EMBO/sd-character-level-ner huggingface dataset:
Figure 6(A) Control or NRF2 siRNA transfected HeLa cells lysates were subjected to western blotting with indicated antibodies. (B, C) RNA isolated from untreated or MG132-treated (20 µM, 2h) control or NRF2 siRNA transfected cells were subjected to qRT-PCR with primers of genes as indicated. The fold induction in MG132 treated samples is calculated relative to untreated samples. Mean ±SD, n=3, *p < 0.05 (Student's unpaired t-test). Data information: Unless otherwise stated, scale bar: 10 µm. WB analysis of control siRNA or NRF2 siRNA transfected lysates of HeLa cells, untreated or treated with MG132 (20 µM, 2h) as indicated and probed with antibodies as shown. L.E, Low exposure; H.E, High Exposure. WB analysis of control siRNA or NRF2 siRNA transfected lysates of HeLa cells, untreated or treated with H2O2 (200 µM, 2h) as indicated and probed with antibodies as shown. L.E, Low exposure; H.E, High Exposure. Left panels: Representative confocal images of control siRNA and NRF2 siRNA transfected cells treated with (F) MG132 (20 µM, 2h), where IF was conducted with Ub and p62 antibodies. Right panels: fluorescence intensity line tracing corresponding to a white line in zoom panel. Left panels: Representative confocal images of control siRNA and NRF2 siRNA transfected cells treated with (G) H2O2 (200 µM, 2h), where IF was conducted with Ub and p62 antibodies. Right panels: fluorescence intensity line tracing corresponding to a white line in zoom panel. (H) Hela cells, TRIM16KO cells, and TRIM16KO cells complemented with Myc-NRF2 or GFP-NRF2 were treated with MG132 (20 µM, 2h) and subjected to WB analysis with Myc, GFP and indicated antibodies. (I) Representative confocal images of Hela cells, TRIM16KO cells, and TRIM16KO cells complemented with Myc-NRF2 which were treated with MG132 (20 µM, 2h) and subjected to IF with Ub and p62 antibodies. (J) Graph represent % of cells with Ub-p62 dots, data from ≥ 10 microscopic fields (40X), n=3, mean ±SD, *p < 0.005 (one-way ANOVA test). Data information: Unless otherwise stated, scale bar: 10 µm. (K) Graphical representation of results
Figure 2A,B Tumor growth curve (left) and summary graph of end-point tumor masses (right) of wildtype (WT) and Peli1-TKO (TKO) mice inoculated s.c. with B16-OVA (A; WT, n=5; KO, n=5) or E.G7 (B; WT, n=6; KO, n=4) tumor cells. C Flow cytometry analysis of IFNγ-producing CD4+ and CD8+ T cells in TILs and dLN cells from B16OVA tumor-bearing wildtype and Peli1-TKO mice, presented as a representative plot (upper) and summary graph (lower). (WT, n=5; KO, n=5) D Flow cytometry analysis of GZMB+CD8+ T cells in TILs from B16-OVA tumor-bearing wildtype and Peli1-TKO mice, presented as a representative plot (upper) and summary graph (lower). (WT, n=5; KO, n=5) E Tumor growth curve (left) and summary graph of end-point tumor masses (right) of B16-OVA tumor-bearing wildtype and the indicated Peli1 conditional KO mice. (Treg KO model: WT, n=5; Treg KO, n=6; MKO model: WT, n=3; MKO, n=5; BKO model: WT, n=9; BKO, n=9)
a) Mosaic clones of dUTX1 (hsFLP; ubi-GFP FRT40A/dUTX1 FRT40A) from third instar larval salivary glands with wild-type cells marked by nuclear-localized GFP (green).f7(a) Mosaic clones of dUTX1 (hsFLP; ubi-GFP FRT40A/dUTX1 FRT40A) from third instar larval salivary glands with wild-type cells marked by nuclear-localized GFP (green). dUTX1 clones have high levels of H3K27me3 (red) compared with the neighbouring control cells marked by GFP. Scale bar represents 50 μm. (b) Western blot analysis of the level of H3K27me3 in control and dUTX1 larval salivary glands. Alpha-tubulin and H3 serve as loading controls. (c) Expression of dUTX wild-type (dUTXWT) and dUTX catalytic mutant (dUTXJmj) in the salivary glands (using Sgs3-GAL4 driver) of dUTX1. Histology analysis of paraffin sections at 24 h RPF shows intact salivary glands present in dUTX1 and in dUTX1; dUTXJmj* compared with control and dUTX1; dUTXWT at 24 h RPF. Ovals indicate the position of salivary glands and fragments. Scale bar represent 50 μm. Quantification of the salivary gland phenotypic data at 24 h RPF is shown on the right. (d) A schematic representation of the role of dUTX in ecdysone-mediated transcriptional regulation of cell death genes, directly or via E93.
