shown for aggregators with the bottom 10%, middle 80% and top 10% of disaggregation rates. Proteins with bottom 10% of disaggregation rates and related to DNA repair are highlighted with red. Proteins with top10% disaggregation rates and directly related to transcription are highlighted with blue. D Protein localization upon heat shock and during recovery as analyzed by immunofluorescence microscopy. HeLa cells were fixed and immunostained with antibodies against indicated target proteins (green) at different conditions [control, after heat shock (10 minutes at 44°C) and after five hours of recovery from the heat shock]. DNA staining (Hoechst) is shown in blue. E-H Comparison of disaggregation rates with different protein characteristics enriched in aggregators (presented in Fig 2B-C and Fig 2E-G). Volcano plot presenting correlation coefficients and Benjamini-Hochberg adjusted p-values (E; horizontal dashed line shows a p-value of 0.05). Scatterplots comparing disaggregation slopes and solubility change after heat shock (F), fraction of intrinsically disordered regions (G) and fraction of (random) coil-like secondary structure (H). In F-H, scatter plots are shown for correlations with a p-value lower than 0.05. Correlation coefficients (r) with p-values are shown for Spearman's rank-order correlation.
Fs can activate APC/C by repressing transcription of Emi1 and CDK2-associated cyclins.Figure 5A Immunoblots showing Emi1 protein levels in cell lines with stable inducible expression of E2F7 or E2F8 after 16 hours of doxycycline treatment. B Quantitative PCR of FBXO5 transcripts in FACS-sorted cells with moderate or high levels of E2F7/8 expression, after 8 hours of doxycycline treatment. To avoid bias from cell cycle defects, cells were released from HU arrest at the onset of doxycycline treatment, resulting in a strong enrichment of cells in late S or G2 of both vehicle and doxycycline-treated cells (see Fig EV3C). C Quantitative PCR of FBXO5 transcripts in HeLa cells treated with siRNA against E2F7 or E2F8. Cells were harvested 6 hours after release from HU block to enrich for cells in mid- to late S phase. Bars in (B) and (C) indicate mean ± sem (n=3); asterisks indicate P<0.05 versus vehicle and scrambled siRNA, respectively. D Overlap between E2F target genes and APC/C substrates. The APC/C substrates are taken from a curated list [32, 33] with the addition of E2F7 and E2F8 based on the current paper; E2F7/8 targets from [8, 10]. E Simplified working model of interaction between atypical E2Fs and APC/C to coordinate DNA replication, based on the work described in this paper. Asterisks indicate indirect effect via transcriptional regulation of Emi1 and Cyclin A/E.