EMBO/sd-character-level-ner huggingface dataset:
Figure 7A. Splenocytes were isolated from Ambragt/+ and wt C57/BL6mice and activated as indicated in materials and methods. At day 7 after activation, AICD was induced with 0.1μg/ml of plate-bound anti-mCD3 (Clone 17A2). Apoptotic cells were detected 12h after AICD induction by flow cytometry as AnnexinV/PI double positive cells. B. Splenocytes were isolated from ATG7 fl/fl and wt C57/BL6mice. Activation of splenocytes and AICD were performed as previously described. At day 5 after activation, cells were infected with retroviral constructs expressing CRE recombinase. Where indicated, cells were pre-incubated with 100nM rapamycin for 24h before AICD induction. Apoptotic cells were detected 2h after AICD induction by flow cytometry as AnnexinV/PI double positive cells. C. Jurkat cells were transfected with scramble or ATG7 siRNA 24h before AICD induction. AICD was induced as described in materials and methods. Where indicated 100nM rapamycin was added at time 0h of AICD induction. Apoptotic cell death analysis was carried out as in (A), except that cells were analysed 32h after AICD induction. Data represent mean ± SE of five independent experiments. D. Mitochondrial depolarization upon AICD induction was analysed in cells treated as indicated. Data represent mean ± SE of five independent experiments. E. Protein samples (20μg) from Jurkat cells treated as indicated were separated by SDS-PAGE and immunoblotted with the indicated antibodies. WB are representative of three independent experiments.F. Quantitative analysis of the ratio between the protein levels of mitochondrial markers and GRP75 during AICD is shown. Data represent mean ± SE of three independent experiments.
Fig.Figure 4(A) Representative electron microscopy images of liver samples harvested from wild-type (WT) and AslNeo/Neo mice treated with TB-1 or vehicle. False color on the images indicates glycogen within the nuclei. Scale bar: 1 μm. (B) Quantification of nuclei containing glycogen (approx. 180 nuclei were analyzed in total per condition, n=3 mice/group).
a, spr-4(by105) worms incubated continuously with paraquat (5 mM) exhibit increased mortality rescued by wild-type SPR-4 or human REST.f4a, spr-4(by105) worms incubated continuously with paraquat (5 mM) exhibit increased mortality rescued by wild-type SPR-4 or human REST. Shown is a representative experiment replicated three times. b, Quantitative analysis of survival in wild-type worms expressing mCherry or SPR-4 (WT+mCherry and WT+SPR-4), spr-4(by105) mutants, and spr-4(by105) mutants expressing SPR-4 or human REST (spr4(by105)+SPR4 and spr4(by105)+REST). Shown is the percentage change in mean survival relative to wild-type. Values represent the mean ± s.d., n = 3 independent replicates of at least 30 animals per genotype; **P 0.01 relative to wild type by the log-rank test. c, REST and SPR-4 reduce levels of reactive oxygen species (ROS). Shown are representative confocal images of paraquat-treated worms labelled with the ROS-sensitive dye DCFDA. WT, wild type. Scale bar, 30 µm. d, Quantitation of ROS levels by DCFDA labelling. Horizontal bars indicate the median; boxed areas represent the second and third quartiles. *P 0.05 relative to wild type by analysis of variance with post-hoc Tukey test; n = 30 worms. AFU, arbitrary fluorescence units. e, SPR-4 protects against Aβ neurotoxicity. Shown are Aβ worms (expressing a stably integrated Aβ1-42 transgene) and Aβ spr4(by105) worms. Neuronal degeneration does not occur in wild-type worms or spr4(by105) mutants in the absence of the Aβ transgene. Values represent the percentage of worms that retain five glutamatergic tail neurons at the indicated age (days), and are the mean ± s.d., n = 3. *P  0.05 by unpaired t-test.
