EMBO/sd-character-level-ner huggingface dataset:
Figure 3A. Rpp3018.2 homozygous early oogenesis arrest is partially rescued by p53 mutation. Scale bar,100μm. B. Early oogenesis arrest found in Rpp3018.2 homozygous ovaries is rescued by chk2 mutation (Rpp3018.2 , mnkp6 and Rpp3018.2/ Rpp30PE, mnkp6). Scale bar, 100μm. Star: example of one rescued stage 9 egg chamber. Arrow: example of non-rescued egg chambers. C. Germline clones mutant for Rpp3018.2 were immunostained for PCNA (red). DAPI is in blue. Magnifications: PCNA signal in a control or mutant nurse cell. Scale bar, 10μm. D. Germline clones mutant for Rpp3018.2 were immunostained for Brf (pol-III) (red). DAPI is in blue. A Z-projection of Brf staining is shown. The arrow points Brf aggregates in a mutant chamber. Scale bar, 10μm. E. Early oogenesis arrest found in Rpp3018.2 homozygous ovaries is partially rescued by claspin mutation (Rpp3018.2; claspinEP/TM3). Scale bar, 100μm. F. Quantification of rescued ovaries harboring at least one stage 9 egg chamber in genotypes numbered from 1 to 11 after one day or several days on yeast. Grey bars (1-4) are control flies. n= number of ovaries. G. Top panel. Early oogenesis arrest found in Rpp30182./PEtransheterozygous ovaries is rescued by sqd-S-HAoverexpression. Scale bars: 100µm. Low panel. Rpp3018.2/PE ; sqd-S-HA/sqd-S-HA ovaries were dissected and stained for Orb (green), HA (squid, red) and DAPI (blue). Scale bar, 10µm.
Figure 2A) Cells were treated for 1.5 hours with 5µM CPT, before sequential addition of IdU and CldU for 20 minutes each. Frequency of new firing origins (relative to total structures counted) from Mybl2+/+ and Mybl2Δ/Δ ESCs treated or not with CPT (n=3 independent experiments; Error bars indicate SEM). Statistical analysis using two- tailed unpaired t-test. (B) Distribution of replication rate in Mybl2+/+ and Mybl2Δ/Δ ESC treated or not with CPT as above. Statistical analysis was carried out using unpaired Mann Whitney U tests. A minimum of 200 forks were quantified for each of the genotypes and conditions shown from 3 independent repeats (*p < 0.05; **p< 0.01; ***p< 0.001; **** p< 0.0001). C) ESCs were treated with or without CPT for 4 hours before 0.1ug/ml colcemid was added during the final 3.5 hours of treatment. Representative image of H3-pS10 positive cells (turquoise). (Scale bar 10μm). Frequency of H3-pS10 positive cells in Mybl2+/+ and Mybl2Δ/Δ ESCs. Data from three independent experiments. Error bars represent SEM. Statistical analysis was carried out using a two-tailed unpaired t-test (*p < 0.05; **p< 0.01; ns= no significant). D) P-CDK1 (Tyr15), CDK1 and Beta-actin expression levels of Mybl2+/+ and Mybl2Δ/Δ ESCs with or without CPT treatment analyzed by immunoblotting. Bar graph represents average band density of P-CDK1 in Mybl2Δ/Δ ESCs relative to loading control and relative to the MYBL2+/+ from 6 repeats. Error bars represent SEM. Statistical analysis was carried out using a two-tailed unpaired t-test (**p< 0.01).
a) Immunofluorescence images of U2OS cells stained for LAMP1 and the Golgi marker RCAS1 after 24 h glutamine deprivation with or without treatment with 1 μM BrefA or 100 nM WYE-125132.f5(a) immunofluorescence microscopy images of U2OS cells stained for LAMP1 and the Golgi marker RCAS1 after 24 h glutamine deprivation with or without treatment with 1 μM BrefA or 100 nM WYE-125132. Scale bar, 15 μm. (b) Cytokine array analysis of conditioned medium from U2OS cells subjected to glutamine deprivation in the presence of 1 μM BrefA or 100 nM WYE-125132. (c) IL-8 enzyme-linked immunosorbent assay in U2OS cells after 24 h glutamine deprivation with or without CCI-779 or WYE-125132. (d) RT-PCR analysis of IL-8 mRNA expression in U2OS cells after 24 h glutamine deprivation with or without CCI-779 or WYE-125132. Error bars in all figures represent s.d. of three biological replicates. OD, optical density.