Figure 5Schematic of the experiment. K and KM animals were set on doxycycline food until mammary tumours developed. Then switched to a normal diet and non-regressed tumours collected. Single cells were then injected into Rag2-/- animals to recapitulate the non-regressed tumours followed by drug treatment or vehicle control treatment. Relative volume of tumours grown in Rag2-/- animals after treatment with Tepotinib or vehicle control. One-way ANOVA, Tukey's multiple comparison. Ns: not significant; ** p<0.024. Mean +/- SEM. A total of 19 Rag2-/- mice injected with cMet expressing tumours were treated with Tepotinib and 14 used as controls. For tumours with no cMet, 13 Rag2-/- mice were treated with Tepotinib and 9 were used as control. Immunostaining of phospho-cMet in the same tumours grown in Rag2-/- mice after treatment with Tepotinib or vehicle control. Scale bar 200 µm. Quantification of pH3 in tumours grown in Rag2-/- animals after 5 days treatment with the cMet inhibitor Tepotinib or vehicle control. Quantification of caspase 3 in tumours grown in Rag2-/- animals after 5 days treatment with the cMet inhibitor Tepotinib or vehicle control.
Figure 2(A) Representative images of P4 and P6 mice of the indicated IκBα genotypes. Quantification of the animal weight (B) from 50 WT/HET and 15 KO mice analyzed. Data information: bars represent mean values ± standard error of the mean (s.e.m); p values were derived from an unpaired t-test, two-tailed, ****p-value<0.0001, ***p-value<0.0005, * p-value<0.05, n.s. no significant. Quantification of the intestinal length (C) from 50 WT/HET and 15 KO mice analyzed. Data information: bars represent mean values ± standard error of the mean (s.e.m); p values were derived from an unpaired t-test, two-tailed, ****p-value<0.0001, ***p-value<0.0005, * p-value<0.05, n.s. no significant. D-G (D) Immunohistochemical (IHC), IF ­and Alcian Blue staining with quantification of P6 IκBα WT and KO intestines (D), 2 month-old IκBβ;IκBε KO (E) and 2 month-old IκBαNES mice (F and G). A minimum of 50 crypts/villus was counted per genotype (3 mice). Data information: bars represent mean values ± standard error of the mean (s.e.m); p values were derived from an unpaired t-test, two-tailed, ****p-value<0.0001, ***p-value<0.0005, * p-value<0.05, n.s. no significant. Scale bars in D, E, F and G, 50 μm.