Figure 6A) GSEA plot using BroadC2 database showed that genes differentially expressed in young control compared to young Mfn2KO mice and old control mice are enriched in genes related to hypoxia response. B) HIF1α protein expression was measured in nuclear extracts from young and old control and Mfn2KO mice (n=4-6 mice per group). Data were normalized by tubulin as a loading control and expressed as a fold change compared to young control mice. C) HIF1α and BNIP3 protein expression were measured in control and Mfn2KD C2C12 myotubes untreated or treated with the HIF1α specific inhibitor NSC-134754 (n=3 independent experiments performed in triplicate). Data were normalized by tubulin as a loading control and expressed as a fold change compared to control C2C12 myotubes. D) Levels of H2O2 were measured in control and Mfn2KD C2C12 myotubes untreated or treated with NSC-134754 (n=5 independent experiments performed in duplicate). Data were expressed as a fold change compared to control C2C12 myotubes. E) and F) Respiration was measured in control and Mfn2KD C2C12 myotubes untreated or treated with NSC-134754 and ATP-coupled respiration (E) and proton leak (F) were calculated (n=8 independent experiments). Data were expressed as a fold change compared to control C2C12 myotubes. G) Autophagic protein abundance (BNIP3, LC3II and Parkin) was measured in mitochondrial-enriched fractions from control and Mfn2KD C2C12 myotubes untreated or treated with NSC-134754 (n=4 independent experiments). Data were normalized by TIM44 as a loading control and expressed as a fold change compared to control C2C12 myotubes. H) and I) Representative TEM images of control and Mfn2KD C2C12 myotubes untreated or treated with NSC-134754 showing the accumulation of autophagosomes (H) and the presence of mitochondrial alterations (I) in Mfn2KD or NSC-134754 treated cells (Scale bar: 2 µm).
Low dose interferon lambda 1 treatment dramatically reduces SARS-CoV-2 replication in bronchioalveolar-like cultures.Figure 7(A, B) SARS-CoV-2 replication shown as infectious virus titers (A) and viral RNA copies (B) in apical washes at 2, 24, 48, and 72h after infection at MOI 0.1. Cultures were either pretreated with 50 ng/ml IFN lambda 1 in the basal compartment two hours prior to the infection (green), at 24 h p.i. or left untreated. The dotted line indicates the lower limit of detection. Error bars represent SEM. N=2. H p.i. = hours post infection. C) Whole well immunofluorescent staining of representative cultures in (A) and (B) using an anti-NP antibody (red). Nuclei are stained with Hoechst (blue). Scale bar indicates 1mm.
Fig.Figure 1(A) Protein-Protein Interaction enrichment analysis of differentially expressed host factors in HBMEC with and without GBS infection revealing signaling network involving EGFR. (B) Wild type mice received strain GFP-K79 via the tail vein and 1 hr later, animals were perfused and the meninges, cortex and choroid plexus were obtained for demonstration of intravenously injected bacteria. The arrows represent bacteria co-localized with capillaries (e.g., within capillaries) and arrowheads represent bacteria outside the capillaries (e.g., exited from the capillaries). A few bacteria were successfully passed through the meningeal and cortex capillaries, but no bacteria were demonstrated in the choroid plexus at this time. (C) Sulfo-NHS-biotin was administered via intraperitoneal injection 10 min before perfusion for assessing the blood-brain barrier permeability, i.e., extravasation is indicative of leakage from the intravascular lumen. Sulfo-NHS-Biotin was confined to the capillaries of the meninges and cortex (as shown with arrows), and there was no evidence of extravasation. Scale bar = 100 μm. (D) Relative invasion frequency of GBS strain K79 in HBMEC with or without EGFR inhibitor (gefitinib). (E) Bacterial counts recovered from the blood and brain in wild type mice receiving vehicle control (n = 5) or gefitinib (10 mg/kg) (n = 6), infected with strain K79 for 1 h. (F) Tyrosine phosphorylation of EGFR in HBMEC in response to GBS strain K79 infection at 15 min and 60 min post infection. (G) EGFR protein expression in EGFR knockout HBMEC using CRISPR/Cas9 (left panel) and relative invasion frequency of 8 meningitis isolates of GBS strains in EGFR knockout and control HBMEC (right panel). From left to right, p value of K79 is 0.0059, p value of K160 is 0.00024, p value of K161 is 0.00080, p value of K181 is 0.00042, p value of K226 is 0.00011, p value of K237 is 0.00072, p value of K238 is 0.0093, p value of P539 is 0.00013 (right panel). (H) GBS strain K79 traversal of HBMEC monolayer was significantly decreased in EGFR knockout HBMEC compare to control HBMEC. (I) Bacterial counts recovered from the blood and brain of EGFR conditional knockout (n = 5) and control mice (n = 5) 1 h after intravenous inoculation with strain K79.
Figure 3Schematic diagram showing Npr3-sCreER or Npr3-CreER mouse crossed with a newly generated Rosa26 reporter R26-LZLT. Schematic diagram showing experimental strategy. Whole mount fluorescent images of E14.5 or E15.5 Npr3-sCreER;R26-LZLT and Npr3-CreER;R26-LZLT mouse hearts. No tam is as control for detecting leakiness of sCreER or CreER. Representative Immunostaining images for tdTomato, ESR and CDH5 on heart sections. Quantification of the percentage of endocardial cells expressing tdTomato in Npr3-sCreER;R26-LZLT (sCreER) or Npr3-CreER;R26-LZLT (CreER) hearts. Data are mean ± s.e.m.; n = 5; *P < 0.05. Scale bars, 500 µm. Each figure is a representative of 5 individual biological samples.
