EMBO/sd-character-level-ner huggingface dataset:
XIAP supports tumour growth by promoting neutrophil infiltration in Hgf-Cdk4R24C mouse melanoma A Sum tumour size in Hgf-Cdk4R24CXiapfl/fl and Hgf-Cdk4R24CXiapKO mice at 38 weeks of age.Figure 5A Sum tumour size in Hgf-Cdk4R24CXiapfl/fl and Hgf-Cdk4R24CXiapKO mice at 38 weeks of age. Dots represent sum of all tumours size in an individual mouse. B Representative images of back skin of the DMBA-treated Hgf-Cdk4R24CXiapfl/fl and Hgf-Cdk4R24CXiapKO mice at 8 weeks of age. C Number of tumours in the DMBA-treated mice at 8 weeks of age (n=3). Dots represent tumours having a range size of 0-10, 10-40 or >40 mm2 in an individual mouse. D Tumour size in the DMBA-treated Hgf-Cdk4R24CXiapfl/fl and Hgf-Cdk4R24CXiapKO mice at 8 weeks of age (n=3). Dots represent an individual tumour. E qRT-PCR analysis (PrimePCRTM Assay) of different cyto-/chemokines in Hgf-Cdk4R24CXiapfl/fl mice plotted vs 3 independent Hgf-Cdk4R24CXiapKO mice. Samples that are at least 2-fold up-regulated are shown in red, samples that are at least 2-fold down-regulated are shown in green. F and G Representative IHC staining of neutrophils in small (early time points) (F) and large (late time points) (G) tumours. Scale bar 100 µm (upper panel) and 10 µm (lower panel). Arrows show neutrophils.
Figure 3A Representative images and quantification of vacuolar morphology of late meristematic cells. PI (green) and MDY-64 (yellow) staining depict cell wall and vacuolar membrane, respectively. Seedlings were treated with DMSO solvent control or 50 µM EGCG for 22 h on solid medium (n=40). Mann Whitney U test (***p < 0.001). Data information: Scale bars: 5 µm. B 3-D reconstructions of PI-stained cell wall (red) and BCECF-stained vacuole (green) of late meristematic cells. Boxplot depicts vacuolar occupancy of the cell. Seedlings were treated with DMSO (n=16) or 50 µM EGCG (n=15) for 22 h on solid medium. Student´s t-test (**p < 0.01). Data information: Boxplots: Box limits represent 25th and 75th percentile, horizontal line represents median. Whiskers display min. to max. values. C Representative images and quantification of vacuolar morphology of late meristematic cells. PI (green) and MDY-64 (yellow) staining depict cell wall and vacuolar membrane, respectively. Seedling roots were grown on the surface (n=28) or into the matrix (n=28) of 2 % agar-containing solid medium. Mann Whitney U test (***p < 0.001). Data information: Scale bars: 5 µm. D 3-D reconstructions of PI-stained cell wall (red) and BCECF-stained vacuole (green) of late meristematic cells. Boxplot depicts vacuolar occupancy of surface (n=10) and into the medium (n=11) grown seedling roots. Student´s t-test (**p < 0.01). Data information: Box limits represent 25th and 75th percentile, horizontal line represents median. Whiskers display min. to max. values.
Figure 1Left panel, a scheme of chromosome 18 copy number alterations depicting the distal long arm loss across TCGA Pan-Cancer dataset. Vertical red line indicates the localization of VPS4B. Right panel, enlarged fragment of chromosome 18 showing frequent deletions of VPSB locus in cancer samples. Deletions are marked in blue, amplified regions are marked in red. Analysis of VPS4B copy number alterations in TCGA Pan-Cancer dataset. Cancer types were sorted according to the mean VPS4B copy number after removing germline values. Scatter plot, analysis of VPS4B mRNA expression (number of transcripts per million) plotted against VPS4B copy number from TCGA CRC patient samples (n=376); plot generated using cBioPortal Gao et al, 2013(). Pie chart, summary of all types of VPS4B copy number alterations based on the analysis of data from 615 CRC samples deposited in the Colorectal Adenocarcinoma (TCGA, Provisional) qPCR analysis of VPS4B mRNA abundance in normal colon, adenoma and CRC samples. Adenocarcinoma (n=26); adenoma (n=42); normal colon (n=24). Green horizontal bars indicate means and red whiskers indicate SD. Examples of immunohistochemical staining of VPS4B in normal colon and matched CRC samples as an illustration of the scoring system used for the evaluation presented in (F). 3+ - very intensive staining, 2+ - medium-intensive staining, 1+ - weak staining, 0 - no staining. Scale bar, 50 µm. Comparative analysis of VPS4B staining performed in 100 pairs of normal colon and matched CRC samples.