AAV2.Figure 4Vector mediated expression after intravitreal injection in ~3-year old cynomolgus macaque eyes. (A-B) In vivo cSLO examination images collaged to provide NHP fundus overview at 8 WPI of E12 total vg (in 38.9-100 µl) of sc-CMV-eGFP packaged with AAV2.GL and AAV2.NN capsid variants, respectively. (C-D) Confocal scans of immunolabelled retinal cross sections (14 μm) of the fovea centralis from AAV2.GL and AAV2.NN treated animals, respectively. (E) Percentage quantification of eGFP+ photoreceptors in the ONL of foveal cross sections. AAV2.GL technical repeats n=4, AAV2.NN technical repeats n=3. (F-G) Confocal scans of immunolabelled retinal cross sections (14 μm) of the periphery from AAV2.GL (F) and AAV2.NN (G) treated retinae. (Η-Ι) Higher magnification confocal scans of immunolabelled peripheral retina. ONL, outer nuclear layer; INL, inner nuclear layer
Figure 4A. 293T cells were transfected with USPs cDNA or control plasmid. After 36 hr of transfection, cells were treated with 50 ng/ml Wnt-3a for 4 hr. FoxM1 protein levels in the cells were then determined by Western blotting. B. 293T cells were transfected with 1 µg control plasmid or USP5 cDNA in 6-well plates. After 36 hr of transfection, cells were treated with 100 µg/ml CHX for 0, 2, 4, or 6 hr. FoxM1 protein levels in the cells were then determined by Western blotting. C. GSK3-β CA, USP5, or control plasmid was co-transfected with Flag-FoxM1 into 293T cells. After 36 hr of transfection, cells were treated with 50 ng/ml Wnt-3a or control PBS for 4 hr. Cell lysates were subjected to IP with USP5 antibody and followed by IB with Flag or USP5 antibody. D. GSK3βCA, USP5, or control plasmid was co-transfected with Flag-FoxM1 into 293T cells. After 36 hr of transfection, cells were treated with 50 ng/ml Wnt-3a or control PBS for 6 hr. Cell lysates were subjected to USP5 with Flag antibody and followed by USP5 with Flag or USP5 antibody. E. USP5 and Flag-FoxM1 (WT) or mutants Flag-FoxM1S474A or Flag-FoxM1S474E plasmids were co-transfected into 293T cells for 36 hr. Cell lysates were subjected to IP with Flag antibody and followed by IB with Flag or USP5 antibody. F. GSK3β CA, USP5, or control plasmid was co-transfected with Flag-FoxM1 (WT) or mutants Flag-FoxM1S474A into 293T cells for 36 hr. Cell lysates were subjected to IP with Flag antibody and followed by IB with Flag or USP5 antibody. G. HA-ubiquitin and Flag-FoxM1 were co-transfected with or without GSK3β CA into 293T cells. After 36 hr, cells were treated with 25 nM MG132 for 6 hr. Cell lysates were subjected to IP with Flag followed by IB with HA or FoxM1 antibody. H. HA-ubiquitin and Flag-FoxM1 were co-transfected with or without siUSP5 into U87 cells. After 36 hr, cells were treated with 50 ng/ml Wnt-3a and 25 nM MG132 for 6 hr. Cell lysates were subjected to IP with Flag antibody followed by IB with HA or FoxM1 antibody.
a) [3H]thymidine incorporation at 72 h by wild-type (WT) and Irgm1−/− CD4+ T cells activated with soluble mAb to CD3 and irradiated syngenic wild-type splenocytes.figf1(a) [3H]thymidine incorporation at 72 h by wild-type (WT) and Irgm1-/- CD4+ T cells activated with soluble mAb to CD3 and irradiated syngenic wild-type splenocytes. (b) [3H]thymidine incorporation at 72 h by OT-II and Irgm1-/- OT-II CD4+ T cells stimulated with OVA protein (OVA) or OVA peptide of amino acids 323-339 (OVA(323-339)) in the presence of syngenic splenic DCs. (c,d) Kinetic analysis of [3H]thymidine incorporation (c) and cell numbers (d) in cultures of OT-II and Irgm1−/− OT-II spleen cells stimulated with OVA peptide. (e) Flow cytometric analysis of the activation and proliferation of OVA peptide-stimulated OT-II and Irgm1-/- OT-II CD4+ T cells pulsed with BrdU during the final 4 h of 72 h of culture and then stained with mAb to CD25 and BrdU. Numbers in quadrants indicate percent viable CD4+ cells. (f) Enzyme-linked immunosorbent assay of IL-2 produced by OT-II and Irgm1-/- OT-II CD4+ T cells stimulated with OVA peptide in the presence of splenic DCs, measured at 72 h. Data are representative of more than five (a,b) or three (c-f) independent experiments with similar results (mean and s.d. of triplicate cultures, a-d,f).