Figure 4(A) Diagram of dosing regimen for the long-term study of PMO-M in mdx mice. i.v. refers to intravenous injection. Immunohistochemistry of dystrophin-positive fibres in body-wide muscles from mdx mice treated with repeated doses of PMO-M (n=4) or PMO alone (n=3) (scale bar: 100 μm) (*p<0.05, **p<0.001; two-tailed t test). quantitative analysis of dystrophin-positive fibres in body-wide muscles from mdx mice treated with repeated doses of PMO-M (n=4) or PMO alone (n=3) (scale bar: 100 μm) (*p<0.05, **p<0.001; two-tailed t test). (D) Western blot for dystrophin expression in body-wide muscles from mdx mice treated with PMO-M or PMO alone. 0.5 µg, 2.5 µg, 5 µg, and 15 µg total protein from C57BL/6 and 50 µg of muscle samples from untreated and treated mdx mice were loaded. (E) Quantitative analysis of dystrophin expression in body-wide muscles from mdx mice treated with PMO-M (n=4) or PMO alone (n=3) (*p<0.05; two-tailed t test). (F) Muscle function was assessed to determine the physical improvement with grip strength test for mdx mice treated with PMO-M (n=4) or PMO alone (n=3), untreated mdx controls (n=3) and C57BL/6 (n=3) (*p<0.05, **p<0.001, One way-ANOVA post hoc Student-Newman-Keuls test). (G) Measurement of muscle endurance with the running wheel test for wild-type C57BL/6 (C57) (n=3), untreated mdx controls (mdx) (n=3) and mdx mice treated with PMO-M (n=4) or PMO alone (n=3) (*p<0.05, One way-ANOVA post hoc Student-Newman-Keuls test). Muscle functional tests were conducted two weeks after last injection.
Figure 4A HCT116 cells were transfected with increasing amount of DNA expressing NAT10. Forty-eight hours later, whole cell lysates were prepared and proteins from lysates were analyzed by western blot using the indicated antibodies. B HCT116 cells were transfected with Flag-NAT10 and harvested at indicated time points after treated with CHX. Proteins from cell lysates were immunoblotted with the antibodies as indicated (left panel). Relative Mdm2 protein levels standardized by β-actin are shown in the right. Mdm2 half-life (t1/2) obtained from three independent experiments was presented as mean ± SEM at the bottom. C HCT116 cells transfected with the indicated constructs were harvested after treatment with MG132 for 4 hours. The Mdm2 protein was immunoprecipitated with anti-Mdm2 antibody and immunoblotted with anti-HA antibody. D In vitro ubiquitination was performed with recombinant GST-Mdm2 (left) or GST-Mdm2 C464A mutant (right) and Flag-NAT10 or Flag-NAT10 GE in the presence of E1, E2 and Ub in vitro as described in Experimental Procedures. The reaction products were resolved by SDS-PAGE, followed by western blot for evaluation of the indicated proteins. E Mdm2 is ubiquitinated by NAT10 in a dose-dependent manner. In vitro ubiquitination assay were carried out with Mdm2 and different doses of purified Flag-NAT10 as described in D. F Schematic representation of NAT10 deletion mutant constructs. G U2OS cells transfected with NAT10 siRNA-3 (targeting 3'-UTR of NAT10 mRNA) or control siRNA were co-expressed different NAT10 deletion mutants. Cell lysates were extracted and proteins from lysates were subjected to western blot using the indicated antibodies. Relative p53 protein levels standardized by β-actin are shown in the lower. H In vitro ubiquitination reaction was conducted using recombinant GST-Mdm2 C464A and full-length Flag-NAT10 or Flag-NAT10-D5. Ubiquitinated Mdm2 was detected as in D. I HCT116 cells were transfected with the indicated constructs. Cells were harvested after CHX treatment for the indicated times and cell lysates were prepared. Mdm2 protein levels were evaluated by western blot. Relative Mdm2 protein levels standardized by β-actin are shown in the right.