Smaducin-6 reduces sepsis-induced apoptosis.Figure 7A, B Quantitative analysis of (A) TUNEL-positive cells and (B) caspase-3-positive cells in IHC in the spleens of CLP + PBS, CLP + Pal-Scram #1, and CLP + Smaducin-6 mice. Three independent experiments (n = 3 mice per group per experiments) were performed. At least five hot spots in a section of TUNEL and IHC per experiment were selected, and average count was determined. The data were expressed as a mean percentage of total cell numbers and statistically analyzed by a t-test and show the mean ± SD of three independent experiments. **P < 0.005, ***P < 0.001 compared to sham or vehicle control (CLP + Pal-Scram #1). C ELISA showing that Smaducin-6 treatment reduced IFN-β1 levels in peritoneal lavage fluids compared to Pal-Scram #1-treated CLP mice. n = 10 mice per group per experiment. Data were statistically analyzed by the Dunnett's multiple comparison test (one-way ANOVA). ***P < 0.001 compared to sham or vehicle control (CLP + Pal-Scram #1). D, E IFN-β1 (D) and TRAIL (E) expression detected by FACS in splenocytes is decreased in Smaducin-6-treated CLP mice compared to scrambled peptide-treated CLP mice. Data are representative of three independent experiments. F Decreased expression of TRAIL was detected by IHC in the spleen of CLP-induced mice treated with Smaducin-6 compared to mice treated with Pal-Scram. Scale bar, 100 μm (magnification, 200×). Data are representative of three independent experiments.
Fig.Figure 4A. Parental HEK293T, ACVR2A KO, ACVR2B KO, ACVR2A/B dKO, HOM1, HOM1 ACVR2A KO, HOM1 ACVR2B KO, HOM1 ACVR2A/B dKO cells were treated with Activin A or BMP2 for 1 h. Data information: In all panels, Western blots of whole cell lysates were probed with the antibodies indicated. Representative experiments are shown. Par, parental HEK293T cells. B. ACVR2A/B dKO cells were transfected with increasing concentrations of either wildtype or kinase-dead ACVR2A (ACVR2AWT or ACVR2AKR respectively) and treated with Activin A for 1 h. Parental HEK293T cells served as a control. Data information: In all panels, Western blots of whole cell lysates were probed with the antibodies indicated. Representative experiments are shown. Par, parental HEK293T cells. C. HOM1 ACVR2A/B dKO cells were transfected with increasing concentrations of either wildtype or kinase dead ACVR2A (ACVR2AWT or ACVR2AKR) and treated with Activin A for 1 h. HOM1 cells served as a control. Data information: In all panels, Western blots of whole cell lysates were probed with the antibodies indicated. Representative experiments are shown. Par, parental HEK293T cells.
Figure 7a. mRNA expression on ex vivo isolated Tregs from MS patients and healthy controls (n=15). b. Representative example of the expression of AKT3, Foxo3 and PTEN on ex vivo sorted Tregs from MS patients (black open histogram) and HC (grey open histogram). Grey histograms represent isotype. c. Ex vivo expression of pAKT, pFoxo1 and pFoxo3 in Tregs from RRMS patients (black open histogram) and healthy controls (white open histograms). Grey histograms represent isotype. d. Representative example of the frequency of IFN+Foxp3+ Tregs isolated ex vivo from RRMS patients (lower row) and healthy controls (HC, upper row) and stimulated in the presence of vehicle (left histogram), LY294002 (middle histogram) or MK2206 (right histogram) for 3 days. Staining was performed after a 4-hour stimulation with PMA and ionomycin. e. Statistical analysis of the frequency of IFN+Foxp3+ Tregs in HC (white bars) and RRMS patients (black bars) stimulated in the presence of either vehicle, LY294002 or MK2206 (n=10). f. Representative example of the proliferation of Tresp cells from HC (upper row) and RRMS patients (lower row) either alone (first column) or in co-culture with Tregs that had been previously incubated with either vehicle, LY294002 or MK2206 for one hour and washed to remove drug traces. g. Statistical analysis of the percentage of suppression as in h at different Treg:Tresp ratios of n=8 experiments performed. *p < 0.05, **p < 0.005, ***p < 0.0005.