Figure 4A. Scramble and Rab22a KD JAWS-II DCs were infected with OVA-YFP-expressing T. gondii (TgRHYFPSAG1-OVA) for 8 h and confocal images detecting the parasite (green), endogenous Rab22a (red) and GRA6 (magenta) were taken. Top panels: Scramble cells; bottom panels: Rab22a KD cells. White boxes are shown at higher magnification in the insets. The nuclear marker DAPI (blue) and DIC images are shown in the left panels. Overlays are shown in the right panels. Scale bars: 5 µm. Images are representative of at least 30 analyzed from three independent experiments. B. The cross-presentation of OVA secreted by T. gondii (TgRHYFPSAG1-OVA) after 8 h of infection at the indicated MOI was evaluated by B3Z T cell activation. Data represent mean ± SEM of triplicate values and are representative of three independent experiments. ***P < 0.001. A two-way ANOVA and the Bonferroni post-test were performed. C. The efficiency of infection (8 h) of TgRHYFPSAG1-OVA in Scramble and Rab22a KD JAWS-II DCs was measured by FACS analysis at the indicated MOI. Data represent mean ± SEM of duplicate values and are representative of three independent experiments. D. Immunoblotting showing the total amount of OVA in Scramble and Rab22a KD JAWS-II DCs after 90 min and 7 h of infection with TgRHYFPSAG1-OVA parasite at MOI 2 and 6. Fifty µg of total cell lysates were loaded onto each lane. Data are representative of two independent experiments.
Figure 3Chromogenic WM-ISH of miR-9 using miR-9 LNA 5'-Dig observed at different stages during development; longitudinal view, anterior to the left. Transverse section of double fluorescent WM-ISH for her6 (green) and mir-9-4 (magenta) imaged in fixed embryo at 31hpf; scale bar 30μm. Schematic representing the miR-9 binding site (MBS) of her6::venus mutated by CRISPR-Cas9nls protein; MBSm refers to specific sgRNA to mutate the MBS as opposed to sgRNA that does not produce mutation (control-CTRL). Confocal imaging of CTRL and MBSm embryo showing Her6::Venus expression in the hindbrain, rhombomeres 3 to 6 (r3-r6) over the course of development; longitudinal view, scale 30μm, otic vesicle (ov). Images are representing 2D maximum projection. Her6::Venus expression (red arrows) in hindbrain, rhombomere 6 of live CTRL and MBSm embryos at 52hpf; edge of ventricular zone shown in yellow; transversal view; scale 30μm. Her6::Venus protein abundance in CTRL versus MBSm homozygous embryos observed in hindbrain, r6 between 34hpf and 69hpf; bars indicate median and interquartile range of (34hpf CTRL: 36 cells, 1 embryo), (34hpf MBSm: 35 cells, 1 embryo), (48±2hpf CTRL: 172 cells, 7 embryos), (48±2hpf MBSm: 124 cells, 5 embryos), (56±2hpf CTRL: 69 cells, 4 embryos), (56±2hpf MBSm: 31 cells, 2 embryos), (69hpf CTRL: 12 cells, 1 embryo), (69hpf MBSm: 6 cells, 2 embryos); Kruskal-Wallis with Dunn's multiple comparison test, significance: p<0.05*, p<0.001***, p<0.0001****. Comparative profiles of Her6 protein abundance over development in CTRL and MBSm embryos quantified as median of single cell abundance data in (f).
