EMBO/sd-character-level-ner huggingface dataset:
Figure 4A Quantitative RT-PCR evaluation of KDM6A mRNA expression in kidney glomeruli isolated from wild-type and KDM6A-knockout (KO) mice. *P < 0.05, significant difference versus wild-type controls (Wilcoxon two-sample test; n = 8). B Western blot analysis of KDM6A expression in glomeruli and podocytes isolated from wild-type and KDM6A-KO mice. Presented experiments were performed at least three times independently. C Changes in body weights and levels of blood glucose in wild-type or KDM6A-KO mice with or without STZ treatment. No significant differences in body weights or blood glucose levels between wild-type and KDM6A-KO mice with diabetes were observed during the 8-week experimental period (Wilcoxon two-sample test; n = 8). D Levels of urinary protein excretion, weights of kidney and levels of HbA1c in wild-type and KDM6A-KO mice with or without STZ treatment (at 8 weeks after the onset of diabetes). The mean relative kidney weight (%) shown in the study is determined as the percent of kidneys out of total body weight, and the HbA1c level is defined as the ratio of HbA1c to the total hemoglobin (% HbA1c; DCCT unit). *P < 0.05 versus untreated wild-type controls, #P < 0.05 versus STZ-treated wild-type mice (Parametric ANOVA and a Bonferroni post hoc test; n = 8). E Immunofluorescence images of kidney sections stained with KDM6A and nephrin in wild-type (WT-NC), KDM6A-KO (KO-NC), STZ-treated wild-type (WT-DM), or STZ-treated KDM6A-KO (KO-DM) mice. Scale bars, 20 μm. *P < 0.05 versus untreated wild-type controls, #P < 0.05 versus STZ-treated wild-type mice (Parametric ANOVA and a Bonferroni post hoc test; n = 3). F Immunofluorescence images of kidney sections stained with KDM6A and H3K27me3 in wild-type (WT-NC), KDM6A-KO (KO-NC), STZ-treated wild-type (WT-DM), or STZ-treated KDM6A-KO (KO-DM) mice. Scale bars, 20 μm. *P < 0.05 versus untreated wild-type controls, #P < 0.05 versus STZ-treated wild-type mice (Parametric ANOVA and a Bonferroni post hoc test; n = 3). G Immunofluorescence staining of F-actin and KDM6A in primary podocytes isolated from the above treated mice. Presented experiments were performed at least three times independently. Scale bars, 20 μm. H Western blot analysis of nephrin, KDM6A and WT-1 expressed in primary podocytes isolated from the above treated mice. *P < 0.05 versus untreated wild-type controls, #P < 0.05 versus STZ-treated wild-type mice (Parametric ANOVA and a Bonferroni post hoc test; n = 3). I Effect of high glucose on the expression of podocyte-related markers in primary cultured podocytes isolated from wild-type or KDM6A-KO mice. The primary podocytes isolated from wild-type or KDM6A-KO mice were cultured in normal or high glucose (30 mM) for 48 hours. Protein lysates from the cultured podocytes were subjected to Western blot analysis with the indicated antibodies. *P < 0.05 versus wild-type podocytes in normal glucose, #P < 0.05 versus wild-type podocytes in high glucose (Parametric ANOVA and a Bonferroni post hoc test; n = 3).
Tap-1 is required for TseL-mediated killing.Figure 2Tap-1 is required for the secretion of TseL. Pellet (P) and supernatant (S) of bacterial cultures were analyzed by SDS-PAGE. The result of immunoblotting with antisera against TseL and Hcp and purified antibodies against DnaK is shown. The asterisk marks an unspecific band, detected by the antiserum against TseL, present in the pre-immunized serum. One representative experiment of three independent experiments is shown. Tap-1 is required for TseL-mediated killing. Wild-type C6706 or C6706 lacking tsiV1 were exposed to the indicated V52 mutants in a killing assay. The y-axis indicates the number of surviving C6706. Tap-1 is required for TseL function. Killing assays in which V52Δtap-1 was mixed with wild-type C6706 or mutants lacking tsiV1, tsiV2, or tsiV3. The y-axis indicates surviving C6706.