Figure 6(A) Mouse experiment protocol. Adult (8-12 weeks of age) IFNAR1-/- pregnant mice were infected i.p. with 1*105 pfu ZIKV or 1% FBS DMEM/F12 media (vehicle) on embryo day 8.5 (E8.5). 1 h after ZIKV/vehicle injection, treatment with rISG20-Fc (1mg/kg) or PBS (control) was administrated i.p. to the pregnant mice. On E9.5 and E10.5, same protein/PBS injection was performed on the pregnant mice. On E14.5, the mice were sacrificed and organs were collected for viral titer quantification. N=3 for control PBS group, and n=4 for treatment groups. (B) Macroscopic evaluation of the pregnant uterus and fetus. Compare to the fetal death and reabsorptions in ZIKV-infected mice, rISG20-Fc treatment rescued this phenotype by decreasing pregnancy loss (resorption). (C) Effect of rISG20-Fc treatment on pregnancy loss. rISG20-Fc treatment significantly decreased the pregnancy loss caused by ZIKA infection. The pregnancy loss rate (number of dead fetus + resorptions/ total number of implantations x100) was calculated. Data represent as mean ± SEM; n=3-4 mice per group; *p < 0.05 and **p < 0.01 by One-way ANOVA. (D) rISG20-Fc treatment improves fetal development. Measurement of crown-rump length (CRL) and occipitofrontal diameter (OFD) were determined on E14.5. n=3-4 mice per group and 6-8 fetuses from each pregnant mouse were evaluated; *p < 0.05, **p < 0.01 by One-way ANOVA.
| Hypothetical model explaining the role of AP-2 in intracellular trafficking of BACE1.Figure 6In WT neurons, AP-2μ is required for transport of BACE1- containing autophagosomes and late endosomes en route to lysosomes. Absence of AP-2μ causes defective axonal trafficking of BACE1 leading, which results in increased amyloidogenic processing of APP.
cKO heart.Figure 3(A) LC-MS/MS metabolomic analysis of NAD+, NADH, NADP+, nicotinamide and NAAD in the 6 months old WT and p32cKO hearts. NAD+ and NADP+ showed significantly different levels between WT and p32cKO hearts (n = 12). Error bars are presented as mean ± SD. Student's t-test was performed on WT vs p32cKO, **p < 0.001, ***p < 0.0002. (B) Real-time PCR analysis of RNA expression of NAD-synthesizing enzymes in the 6 months old WT and p32cKO hearts. Error bars are presented as mean ± SD. Student's t-test was performed on WT mice vs p32cKO mice (n = 6), ***p < 0.005, *p < 0.05. The right panel shows the NAD synthesis pathway and mRNA expression levels are indicated by red arrows. NAM: nicotinamide, NA: nicotinic acid, QA: quinolic acid, NMN: nicotinamide mononucleotide. (C) Reduced Nmnat3 expression in 9 months old p32cKO heart. Quantification is shown on the right (n = 4 mice per group). Error bars are presented as mean ± SD. Student's t-test was performed on WT vs p32cKO, **p < 0.01. (D) NAD+ levels without or with NMN addition in 9 months old WT and p32cKO heart lysates (n = 3). Error bars are presented as mean ± SD. Student's t-test was performed on WT vs p32cKO, **p < 0.002. (E) Western blot analysis of HIF1α in 9 months old WT and p32cKO heart. Quantifications is shown on the right (n = 4 mice per group). Error bars are presented as mean ± SD. Student's t-test was performed on WT vs p32cKO, **p < 0.01. GAPDH was used as an internal control. (F) Immunoblot analysis of HIF1α; the expression in 3T3-L1 cells was increased after treatment with 1 mM chloramphenicol (CAP), which inhibits mitochondrial translation. β-actin was used as an internal control. One representative experiment out of three shown. (G) Nmnat3 mRNA expression in 3T3-L1 cells after 1 mM CAP treatment for 72 h. The HIF1α inhibitor (20 or 50 µM) was added 2 h before the CAP treatment. Error bars are presented as mean ± SD of three independent experiments. Student's t-test was performed on WT cells vs WT cells treated CAP and (or) HIF1α inhibitor,**p < 0.01, *p < 0.05. (H) Immunoblot analysis of HIF1α after treatment with 150 µM CoCl2 in 3T3-L1 cells, which stabilized HIF1α. β-actin was used as an internal control. (I) Nmnat3 mRNA expression in 3T3-L1 cells after 150 µM CoCl2 treatment for 72 h (n = 3). Error bars are presented as mean ± SD of three independent experiments. Statistical significance was assessed by Student's t-test, **p < 0.01.