Enhancer-promoter pairing based on transcription correlation.Figure 6(A) Schematic diagram describing enhancer-promoter pairing methods. With the 'nearest' (N, top), 'correlated neighboring' (CN, middle), and 'correlated window' (CW, bottom) approach, enhancers are paired to the nearest transcribed promoter, the neighboring upstream and downstream transcribed promoters, and all transcribed promoters within ± 500 kbp distance, respectively. For CN and CW only pairs with correlated transcription rates over all cell lines (Pearson correlation coefficient r > 0.6) were analyzed further (B) Number of enhancers per promoter, for all promoters that were paired to at least one enhancer with the CN approach. Fractions are shown for enhancers per promoter per cell line (light grey) and over all cell lines (dark grey). (C) As in (B) for pairs obtained with the CW approach. (D) Distance distribution between enhancer midpoints and promoter TSSs for E-P pairs obtained with the CN (light grey) and CW (dark grey) approach. The histogram shows the 500 kbp distance range in 20 kbp bins. (E) Left: stacked barplot showing the fraction of enhancers that pair to neighboring transcribed promoters (neighbor, dark grey), to transcribed promoters further away thereby skipping the transcribed neighboring promoters (skipped, white), or both (light grey) using the CW approach. Right: stacked barplot showing the fraction of E-P pairs between enhancers and their neighboring transcribed promoters (dark grey) as well as between enhancers and promoters skipping a transcribed neighboring promoter (white). (F) Barplot showing the percentage of E-P pairs supported by promoter capture Hi-C interactions for different pairing approaches (dark grey). Expected interactions based on 100 sets of randomly sampled matched E-P pairs (light grey, Materials and Methods). Light grey bars depict the mean and error bars represent 95% confidence intervals. (G) Boxplots showing promoter cell type specificity score for different numbers of paired enhancers using the CN approach (from left to right: n=124,767; 12,347; 3,754; 1,619; 2,373). Indicated P-value by Kruskal-Wallis test. All pairwise comparisons are significantly different (P-values < 1e-10, two-sided Mann-Whitney U-test). Specificity scores and number of enhancers were assessed for each cell line (Materials and Methods). Box limits are the first and third quartiles, the band inside the box is the median. The ends of the whiskers extend the box by 1.5 times the interquartile. Notches represent 95% confidence intervals for the median values. Outliers not shown. (H) As in (G) showing promoter transcription. Indicated P-value by Kruskal-Wallis test. All pairwise comparisons are significantly different (P-values < 2.2e-16, two-sided Mann-Whitney U-test). (I) Barplots showing the average number of paired enhancers per promoter per cell line (n=14, mean ± SD) for promoters paired to at least one enhancer and being classified as housekeeping genes ((Eisenberg & Levanon, 2013), left), transcription factors ((Abugessaisa et al., 2016), middle) or cancer-associated genes (Cancer Gene Census, (Sondka et al., 2018), right).
Figure 5A. Schemes of the five different vectors used for the in vivo comparison. ss, single-stranded; sc, self-complementary; i.v., intravenously. B to D. Determination of anti-HBV efficacy and safety by measuring HBsAg in the serum of treated mice (B). B to D. Determination of anti-HBV efficacy and safety of treated mice by measuring their body weight (C). B to D. Determination of anti-HBV efficacy and safety of treated mice by measuring serumALT levels (D). E. Hematoxylin/Eosin (HE) stains of livers harvested on day 84. Scale bar at bottom right represents 100 µm.
Autophagic degradation is impeded in Tsg101- and Vps24-depleted cells.Figure 3.Autophagic degradation is impeded in Tsg101- and Vps24-depleted cells. HeLa cells stably expressing GFP-LC3 (green) were transfected with control (A), Vps24 (B), or Tsg101 (C) siRNA for 5 d and processed for immunofluorescence analysis. Cells were labeled with antibodies against p62 (red) and analyzed by confocal microscopy. Colocalization is indicated in yellow. Bar, 10 μm. (D) The total level of p62 and GFP-LC3 (normalized to control cells) and their degree of colocalization (% of total) in control and Tsg101- and Vps24-depleted cells were quantified using the Zeiss LSM 510 Meta software. 30 cells from three independent experiments were used for quantification. Error bars = SEM. (E) Western blot showing accumulation of LC3-II and p62 in HeLa cells depleted of Vps24 and Tsg101. p62 also accumulates in the insoluble fraction (bottom picture), indicating a shift to a more aggregated form upon depleted of Vps24 and Tsg101. (F) Western blot showing that LC3-II levels were similar in control and Vps24-depleted cells treated with Bafilomycin A.
