EMBO/sd-character-level-ner huggingface dataset:
Process length and AV distribution in neurons exposed to different treatments.Figure 3(A) The number of AV in neurites decreased significantly in neurons exposed to the SIV-infected microglia supernatant for 3 or 24 hr, however; pretreatment with rapamycin blocked this effect. *** P<0.001, for n = 6 experiments. Scale bar, 20 µm. (B) The AV number is unchanged in neuronal soma after exposure to the SIV-infected microglia supernatant for 3 or 24 hr. (C and D) Both the total length of neuronal processes (C) and number of brunch points (D) are decreased after exposure to after SIV-infected microglia supernatant treatment. *** P<0.001, for n = 6 experiments. All values are mean±SEM.
Figure 2A. Cytosolic iron in H9c2 cells preloaded with radioactive 55Fe and treated with indicated iron chelators. ANOVA followed by post-hoc Tukey test was performed. * P=7.9E-8 PBS vs. DFO. * P=9.4E-6 BPD vs. DFO. * P=0.003 PBS vs. BPD. N=4 independent samples for PBS group and N=6 independent samples for other groups.B. Nuclear iron in H9c2 cells preloaded with radioactive 55Fe and treated with indicated iron chelators. ANOVA followed by post-hoc Tukey test was performed. * P=0.0003 PBS vs. BPD. * P=5.58E-5 PBS vs. DFO. N=4 independent samples for PBS group and N=6 independent samples for other groups. C. Representative RPA fluorescence staining for labile mitochondrial iron in H9c2 cells with indicated iron chelator treatment. Scale bar = 100μm.D. Labile mitochondrial iron measured by RPA fluorescence in H9c2 cells with indicated iron chelator treatment. ANOVA followed by post-hoc Tukey test was performed.* P=0.016 PBS vs. BPD. * P=0.04 DFO vs. BPD. N=8 independent samples for each group. E. H2O2-induced cell death in H9c2 cells with indicated treatments. ANOVA followed by post-hoc Tukey test was performed.* P=5E-8 PBS-PBS vs. PBS-H2O2. * P=4E-6 PBS-H2O2 vs. DFO-H2O2. * P=5E-8 PBS-H2O2 vs. BPD-H2O2. * P=1E-7 BPD-H2O2 vs. DFO-H2O2. N=6 independent samples for each group. F. Labile mitochondrial iron in H9c2 cells with indicated treatment. PBS with and without H2O2 data were copied from Fig 1C. ANOVA followed by post-hoc Tukey test was performed. * P=1E-8 PBS-PBS vs. PBS-H2O2. * P=3E-9 PBS-H2O2 vs. BPD-H2O2. N=8 independent samples for PBS-PBS, and N=12 for the other groups. All data are expressed as mean ± SEM. N.S. = not signigicant.
Figure 6(A) Heatmap of differentially expressed genes (adjusted P-value < 0.05 and fold change > 1.5) from RNA-seq (n=3 different WT and iKO cell lines). (B) qRT-PCR validation of candidate genes selected from RNA-seq analysis using RNA from cultured cells (average SD, n=4). *Unpaired t-test P-values: Edaradd=0.03, Apcdd1=0.005, Gls2=7x10-5, Ppargc1b=3x10-4, Rab17=2x10-5, Foxn1=0.03. Mann-Whitney U test P-values: Edaradd=0.06, Apcdd1=0.03, Gls2=0.03, Ppargc1b=0.03, Rab17=0.03, Foxn1=0.03. (C) Table summarizing selection of candidate genes from (B). Genes were selected based on known function or skin phenotype related to Gata6 iKO phenotype. (D) Staining of ROS by DHE. LY83583 treated cells served as positive control with elevated ROS. (E) Quantification of fold change in ROS at 6 and 24 hours after TM compared to WT, based on fluorescence intensity of DHE (average SD, n=2). (F) Analysis of transcription factor binding site overrepresentation (oPOSSUM) in differentially expressed genes from RNA-seq. Points represent transcription factors plotted as Z.score of the frequency of transcription factor binding site occurrences in the promoters of downregulated genes compared to the background rate of occurrences in all gene promoters and Fisher P-value of the proportion of downregulated gene promoters containing each transcription factor binding site compared to the frequency in all gene promoters. (G) Comparison of Gata6 iKO downregulated genes with genes upregulated in Eda transgenic (Tg) skin. Genes related to DNA replication and repair are found in the set of genes shared between the two datasets. (H) qRT-PCR of selected genes found to be overlapping between Gata6 iKO and Eda transgenic analyzed in Gata6 WT and iKO keratinocytes (average SD, n=4). *Unpaired t-test P-values: Mcm3=0.001, Mcm6=0.02, Mcm10=0.04, Exo1=0.02, Pold1=0.02. Mann-Whitney U test P-values: Mcm3=0.03, Mcm6=0.03, Mcm10=0.03, Exo1=0.03, Pold1=0.06.