RNF26 interacts with MITA.Figure 2(A) RNF26 interacted with MITA. The 293 cells (5×106) were transfected with the indicated plasmids (5 µg each). Twenty-four hours later, cells were lysed and cell lysates were immunoprecipitated with anti-Flag or control IgG. The immunoprecipitates and whole cell lysates were analyzed by immunoblots with anti-HA or anti-Flag. (B) RNF26 was physiologically associated with MITA. THP-1 cells (2×107) were infected with SeV or HSV-1 for the indicated time points or left uninfected. Cells were lysed and immunoprecipitation and immunoblot analysis was performed with antibodies against the indicated proteins. NC, negative control. (C) RNF26 is localized to the ER and mitochondria. HeLa cells (2×105) were transfected with the indicated plasmids (0.5 µg each). Twenty-four hours after transfection, the cells were stained with Mito-Tracker Red for 15 minutes or left untreated. The cells were fixed with 4% paraformaldehyde and subjected for confocal microscopy analysis. D) Subcellular distribution of RNF26. The 293 cells (5×107) were infected with SeV for the indicated time points or left uninfected. Subcellular fractionation assays were performed and the cellular fractions were analyzed by immunoblots with antibodies against the indicated proteins. (E) RNF26 and MITA were colocalized to the ER. HeLa cells (2×105) were transfected with the indicated plasmids (0.5 µg each). Eighteen hours after transfection, cells were infected with SeV or HSV-1 for 6 hours or left uninfected, followed by ER-Tracker Blue/White staining for 30 minutes. Cells were fixed with 4% paraformaldehyde and subjected for confocal microscopy analysis. All experiments were repeated for at least three times with similar results.
Fig.Figure 5(A,B) The blocking of the binding of WT PD-1-mFc (A) or N58A mutated PD-1-mFc (B) to PD-L1 transiently expressed on the surface of 293T cells are analyzed with varied concentrations of camrelizumab. The PD-L1 expressing 293T cells stained with PBS alone is used as negative controls, whereas the cells stained with WT or N58A mutated PD-1-mFc proteins are used as a positive control. The frequency of PD-1-mFc staining positive cells in PD-L1-GFP positive cells is labeled in the upright corner. The reduced frequency of the PD-L1-mFc-staining-positive subpopulation compared with positive control indicates the blockage of the PD-1/PD-L1 interaction. The density of events at a given position in the plot is color-coded, with red representing the highest number of events at that point in the graph, while yellow, green, and blue represent progressively lower event densities. The results presented here are representative of three independent experiments with similar results.