Figure 5A, B Western blot analysis (A) and quantification (B) of NB-3 expression in cultured pyramidal neurons infected with lentivirus encoding negative control (LV-nc-GFP) and that encoding NB-3 shRNA (LV-NB-3 shRNA-GFP). NB-3 expression levels were normalized by b-tubulin. ***p < 0.001; one sample t-test. Data were analyzed from 5 independent experiments. C-E Sagittal sections showing the BDA-labeled corticospinal axons in injured spinal cords following cortical injection of LV-nc-GFP (C) and LV-NB-3 shRNA-GFP (D). High-magnification image (E) of the area in (D) (asterisk).F, J Quantification of the intensity index of BDA-labeled axons at certain distances from the lesion border in (C, D, G, and H). *p < 0.01; two-way ANOVA followed by Fisher's LSD. n= 19 mice per group in (F), and n= 17 mice per group in (J). G-I Sagittal sections showing the corticospinal axons in injured spinal cords following injection of LV-nc-GFP (G) and LV-NB-3 shRNA-GFP (H) into the lesion sites. High-magnification image (I) of the area in (H) (asterisk). The arrows in (C-E and G-H) indicate the lesion sites; the asterisks in (D and H) indicate the lesion epicenters; the white dashed lines in (D and H) indicate the regenerative corticospinal axons extending into the distal spinal cord.F, J Quantification of the intensity index of BDA-labeled axons at certain distances from the lesion border in (C, D, G, and H). *p < 0.01; two-way ANOVA followed by Fisher's LSD. n= 19 mice per group in (F), and n= 17 mice per group in (J).
Figure 8Live-cell Airyscan images of laser-induced depolarization time series of Mic10 KO Hap1 cell stained with TMRE. Mitochondrion exposed to rapid, high 2-photon laser power in region of white box, arrow (~0.3 sec); large, hyperpolarized vesicle (arrowhead) remains polarized after the rest of the mitochondrion loses ΔΨm, indicating electrochemical discontinuity between hyperpolarized vesicles and the rest of the mitochondrion, including the IBM. Scale bar = 500 nm. N = 2 independent experiments. Live-cell Airyscan image of time series showing hyperpolarized vesicles (arrowheads; region cropped from dashed white box) inside mitochondria lacking both Opa1 and Drp1. Scale bar = 500 nm. N = 2 independent experiments. Free movement of hyperpolarized vesicles within the matrix indicate their detachment from the IBM, representing an extreme case of cristae independency within one mitochondrion (see Movie EV 1 for time series).
Figure 7A. Bar graph representing relative expression of VEAL2 (in fold change) in choroid tissue isolated from control and diabetic retinopathy (DR) patients. Data obtained from 8 biological replicates (patients) and represented as individual values with mean fold change ± standard deviation. B. Bar graph representing relative fold change of VEAL2 expression in blood samples of patients with different diabetic stages with aggravating vascular dysfunctions from diabetic melitus (DM) to non proliferative diabetic retinopathy (NPDR) to proliferative diabetic retinopathy (PDR) compared to control patients. Data was collected from 50 different patients in each condition and represented as individual values with mean fold change ± standard deviation. C. ROC curve shows sensitivity and specificity of VEAL2 as a diagnostic biomarker for proliferative diabetic retinopathy with endothelial dysfunction. D. Complementation of VEAL2 and veal2 reverted increased permeability levels in the HUVEC monolayer model for hyperglycemia. Bar graph representing the effect of overexpression of VEAL2 and veal2 on permeability levels in hyperglycemia disease model, measured as efflux of dextran conjugated FITC. Data obtained from 3 different biological replicates and plotted as mean percentage fold change values ± standard deviation. E-S. Modeling hyperglycemia in HUVEC resulted in dysregulation of junctional assembly of CDH5 and CTNNB1 proteins and increased membrane localization of PRKCB protein. Complementation of VEAL2 and veal2 in hyperglycemic conditions reverted junctional disassembly of CDH5 and CTNNB1 and also kept PRKCB in cytoplasm to mitigate pathological conditions associated with hyperglycemia. (E-H,Q) CDH5 protein, (I-L,R) CTNNB1 protein, (M-P,S) PRKCB protein. (E-P) Magnification-60X, Scale bar-15μm. Arrowheads indicate representation of signals of proteins in HUVECs. (Q-S) Data from cells of different fields of 3 technical replicates of 1 biological replicate is presented as representation. Data is shown as individual values; the middle bar represents the mean and the error bar represents ± standard deviation.
