EMBO/sd-character-level-ner huggingface dataset:
Figure 3(A) Representative TEM recordings of WT cells and MICOS-KO mutants, as indicated. (B) Quantification of the overall cristae morphology on TEM recordings. Data information: n: number of mitochondrial sections (C) Quantification of detailed morphology of CMs in MICOS mutants (left). TEM recordings were assigned to 8 classes (right). Data information: n: number of mitochondrial sections (D) CJ frequency on TEM recordings. The number of CJs was manually determined and normalized to the length of the OM. Samples were compared to WT by a One-way ANOVA test. Error bars: SEM. Data information: n: number of CJs (E) Diameter of CJs. CJs were manually measured on TEM recordings. Red line: Average. Error bars: SD. Samples were compared to WT by a One-way ANOVA test. Data information: n: number of mitochondrial sections (F) Representative ET reconstructions of mitochondria from a WT cell (left), a Mic10-KO cell (center) and a Mic60-KO cell (right). The OM is displayed in clear grey, the side of the IM that faces the matrix is shown in dark blue. The IM side that faces the IMS is shown in light blue.
Figure 7A Oxygen consumption rate (OCR) of cardiomyocytes from patient TAZ10 and control at basal conditions and after the administration of the indicated compounds. B Mitochondrial membranes analyzed by BN-PAGE and Western-blotting using indicated antibodies. For a loading control samples were separated on SDS gel and analyzed by Western-blotting (lower panel). C Western blot analysis of SDHA protein levels compared to VDAC3. Quantification of SDHA relative to VDAC3 in percent with control set to 100%. Error bars indicate range of values of two independent experiments. D Analysis of SDHA gene expression in triplicates by qPCR in cardiomyocytes, analyzed in (C).
Histological analyses of the FBXO7-/- mice.Figure 2A. B. C. and D. Striatal tissue from P18 FBXO7+/+ and FBXO7-/- mice was subjected to HPLC analyses to determine dopamine (A), 3,4-dihydroxyphenylacetic acid (DOPAC) (B) and homovanillic acid (HVA) (C) concentrations along with the metabolite ratio (D). n=12 and 13, respectively (unpaired t-test, *p<0.05, mean +s.e.m.). E. Representative images from FBXO7+/+ and FBXO7-/- midbrains. Insets display higher magnification of TH+ neurons of substantia nigra. Scale bar = 500 µm. Stereological analysis of the ratio of TH+Nissl+ cells/Nissl+ cells in the substantia nigra. Three independent litter pairs were analyzed (paired t-test, * p<0.05, mean +s.e.m.) F. Sagittal paraffin sections of brains from P18 FBXO7+/+ and FBXO7-/- mice were subjected to immunohistochemistry using the GFAP antibody. Five independent litter pairs were analyzed (paired t-test, * p<0.05, mean +s.e.m.). Scale bar = 40 µm. G. Representative images of sagittal paraffin sections of cortices from P18 FBXO7+/+ and FBXO7-/- mice that were subjected to TUNEL staining. Three independent litter pairs were analyzed (paired t-test, * p<0.05, mean +s.e.m.). Arrows indicate TUNEL+ cells. Scale bar = 40 µm. H. Analyses of TUNEL staining of cerebellum and hippocampus. Three independent litter pairs were examined (paired t-test, mean +s.e.m.). I. Cortical neurons were transfected at day in vitro (DIV) 3 with control vector, a functional or a non-functional FBXO7 shRNA plasmid together with a transfection marker. At DIV6 apoptotic neurons were counted. Four independent experiments were included in the analysis (ANOVA, ** p<0.01, mean +s.e.m.).
