EMBO/sd-character-level-ner huggingface dataset:
Figure 2(A) qPCR was conduced to measure the expression of RAD21. The indicated cells were stably transduced with control GFP-shRNA or two different RAD21-specific shRNA (shR#1 or shR#2, day 30). Data are presented as the mean ± S.D. for three independent experiments *P < 0.01 and; **P < 0.05. Statistical significance was validated by Student's t-tests. (B) Results of the Western blot analysis for RAD21, E-cadherin, and VIM in RAD21KD- MCF7, -T47D, -SNU16, and -SNU620cells are presented. (C) qPCR was performed to measure the expression of EMT markers ZEB2, CDH2, ZEB1, FN1, TWIST, and SNAIL2 in RAD21KD-MCF7 (upper graph) or -SNU16 cells (lower graph). Data are presented as the mean ± S.D. for three independent experiments *P < 0.01 and; **P < 0.05. Statistical significance was validated by Student's t-tests. (D) Up-regulated genes by RAD21KD (shR#1) in MCF7 or SNU16 cells, each of which showed more than a 2-fold change in RAD21KD cells compared to the shCont. (E) Induction of TGFB1 and ITGA5 transcription in RAD21KD-MCF7, -T47D, -SNU16, and -SNU620 cells was evaluated using qPCR. Data are presented as the mean ± S.D. for three independent experiments. *P < 0.01. Statistical significance was validated by Student's t-tests. (F) Expression of TGFB1 and ITGA5 mRNA normalized to the 18S transcript in luminal and basal-like breast cancer cell lines, E: epithelial-like, M: mesenchymal-like breast cancer cell lines. Data are presented as the mean ± S.D. for three independent experiments.
Fig.Figure 1Western blot analysis of lysates from cerebellum of control and FXS patients ''FXS'' (n = 4). Extract of pCI-hDGKk-transfected Hela cells (hDGKk) was used as antibody specificity control and size marker. Representative images of immunoblots probed with antibodies against the indicated proteins are shown. GAPDH was the loading control. Quantification of Western blots is shown on right. Protein amounts of DGKk are normalized to GAPDH and presented as fold change relative to control. B) Schematic map of DGKk constructs used for the transfection experiments and subsequent vector preparations. The different domains of the protein are indicated and represented at scale (repeated EPAPE, Pleckstrin Homology PH domain, phorbol ester/diacyl glycerol binding C1 domain, catalytic split C4 a and b domains, putative PDZ binding motive, HA-tag, grey bars interval 1kB), 5' and 3' UTR regions are represented with blue line, 3'UTR not at scale (3.8 kB). C-D) Immunoblots and quantification of lysates from Cos-1 cells transfected with plasmid pCI-mDGKk-HA or non-transfected (NT) and pre-transfected 24h before with siRNA control (siC) or against FMRP (siFMR1). GAPDH was used as a loading control. For quantification, the DGKk and FMRP signals were normalized against GAPDH signal and presented relative to the signal for siC treated cells (C). Immunoblots and quantification of lysates from Cos-1 cells transfected with pCI (empty plasmid), pCI-HA-∆N-DGKk, or pCI-mDGKk-HA, and pre-treated with siRNA control (siC) or against FMRP (siFMR1) (D). Quantifications as in C.
Figure 5Designing strategy of the Ki67-Fucci cell cycle indicator system. Representative image of CLESCs harbouring the Ki67-Fucci system showing colored cells at different cell cycle phases. Flow cytometry analysis of Ki67-Fucci CLESCs at different cell cycle stages in cells transfected with empty vector or full length Prom1. As a result, >95% of the events have been included in the analysis. The accuracy of the strategy has been validated by two independent researchers for IF quantification using Fiji 1.0 software. Representative IF staining of AcTub (blue) on Ki67-Fucci CLESCs without and with the presence of SHH. Note that the cilium could be visualised (arrows) solely when SHH is present. Representative IF double staining of either AcTub (D, red) or Arl13b (E, red) with Sox2 (green) or Ki67 (green) in CLESCs in the presence of SHH. Note the expression levels of Sox2 and Ki67 distinguished stem cells and transit amplifying cells, respectively.
Figure 1(A) Immunofluorescent staining of ABCA12 (green) and Insulin (red) in mouse islets showing extensive co-localisation in β-cells as well as expression in other islet populations (arrowheads, scale bar=25μm). (B) Co-expression of Abca12 (green) and Insulin (red) in MIN6 cells (scale bar = 10μm). (C) LacZ staining (blue) to detect gene expression of the knocked-in LacZ cassette from the Abca12 locus in islets (islet outlined, scale bar = 25μm). (D,E) ABCA12 immunostaining and the development of hyperkeratotic skin disease in E18.5 Abca12tm1a/tm1a mouse embryos (brackets indicate thickened epidermis, dashed line indicates epidermal basement membrane, scale bar = 50μm). (F, G) Immunohistochemical detection of cre expression (green) in pancreatic islets from conditional mouse lines showing co-expression with Insulin (red)(scale bar = 50μm). (H) Detection of recombination of the Abca12 locus by PCR, indicating different alleles (tm1c, wild type and tm1d). (I-K) Detection of ABCA12 expression (green) and deletion in pancreas sections from mice of indicated genotypes (islets outlined, scale bar = 25μm). (L) Quantitation of ABCA12 expression by densitometry of Western blots from purified pancreatic islets (values relative to wild type, n=3-8 animals per genotype, mean±SEM, *p=<0.05, Students t-test), showing representative blot with Actin loading control (right). Average daily food intake of conditionally modified mice (n=4-10 animals/genotype, mean±SEM). (N) animal body weight (n=4-10 animals/genotype, mean±SEM).