Proposed model on GBS pigment-mediated host cell lysis and preterm birth.Figure 8The GBSpigment intercalates into host cell membranes leading to membrane permeability and ion flux. In RBC, this leads to colloidal osmotic lysis. In macrophages, the loss of intracellular potassium serves as a trigger for NLRP3 inflammasome formation. The activated inflammasome then activates caspase 1, which cleaves pro-IL-1β and pro-IL-18 to their active forms. Finally, caspase 1 activation causes cell death by pyroptosis. While NLRP3-dependent pyroptosis is the major form of cell death observed in macrophages exposed to purified pigment and hemolytic/pigmented GBS strains, hyperpigmented GBS strains induced low levels of cell death in NLRP3-deficient and ASC-deficient macrophages that were independent of inflammasome and caspase 3/7. In host cells that are either resistant or recover from pigment-induced membrane damage (e.g., NLRP3-deficient macrophages), we predict that lysosomal degradation of pigment or shedding of vesicles that contain pigment-membrane complexes and the turnover of plasma membrane prevents cell death. The GBSpigment exacerbates fetal injury and preterm birth through the combined action of colloidal osmotic lysis and pyroptosis.
Figure 4 A) Representative western blot of autophagy markers at steady state in skeletal muscle. GAPDH was used as a loading control. B) Densitometric analysis of immunoblots similar to that shown in A (n=4/group). Black: untreated WT; grey: rapamycin-treated WT; red: untreated Cox15sm/sm; blue: rapamycin-treated Cox15sm/sm. Error bars represent SEM. The asterisks represent the significance levels calculated by one-way ANOVA with Tukey post-hoc multiple comparison test **p=0.0068 (LC3-II WT vs. Cox15sm/sm), **p=0.0063 (P62 WT vs. Cox15sm/sm) *p=0.025 (P62 Cox15sm/sm vs. Cox15sm/sm+rapamycin), **p=0.0098 (S6 WT vs. Cox15sm/sm), *p=0.034 (P-S6 WT vs WT+rapamycin), ***p=0.0002 (Cox15sm/sm vs. Cox15sm/sm+rapamycin), *p=0.014 (P-S6 WT vs. WT+rapamycin) ***p=0.0007 (Cox15sm/sm vs. Cox15sm/sm+rapamycin)  C) Analysis of the autophagic flux (n=3/group). Upper panel: representative western blot of Cox15sm/sm and WT muscles treated with colchicine, rapamycin, or rapamycin plus colchicine. Note that LC3-II was increased in Cox15sm/sm vs. WT samples. Colchicine did not increase LC3-II levels suggesting a block in the autophagic flux; rapamycin plus colchicine treated Cox15sm/sm mice showed higher levels of LC3-II, suggesting that rapamycin increased the autophagic flux in Cox15sm/sm muscles. Black: WT; dark grey: colchicine-treated WT; middle grey: rapamycin-treated WT; light grey: colchicine plus rapamycin treated mice; red: untreated Cox15sm/sm; orange: colchicine-treated; blue: rapamycin-treated Cox15sm/sm; purple: colchicine plus rapamycin treated Cox15sm/sm. Error bars represent SEM. The asterisks represent the significance levels calculated by one-way ANOVA with Tukey post-hoc multiple comparison test **p=0.0049 (WT vs. WT+colchicine), **p=0.0046 (WT+rapamycin vs. WT+rapamycin+colchicine), *p=0.0225 (WT vs. Cox15sm/sm), **p=0.0057 (Cox15sm/sm colchicine vs. Cox15sm/sm+rapamycin+colchicine), ***p=0.0003 (Cox15sm/sm rapamycin vs. Cox15sm/sm+rapamycin+colchicine)
Figure 1Treatment of HeLa cells with DFP (1mM for 24 h) reduces the conjugation of SUMO-2/3 (n=5, biological replicates; **, p<0.01; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) does not change global SUMO-1-ylation (n=5, biological replicates; p>0.05; N.S., not statistically significant; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) leads to increased levels of SENP3 (n=8, biological replicates; **p<0.01; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) does not change levels of SENP5 (n=5, biological replicates; p>0.05; N.S., not statistically significant; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) leads to decreased levels of SENP3 mRNA (n=3, biological replicates; *p<0.05; unpaired t-test). Treatment of DFP (1mM for 24 h)-treated HeLa cells with 50μM MG132 for the final 6 h prevents further increase in SENP3 levels induced by DFP. DFP treatment increases SENP3 stability. DFP (1mM for 24 h)-treated HeLa cells were treated with cycloheximide (CHX; 50μg/ml) for the indicated durations in a chase experiment. Lysate samples were separated on different lanes in the same gel and immunoblotted as indicated.