Figure 6(A) IHC staining of Nfatc1 expression in HFs on P84 in Sirt7+/+ and Sirt7-/- mice with or without Cyclosporin A (CsA) treatment. The black dashed line indicates bulge (Bu) of hair follicle. (B) Quantitative analysis of Nfatc1 levels in (A, n=10 hair follicles) using ImageJ plus 6.0. (C) Immunofluorescent staining of Ki67 and Pcad at P84 in Sirt7+/+ and Sirt7-/- HFs with or without CsA treatment. (D) The percentage of Ki67-positive cells per HG in (C, n=10 hair follicles). (E) H&E staining of the skin in the indicated treatment groups at P100. (F) Representative images showing haircoat recovery after the indicated treatments. The dashed circle indicates the shaved area; the dotted line indicates the hair growth boundary.
BRCA1 and CtIP are required for telomeric end‐to‐end fusions induced by TRF2 inactivationImmortalized Brca1F/− MEFs were infected with retroviruses expressing the indicated shRNAs and/or Cre recombinase, followed by selection with puromycin for 72 h.Figure 2A Immortalized Brca1F/−MEFs were infected with retroviruses expressing the indicated shRNAs and/or Cre recombinase, followed by selection with puromycin for 72 h. Cells were arrested in mitosis with colcemid and mitotic chromosomes were processed for CO‐FISH analysis. Metaphase chromosome spreads were stained with Cy3‐conjugated leading strand telomeric PNA probe (red) and FITC‐conjugated lagging strand telomeric PNA probe (green). DNA was counter‐stained with DAPI (blue).B The frequency of chromosome‐type telomeric fusions in cells treated as in (A) was quantified as a percentage of total number of chromosomes (illustrated in Supplementary Fig S2A). A minimum of 2,000 chromosomes were scored for each treatment. Error bars represent SD of at least two independent experiments. P‐values were calculated using an unpaired two‐tailed t‐test. *P ≤ 0.05; **P ≤ 0.01; NS, P > 0.05.
Figure 71x106 HCT116 colon carcinoma cells were implanted subcutaneously. Animals were treated intraperitoneally with vehicle, LXR623 (100-200 mg/kg), ABT263 (100 mg/kg) or both agents (3 days per week for 1.5 weeks. Tumor growth curves show the development of tumor size for each treatment group. Scatter plots display the quantitative representation of the tumor size among the different treatments toward the end of the experiment. Shown are means and SD (n ≥ 5). *p=0.022, **p=0.0063, ****p<0.0001. Statistical significance was determined by one-way ANOVA. B, E, GBM43 PDXs were implanted subcutaneously. Animals were treated intraperitoneally with vehicle, LXR623 (100 mg/kg), ABT263 (75 mg/kg) or both agents (3 days per week for 1.5 weeks). Tumor growth curves show the development of tumor size for each treatment group. Scatter plots display the quantitative representation of the tumor size among the different treatments toward the end of the experiment. Shown are means and SD (n ≥ 5). *p=0.022, **p=0.0031. Statistical significance was determined by one-way ANOVA. C, F, A375 BRAF V600E mutated melanomas were implanted subcutaneously. Animals were treated intraperitoneally with vehicle, LXR623 (100 mg/kg), ABT263 (75 mg/kg) or both agents (3 days per week for 1.5 weeks). Tumor growth curves show the development of tumor size for each treatment group. Scatter plots display the quantitative representation of the tumor size among the different treatments toward the end of the experiment. Shown are means and SD (n ≥ 5). *p=0.0109 (GW3965 vs Combination), *p=0.0166 (ABT263 vs Combination), **p=0.0047. Statistical significance was determined by one-way ANOVA. Related to the HCT116 colonic xenograft model, representative histopathological images (hematoxylin and eosin stain) are shown from each treatment group. Scale bar: 50 µm. Sections from the same treatment groups as in G were stained with TUNEL and SDHB, repectively. Displayed are representative images of the staining. Scale bar 50 µM. Sections from the experiment in 7A were quantified for apoptotic bodies , Sections from the experiment in 7A were quantified for , number of TUNEL positive cells Sections from the experiment in 7A were quantified for number of mitosis per multiple high-power fields.