Mitochondrial respiration of Coq9+/+, Coq9Q95X and Coq9R239X mice.Figure 7A-D Measurement of phosphorylating respiration (represented as State 3o, in the presence of ADP and substrates) in kidney (A) and skeletal muscle (C) from male and female Coq9+/+, Coq9Q95X and Coq9R239X mice at 3 months of age. Representative O2 consumption graphic in kidney (B) and skeletal muscle (D) from female Coq9+/+, Coq9Q95X and Coq9R239X mice.Data information: All values are presented as mean ± SD. (A, C) *P < 0.05; **P < 0.01; ***P < 0.001; Coq9Q95X and Coq9R239Xmice versus Coq9+/+mice. #P < 0.05; Coq9Q95X versus Coq9R239Xmice. One-way ANOVA with a Tukey's post hoc test. Numbers above columns indicate P-values of the one-way ANOVA test (n = 3 for each group).
Depletion of FYCO1 leads to the accumulation of perinuclear clusters of autophagosomes.Figure 9.(A) Western blot (WB) of cell lysates from HeLa cells stably expressing GFP-LC3 48 h after transient transfection with siRNA against FYCO1 (siRNAs: A, Applied Biosystems; D, Thermo Fisher Scientific). MM, molecular mass. (B) Representative images of HeLa cells with or without the perinuclear clusters of GFP-LC3-positive vesicles (right) and quantification of the number of cells with this phenotype 48 h after transient transfection with siRNAs against FYCO1 (left). Error bars represent SDs based on three independent experiments. (C, D, and E) The perinuclear clusters of GFP-LC3-positive vesicles colocalize with mCherry-Rab7 but not with Golgi protein marker 58K or LysoTracker red. GFP-LC3 HeLa cells were transiently transfected with siRNA against FYCO1 (Thermo Fisher Scientific) 48 h before imaging or cotransfected with mCherry-Rab7. Cells were either fixed and stained with anti-Golgi 58K antibody or incubated with 50 nM LysoTracker red for 1 h. Insets show an enlarged field of interest. Bars, 10 µm.
Comparison of the Atg14-binding capacities and autophagy-induction activities of CbpC family proteins.Figure 6A Comparison of the amino acid sequences of the dp3 domains from CbpCT4 orthologs of other Streptococcus strains. The identical amino acid residues in all sequences in (A) and (B) are indicated by asterisk. B Comparison of the amino acid sequences of the dp3 domains in CbpCT4 orthologs CbpJ, CbpFT4, CbpFR6, and PcpC. C Full-length structures of CbpCT4 (PDB ID: 2V04), CbpJ (PDB ID: 6JYX), and PcpC (homology model). D GST-pulldown assays using the indicated GST-fusion proteins or GST and lysates from 293T cells expressing GFP-Atg14 CCD or GFP. Bound proteins were analyzed by immunoblotting using an anti-GFP antibody. E Lysates from 293T cells expressing GFP-fusion proteins or GFP were subjected to SDS-PAGE and analyzed by immunoblotting using antibodies against LC3, GFP, or actin. F Confocal images of HeLa cells transiently expressing GFP-fusion proteins. Scale bars, 10 µm. G Diagram of CbpCT4/CbpFR6, CbpJ, and PcpC dp3 domain swaps with CbpJ or CbpCT4/CbpFR6. H, I GST-pulldown assays using the indicated GST-fused proteins or GST and lysates from 293T expressing GFP-Atg14 CCD or GFP. Bound proteins were analyzed by immunoblotting using an anti-GFP antibody.