Fig.Figure 3A: Brightfield images of a NTH-KO and NTH/GFP ookinete, showing normal ookinete morphology, but lack of pigment clusters associated with crystalloids (arrows) in the NTH null mutant. Scale bar = 5μm. B: Aligned dot plot of oocyst numbers in Anopheles stephensi infected with parasite lines NTH/GFP, NTH-KO and NTH∆PP. Horizontal lines mark mean values (n=20); ns, not significantly different (Mann-Whitney). C: Brightfield and fluorescence images of oocysts of parasite lines NTH/GFP and NTH-KO at 15 days post-infection, showing lack of sporulation in the NTH null mutant. Hoechst DNA staining (artificially depicted red) labels parasite nuclei. The large nuclei of the epithelial midgut cells also show. Scale bar = 20μm. D: Aligned dot plot of oocyst diameters in Anopheles stephensi at 15 days post-infection with parasite lines NTH/GFP, NTH-KO and NTH∆PP. Horizontal lines mark mean values (n=20). ** mark statistically significant differences (P<0.0001, Mann-Whitney); ns, not significant. E: Images of ookinetes of parasite lines NTH∆PP showing dispersed NTH∆PP::GFP fluorescence; LAP3/mCherry showing focal LAP3::mCherry fluorescence in crystalloids; Cross (carrying modified alleles encoding NTH∆PP::GFP and LAP3::mCherry), showing dispersed LAP3::mCherry fluorescence. Arrow marks pigment cluster associated with crystalloids. Scale bar = 5μm. F: Schematic diagram of a genetic cross between parasite lines NTH∆PP and LAP3/mCherry and the resultant ookinete population after mosquito transmission and drug selection (oocysts, liver and blood stage parasites not shown). Colours represent NTH::GFP (green), LAP3::mCherry (red) and their co-localisation (orange). Spots in ookinetes depict crystalloids. The red cross indicates that sporozoite formation is blocked. Self-fertilization events are omitted. G: PCR diagnostic for the presence of modified nth alleles in blood stage infections obtained after sporozoite transmission of a NTH∆PP x LAP3/mCherry genetic cross (lane NTH∆PP), amplifying an approximately 4.4kb fragment. NTH/GFP parasites are used as a PCR control, amplifying approximately 4.6kb. Parental parasites provide a negative control. H: Image of a NTH/ND500LK ookinete, showing fluorescent spots co-localizing with pigment clusters (arrows) indicative of normal crystalloid formation. Scale bar = 5μm.
Prmt1 regulates similar and overlapping transcriptional programs mediated by β-catenin in LSK-MLL-ENL cells.Figure 6Figure 6: Prmt1 regulates similar and overlapping transcriptional programs mediated by β-catenin in LSK-MLL-ENL cells. (A) Heatmap analysis and Venn diagram showing the commonly regulated genes after the loss of function of Prmt1 and β-catenin in Hoxa9-/- LSK-MLL-ENL cells. (B) The FDR q-values of the top GO:Molecular function (top panel) and GO:Biological process (bottom panel) (https://toppgene.cchmc.org/enrichment.jsp) for the indicated up- and down-regulated genes is shown. (C) Venn diagrams showing the total, positively and negatively regulated genesets identified by GSEA upon the loss of function of Prmt1 and β-catenin in Hoxa9-/- LSK-MLL-ENL cells. (D) Examples of stem cell and stemness signatures which were commonly higher expressed in the single knockout Hoxa9/- LSK-MLL-ENL cells compared to Prmt1 down-regulated or β-catenin inactivated Hoxa9-/- LSK-MLL-ENL cells.