Figure 3A Number of cell divisions in activated mature B lymphocytes. Sorted B220+GFP+ spleen cells from HET-cd19 (n=5), MaxKO-cd19 (n=5), MycKO-cd19 (n=3), and DKO-cd19 (n=3) mice were stained with a cell tracer, activated in vitro with LPS plus IL-4 for 72 h and analyzed by flow cytometry. Experiment representative of at least three independent experiments B Cell counts from the cultures C Photographs of B cell cultures Scale bar, 500 µm. 4× magnification. D Forward scatter (FSC) histograms of activated B lymphocytes An arbitrary dashed line is shown for comparison. Cell cycle analysis of sorted B220+GFP+ B lymphocytes activated in vitro. Cells were activated and EdU was added 4 h before harvesting. Cells were stained with propidium iodide to measure DNA content. Experiment representative of at least three independent experiments. An arbitrary dashed line shows the difference of EdU MFI. S phase mean fluorescence intensity (MFI): 7871 HET, 6512 MaxKO, 5860 MycKO, 6420 DKO Representation of labeling with nucleotide analogues CldU (red) and IdU (green) Representation of labeling with nucleotide analogues CldU (red) and IdU (green) and photographs of DNA fibers from sorted B220+GFP+ B lymphocytes activated for 48 h. Scale bar, 10 µm. H Fork rate of DNA fibres from primary B lymphocytes sorted and activated Data is pooled from three replicate experiments. Horizontal bars represent median values. 150-250 forks were measured per condition. ***, p<0.001; **, p<0.01 I Western blotting of DNA replication proteins in activated B lymphocytes Sorted B220+GFP+ spleen cells were activated in vitro with LPS plus IL-4 for 72 h. MEK2, SMC1 are shown as loading controls. The blots shown are representative of three independent experiments.
Figure 5(A) Immunoblots indicate miR-124-induced SIX4 depletion. siSIX4 is shown as an antibody control. (B) Consequence of SIX4 depletion on EOCcell viability (as in 2A, N, normal cell lines: HOSE, HBEC30). (C) in vivo knockdown of SIX4 using neutral liposome incorporated siRNA reduced tumor burden and ascites volume in an orthotopic xenograft mouse model. Box-and-whisker plot from n=8. p-value from Student's T-Test. (D) Consequence of SIX4 depletion in PEO1 cells on E-cadherinplasma membrane accumulation 48 hours after transfection. Cells were counterstained with phalloidin and DAPI. (E) Immunoblots indicate consequence of SIX4 depletion on LKB1 accumulation and AMPK pathway activation.
Loss of the insulin and IGF-1 receptor from Phb2pko mice does not improve mitochondrial ultrastructure.Figure 5A Serum creatinine and urea levels in the fifth week of life (left graph: Phb2pko 74.73 ± 2.20 μmol/l versus Phb2pko/Insrpko/Igf1rpko 37.36 ± 8.83 μmol/l, n = 3 for Phb2pko and n = 4 for Phb2pko/Insrpko/Igf1rpko, *P = 0.0169; right graph: Phb2pko 493.70 ± 45.34 mg/dl versus Phb2pko/Insrpko/Igf1rpko 272.80 ± 49.29 mg/dl, n = 3 for Phb2pko and n = 4 for Phb2pko/Insrpko/Igf1rpko, *P = 0.0247). B Abundance of deleted Phb2 gene in isolated glomeruli of Phb2fl/fl, Phb2het, Phb2pko and Phb2pko/Insrpko/Igf1rpko mice compared to the floxed Phb2 gene (left graph; quantified by qPCR: n = 3 for all groups, P = 0.83 for Phb2pko/Insrpko/Igf1rpko versus Phb2pko) and compared to a reference gene (right graph; quantified by qPCR: n = 3 for all groups, P = 0.58 for Phb2pko/Insrpko/Igf1rpko versus Phb2pko). C Analysis of mitochondrial morphology in electron micrographs (left image: scale bar, 1 μm; right image: scale bar, 0.1 μm).
PAT1 is associated with MPK4 and SUMM2 in N.Figure 7Proteins were transiently co‐expressed (PAT1‐GFP + MPK4‐HA or SUMM2‐HA) in N. benthamiana and tissue harvested 2 days post‐infiltration. Anti‐HA immunoblots of inputs (20 μg each) are shown in the bottom panel. Immunoblots of GFP IPs were probed with anti‐HA antibodies, then stripped and probed with anti‐GFP antibodies. Molecular weights are shown in kDa on the right. Confocalimages of yellow fluorescent protein complementation. PAT1‐Nc was transiently co‐expressed with either SUMM2‐Nn or At4g12010 in N. benthamiana. Confocalmicroscopy on leaf disks was conducted 2 days post‐infiltration. Merged pictures show overlay of brightfield, chlorophyll and eYFP. The scale bar corresponds to 25 μm.