Fig.Figure 3(A) Representative images of Sca-1 (green), Laminin (white), DAPI (blue) immunofluorescence, and miR-206-3p (violet) immunohistochemistry on 10 uM sequencial cryosections of Tibialis Anterior (TA) of young mdx mice (CTR) and after TSA treatment (TSA) (0.6 mg/kg/day for 15 days by i.p.) (n=6). Scale bar = 25 μm.(C) Graph representing the miR-206-3p relative expression in EVs isolated from muscle interstitium of tibialis anterior from wild type mice (CTR), mdx young and old mice (control -CTR- and TSA treated) (n=4). Star (*) indicates Tukey analysis compared to wild type mice (CTR), ****p < 0.0001; hash (#) indicates Tukey analysis compared to mdx young CTR, #### p<0.0001. § represents statistical analysis by Anova test. §§§§ p<0,0001. (D) Representative images of CD90 (green), Dapi (blue) and miR-206-3p (violet) immunohistochemistry in brachial biceps bioptic samples of DMD patient before (CTR) and after Givinostat treatment (GIV). Scale bar = 50 μm.(F) Graph showing the relative expression of miR-206-3p in EVs isolated from a human population of muscle resident cells enriched in FAPs (ehFAPs) isolated from biopsies of two DMD patients (DMD-1 and DMD-2) before (CTR) and after (GIV) Givinostat treatment, Two technical replicates are shown.
Figure 2Quantitative RT-PCR and immunoblots of the expression of CD38 and SARM1 in DC and age-matched health control fibroblasts. All values are presented as mean ± SD of three replicates in quantitative RT-PCR. Protein levels are normalized to GAPDH, and mean (± SD) quantification values of 4 controls vs 5 DC samples are shown. Student's t test was performed on DC cells vs controls. Irrelevant intervening lanes have been removed for clarity (full blots are available online as Source Data). Immunoblots and quantification of indicated proteins in DC fibroblasts treated with DMSO and with 2 μM ATM inhibitor, KU-55933 for 48 hours. Mean (± SD) quantification values of three DC lines with and without ATMi treatment are shown. P values on the basis of Student's t test. The efficacy of CD38 shRNA (sh-1 and sh-2) was verified by quantitative RT-PCR in DC fibroblasts and by RT-PCR and immunoblots in A549 cells. The relative CD38 mRNA values in the CD38 knockdown cells were normalized to the scrambled shRNA. All values are presented as mean ± SD of three replicates. Student's t test was performed on DC and A549 cells in indicated conditions. Intracellular NAD levels in the scramble and CD38 knockdown DC fibroblasts. The CD38 knockdown DC fibroblasts were treated with the PARP1 inhibitor Olaparib (400nM) and the SIRT1 inhibitor EX 527 (1 μM) for 24 hours. Data are representative four replicates. All values are presented as mean ± SD of four replicates. One-way ANOVA was performed on DC cells in indicated conditions. immunoblots of the expression of indicated proteins in the scramble and CD38 knockdown DC fibroblasts. The CD38 knockdown DC fibroblasts were treated with the PARP1 inhibitor Olaparib (400nM) and the SIRT1 inhibitor EX 527 (1 μM) for 24 hours. Data are representative four replicates. All values are presented as mean ± SD of four replicates. One-way ANOVA was performed on DC cells in indicated conditions.
Figure 1(A) Heat map representation of the TNF-α-up-regulated genes. Genes were classified based on the TNF-α-treated time points, using the algorithm "HOPACH". (B) Heat map showing down-regulated miRNAs at 4 hrs, but not 24 hrs, after TNF-α treatment. The middle column indicates the fold-change values (control/TNF) at 4 hrs. The right column indicates the relative expression level of each miRNA from the FANTOM5 database (Consortium et al, 2014). Asterisks indicate the top three most down-regulated miRNAs which were reported to be expressed in human endothelial cells (ECs) according to the FANTOM5 database. (C) Bar plots showing mean mRNA levels of vascular cell adhesion molecule-1 (VCAM1), intracellular adhesion molecule-1 (ICAM1), and E-selectin (SELE), measured 4 hrs after stimulation of human ECs with or without TNF-α ± miR-3679-5p. Data are shown as means ± SE (n=3). *P < 0.05 compared to TNF-α (-) mimics (-). † P < 0.05 and ‡ P = 0.203 compared to TNF-α (+) mimics (-). Statistical differences were analyzed by the Tukey-Kramer test. (D) Western blots for VCAM1, ICAM, SELE, and β-actin (ACTB) in lysates from ECs treated with or without TNF-α (4 hrs) ± miR-3679-5p. (E) Representative images (top) and bar plot quantification (bottom) showing the adhesion of calcein-labeled U937 monocytes to ECs treated with or without TNF-α ± miR-3679-5p. Scale bar represents 250 μm. Graphs are representative of three independent experiments. Data are shown as means ± SD. *P < 0.05 compared to TNF-α (-) mimics (-). † P < 0.05 compared to TNF-α (+) mimics (-). Statistical differences were analyzed by the Tukey-Kramer test. (F) The scatter plot shows the correlation of RNA immunoprecipitation (RIP)　followed by microarray data from two replicate screens for the identification of miR-3679-5p target genes. Genes up- and down-regulated by miR-3679-5p treatment (|average Log2FC|>1) are indicated in red and blue, respectively. (G) Venn diagram showing the overlap of the genes selected by RIP-ChIP (1,334) and TargetScan7.0 (1,224) (http://www.targetscan.org/vert_70/). (H) Gene ontology (GO) analysis of the 253 potential target genes for miR-3679-5p, as determined in Fig 1G. The P-values of each category analyzed from DAVID (https://david.ncifcrf.gov/) are shown in the bar graphs. (I) Seed sequence of miR-3679-5p and complementary 3′ UTR sequence of lysine demethylase 7A (KDM7A). The letters in red indicate matched bases. (J and K) Effect of miR-3679-5p mimics (J) or miR-3679-5p inhibitor (K) on luciferase activity in pmirGLO-transfected ECs expressing the 3′ UTR of KDM7A. Data are shown as means ± SE (n=3). *P < 0.05 compared to control. Statistical differences were analyzed by the Student's t test. (L) Seed sequence of miR-3679-5p and complementary 3′ UTR sequence of lysine demethylase 6A (UTX). The letters in red indicate matched bases. (M and N) The effect of miR-3679-5p mimics (M) or miR-3679-5p inhibitor (N) on luciferase activity in pmirGLO-transfected ECs expressing the 3′ UTR of UTX was measured. Data are shown as means ± SE (n=3). *P < 0.05 compared to control. Statistical differences were analyzed by the Student's t test.
a) Haematoxylin and eosin staining of cross-sections from Dia showing central nuclei (arrow head) and smaller fibres (arrow).f4(a) Haematoxylin and eosin staining of cross-sections from Dia showing central nuclei (arrow head) and smaller fibres (arrow). (b) Immunoblot analysis of macrophage content (CD68). (c) Immunofluorescent and bright field images of Dia cross-sections showing fibre-type distribution. Type I (red), IIA (green), IIB/IIX ( white x, unstained and viewed from bright field overlay). (d) Serum creatine kinase activity. (e) Force-frequency relationship in Dia muscle strips from WT (black), mdx (red) and p47−/−-mdx (blue) mice. Scale bars represent 55 μm. For e, #P0.01 p47−/−-mdx versus mdx.##P0.01 p47−/−-mdx versus WT and mdx. Mdx was statistically different than WT at all frequencies of stimulation. Statistical differences between groups were determined using ANOVA with Tukey's post hoc test. *P0.05 and **P0.01.
Figure 1A Analysis of Dusp5 mRNA expression by quantitative PCR analysis. Splenic innate and adaptive immune cells were enriched by FACS sorting to >95% purity and RNA harvested for Taqman gene expression analysis. Gene expression levels were normalized to Gapdh using the 2-delta Ct method. B Percent eosinophils in blood of Dusp5+/+ Rag2-/- (open circles) and Dusp5-/- Rag2-/- (grey squares) mice prior to and at days 3, 6 and 13 following N. brasiliensis infection. C-F Eosinophils numbers in (C) blood, (D) BALF, (E) spleen and (F) BM of Dusp5+/+ Rag2-/- (open circles) and Dusp5-/- Rag2-/- (grey squares) mice 13 days following infection with N. brasiliensis. C, E and F are from the same experiment and are representative of three independent experiments. D is cumulative of 3 independent experiments. G Intestinal worm burden 13 days post infection in Dusp5+/+ Rag2-/- (open circles) and Dusp5+/+ Rag2-/- (grey squares) mice. Data shown in (G) are cumulative of three independent experiments.