Figure 3A. Representative intracellular Ca2+ signals in hTRPA1-expressing HEK293 cells in response to different concentrations of iodine. 2-APB, a TRPA1 agonist, was subsequently applied to fully activate TRPA1. RFU: relative fluorescence unit. B. Concentration-response relationships of iodine-induced intracellular Ca2+ increase in HEK293 cells expressing WT or mutant TRPA1 channels. Data are presented as mean ± s.e.m. n ≥ 8 for each construct at each concentration. The smooth curves are fits to the Hill equation. C. Time course of iodine-induced currents in hTRPA1-expressing Xenopus oocytes. HC: HC030031 D. Concentration-response relationship of iodine-induced currents in hTRPA1-expressing Xenopus oocytes. Data are presented as mean ± s.e.m. n ≥ 8 for each concentration. The smooth curve is a fit to the Hill equation. E. Time course of intracellular iodine-induced macroscopic currents in an inside-out patch from hTRPA1-expressing HEK293 cell (n = 3). F. Time course of iodine-induced currents in a hTRPA1-expressing Xenopus oocyte, which presents the current reduction upon DTT treatment (n = 3). G, H. Comparisons of the iodine-induced whole-cell current in HEK293 cells expressing WT hTRPA1 (G) or the triple mutant channel (C421S/K710R/C856S) (H) (n ≥ 4). I. Representative whole-cell currents in HEK293 cells expressing human TRPV1 in response to iodine and subsequently applied capsaicin (n = 3).
Figure 3Schematic diagram for generating PAX6D knockout (PAX6D KO). The exon α was replaced by PGK-Puromycin. Primers indicated in this panel are used to detect the PAX6D KO. qPCR verification of PAX6D KO efficiency. PAX6A&B and PAX6D isoforms were also evaluated by qPCR with isoform specific primers (n=3 biological replicates for each group). Western blot detection of PAX6 isoforms in wildtype and PAX6D KO cells. qPCR quantification of retinal gene VSX2 in PAX6D KO cells (n=3 biological replicates for each group).   qPCR quantification of retinal gene SIX6 in PAX6D KO cells (n=3 biological replicates for each group).   Immunostaining of VSX2 in wildtype and PAX6D KO cells. Scale bars:100 µm. Samples were from day 30 differentiating retinal cells. GO analysis of differentially expressed genes in PAX6 KO cells. Up-regulated genes and down-regulated genes are analyzed separately. The biological processes are ranked by p-value. Only top 15 terms were shown. GO analysis of differentially expressed genes in PAX6D KO cells. Up-regulated genes and down-regulated genes are analyzed separately. The biological processes are ranked by p-value. Only top 15 terms were shown. Venn diagram shows the overlap of differentially regulated genes in PAX6 KO and PAX6D KO cells. The numbers indicate the number of genes in each area. Also see Figure EV3C for detailed GO analysis of genes unique in PAX6 KO cells. GO analysis of differentially expressed genes in PAX6D KO cells. Up-regulated genes and down-regulated genes are analyzed separately. The biological processes are ranked by p-value. Only top 15 terms were shown. GO analysis of biological process for overlap genes in panel I. The numbers in the cycle are the ratio of genes. The GO terms are ranked by p-value in the clockwise direction.
Tfcp2l1 deletion impairs fatty acid oxidation in 2i-ESCs.Figure 3A. Real-time fatty acid oxidation (FAO) profile of WT and Tfcp2l1_KO 2i-ESCs using the FAO stress test kit in Seahorse XF. Oxygen consumption rate (OCR) were measured. Error bars indicate SEM; n = 5 biological repeats. Palm-BSA, palmitate conjugated BSA, a substrate of FAO; Oligomycin, an inhibitor of ATP synthase (complex V); FCCP, Carbonyl cyanide-4 (trifluoromethoxy) phenylhydrazone, a proton gradient uncoupler; Rotenone and Antimycin, a complex I inhibitor and a complex III inhibitor, respectively. B. Basal respiration, maximal respiration, and spare respiration capacity were calculated from A. n=5 biological repeats; error bars indicate SEM; p values were calculated using student t-test. C, D. Mito Stress Assay (C) and Glycolysis Stress Assay (D) in Seahorse XF. Error bars indicate SEM; n=3 biological repeats; no time point shows a statistically significant difference between WT and KO 2i-ESCs. E. Real-time OCR profile using FAO substrate (Palm-BSA) or BSA control followed by FAO inhibitor, etomoxir. n=5 biological repeats; error bars indicate SEM. F. Quantification of OCR changes after etomoxir addition. n=5 biological repeats; error bars indicate SEM; p-values were calculated using two-sample t-test. G. Mitochondrial membrane potential (left) and amount (right) were measured by TMRM and Mitotracker Green, respectively. Representative histograms were shown. The numbers shown are mean fluorescence intensity; p-values were calculated using two-sample t-test.
