EMBO/sd-character-level-ner huggingface dataset:
Abolishing TREM2 signaling does not rescue lysosomal dysfunction in Grn-/- miceA.Figure 6A. Western blot of CatD in total brain lysates from 14-month-old female WT, Grn-/-, Trem2-/-, and Double-/- mice. CatD maturation variants are indicated (sc: single chain; hc: heavy chain; n=3). B., C. Quantification of CatD variants in A normalized to WT (n=3 per genotype). Data information: Data represent mean ± SEM. For statistical analysis one-way ANOVA with Tukey's post hoc test of Grn-/-, Trem2-/- and Double-/- was used. Statistical significance was set at *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001, and ns, not significant. D., E. Catalytic activity of CatD in brain lysates from female 6-month-old (n=4 per genotype) (D) or 14-month-old (n=3 per genotype) (E) Grn-/-, Trem2-/-, Double-/- mice normalized to WT. Data information: Data represent mean ± SEM. For statistical analysis one-way ANOVA with Tukey's post hoc test of Grn-/-, Trem2-/- and Double-/- was used. Statistical significance was set at *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001, and ns, not significant. F. Immunohistochemical analysis of lipofuscin (green) in coronal brain sections. Representative images of thalamus are shown. Scalebars = 50 μm. G. Quantification of lipofuscin autofluorescence. Five images per mouse were taken, means were normalized to WT samples (n=3 per genotype, female). Data information: Data represent mean ± SEM. For statistical analysis one-way ANOVA with Tukey's post hoc test of Grn-/-, Trem2-/- and Double-/- was used. Statistical significance was set at *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001, and ns, not significant.
a) THPN induces TR3 translocation from the cytoplasm to the mitochondria, detected by western blotting for the various cell fractionations (top) and by confocal microscopy (bottom).figf2(a) THPN induces TR3 translocation from the cytoplasm to the mitochondria, detected by western blotting for the various cell fractionations (top) and by confocal microscopy (bottom). The cells were treated with THPN at the times indicated and detected by TR3 (red) and Hsp60 (green). The nuclei were stained with DAPI (blue) (scale bars, 10 μm). PARP, Tubulin and Hsp60 were used as loading controls for nuclear extracts (NE), cytoplasmic extracts (CE) and mitochondrial fractions (Mito), respectively. IB, immunoblotting. (b) TR3 mutant Flag-ΔNLS but not Flag-NLS promotes the induction of autophagy in response to THPN treatment. The siRNA-resistant versions of Flag-ΔNLS and Flag-NLS were transfected into TR3-knockdown A375 cells. Top, the cells cotransfected with GFP-LC3 were further treated with THPN for 24 h before microscopic analysis (scale bars, 10 μm). Middle, the conversion of LC3-I to LC3-II was analyzed by western blotting. The rate of cell survival was determined after THPN treatment for 48 h (bottom). '-' represents vehicle treatment only. (c) Nix assists the TR3 mitochondrial targeting. Cells with Nix, Pink1 and Bnip3 knockdown were treated with THPN (20 μM, 6 h). Top, mitochondrial TR3 was assayed by western blotting. Nix knockdown abolishes the THPN-induced (20 μM, 24 h) formation of LC3 foci (middle and bottom; scale bars, 10 μm). Images of full blots are shown in Supplementary Figure 7. All of the data are presented as the mean ± s.e.m. of three independent experiments. **P 0.01; NS, not significant; ctrl, control.
Figure 4(A) The carotenoid pigment of different S. aureus strains was extracted by methanol and quantified by the optical density at 465 nm, and the values were normalized to WT Newman strain, which was set at 100. Data are expressed as the average ± SD of five independent experiments. ** Denotes p < 0.005 and *** denotes p < 0.001 compared to WT via one-way ANOVA with Dunnett's posttest. (B) Hydrogen peroxide sensitivity was measured by incubating WT and isogenic strains, previously grown for 20 h in MHB-ca, with different concentrations of hydrogen peroxide. Serial dilutions were plated in TSB medium for measuring CFU. Data are expressed as the average ± SD of three independent experiments.