Phosphorylation of TIP60 S90 and S86 are required for TIP60 HAT activity (A) U2OS cells were transfected with a siRNA pool targeting human TIP60 (Dharmacon) or luciferase (Dharmacon).Figure 5 (A) U2OS cells were transfected with a siRNA pool targeting human TIP60 (Dharmacon) or luciferase (Dharmacon). The nuclear lysates were analysed by Western blotting 24h after transfection  (B) U2OS cells stably expressing FLAG-TIP60WT, FLAG-TIP60S90A, FLAG-TIP60S86A or empty vector (vec) were analysed by Western blotting 60 hours after viral transduction  (C) 48 hours after viral transduction, U2OS cells stably expressing FLAG-tagged TIP60WT, TIP60S90A, TIP60S86A or empty vector (vec) were subjected to real time cell analysis (RTCA) for 42 hours. Real-time resistance measurement data were recorded every hour and presented as Cell Index. Curves and arrow bars represent mean and standard deviation of four technical replicates of one experiment (D) The experiment as shown in (C) was repeated three times. Each dot represents the slope of the Cell Index of the first 30 hours of one technical replicate (one-way ANOVA, Bonferroni"s multiple comparison test, vec vs. WT p>0,9999, S90A vs S86A p=0,1276, ****p < 0,0001;; n.s. not significant, p > 0,05)  (D) The experiment as shown in (C) was repeated three times. Each dot represents the slope of the Cell Index of the first 30 hours of one technical replicate (one-way ANOVA, Bonferroni"s multiple comparison test, vec vs. WT p>0,9999, S90A vs S86A p=0,1276, ****p < 0,0001;; n.s. not significant, p > 0,05)  (E) Proposed model: The interaction of TIP60 with chromatin is regulated by CDK9 and Brd4. The phosphorylation of S90 stabilises the interaction of TIP60 and H3 via the chromodomain, and BRD4 and CDK9 positively regulate the recruitment of TIP60 to chromatin. In addition, the phosphorylation of S90 is the priming-phosphorylation required for GSK3-mediated phosphorylation of S86. Double phosphorylated TIP60, on S90 and S86, exhibits increased HAT activity
TLR8-mediated inhibition of HIV is autophagy dependent.Figure 7TLR8-mediated inhibition of HIV is autophagy dependent.(A-B) Macrophages transduced with BECN1 shRNA (shBECN1) (A) or ATG5 shRNA (shATG5) (B) were incubated with CL097 for 24 h before infection with HIV with continuous CL097 treatment for 10 d. Right, immunoblot analysis performed using antibodies raised to BECN1, ATG5 and β-actin. Left, ELISA performed for extracellular release of HIV p24 antigen over 10 d. Results are reported as mean ± s.e.m., n = 4. (C) Macrophages were pretreated with bafilomycin A1 (Baf A1), SID 26681509 (SID) or vehicle control before incubation with CL097 or ssRNA40 for 24 h before infection with HIV. Cells were then washed and incubated with TLR8 agonists and inhibitors for 10 d and ELISA performed for extracellular HIV p24 antigen release over 10 d. Results are reported as mean ± s.e.m., n = 5. * p<0.05; # p<0.001.
Losartan normalizes Tgf-β activity in C7-hypomorphic mice.Figure 3A-D C7-hypomorphic mice were treated with losartan for 7 weeks, and the forepaws of age-matched untreated, losartan-treated, and wild-type mice were subjected to immunofluorescence staining with antibodies to active thrombospondin 1 (Tsp1, green) (A), active Tgf-β1 (green) (B), TGF-β receptor II (Tgfbr2, red) (C), and phospho-Smad2 and 3 (P-Smad2/3, red) (D). The nuclei were counterstained with DAPI (blue). Images were acquired with a 20× objective (scale bar = 100 μm) (A), with a 4× objective (scale bar = 200 μm) (B, D), and with a 40× objective (scale bar = 50 μm) (C). The bar graphs on the right show quantification of the stainings in the left panel of (B) and (D). Paired values were normalized to the staining intensity of untreated C7-hypomorphic paws, which were set to 100%. Values represent mean ± S.E.M., n = 6, paired Student's t-test, ***P-value wild-type vs. C7-hypomorph receiving no treatment = 0.0004; ***P-value losartan treatment vs. no treatment = 0.0005 (B). Values represent mean ± S.E.M., n ≥ 7, unpaired t-test with Welch's correction used, ***P-value wild-type vs. C7-hypomorph < 0.0001; ***P-value C7-hypomorph + losartan vs. C7-hypomorph = 0.0002; **P-value wild-type vs. C7-hypomorph + losartan = 0.0016 (D).