a) The basal flux of autophagy is affected by IRGM expression level.figf3(a) The basal flux of autophagy is affected by IRGM expression level. HEK293 cells transfected with an IRGM-expressing plasmid, miR-196 or siIRGM were treated with bafilomycin A1 for 2 h and processed for immunoblotting with anti-LC3B. (b) Quantification of LC3-II relative to actin (mean of three independent experiments ± s.d.). (c) Downregulation of IRGM expression by miR-196 abrogates AIEC-mediated autophagy in cells treated with autophagic inhibitors (Inh) or transfected with miR-196B and infected for 4 h with AIEC LF82 (mean of three independent experiments ± s.d.). (d) Confocal microscopic examination of LC3 revealed a significant decrease in the percentage of LC3-associated (red) LF82 bacteria (green) in miR-196 transfected cells compared to control cells (mean ± s.d.). (e) miR-196 transfection leads to increased intracellular LF82 replication. Results are expressed as a fold increase ± s.e.m. of intracellular bacteria. (f) IRGM overexpression did not inhibit autophagic flux and it increased LC3-II accumulation slightly in response to AIEC infection. HEK293 cells were transfected with IRGM-expressing plasmid, treated with autophagic inhibitors and infected with AIEC bacteria for 4 h (mean of three independent experiments ± s.d.). (g) Confocal microscopic examination showed an increased percentage of LC3-associated AIEC bacteria in IRGM cells compared to control cells (mean ± s.d.). (h) IRGM overexpression led to a high rate of intracellular replication of LF82 bacteria. (i) Most of the bacteria reside in non-acidic vacuoles, as shown with lysotracker at 8 h post infection (means of three independent experiments ± s.d.).
Figure 2A Transcriptional activity associated with key oncogenic traits and signals, by tumor-specific cell type and patient, as indicated. Red: High activity, blue: low activity. Significance was assessed by Kruskal Wallis Test (FDR corrected p<0.05), followed by a post-hoc analysis using a Wilcox-test of each group against all other groups, FDR corrected significance levels are shown (*=p<0.05; **=p<0.01; ***p<0.001). B Visualization of signatures corresponding to oncogenic traits and signals in the tumor cell transcriptome UMAP. C Cell cycle distribution of TC1-4 epithelial tumor cells, as inferred from single cell transcriptomes. D Immunofluorescence of DNA-damage-associated nuclear protein PARP. Images show adjacent normal and tumor crypts of tissue P009T, marked by N and T, respectively, and tumor tissue of patients P008 and P009, as indicated. Scale bar 100µm. Significance was assessed by an unpaired t-test, after blinded analysis of 10 random images per tumor.
a) Coimmunoprecipitation (IP) of PINK1-Myc and Flag-Fbxo7 in whole-cell lysates from U2OS cells overexpressing both proteins.figf4(a) Coimmunoprecipitation (IP) of PINK1-Myc and Flag-Fbxo7 in whole-cell lysates from U2OS cells overexpressing both proteins. IB, immunoblot. (b) Coimmunoprecipitation of PINK1-Myc with full-length and two N-terminally deleted Flag-Fbxo7 forms. PINK1-Myc is detected at low levels in complex with Flag-Fbxo7(89-522) but not with Flag-Fbxo7(129-522). (c) As in b, using lysates from U2OS cells expressing PINK1-Myc, and Flag-Fbxo7 containing either N- or C-terminal truncations. (d) In vitro coprecipitation experiments were performed using in vitro-translated (IVT) PINK1-Myc and either GST or GST fusions of Fbxo7(1-398) or Fbxo7(129-398) immobilized on glutathione beads. (e) Competitive binding assays using immobilized GST-Fbxo7(1-398) incubated with IVT Flag-Parkin and/or full length (top panel) or N-terminally truncated (bottom panel) PINK1-Myc. Input and bead-bound proteins were analyzed by immunoblotting as indicated. (f) As in e, immobilized GST-ΔN-PINK1 (top panel), GST-Parkin (middle panel) and immobilized GST alone (bottom panel) were incubated with combinations of IVT Flag-Parkin, T7-ΔN-PINK1 and Fbxo7-HA as indicated. Input and bead-bound proteins were analyzed by immunoblotting with the indicated antibodies. All western blots were performed a minimum of three times. Full-length blots are presented in Supplementary Figure 9.
Figure 9Nitrate treatments increase cytosolic calcium levels which interact with calcium binding proteins such as kinases in the nitrate signaling pathway. Kinases phosphorylate protein targets such as transcription factors that can mediate changes in gene expression in response to nitrate treatments. Nitrate can also cause dephosphorylation of specific proteins. We showed nitrate treatments promote PIN2 dephosphorylation at S439 which impact membrane localization and polarity of the PIN2 protein. This post-translational regulatory mechanism is important for modulation of primary root growth and lateral root density in response to nitrate treatments. Arrows and bar-headed solid lines represent activation or inhibition in response to nitrate, respectively. Dashed arrows indicate proposed connections. A working model (depicted in synopsis image) was constructed with the BioRender.com software.