Figure 8A) Pictures of YES plates with G418 showing growth of G418 resistant subclones in fft3D (PP3) and fft2Dfft3D (PP4) mutant strains with new integrations of Tf2-12-neoAI. After 3 days at 32o C. B) A Table showing the Tf2 transposition frequencies and rates calculated with fluctuation tests using FALCOR. C) Southern blot probed for neoA fft3D (PP3) and fft2Dfft3D mutant strains (PP4) and derived G418 resistant subclones.
Figure 7 A) Real-time PCR analysis of Lamp1 transcript (n=3/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 Student's t test **p=0.008, *p=0.038  B) Anti-LAMP1 staining in rapamycin- and rilmenidine-treated WT and Cox15sm/sm muscles vs. untreated (UT) samples. Rapamycin, but not rilmendine, increases the number of LAMP1-positive vesicles in Cox15sm/sm samples (inset). The scale bars correspond to 30µm  C) Quantification of LAMP1-positive vesicles (n=3/group). Error bars represent SEM. The asterisks represent the significance levels calculated by one-way ANOVA with Tukey post-hoc multiple comparison test ****p<0.0001  D) Cathepsin L specific activity (n=3/group). Error bars represent SEM. The asterisks represent the significance levels calculated by Student's t test *p=0.0133
Figure 6(A) The heat map shows the gene profiling expression in E13 forebrain. Gene analysis revealed that the transcript levels of genes were upregulated or downregulated by more than two-fold between the KO and the WT cells. n= 2 biological replicates. (B) RNA-seq analysis showing that the NKX2.2 mRNA expression level is dramatically reduced in the KO cells. (C) Western blot analyses of the NKX2.2 expression levels in the WT and KO cortices at E13.5. (D) NKX2.2-shRNA or control plasmid was electroporated into the brains of E13.5 embryos. At E16.5, the brains were harvested to investigate the cell distribution. Date information: Representative images from at least three independent experiments. (E) Graph shows the percentage of GFP+ cells in the VZ/SVZ, IZ and CP. The number of GFP+ cells in the VZ/SVZ region increased significantly, and the number of GFP+ in the CP layer decreased significantly. n =5 mice, independent replicates. Date information: Error bars represent the means ± S.E.M.; Two-tailed unpaired t-test, P < 0.05 (*), P < 0.01(**). n.s., not significant. (F) ChIP analysis revealed that mH2A binds to the NKX2.2 promoter region in NPCs. Primary NPCs were infected with control, or Flag-mH2A overexpression lentivirus. Six different regions (from −5k to CDS) of the NKX2-2 sequence were used for the analysis. n =4 independent replicates. Date information: Error bars represent the means ± S.E.M.; Two-tailed unpaired t-test, P < 0.05 (*), P < 0.01(**). n.s., not significant. (G) Western blot analyses of H3K27ac, H3K56ac, H3K36me3, H3K4me3 and H3K27me3 expression protein levels in WT and KO cortices at E13.5. (H) Quantification of H3K27ac expression in WT and KO cortices. The levels of H3K27ac decreased. n=5 mice, independent replicates. Date information: Error bars represent the means ± S.E.M.; Two-tailed unpaired t-test, P < 0.05 (*), P < 0.01(**). n.s., not significant. (I) Representative images of E16.5 cortices that had been electroporated with GFP or GFP+NKX2.2 into WT and KO brains at E13.5. Scale bar represents 50 μm. (J) Graphs of the percentage of GFP+ cells in the VZ/SVZ, IZ and CP. Overexpression of NKX2-2 restored the distribution of GFP+ cells to the normal state. n=5 mice, independent replicates. Date information: Representative images from at least three independent experiments. Error bars represent the means ± S.E.M.; Two-tailed unpaired t-test, P < 0.05 (*), P < 0.01(**). n.s., not significant.