Figure 1Kaplan-Meier curves for overall survival (OS) in lung adenocarcinoma TCGA_LUAD (RASSF1 mRNA high/low cutoff FKPM 5.85) and squamous cell carcinoma patients TCGA_LUSC (RASSF1 mRNA high/low cutoff FKPM 6.52). Significance derived from log-rank test. Western blot with indicated antibodies of isogenic H1299 cells stably transfected with either empty vector pcDNA3 (H1299control) or RASSF1A (H1299RASSF1A). Bottom: cell proliferation resazurin assay. (n=2). Error bars represent mean ± S.E.M. Cartoon of lung adenocarcinoma orthotopic injection, with sites of primary and metastatic tumours indicated (ipsil = ipsilateral, same lung as primary tumour; contra = contralateral in the opposite lung). Mice were euthanized to collect the lungs at day 17 or day 30 after inoculation with tumours cells into the left lung. Size of lung primary tumours measured at day 30 after lung orthotopic injection either with H1299control (n=14 mice per group; experimental groups of 6 and 8 different shading within the groups represents two independent experiments) or with H1299RASSF1A cells stably re-expressing RASSF1A, (n=12 mice per group; experimental groups of 6 and 6). The size of primary tumours was measured by MRI software ITK-SNAP. Statistical significance via 2-tailed Student's t-test. Error bars represent mean ± S.E.M. Visual assessment quantification of metastatic events on the ipsilateral (left) and or contralateral (right) lungs, generated by H1299control or H1299RASSF1A at day 30. Result was calculated from 2 independent in vivo experiments Graph shows significant decreasing of metastases when lungs were injected with H1299RASSF1A. Statistical significance via 2-tailed Student's t-test. Error bars represent mean ± S.E.M.
Figure 4Coomassie blue stained SDS-PAGE gel (15%) of purified human MRE11. (i) Scheme shows the nuclease activity of MRE11 in degrading 5' Cy3-labeled substrates (25 nt + 60 nt with phosphorothioate bonds on both ends). (ii) The 5' Cy3-labeled substrates (80 nM) were incubated with the indicated concentration of MRE11 with or without the Mn2+ cofactor at 37°C for 20 min. Reactions were stopped by SDS and proteinase K and resolved on 27% denatured polyacrylamide gel. (iii) The results are graphed, and error bars represent the standard deviation (± SD) calculated from at least three independent experiments. Coomassie blue stained SDS-PAGE gel (15%) of purified human CST complex (CTC1 wild- type and ∆700N). DNA binding activity of wild-type CST complex and Δ700N mutant complex. The 5' Cy3- labeled substrates (80 nM) were incubated with the indicated concentrations of CST. Samples were analyzed by 0.8 % agarose gel. MRE11 degradation analysis. 5' Cy3-labeled substrates (80 nM) were pre-incubated with indicated concentrations of CST wild-type (i) or ∆700N mutant variant (ii) at 37°C for 5 min. Reactions were completed by adding MRE11 (200 nM) for an additional 20 min incubation and then stopped by SDS and proteinase K. Samples were resolved in 27% denatured polyacrylamide gel. The results are graphed and error bars represent the standard deviation (± SD) calculated from at least three independent experiments.
Figure 8A-C. Graph depicting a direct correlation between MT-flux and mitotic spindle where mean spindle length is plotted over mean MT-flux rates for all indicated conditions (A), mitotic conditions containing stable end-on attachments (B), and NDC80 depleted conditions lacking stable end-on attachments (C). Solid and dotted lines represent the linear regression and 95% confidence interval, respectively. Pearson correlation coefficient (r), coefficient of determination (r2) and corresponding p values are indicated. The error bars over x and y axis represent the standard deviation of MT-flux and spindle length, respectively. D. Representative images from spinning disk confocal live-cell time series of U2OS PA-GFP-α-tubulin/mCherry-α-tubulin cells under indicated treatments, with SiR-DNA-labelled DNA. Microtubules are shown in red and DNA in cyan in the merged image. Scale bar: 10 µm. E. Quantification of spindle lengths at metaphase following indicated treatments. Graph represents the spindle length of individual cells with mean ± SD. N (number of cells, number of independent experiments): siControl (49, 5), siMCAK (34,3), siKIF4A + siKIF15 + STLC (46, 6), siKIF4A + siKIF15 + STLC + siMCAK (43, 3), siKIF4A + siKIF15 + STLC + siKIF2A (22, 3), siCLASPs (64, 6), siCLASPs + siMCAK (33, 3), siCLASPs + siKIF2A (32, 3).