Figure 5A, B. The protein level of STAT1 was negatively correlated with METTL3 and METTL14 in human pMMR-MSI-L CRC colon tissue (r2=-3.2477 for METTL3, r2=-2.7491 for METTL14). Each dot represents one tumor tissue. C. Schematic showing the functional and molecular mechanisms of Mettl3 and Mettl14 in antitumor immunotherapy.
Figure 7Parental MDCK cells or those transfected with Prom1 or K138Q mutant were grown for 7 dpc Cells were observed by phase-contrast microscopy (A) Box-whisker plots show the number of domes per 600 mm2 (A). Three images were analyzed per experiment (n = 4). 4X magnification (inset in panel A). Parental MDCK cells or those transfected with Prom1 or K138Q mutant were grown for 7 dpc Cells were observed by SEM (B, top panels) or confocal laser scanning microscopy after DAPI staining (B, bottom panels, Individual cells within a given dome are indicated (B, top panels) and were quantified using three different planes (bottom, centre and top) of DAPI-labeled nuclei (B, bottom panels). Parental MDCK cells or those transfected with Prom1 or K138Q mutant were grown for 7 dpc (A-C) or as indicated (D). confocal laser scanning microscopy after DAPI staining Domes of similar size were evaluated (C). Means and standard deviations are displayed. 15 domes per independent experiment were examined (n = 3). Cell density at a given time of culture (7, 10 and 14 days) was determined by counting DAPI-labeled nuclei per 0.1 mm2 (D, n = 4). Cilia with a length of >4 µm were rarely observed in K138Q mutant and displayed an irregular shape with constrictions along their distal part as observed by SEM (arrows). Mutation in the Hdac6-binding site of Prom1 increases the release of Prom1+ vesicles. 16 h-conditioned medium was subjected to differential centrifugation for 5 min at 300 x g (P1); 20 min at 1,200 x g (P2); 30 min at 10,000 x g (P3) and 1 h at 200,000 x g (P4). In parallel, detergent cell lysates were prepared. The entire pellets and 1/40 of cell lysates were analyzed by immunoblotting using antibodies against human Prom1, AcTub and β-actin (F). Molecular mass markers are indicated. All three proteins were detected in the vesicle fractions (P3 and P4) suggesting Prom1+ vesicles are released from primary cilia and microvilli. Mutation in the Hdac6-binding site of Prom1 increases the release of Prom1+ vesicles. 16 h-conditioned medium was subjected to differential centrifugation for 5 min at 300 x g (P1); 20 min at 1,200 x g (P2); 30 min at 10,000 x g (P3) and 1 h at 200,000 x g (P4). In parallel, detergent cell lysates were prepared. The release of Prom1+ vesicles was quantified using its immunoreactivities associated with cell and vesicle fractions (G)
Induction of autophagy impairs the survival of Mfa1+Pg strain within MoDCs.Fig 4A) Survival of P. gingivalis strains within MoDCs after 6, 12, 24 and 48 hours. Blue lines show P. gingivalis strains survival within MoDCs and their survival in anaerobic condition in the absence of DCs are showed in black lines. The effect of rapamycin on Mfa1+Pg, Pg381 and FimA+Pg survival within MoDCs are shown in figures B, C and D respectively. A three-factor repeated measures ANOVA using mixed models was used to test the effect of strain and rapamycin treatment over time on OD reading. The survival curves for the strains are showed in blue, while the effect of rapamycin treatments are in red. Bacterial survivals in the absence of MoDCs with and without rapamycin are plotted in grey and black, respectively. Statistical analysis showed that the strain by rapamycin treatment overtime interaction indicates the pattern of means in each strain (Mfa1+Pg, Pg381 and FimA+Pg) between treated (rapamycin) and untreated were significantly different overtime (p-value <0.=001).