Figure 7A. Representative Per2::Luciferase traces before (black) and after vehicle (grey) or Prok2RA (pink) topical application of SCN slices. B. Comparison of the Per2::Luciferase rhythm amplitude of the cycle preceding and the first and second cycles succeeding vehicle (applied CT23-20 dark grey n=12 SCN slices, CT-20-23 light grey n=6 SCN slices) or Prok2RA (applied CT23-20 purple n=13 SCN slices, CT20-23 pink n=11 SCN slices) treatment (interaction, p<0.0001). A significant decline in amplitude occurred when Prok2RA signalling was blocked between CT20-23 (****p<0.0001). C. Change in SCN period after treatment with vehicle (grey) or Prok2RA (pink) between CT23-20 and CT20-23 (vehicle CT23-20 vs. Prok2RA CT20-23, **p=0.009; vehicle CT20-23 vs. Prok2RA CT20-23, **p=0.001) (n = 6, 12, 13 and 11 independent SCN slices, respectively). D. Prok2RA application resulted in a significant period lengthening effect of ongoing SCN ensemble Per2::Luciferase rhythms, which was reversed upon drug washout (p=0.0002, ***p=0.0009, *p=0.01) (n = 22 independent SCN slices with repeated measure).
Figure 5Figure 5. ΗHelix-7 Ago2 is sensitive to inhibition by off-target RNAs. A) Ago2-miRNA122 (1 nM) was incubated with a 32P-labeled, seed-matched target RNA (0.1 nM) in the presence of increasing concentrations of unlabeled a competitor RNA (top panel). Fraction target RNA bound to Ago2 is plotted as a function of off-target competitor RNA concentration. B) Fraction of target RNA bound to Ago2 in the presence of increasing concentrations of unlabeled total cellular RNA. For A-B, average values from at least three independent experiments ± SD are plotted.
Figure 1(A) Alignment of the extracellular region of human TAPBPL (hTAPBPL) with that of some known human B7 family members. Identical amino acids are shaded black. Amino acids with strong homologies are shaded in gray. (B) Alignment of hTAPBPL with mouse TAPBPL (mTAPBPL). Predicted signal peptide, IgV- and IgC-like, and transmembrane (TM) regions for hTAPBPL are marked.
Figure 6(A) Stacked bar chart depicting the percentages of [231+/235+], [231+/235-], [231-/235-] and [231-/235+] cases in each preclinical group. Table below indicates the exact percentages of participants in each group (exact number of participants in parenthesis). Discordant cases with the sequential phosphorylation hypothesis [231-/+235] shown in grey. (B) Correlation between CSF p-tau235 and p-tau231 in the whole ALFA+ cohort (Spearman's rank correlation: rS = 0.80, P˂0.0001). Assay cut-offs were determined as the mean + 2 SD of the A-T- group (defining p-tau231 and p-tau235 positivity or negativity in each participant). Cut-off values are indicated in red (19.92 and 9.59 pg/mL for CSF p-tau235 and p-tau231 respectively) and displayed with black dashed lines, resulting in four quadrants, each of them representing the four different positive or negatively status for each biomarker. Discordant cases with the sequential phosphorylation hypothesis ([231-/+235]) showed on the lower right quadrant (2.35% of the total, 9 participants out 383). All samples were run in singlicates.
Figure 1(A) Lentiviral construct used to permanently express S protein in Jurkat. The full length mature S protein is preceded by a leader sequence of GM-CSF and followed by a T2A sequence which is followed by a tail-less truncated human EGFR construct. (B) Expression of S protein on the plasma membrane of Jurkat-S cells assessed by surface biotinylation and followed by immunoprecipitation with anti-T2A, SDS-PAGE under reducing conditions and western blot with streptavidin-peroxidase. Biotinylation of the parental Jurkat cells was carried out in parallel as a negative control. The arrow indicates the position of the reduced S protein.(D) Formation of syncytia between the CTFR-labelled ACE2+ human cell line and the CFSE-labelled Jurkat-S cells was measured by flow cytometry by analyzing the percentage of cells that become double positive for CTFR and CFSE markers. The Bar plot to the right shows the effect of different doses of HepG2 cells on the formation of syncytia with a fixed number of Jurkat. Parental Jurkat cells (not expressing S protein) are considered negative controls. Data represent the mean±SD of triplicated datasets. (E) Formation of syncytia between the CTFR-labelled ACE2+ human cell line and the CFSE-labelled Jurkat-S cells, in an experiment as in D, was confirmed by confocal microscopy after sorting cells double positive for CTFR and CFSE. Nuclei were identified by DAPI staining. Red arrowhead in the DAPI image indicates a nucleus of Hep-G2 origin; the green arrowhead the nucleus of Jurkat-S origin. (F) Overlay plot of Jurkat-S cells that were incubated with anti-EGFR mAb conjugated to Bv421 and serum from either donor #15 or from a pre-COVID-19 donor and followed by a secondary anti-human IgG1 antibody conjugated to PE.