Non-ubiquitinated Orm2 accumulates in Golgi and endosome structures.Figure 3(A) Epifluorescence and phase contrast microscopy of living WT yeast cells and the indicated mutants expressing GFP-Orm2 or GFP-Orm2-K25,33R (green) with dsRed-HDEL (red). Arrowheads point to GFP-Orm2 accumulations outside the endoplasmic reticulum (ER). Data information: Scale bars = 5 µm. (B) Epifluorescence and phase contrast microscopy of tul1∆ mutants expressing GFP-Orm2 (green) with Vps4-mCherry (red, endosomes) or with Sec7-mCherry (red, Golgi) or Vps4-eGFP (green) with Sec7-mCherry. Data information: Scale bars = 5 µm. (C) Left panel: Epifluorescence and phase contrast microscopy of the indicated mutants expressing GFP-Orm2 (green) with mCherry-Cps1 (red, MVB cargo). E indicates class E compartments; Right panel: Equal amounts of the indicated mutants in serial dilutions were incubated on agar plates at the indicated temperatures. Data information: Scale bars = 5 µm.
Apoptotic induction of bortezomib.Fig 2A,C) Bortezomib treated chondrosarcoma cells showed a significantly higher level of caspase 3/7 activity than untreated cells. Untreated control cells served as reference value (ratio = 1). B,D) Cleavage of caspase-3 was detected after 24 h of bortezomib treatment by flow cytometry. The y-axis denotes cell counts and the x-axis represents fluorescence intensity of the APC antibody. The black filled histogram represents untreated control cells, the striated histogram represents 5 nM, and the checkered histogram shows 10 nM bortezomib treated cells.
Lnc-SMaRT controls the translation of Mlx-γ through direct base pairing with the G4-containing region.Figure 5A Left panel: schematic representation of the luciferase constructs produced for Mlx. Mlx 5'- and 3'UTR-interacting regions were cloned respectively upstream and downstream the Renilla luciferase coding region (RLuc-Mlx 5'-3') and mutant derivatives devoid of the 5' interacting region (RLuc-Mlx-3') or the 3' UTR interacting region (RLuc-Mlx-5') were obtained. These constructs were co-transfected in C2C12 myoblasts in growth conditions together with plasmids expressing lnc-SMaRT (+) or with a control vector (-). Right panel: Luciferase activity data, presented as the mean ± s.e.m. of three biological replicates (dots), are shown with respect to each RLuc control vector (RLuc-Mlx 5'-3', RLuc-Mlx-3', RLuc-Mlx-5') set to a value of 1. Statistical analysis was performed with ordinary analysis of variance (ANOVA) followed by Dunnett's multiple comparison test. *P < 0.05. B The RLuc-Mlx-5' construct was co-transfected in N2a cells together with plasmids expressing lnc-SMaRT (+) or with a control vector (-) and treated with control siRNA (+) or siRNA against DHX36 (-). Luciferase activity data, presented as the mean ± s.e.m. of three biological replicates (dots), are shown with respect to RLuc-Mlx-5' vector set to a value of 1. Statistical analysis was performed with ordinary analysis of variance (ANOVA) followed by Dunnett's multiple comparison test. **P < 0.01, ***P < 0.001. C Left panel: schematic representation of the luciferase constructs used. The RLuc-Mlx-5' construct and its derived mutants (RLuc-Mlxmut30, RLuc-Mlx Δ75) were transfected in N2a cells in growth conditions together with lnc-SMaRT-expressing plasmids (lnc-SMaRT) or with its derived mutant (lnc-SMaRTmut) in the indicated combinations. Right panel: Luciferase activity data, presented as the mean ± s.e.m. of three biological replicates (dots), are shown with respect to each RLuc control vector (RLuc-Mlx-5', RLuc-Mlxmut30, RLuc-Mlx Δ75) set to a value of 1. Statistical analysis was performed with ordinary analysis of variance (ANOVA) followed by Dunnett's multiple comparison test. ***P < 0.001.