Figure 6(A) A schematic diagram of intravitreal AAV2 injection to 1-month-old mice followed by CTB488 and CTB-647 injection after 29 days. The brains were collected one day later. (B) Confocal images of RGC inputs traced by CTB488 and mCherry. CTB488+ RGC inputs in the Cdc50a-RNAi area overlapped with vGluT2. Scale bar, 20 µm. (C) Quantification of colocalization among CTB488, mCherry and vGluT2 described ~98% CTB488+/mCherry+ RGC inputs were vGluT2+. N = 7 for biological replicates, mean ± SD. (D) Images of CTB488 and CTB647 labeling of RGC inputs in the dLGN. Scale bar, 20 µm. (E) Quantification of CTB488+ inputs (non-infected areas) versus CTB647+ inputs (infected areas) in Cdc50a RNAi mice and controls. N = 3 for biological replicates, mean ± SEM, **** p < 0.0001, two-way ANOVA with Sidak's multiple comparisons test.
Figure 5HeLa and MDA-MB-231 cells were transfected with either scrambled or RNF146 siRNAs (48 h). Protein levels were detected by Western blot. PTEN, which is a known RNF146-interacting protein, was included as a positive control (n=3). HeLa cells were transfected with either scrambled or RNF146 siRNAs for 48 h, followed by incubation with 10 μg/ml cycloheximide (CHX) for the indicated periods of time. Lysates were harvested and analysed by Western blot (n=3). Quantification of BRD7 protein levels from B, n=3. Error bars indicate SEM. Relative amounts normalized to the BRD7 protein level at 0 h. Control and RNF146 stably overexpressing HeLa cells were transfected with Myc-BRD7-WT for 24 h, followed by incubation with 10 μg/ml cycloheximide (CHX) for indicated periods of time. Lysates were subjected to Western blot analysis (n=3). Quantification of BRD7 protein levels from D, n=3. Error bars indicate SEM. Relative amounts normalized to the BRD7 protein level at 0 h. HeLa cells transfected with either RNF146 or control siRNA for 24 h followed by transfection of HA-ubiquitin for another 24 h; MG132 (10 μM) was added for 4 h and lysed with RIPA, followed by anti-BRD7 IP and analysed by Western blot with the indicated antibodies (n=3). HeLa cells stably expressing RNF146 were transfected with HA-ubiquitin for 24 h; MG132 (10 μM) was added for an additional 4 h, followed by anti-BRD7 IP and Western blot (n=3).
Figure 4A) Proteins that differ significantly differ between pathogenic LRRK2 carriers and controls using an ANCOVA analysis with sex, age, PD status and GBA status as confounders and an FDR of 5%. B) Mean fold-changes for each of the 33 proteins that were LRRK2-dependently regulated in both cohorts using a pairwise t-test comparing the four subgroups (HC, NMC, iPD and LRRK2 PD). C) Correlation of mean fold-changes of the proteins changing LRRK2-dependently in the Columbia and LCC cohorts. Only proteins identified in both cohorts are shown (n=237). All proteins associated with the GO-term 'glycosphingolipid metabolic process' labeled in purple are also associated with the lysosomal-related GO-terms. Proteins overlapping between the two cohorts are labeled with their name. D) Fisher exact test was performed to identify significantly enriched GO-terms in the LRRK2-dependently regulated proteins in urine. All GO-terms that were significant in either cohort are displayed (FDR < 5%). E) Proteins that differ significantly between pathogenic GBA carriers and controls or pathogenic LRRK2 carriers and controls using an ANCOVA analysis with sex, age, PD status and LRRK2/GBA status as confounders and an FDR of 5%. In total, 237 proteins were differentially expressed in these two comparisons with 166 and 74 regulated proteins in the LRRK2 carriers and GBA carriers, respectively, only three of which were common between both mutations. F) Correlation of mean fold-changes of the proteins changing LRRK2-dependently and GBA-dependently (n=237) in the Columbia cohort. Carriers of pathogenic variants in both GBA and LRRK2 were excluded from the analysis.