dBruce is required for nurse cell survival by controlling DNA fragmentation during oogenesis.Figure 5.dBruce is required for nurse cell survival by controlling DNA fragmentation during oogenesis. (A) Confocal micrographs of dBrucee00984 mutant ovarioles stained for TUNEL and DNA. Arrows point to degenerating stage 8/9 egg chambers that are TUNEL positive. (B and C) Confocal micrographs of vps34/dBruceE81 and atg, dBruceE81 double mutant egg chambers stained for TUNEL (green and red, respectively) and DNA. (B) vps34−/−GLCs; dBruceE81/dBruceE81 stage 14 mutant egg chamber contain persisting TUNEL-positive nurse cell nuclei (outlined; arrow). (C) atg1−/−GLCs, dBruceE81/dBruceE81 stage 14 mutant egg chamber contain persisting TUNEL-positive nurse cell nuclei. atg1 mutant nurse cells are identified by the lack of GFP staining (outlined; arrow). Draq5/Hoechst staining (blue) was performed to visualize the nuclei. PhC, phase contrast. Bars, 50 µm.
Figure 3(A) Box-and-whiskers plot showing CSF p-tau235 concentrations in the different groups: amyloid-negative cognitively unimpaired (CU-) participants (n=50), participants across the Alzheimer's disease (AD) continuum (amyloid-positive cognitively unimpaired (CU+, n=32); amyloid-positive mild cognitive impairment (MCI+, n=20); AD (n=20) and non-AD cases (n=19). CSF p-tau235 is highly specific for AD pathology. Cut-off value for CSF p-tau235 positivity is displayed with black dashed line (19.92 pg/mL). (B) ROC analysis showing the high diagnostic accuracy of CSF p-tau235 discriminating AD from CU-, CU+, MCI+, and non-AD groups. (C) ROC analysis comparing AUC values of CSF p-tau235, p-tau231 and p-tau217 discriminating CU+ from CU-. CSF p-tau235 discriminating accuracy was statistically lower to that of CSF p-tau231, but not CSF p-tau217. (D) ROC analysis comparing AUC values of CSF p-tau235, p-tau231 and p-tau217 discriminating amyloid-negative cognitively impaired (CI-, non-AD) from amyloid positive cognitively impaired (CI+, MCI+ and AD). CSF p-tau235 performance was statistically similar to that of CSF p-tau217 and p-tau231.
Figure 3A Diagram showing the renaturing assay. B Effect of RNA/protein ratio on protein aggregation after renaturing. C Coomassie-stained gel of soluble (Sup 1/2) and aggregated (Pel 1/2) proteins after removal of GuHCl in the presence (+) or absence (-) of total RNA. Asterix (*) denotes added RNase A. soluble (Sup 1/2) and aggregated (Pel 1/2) proteins after removal of GuHCl in the presence (+) or absence (-) of total RNA. D As in (c) but analysed by Western blot. E ATP-hydrolysing activity of proteins renatured with total RNA or buffer (Ve-) after removal of GuHCl.