Figure 6A. TCs were assembled on DNA-RNA scaffolds (DNA was biotinylated and attached to streptavidin coated magnetic beads); TCs were then divided into three tubes and challenged with buffer ("-", mock treatment) or RNase J1 of Xrn1. Dissociation of RNAP from DNA was monitored by detecting RNAP with anti-β antibody in two fractions: B - beads (RNAP still bound to DNA), R - released (free in buffer). The gel strip shows representative primary data (Western blot). The graph shows averages (the bars) of two experiments (biological replicates) and the error bars show the range. The combined signal for B+R for each treatment was set as 100 %. B. Primary data - polyacrylamide gel. RNAP from E. coli. Lane 1, the full length (33 nt) labeled RNA; lane 2, the same as lane 1 but it included also incubation with RNase J1 (J1), lane 3, the same as lane 1 but included also incubation with Xrn1. C. Quantitation of three independent experiments. "Full length" indicates the remaining undigested RNA. "OUTSIDE", "INSIDE" are fragments The bars represent 100 % (all fragments). The black-grey-white boxes indicate the percentage of each fragment group (in %). The error bars indicate ± SEM for each group of fragments
Figure 1A Transmigration of mouse neutrophils towards the chemokine CXCL-1 through TNF-α-stimulated VE-cadherin WT (WT) or VE-cadherin Y731F (Y731F) primary endothelial cells, each transfected with control or SHP2-specific siRNA. The transmigration rate is presented relative to that of WT cells transfected with control siRNA, set as 100%. On the right, Western blot analysis of lysates from cells used in transmigration assays for the expression of SHP2 and α-tubulin. B Transmigration of human neutrophils towards the chemokine IL-8 through TNFα-stimulated human endothelial cells (HUVEC), transfected with control or SHP2-specific siRNA and were either not transduced (-) or transduced (+) with adenoviruses expressing human SHP2 (Adv-SHP2). The transmigration rate is presented relative to that of cells transfected with control siRNA, set as 100%. On the right, Western blot analysis of lysates from cells used in transmigration assays for the expression of SHP2 and α-tubulin. C Lysates of WT or Y731F primary endothelial cells were submitted to pulldowns with GST, GST‐SHP2‐WT, or GST‐SHP2‐DA. Pulldowns and total cell lysates were analyzed by Western blotting for VE-cadherin and GST. Graph below represents quantification of two independent experiments, presented as fold change of bound VE-cadherin. TNFα-activated bEnd.5 cells were incubated with mouse T cells for 0-30 min. SHP2 (above) or VE-cadherin (below) were immunoprecipitated (IP) from cell lysates and precipitates as well as total cell lysates were analyzed by immunoblotting for indicated antigens. Molecular weight markers are indicated in kDa. Graph below represents quantification of five independent experiments, presented as fold change of bound VE-cadherin.
Cross-regulation and single-cell fluctuations of SOX2 and OCT4 A Endogenous SOX2 levels in single cells (turquoise) normalized to the value at t=0 in SNSF cells upon induction of SOX2-SNAP (n=37, left) or a truncated SOX2 version missing the DNA binding domain (YPET-SOX2-delDBD, n=47, right).Figure 2Endogenous SOX2 levels in single cells (turquoise) normalized to the value at t=0 in SNSF cells upon induction of SOX2-SNAP (n=37, left) or a truncated SOX2 version missing the DNA binding domain (YPET-SOX2-delDBD, n=47, right). Black line: average SOX2-SNAP levels after induction normalized to the maximum level (n=22); Yellow line: average YPET-SOX2-delDBD levels after induction, normalized to the maximum level (n=20). Sox2 and Oct4 mRNA levels upon overexpression of YPet-SOX2 or YPET-SOX2-delDBD (n=3). p-values: two-sided t-test with unequal variance. Changes of OCT4 half-life upon SOX2-SNAP or YPET-SOX2-delDBD overexpression (n=20 cells each). p-values: Mann-Whitney U test. SBROS cells harbouring inducible SOX2-SNAP or YPET-SOX2-delDBD were treated with 0 or 500 ng/ml dox for 6-8 hours followed by pulse-labelling of OCT4-HALO and live-cell imaging. Sorting strategy to evaluate the impact of endogenous fluctuations of SOX2 and OCT4 on OCT4 and SOX2 protein levels, respectively. Changes of SOX2 and OCT4 levels after sorting for high and low SOX2 levels in cells expressing intermediate OCT4 levels (n=4). Error bars: SE; p-values: one-sided t-test with unequal variance. Changes of SOX2 and OCT4 levels after sorting for high and low OCT4 levels in cells expressing intermediate SOX2 levels (n=4). p-values: one-sided t-test with unequal variance. Representative traces of the absolute number of SOX2-NLUC molecules (top) and inferred concentration in nM (bottom) in in-silico synchronized cells (n=10). Single cells were ranked according to their SOX2 expression at t=0 and assigned a color code for their initial rank from low (blue) to high (red) levels, and changes in ranks over time are shown (n=59). Rank-based autocorrelation function of the SOX2 ranks (n=59). Error bars: SE estimated by bootstrapping. Representative single cell traces of the integrated intensity of OCT4-HALO in single cells (top) and the corresponding inferred concentration (bottom) in in-silico synchronized cells (n=10). Representation of the ranks as in G. Rank-based autocorrelation function as in I (n=47). Error bars: SE estimated by bootstrapping.