Figure 6Snf1 maintains glycolysis and glucose transporter gene expression in both WT and ρ0 cells, and represses ribosome and Ribi gene expression in ρ0 cells. ρ0 snf1∆ cells were prepared by acute EtBr (25 µg/ml) treatment of snf1∆ cells in SCD for 2 days. RNAseq was performed with three biological replicates per genotype (Dataset EV8). All the transcripts and two categories of transcripts (gene lists in Table EV1B) are shown. Significantly changed genes (P value < 0.05; fold change > 1.5 or < 0.67) are highlighted with dashed lines; numbers of significantly changed genes are indicated. Snf1 represses ribosome biogenesis in WT and ρ0 cells. Polysomes of the indicated strains were fractionated by sucrose gradients (10%-50%), and the gradients were separated and measured at A254. Deletion of SNF1 decreases ∆ψm in both ∆V and ρ0 cells. Cells were cultured to mid-log phase and stained with 125 nM TMRM for FACS analysis Deletion of SNF1 decreases the proliferation of both ∆V and ρ0 cells. Serial dilutions of the indicated strains were analyzed on SCD plates at 30℃ for 2 days. Deletion of MIG1 in ρ0 snf1∆ cells rescues ∆ψm. Cells were cultured to mid-log phase and stained with 125 nM TMRM for FACS analysis Deletion of MIG1 in ρ0 snf1∆ cells partially rescues cell proliferation. A plasmid overexpressing MIG1 under the control of MIG1 promoter was transformed into the indicated strains. Serial dilutions (tenfold dilution) were analyzed on SCD plates at 30℃ for 2 days. V: empty vector. Combining MIG1 deletion and NOG2 overexpression in ρ0 snf1∆ cells fully rescues cell proliferation. A plasmid overexpressing NOG2 under the control of TEF1 promoter was transformed into the indicated strains. Serial dilutions (tenfold dilution) were analyzed on SCD plates at 30℃ for 2 days. V: empty vector. Cartoon illustration that Snf1 signaling in ∆V and ρ0 cells upregulates glycolysis and represses ribosome biogenesis to optimize energy metabolism.
Figure 7Scatter plots representing the associations of CSF PGRN with CSF sTREM2 and each of the AD CSF core biomarkers (T-tau, P-tau181P and Aβ1−42) in healthy controls (blue; A, D, G and J), Alzheimer's continuum (red; B, E, H and K) and 'suspected non-Alzheimer's pathophysiology (SNAP)' groups (green; C, F, I and L). Each point depicts the value of CSF PGRN and the corresponding biomarker of a subject and the solid lines indicate the regression line and the 95% confidence interval (CI) for each of the groups. The standardized regression coefficients (β) and the P-values are shown and were computed using a linear model adjusting for age, gender and APOE ε4. The sample contained some outliers (defined as 3 SDs below or above the group mean) of the CSF core markers of AD, the analysis including these outliers yielded similar results (Appendix Table S8). The Aβ1-42 values used for the associations test are those based on an extrapolation curve since the upper technical limit is 1700pg/ml. We also tested the associations with Aβ1-42 values truncated at the upper technical limit and the result was similar. Abbreviations: Aβ1-42: amyloid-β 42; T-tau: total tau; P-tau: tau phosphorylated at Threonine 181; SNAP: suspected non-Alzheimer's pathophysiology.
SKIP recruits non-mitochondrial TBC1D15.Figure 8Representative confocal images of either control (siC) Hela cells harbouring endogenous TBC1D15 tagged with GFP (Ge1D15, green) or those depleted of FIS1 using a pool of siRNA oligos, fixed and immunolabeled for the mitochondrial marker TOM20 (blue) in combination with either endogenous A. Rab7 Zoom insets (2.5x) highlight select cell regions, arrowheads point to vesicles positive for TBC1D15 and negative for TOM20 (yellow). Representative confocal images of either control (siC) Hela cells harbouring endogenous TBC1D15 tagged with GFP (Ge1D15, green) or those depleted of FIS1 using a pool of siRNA oligos, fixed and immunolabeled for the mitochondrial marker TOM20 (blue) in combination with either endogenous B. CD63 (red). Zoom insets (2.5x) highlight select cell regions, arrowheads point to vesicles positive for TBC1D15 and negative for TOM20 (yellow). Effect of FIS1 on late compartment distribution. C. Representative confocal images of fixed HeLa cells depleted of FIS1 using a siRNA oligo pool and immunolabeled against CD63 (white). D. Plot of CD63 pixel distribution as a function of FIS1 depletion expressed as fractional distance along a straight line from center of nucleus (0) to the plasma membrane (1.0), number of (pixels) plotted given above each scatter, n siC=14, n siFis1=17 cells analysed from 2 independent experiments. E. Colocalization of SKIP with endogenous Rab7 (Mander's overlap) in response to FIS1 depletion, n siC=13, n siFis1=9 images (3≥ cells per image) analysed from 2 independent experiments F. Validation of FIS1 depletion efficiency assayed by qPCR and expressed as fraction FIS1 mRNA remaining relative to siC, n=3 independent experiments. G-J. Recruitment of endogenous TBC1D15 to the SKIP compartment. G. Left panels: representative confocal images of fixed HeLa cells harbouring endogenous TBC1D15 tagged with GFP (G-e1D15, green) and ectopically expressing either empty vector, HA-SKIP or PLEKHM1-HA (red), immunolabeled against HA. Zoom insets (2.7x) highlight select peripheral (PP) and perinuclear (PN) cell regions. Right panels: pixel plots of endogenous GFP-TBC1D15 (green line) and HA signals (red line) corresponding to the dashed white lines in G. H-J. Colocalization (Mander's overlap) between the indicated pairs of proteins, n≥5 images, 3≥ cells per image, analysed per condition from 2 independent experiments K. Graphical summary of TBC1D15 recruitment to SKIP.