Figure 2Percent body weight of male Prdx4 WT and KO mice over the 48 h course of LPS (4.5 mg/kg BW) injection (i.p.) and treatment with IL-1 receptor antagonist (IL-1RA) Anakinra (200 µg/mouse) or control. Arrows indicate time point of Anakinra injection. Each circle represents a mean of n=5 mice, vertical lines indicate SEM. Serum concentration of Cxcl1, TNF-α and IL-1β in Prdx4 WT and KO mice injected with LPS, LPS and IL-1RA or control. Horizontal lines indicate mean. Percent body weight of male Prdx4-flox and Prdx4-∆LysMCre mice over the 48 h course of 4.5 mg/kg BW LPS ( i.p.). Each circle represents a mean of n=7 mice, vertical lines indicate SEM. Serum concentration of Cxcl1, TNF-α and IL-1β in Prdx4-flox and Prdx4-∆LysMCre mice injected with LPS. Each dot (B, D) represents an individual mouse. Horizontal lines indicate mean.
Analysis of PARP-1 inhibition on DSB repair.Figure 5XRCC4wt and XRCC4m/m cells irradiated with 0.5 Gy of γ-rays were exposed to the PARP1 inhibitor 3′-AB or to solvent (DMSO). The γ-H2AX foci number is the mean ± SD of three independent experiments. *P < 0.05 (XRCC4m/m + 3′-AB versus XRCC4m/m + DMSO), ***P < 0.001 (XRCC4m/m + 3′-AB versus XRCC4wt + 3′-AB), two-way ANOVA, Bonferroni post hoc test.
Figure 3(A) Wild-type or PINK1 knock out (KO) HeLa cells were transfected with GFP-Parkin wild-type or the C431S mutant, treated with 15 µM CCCP, and then immuno-stained with anti-Flag and anti-catalase antibodies. Wild-type Parkin, but not the catalytically inactive Parkin (C431S) mutant, mediated 3Flag-MITOL translocation to peroxisomes following mitophagy stimulation. In PINK1 KO HeLa cells, the redistribution of MITOL to peroxisomes was not observed even in the presence of wild-type Parkin when mitochondria were damaged. Higher magnification images of the boxed regions are shown in the small panel. Scale bars, 10 µm. Arrowheads indicate representative examples of MITOL-peroxisome co-localization observed only in the presence of wild-type Parkin. (B) The redistribution of MITOL from mitochondria to peroxisomes does not require its own E3 activity. The E3-inactive MITOL Cys65Ser/Cys68Ser (CS) and H43W mutants were transfected into HeLa cells stably expressing HA-Parkin, treated with 15 µM CCCP, and then subjected to immunocytochemistry with anti-Flag and anti-catalase antibodies. After 3 hours of CCCP treatment, both the CS and H43W mutants co-localized with catalase. Higher magnification images of the boxed regions are shown in the small panel. Scale bars, 10 µm. Arrowheads indicate representative examples of 3Flag-MITOL co-localization with catalase. (C) Correlation statistics for the localization of 3Flag-MITOL and catalase in the presence of GFP-Parkin wild-type or an inactive C431S mutant. Dots indicate individual Pearson correlation coefficient data points. In the box-plots, the center lines indicate the medians, the box limits indicate the 25th and 75th percentiles as determined in the R software package, and the whiskers extend 1.5 times the interquartile range from the 25th and 75th percentiles. Means and the number of samples are shown on the box and X-axis, respectively. Statistical significance was calculated using a one-tailed Welch's t-test. MITOL overlapped with catalase only in the presence of wild-type Parkin in wild-type HeLa cells. In PINK1 knockout (KO) HeLa cells, wild-type Parkin was unable to induce the peroxisomal translocation of MITOL. (D) Correlation statistics for the localization of 3Flag-MITOL wild-type or inactive mutants with catalase. Dots indicate individual Pearson correlation coefficient data points. In the box-plots, the center lines indicate the medians, the box limits indicate the 25th and 75th percentiles as determined in the R software package, and the whiskers extend 1.5 times the interquartile range from the 25th and 75th percentiles. Means and the number of samples are shown on the box and X-axis, respectively. Statistical significance was calculated using a one-tailed Welch's t-test.