CRISPR/Cas9 knockout library screening shows NDST3 as a regulator of bafilomycin toxicity.Figure 1(A) A schematic diagram of the CRISPR genetic screen. (B) Generation of a stable NDST3 KO RPE1 cell line. (C) Generation of NDST3 polyclonal antibodies targeting a region of NDST3 unique in the NDST family. (D) Immunoblot analysis of NDST3 in stable NDST3 KO RPE1 cells. (E) Bafilomycin A1 (Baf A1) sensitivity assay for NDST3 KO cells. WT and NDST3 KO RPE1 cell lines were treated with 25 nM Baf A1 for 72 h, and cell survival was measured using calcein AM staining. The cell survival was calculated as the fluorescence intensity (percentage) of the treated cells relative to the untreated cells (n = 4 independent cultures, ****P < 0.0001). Data information: Error bars represent ± standard deviation. Scale bar, 200 μm. (F) Detection of Flag-NDST3 and Flag-EGFP overexpressed in NDST3 KO cells by immunoblotting with anti-Flag antibodies. (G) Baf A1 sensitivity assay for NDST3 KO cells after the NDST3 restoration. The NDST3 KO cells with stable expression of Flag-NDST3 or Flag-EGFP (control) were treated with 25 nM Baf A1 for 72 h, and cell survival was measured using calcein AM staining. The cell survival was calculated as the fluorescence intensity of the treated cells relative to the untreated cells (n = 4 independent cultures, ****P < 0.0001). Data information: Error bars represent ± standard deviation. Scale bar, 200 μm.
Figure 3(A) Immunofluorescent staining of C/EBP β (red) and AEP (green), Iba-1 (red) and AEP (green), cleaved Tau N368 (red) and AT8 (green), cleaved APP C856 (red) and Aβ (green), T22 (red) and AT8 (green) in cerebral cortex of brains from DSS-treated mice. Scale bar: 20 μm. White arrows indicate that C/EBP β and AEP were co-localized in mouse brain. (B) Quantitative analysis of AEP positive cells and C/EBPβ positive cells, respectively. The density of both AEP positive cells and C/EBPβ positive cells were significantly decreased through knocking down C/EBPβ or knocking out AEP in mice. Representative data of five samples, data are shown as mean± SEM. *P =0.0390 (*P < 0.05), ***P =0.0002 (***P < 0.001), ****P < 0.0001, one-way ANOVA. (C) Quantitative analysis of AT8 positive cells (left) and TauN368 positive cells (right), respectively. The density of both AT8 positive cells and TauN368 positive cells were significantly decreased through knocking down C/EBPβ or knocking out AEP in mice. n=5 in each group, data are shown as mean± SEM. *P =0.0194 (*P< 0.05), ***P =0.0001 (***P < 0.001), ****P < 0.0001, one-way ANOVA. (D) Quantitative analysis of Aβ positive cells (left) and APPC586 positive cells (right), respectively. The fluorescence intensity of Aβ positive cells and the density APPC586 positive cells were significantly decreased through knocking down C/EBPβ or knocking out AEP in mice. Representative data of five samples, data are shown as mean± SEM. *P=0.0107 (*P < 0.05), **P=0.0048 (3xTg vs 3xTg AEP KO), **P=0.0044 (3xTg vs WT) (**P< 0.01), ***P=0.0006 (***P< 0.001), ****P < 0.0001, one-way ANOVA. (E) Quantitative analysis of AT8 positive cells (upper) and T22 positive cells (lower), respectively. The fluorescence intensity of both AT8 positive cells and T22 positive cells were significantly decreased through knocking down C/EBPβ or knocking out AEP in mice. Representative data of five samples, data are shown as mean± SEM. **P=0.0014 (3xTg vs 3xTg C/EBPβ+/-), **P=0.0030 (3xTg vs 3xTg AEP KO) (**P<0.01), ***P=0.001, ****P<0.0001, one-way ANOVA. (F) Representative electron microscopy of the synaptic structures in hippocampus of brains from DSS-treated mice. Red stars indicate the synapses. (Scale bar: 1 μm). (G) The dendritic spines from the apical dendritic layer of the cerebral cortex region were analyzed by Golgi staining. (Scale bar: 5 μm). (H) Quantitative analysis of the synaptic densities in DSS-treated mice. Representative data of five samples, Data are shown as mean ± SEM. ****P < 0.0001, one-way ANOVA. (I) Quantitative analysis of the spine density. Representative data of five samples, Data are shown as mean ± SEM. ****P < 0.0001, one-way ANOVA.