Figure 4A Plasmids containing human CD45 (hCD45) genomic DNA from intron 3 through intron 7 with a wildtype (I3-I7) or mutated exon 5 CTCF binding site (I3-I7*CTCF) were methylated with M.SssI and confirmed through BsaHI digest. Methylated and unmethylated minigenes were transfected into CHO cells for stable clone generation. B 5hmC and 5mC MedIP were performed in stable clones derived from methylated I3-I7 and I3-I7*CTCF minigenes. Results from individual clones are plotted as a percent of total methylation (n=32 clones) as determined through input normalized IP values for 5hmC or 5mC divided by the sum of the values for 5hmC and 5mC. Methylation in the I3-I7 and I3-I7*CTCF clones used for downstream analysis are indicated by the red and blue bars (WT-hmC and Mut-hmC clones), respectively. C qRT-PCR for relative exon 5 inclusion in methylated I3-I7 and I3-I7*CTCF minigene-derived mRNA. Results are shown for the average of all the I3-I7 and I3-I7*CTCF clones (n=16 clones of each) as well as individually for the WT-hmC and Mut-hmC clones that were selected for downstream analysis. Values represent mean +/- SEM for the pooled data or the mean +/- SD for the individual clones. p= two-tailed Student's t test comparing the I3-I7 to I3-I7*CTCF values. D qPCR of minigene exon 5 relative to CHO GAPDH for comparison of hCD45 copy number in the WT- and Mut-hmC clones. E Bisulfite sequencing of hCD45 exon 5 DNA from the WT- and Mut-hmC clones to confirm the persistence of methylation at the intact and mutated CTCF binding site following stable insertion into the CHO genome. F CTCF-ChIP in the WT- and Mut-hmC clones and qPCR for hCD45 exons, as indicated. ChIP values represent means +/- SD from three independent IP experiments. p= two-tailed Student's t test comparing WT-hmC and Mut-hmC values. G Pol II-ChIP in the WT- and Mut-hmC clones, and a WT-C CHO clone generated with an unmethylated hCD45 minigene. qPCR for hCD45 DNA as indicated. ChIP values represent means +/- SD from three independent IP experiments. *p=0.013, **p=0.0089, two-tailed Student's t test comparing the Wt-hmC and Mut-hmC values. H Summary of the WT- and Mut-hmC data illustrating elevated detection of CTCF and pol II at the wildtype minigene exon 5, relative to the mutated CTCF binding site.
a) Western blot showing EGFP-HDQ71–induced degeneration of rod photoreceptors at 9 d.figf8(a) Western blot showing EGFP-HDQ71-induced degeneration of rod photoreceptors at 9 d.p.f. as indicated by the decrease in the abundance of of monomeric (∼36 kDa) and dimeric (∼72 kDa) rhodopsin. Rhodopsin expression is further reduced by 3 μM (±)-Bay K8644, and recovered upon treatment with 3 μM verapamil, 100 μM 2′5′ddA, 3 μM clonidine and 1 μM calpastatin. No general toxicity to the zebrafish at these concentrations was observed. (b) Expression of EGFP-HDQ71 significantly reduces the levels of rhodopsin expression when compared to wild-type (WT) nontransgenic controls. This effect is further abrogated by 3 μM (±)-Bay K8644, whereas 3 μM verapamil, 100 μM 2′5′ddA, 3 μM clonidine and 1 μM calpastatin all protected against rod photoreceptor degeneration. Error bars show s.e.m. (c) Magnified bright field and EGFP images of zebrafish eye sections, taken at the ventral marginal zone for each of the indicated treatments (with concentrations as in b), are shown. Scale bar, 10 μm. ***P 0.001; **P 0.01; *P 0.05. (d) The total number of aggregates across 8 × 12 μm sections was counted and compared to the EGFP-HDQ71 DMSO-treated control. 3 μM (±)-Bay K8644 was found to significantly increase the aggregate burden, whereas 3 μM verapamil, 1 μM calpastatin, 3 μM clonidine and 100 μM 2′5′ddA decreased the aggregate load. As in our cell models, these aggregates were insoluble in 4% triton/SDS. Error bars show s.e.m. ***P 0.001; **P 0.01; *P 0.05. (e) Schematic representation of an mTOR-independent autophagy pathway involving cAMP-Ca2+-calpains-Gsα with multiple drug targets. Intracellular cAMP levels are increased by adenylyl cyclase (AC) activity, thereby activating Epac, which subsequently activates Rap2B, a small G protein that activates PLC-ε, resulting in the production of IP3 and consequent Ca2+ release from the endoplasmic reticulum (ER). Intracellular Ca2+ levels are also increased by L-type Ca2+ channel agonists. An increase in intracytosolic Ca2+ activates calpains, which cleave and activate Gsα, which activates AC to increase cAMP levels, thereby forming a loop. Activation of this pathway inhibits autophagy. Multiple drug targets acting at various places in this pathway induce autophagy, including the following: I1R agonists (clonidine and rilmenidine) and the AC inhibitor 2′5′ddA, which decrease cAMP levels, agents lowering inositol and IP3 levels (carbamazepine (CBZ) and valproic acid (VPA)), L-type Ca2+ channel antagonists (verapamil and loperamide), calpain inhibitors (calpastatin and calpeptin) and the Gsα inhibitor NF449.