Figure 4A) Gastrocnemiusmuscle weight in young and old control and Mfn2KO mice (n=6 mice per group). B) Representative image of hematoxylin and eosin staining in transversal sections of gastrocnemius muscle and quantification of cross sectional area (CSA) (n=3 mice per group, 200 fibers per mouse). C) Fiber size distribution in gastrocnemius from young and old control and Mfn2KO mice (n=3 mice per group, 200 fibers per mouse). D) and E) Quantification of fiber type distribution by immunohistochemistry in gastrocnemius muscle from young and old control and Mfn2KO mice (D) and a representative image (E). Fibers were classified as IIa, IIx, IIb and I (n=3 mice per group, 200 fibers per mouse). D) and E) Quantification of fiber type distribution by immunohistochemistry in gastrocnemius muscle from young and old control and Mfn2KO mice (D) and a representative image (E). Fibers were classified as IIa, IIx, IIb and I (n=3 mice per group, 200 fibers per mouse). F) Quantification of CSA according to each type of fiber (IIa, IIx, IIb and I) in gastrocnemius muscle (n=3 mice per group, 200 fibers per mouse). G) Muscle performance was assessed by measuring the total distance and time ran on a treadmill exhaustion test (n=6 mice per group). H) Muscle force was assessed measuring grip strength in in vivo old mice (n=6 mice per group). I-L) Muscle force was evaluated in ex vivo soleus muscles from old control and Mfn2KO mice. Maximal tetanic force (I), twitch to tetanus ratio (J), and half relaxation (K) and contraction times (L) were measured (n=4 soleus per genotype).
Figure 1A. A schematic diagram of the Hog1 MAPK signaling pathway. B. Analyses of Hog1 phosphorylation by immunoblotting with anti-phospho-p38 (Hog1-P) and anti-Hog1 (total Hog1) antibodies. Cells of the indicated genotypes were stimulated with the indicated concentrations of NaCl for the indicated time. Strains used are TM257, KT207 and KY594-1. C. Analyses of Hog1 phosphorylation by Phos-tag band-shift assay. Yeast strain KY594-1 was stimulated with the indicated concentrations of NaCl for 5 min. The percentages of phosphorylated Hog1 [Hog1-P (%)] were calculated as explained in Materials and Methods, and are shown beneath the panel. Data information: Representative results from three independent experiments. D. Analyses of Hog1 phosphorylation by immunoblotting with anti-phospho-p38 (Hog1-P) and anti-Hog1 (total Hog1) antibodies. Yeast strain KT219 was transformed with the indicated STE11 mutant gene carried by a single-copy plasmid that is expressed from the STE11 promoter: vec, vector; WT, wild-type; DDD, S281D/S285D/T286D. Cells were incubated with (+) or without (-) 1 M NaCl for 5 min. E-H. Analyses of Hog1 phosphorylation by Phos-tag band-shift assay. Yeast strains (E) KY603-3; (F) TM142; (G) TM257; and (H) FP54 were stimulated with the indicated concentrations of NaCl for 5 min. Data information: Representative results from three independent experiments. I. Comparison of the NaCl dose-responses of Hog1 activation by various strains. Phos-tag band-shift assays were independently repeated three times, and average values were plotted. Data information: (I) Error bars are SEM (n=3).
Figure 3(A) Accumulation of nuclear H3.1-EGFP and the formation of distinct foci. Scale bar represents 5 μm  (B) Representative cells showing two different subpopulations of released H3.1-EGFP staining and their colocalization with PCNA positive foci. PCNA was detected with a PCNA-VHH-TagRFP chromobody. Scale bar represents 5 μm  (C) Representative cells showing points from a time course of H3.1-EGFP and H4-EGFP release. Scale bar represents 5 μm  (D) Colocalization of PCNA and H3.1/H4-EGFP signals at various time points after release. A Wilcoxon rank sum test was carried out for each time point versus the 30 min time point, with those scoring a p-value of <0.001 indicated by brackets  (E) Released H3.1/H4-EGFP colocalize with mitotic chromosomes 48 hours post release. Scale bar represents 5 μm