Figure 2A. Pooled SCN cells generate a combined dataset of 17,363 cells. The sequenced SCN cells carry explicit barcodes which were used to back-reference sequencing run identity. This yields a clear-cut segregation into two transcriptionally distinct cell groups. Left: 98.6% of cells in the yellow cluster are CT7 5 cells. Right: 98.8% of cells in the grey cluster are CT15.5 cells. B. Volcano plot showing the magnitude of differential gene up-regulation, as measured by the respective log 2-fold change in expression level, at day (positive) or night (negative) versus statistical significance, as quantified by a negative binomial exact test (red dots, dashed line p<0.05 adjusted for multiple testing using Benjamin-Hochberg). Known circadian-regulated genes are labelled.
Figure 1A Range of example RAD51 nuclear expression score (RAD51NES) values with respective unmixed monochrome fluorescent IHC staining images. EpCAM is used as a tumour marker and an internal sample quality control. Scale bar is 50μm. B Correlation of RAD51NES with two independent pathologist H‑scores (top left and top right) and correlation of two pathologist with each other (bottom). Pearson correlation. C Distribution of RAD51 nuclear expression score (RAD51NES) in the BCC cohort. The cohort is divided into RAD51-Low expressing cases (first quartile, Q1 - blue), intermediate cases (interquartile range, IQR - grey) and RAD51-High expressing cases (fourth quartile, Q4 - brown). Dashed line denotes the median RAD51NES in this cohort. D Survival analysis of the BCC cohort. Kaplan-Meier plots for progression-free survival (PFS) (left) and overall survival (OS) (right) stratified according to fourth quartile (Q4) and first quartile (Q1) of RAD51NES. Log-rank test, shading denotes 95% confidence intervals. E Number of cases with progression at 12 and 24 months. Chi-square test.
The human RAB13 3'UTR contains a functionally conserved GA-rich region required for peripheral localization.Figure 4A. Schematic showing %GA content and positions along the human RAB13 3'UTR targeted by the indicated PMOs. Red rectangles indicate the location of GA-rich motifs. Graphs present PDI measurements of RAB13 RNA (upper panel) or NET1 RNA (another APC-dependent RNA; bottom panel) detected in MDA-MB-231 cells treated with the indicated PMOs. PDI=1 indicates a diffuse distribution. p-values: **<0.01, ***<0.001, ****<0.0001 by analysis of variance with Dunnett's multiple comparisons test. n.s.: non-significant. N=30-73 cells observed in 3-5 independent experiments. Bars: mean ± s.e.m.. B. Representative FISH images of cells treated with the indicated PMOs. Arrows point to peripheral RNA. Arrowheads point to perinuclear RNA. Boxed regions are magnified in the insets. Note that RAB13 RNA becomes perinuclear in cells treated with PMOs against the GA-rich region, while NET1 remains localized at protrusions. Scale bars: 10 μm (4 μm in insets). C. RAB13 protein levels were measured by quantitative Western blot and normalized to total α-Tubulin or GAPDH levels (for representative blot, see Figure 5E). Relative levels in RAB13 PMO-treated compared to control are shown. No significant differences were detected by Kruskal-Wallis test with Dunn's multiple comparisons test. N=3-8. Bars: mean ± SD.