Fig.Figure 1A) (left) t-sne plot from scRNAseq data of human organoids and cluster assignment of colonic cell populations. Red color corresponds to undifferentiated cells, light green corresponds to surface colonocytes (enterocytes). t-sne plot for selected genes from single cell RNAseq of human colon epithelial cells revealing expression of ACE2 specifically in differentiated surface colonocytes expressing KRT20 and AQP8, which also express the IFN-γ receptor IFNGR2. B) Immunofluorescence staining for ACE2 (red) and KRT20 (green) of human colon tissue section indicating ACE2 expression in surface colonocytes. Scale bar: 50 µm. C) Human colon organoids cultured in full medium (FM) condition (upper panel) or FM treated with IFN-γ (lower panel). Left: bright field images of 7-day cultured human colon organoids, scale bar: 100 µm. Right: Immunofluorescence for KRT20 (green; note increased expression upon IFN-γ treatment). Scale bar: 25 µm. D) Human colon organoids cultured in basic medium (upper panel) or additionally treated with IFN-γ (lower panel). Left: bright field images of organoids that were cultured for 4 days in basic medium and in addition for 3 days with IFN-γ (lower panel) scale bar: 100 µm. Middle: Immunofluorescence labelling for KRT20 (green) Right: Immunofluorescence labelling for E-cadherin (green) to visualize epithelial cell boundaries; insets with higher magnification indicate elongation of epithelial cells that are grown in basic medium and treated with IFN-γ. Scale bar: 25 µm. Comparison of E) KRT20, F) MUC2, G) CHGA, H) LGR5 mRNA expression in organoids grown in FM or in basic medium and either untreated or treated with IFN-γ (n=3). Data are presented as mean +/- SD ,*: p≤0.05, **: p≤0.001, ***: p≤0.0001, as determined by one-way ANOVA, followed by Tukey's multiple comparisons test ACE2 mRNA expression in organoids grown in FM or in basic medium and either untreated or treated with IFN-γ (n=3). Data are presented as mean +/- SD ,*: p≤0.05, **: p≤0.001, ***: p≤0.0001, as determined by one-way ANOVA, followed by Tukey's multiple comparisons test J) Immunofluorescence staining for ACE2 (red) and KRT20 (green) of organoids grown in full medium (upper) or basic medium (lower) and additionally treated with IFN-γ (right), indicating expression of ACE2 in differentiated enterocytes. White arrows point to ACE2. Scale bar: 10 µm
Figure 3(A) Simultaneous visualization of GLT-1a+ (green) and VGLUT1+ puncta (red) in first somatic sensory cortex (SI) of a WT and a KI mouse (P35). Arrows point to some GLT-1a+ puncta overlaying with VGLUT1+ puncta (i.e., GLT-1a/VGLUT1 related puncta); framed regions (enlarged below) are examples of GLT-1a/VGLUT1 related puncta (arrowheads). All microscopic fields are from layers II/III. Scale bars: 3.5 µm for left and right panels and 1 µm for enlarged framed areas.(B) Percentage and size of GLT-1a+ puncta overlaying with VGLUT1 in P35 WT and KI mice. Left, percentage of GLT-1a+ puncta overlaying with VGLUT1 is comparable in WT and KI mice (data were obtained from 14 and 18 fields of 20 µm x 20 µm from 2 WT and 2 KI mice (4 sections/animal), respectively) (unpaired t-test: P = 0.52). Right, size of GLT-1a/VGLUT1 related puncta is 19 % reduced in KI mice (169 GLT-1a+ puncta analyzed from 2 mice) compared to WT (174 GLT-1a+ puncta analyzed from 2 mice) (Mann-Withney U test: P = 0.016).