a) Flow cytometry of circulating LCMV-specific CD8+ T cells among peripheral blood mononuclear cells from Atg5fl/fl and Atg5fl/flGzmb-Cre mice on days 8, 15 and 50 after infection with LCMV Armstrong strain.figf5(a) Flow cytometry of circulating LCMV-specific CD8+ T cells among peripheral blood mononuclear cells from Atg5fl/fl and Atg5fl/flGzmb-Cremice on days 8, 15 and 50 after infection with LCMV Armstrong strain. Numbers adjacent to outlined areas indicate percent H-2Db-gp33+ T cells, gated on CD8+ T cells. (b) Longitudinal kinetics of the appearance of H-2Db-gp33+ T cells in the peripheral blood of mice as in a (left), and percent H-2Db-gp33+ CD8+ T cells remaining on day 15 (right) (all presented as in Fig. 4h). (c) Flow cytometry of T cells in tissues on day 279 after infection as in a (numbers in plots as in a, among total gated CD8+ T cells). (d) Total T cells specific for H-2Db-gp33 (left) or H-2Db-NP396 (right) in the spleen, liver and lungs of mice as in a. *P ≤ 0.05, **P ≤ 0.005 and ***P ≤ 0.0005 (unpaired t-test). Data are representative of two independent experiments with three to seven mice per group (error bars (d), s.e.m.).
Pemafibrate reduces tissue damage during sepsis.Figure 6Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Plasma was collected 24h post-surgery and (A) Alanine-amino transferase (ALT), were measured as described in methods. P-values were calculated with 2-way ANOVA tests. n = 6-7/group, combined data of 2 independent experiments. Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Plasma was collected 24h post-surgery and creatinine concentrations, were measured as described in methods. P-values were calculated with 2-way ANOVA tests. n = 6-7/group, combined data of 2 independent experiments. Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Plasma was collected 24h post-surgery and IL-6 levels were measured as described in methods. P-values were calculated with 2-way ANOVA tests. n = 6-7/group, combined data of 2 independent experiments. Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Apoptosis in liver paraffin-fixated sections 24h after sepsis, measured with TUNEL staining and depicted as % of PI positive cells/µm² tissue area. P-values were calculated with 2-way ANOVA tests. n = 6-7/group, combined data of 2 independent experiments. Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Apoptosis in kidney paraffin-fixated sections 24h after sepsis, measured with TUNEL staining and depicted as % of PI positive cells/µm² tissue area. P-values were calculated with 2-way ANOVA tests. n = 6-7/group, combined data of 2 independent experiments. Mice were pre-treated with pemafibrate (1mg/kg) or vehicle (0,9% NaCl) for 1 week before being subjected to CLP. Systemic bacterial load (CFU/ml blood) 24h post-sepsis in vehicle or pemafibrate treated mice. P-values were calculated with 2-way Student's t-test. Pemafibrate (1 mg/kg) or vehicle (0,9% NaCl) was administered at different timepoints before and after the induction of sepsis and survival was monitored during 9 days, after which no further deaths occurred. Survival curve was analyzed via Log rank tests Mice were injected with the PPARα antagonist GW6471 (10µg/g) or vehicle (DMSO) 3h pre- and 3h post-CLP. Survival was monitored during 9 days, after which no further deaths occurred. Survival curve was analyzed via a Log rank test. Combined data of 2 independent experiments, n = 16/group.