Figure 2(A) Immunoblot analysis for BIK1-HA in Arabidopsis protoplasts exposed to Pep2 or flg22 for 10 min. Arrowheads indicate phosphorylated BIK1-HA. (B) Immunoblot analysis for Pep2-triggered MAPK activation. Ponceau S-stained loading controls are shown (bottom). (C) Root length (averages ± SD, n ≥ 15) of Arabidopsis seedlings that were exposed to 100 nM Pep2. (*, p < 0.05 in two-tailed tests compared to the differences (± Pep2) from the corresponding values of bak1-3 plants. Two independent experiments were combined for statistical analysis.)
Figure 5(A-C) Transmission electron microscopy of β-cells from Abca12tm1d and control mice at 8 weeks of age. Panels A'-C' depict higher resolution images from the boxed regions in A-C (scale bar = 1μm). (D) Quantitation of the area of 3000 granules from each genotype (mean±SEM, n=4, 3 and 3 animals respectively, *p=<0.05 of cre relative to tm1d, Students t-test). (E) Relative frequency distribution of granule areas from (D). (F) Assessment of total cholesterol (COH) by mass spectroscopy in mice fed normal chow or a high cholesterol diet (HCD)(mean±SEM, n=5 and 4 animals respectively, **p=<0.01, Students t-test). (G, H) Quantification of change in mean granule area in Abca12tm1d on chow diet and Abca12tm1d on high cholesterol diet islets assessed by transmission electron microscopy (n=3000 granules/genotype, mean±SEM, n=4, 3, 3 and 4 animals respectively, *p=<0.05, Students t-test) as an average and as a relative frequency distribution. (I) Correction in abnormal granule shape following high cholesterol feeding (bar = 1μm), with I' showing magnification of the boxed region indicated. (J) Glucose tolerance test results following intraperitoneal glucose challenge in Abca12tm1d and cre control mice fed normal chow or HCD (mean±SEM, n=3 mice per genotype, *p=<0.05, for tm1d versus tm1d+HCD, Students t-test). (K) Serum insulin in Abca12tm1d +/- HCD mice after i.p glucose challenge (mean±SEM, n=3 mice/genotype, **p=<0.01, Students t-test).
Figure 6Brown adipocytes were induced differentiation with or without 1mM asparagine. [U-13C] glucose was added into culture media on differentiation day 6 for 30 min. Then tracing analysis from U-13C glucose was performed. Intracellular abundance of 13C-isotopologue metabolites downstream of glycolysis in brown adipocytes. n=5 independent experiments. Data information: Data represent means±SEM. *P<0.05; **P<0.01; ***P<0.001; unpaired two-tailed Student's t test. C,D. Quantification of glucose (C) and lactate (D) concentration from media. The media was collected after incubated with mature brown adipocytes for 48 h. n=3 independent experiments in (C), n=6 independent experiments in (D). Data information: Data represent means±SEM. *P<0.05; **P<0.01; ***P<0.001; unpaired two-tailed Student's t test.