Figure 6A. Different invasion stages of P. falciparum merozoites expressing Cb-EmeraldFP. DAPI labels the nucleus (DNA), Cb-EmeraldFP labels actin filaments (CB-EME, green) and anti-RON4 labels the junction (red). Left panel: onset of invasion. Middle panel: A merozoite in the middle of invasion; note the constriction of the nucleus (blue) as it passes through the RON4 junction and F-actin colocalising with it (green). Right panel: A merozoite completing invasion; note the F-actin ring beyond the RON4-junction and actin filaments surrounding the nucleus. Data information: Scale bar represents 1 µm for Plasmodium falciparum B. IFA showing various stages of invasion of erythrocytes by P. knowlesi merozoites. DAPI labels the nucleus (DNA), anti-actin Antibody labels actin filaments (anti-actin, green) and the invasion junction is marked with a C-terminally tagged PkRON2 mCherryHA (red). Left panel: a merozoite beginning invasion; note the formation of an F-actin ring (green) behind the PkRON2 junction (red). Middle panel: A merozoite in the middle of the invasion event; note the constriction of the nucleus (blue) as it passes through the PkRON2 junction and F-actin colocalising with it (green). Right panel: A merozoite completing the process of invasion; note the F-actin ring beyond the PkRON2-junction and actin filaments colocalising with the nucleus. Data information: Scale bar represents 3 µm for Plasmodium knowlesi. C. Super-resolution images showing various stages of invasion of erythrocytes by P. falciparum merozoites expressing Cb-EmeraldFP. Left panel: DAPI labels the nucleus (blue), Cb-EmeraldFP labels actin filaments (CB-EME, green) and the invasion junction is marked with an anti-RON4 antibody (red). Brightfield images have been marked with red dots depicting the junction, black dots depicting merozoite boundary still outside the erythrocyte and white dots depicting a merozoite that has penetrated the host cell. Top panel shows an attached merozoite beginning the process of invasion, the second panel during the process of invasion and the bottom two panels show merozoites completing the process of invasion. Right panel: Schematic depicting merozoite invasion into red blood cells. Letters depict the corresponding event in the model (left) and microscopy pictures (right). White arrows depict actin filaments that colocalise with the nucleus. D. Quantification of Plasmodium falciparum invasion events categorising actin-TJ and actin-nucleus association events. 121 events were counted in total. Data information: Error bars represent standard deviation. Scale bar represents 1 µm for Plasmodium falciparum
Hwa degraded through lysosomal pathway.Figure 1A Illustration of experiment design. hwa mRNA (100 pg) was injected into oocytes at stage VI. Embryos were treated with 100 nM Baf-A1 or 20 μM MG132 at 1-cell stage and collected at indicated stages. B Western blot analysis for Hwa performed with lysates from embryos treated as indicated in (A). 1cs, 1-cell stage; 2cs, 2-cell stage; st6, 32-cell stage; st8.5, blastula stage. Blots are representative of 3 independent experiments. C Densitometry analysis of Hwa bands from the western blot in (B) for corresponding lanes normalized to α-tubulin. n = 3 independent experiments, mean ± SEM, *p < 0.05, **p < 0.01 by unpaired t test. D Western blot analysis of exogenous Hwa in HEK293T cells. Treatment of cells with lysosome inhibitors bafilomycin A1 (Baf-A1) and NH4Cl for 24 hours, DMSO was used as control. Dosage of Baf-A1: 10 nM, 50 nM, 100 nM; Dosage of NH4Cl: 2.5 mM, 5 mM, 10 mM. Hwa with modification is indicated by white arrow; Degraded Hwa is indicated by black arrow. Blots are representative of 3 independent experiments. E Densitometry analysis of Hwa bands from the western blot in (D) for corresponding lanes normalized to α-tubulin. n = 3 independent experiments, mean ± SEM, **p < 0.01 by unpaired t test.