UBE2QL1 is essential for efficient lysophagy and cell survival after lysosomal damage.Figure 6A Gal3 puncta assay for damaged lysosomes. HeLa cells were depleted of UBE2QL1 with two siRNAs for 60 h and treated with LLOMe or EtOH alone (untreated) for 1 h. Cells were fixed at indicated times after washout, stained with DAPI and Gal3 and LAMP1 antibodies, and processed for confocal microscopy. Note increased number of Gal3 puncta in untreated UBE2QL1-depleted cells and their persistence 10 h after LLOMe-induced damage. Scale bars: 10 µm, 2 µm for inlays. Arrows indicate Gal3 and LAMP1 colocalizing vesicles. B Automated quantification The percentage of cells with more than three Gal3 puncta is shown. Graph represents data from three independent experiments with ≥ 50 cells per condition (mean ± SD). *P<0.05; **P<0.01 (One-way-ANOVA with Dunnett's multiple comparison test). C Survival assay. Control or UBE2QL1-depleted HeLa cells (48 h) were treated with increasing concentrations of LLOMe as indicated. Cell viability was measured with the MTS assay. Graph represents data from one experiment including three replicates (mean ± SD). *P<0.05; ****P<0.0001 (One-way-ANOVA with Bonferroni's multiple comparison test).
Figure 1A. Measurement of oxygen consumption rate (OCR), extracellular acidification rate (ECAR) and oxidative burst of monocytes from COVID-19 patients (COVID) and healthy controls (CTR). OCR was measured in real time, under basal condition and in response to mitochondria inhibitors: oligomycin (Oligo, 2 μM), cyanide-4-(trifluoromethoxy) phenylhydrazone (FCCP, 0.5 μM), and antimycin A plus rotenone (Rot/AA, 0.5 μM). Oxidative burst was measured in response to PMA/ionomycin (PMA/Iono, 1 μg/mL). Scatter plots show the quantification of basal respiration (indicated as Basal OCR), ATP-linked respiration (ATP-linked), maximal respiration (Max Resp), proton-leak, spare respiratory capacity (Spare), oxidative burst immediate response and basal ECAR. The maximal respiration kinetic range and the oxidative burst kinetic range are also reported and were obtained by analyzing the area under the curve (AUC) from the sixth to the tenth measurement and from the tenth to the thirteenth measurement, respectively. Data represent individual values, mean and standard error of the mean (CTR, n=12; COVID, n=13). Mann-Whitney test was used for statistical analysis. Exact p values are reported in the figure. Representative traces of OCR of monocytes from COVID-19 patients (COVID) and healthy controls (CTR) are also reported. B. Representative electron micrographs of mitochondrial ultrastructural changes observed in COVID-19 monocytes. Panels "a" and "c" are representative electron micrographs of monocytes from healthy controls, whereas panels "b", "d" and "e" are representative electron micrographs of monocytes from COVID-19 patients. In panel "f" the quantification of mitochondrial area, external perimeter, Feret's diameter, aspect ratio circularity and roundness are reported (n=225 mitochondria for each group). Black arrows in panel "d" indicate mitochondria; N refers to nucleus. Exact p values are reported in panel "f".
Figure 5A NIE-115 cells stably expressing mSt-2xML1N or -2xPLCδ1PH were cultured in growth medium with or without 200 nM Torin1. Cells were fixed and stained with anti-LAMP1 or anti-LC3 antibodies, and then analyzed by immunofluorescencemicroscopy. Insets show the boxed areas at high magnification. Bar, 10 µm. B NIE-115 cells stably expressing mSt-2xML1N treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti-LAMP1 antibodies, and then analyzed by immunofluorescencemicroscopy. Insets show the boxed areas at high magnification. Bar, 10 µm. C Quantitation of signal intensities in B showing mSt-2xML1N colocalizing with LAMP1 (means ± s.d.; n > 100 cells from three independent experiments). **, P < 0.01; *, P < 0.05. D N1E-115 cells stably expressing GFP-2xML1N treated with siControl or siINPP5E were transiently transfected with mSt-INPP5E (WT, D477N). Quantitation of signal intensities showing GFP-2xML1N colocalizing with LAMP1 (means ± s.d.; n > 20 cells from three independent experiments). **, P < 0.01; *, P < 0.05. E NIE-115 cells stably expressing mCherry-2xFYVE treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti-LAMP1 antibodies, and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Bar, 10 µm.F Quantitation of signal intensities in E showing mCherry-2xFYVE colocalizing with LAMP1 (means ± s.d.; n > 100 cells from three independent experiments). **, P < 0.01.
Figure 3Figure 3. Key transcriptional-memory gene Hoxa9 may help to overcome β-catenin dependent transformation in LSK-derived MLL-CSCs. (A) Gene set enrichment analysis (GSEA) shows "Targets of Hoxa9/Meis1, down" (Hess et al., 2006) for the indicated comparison. (B) Log10-fold FDR q-values of the indicated gene sets positively enriched in β-catenin deleted LSK-MLL-ENL compared to β-catenin deleted GMP-MLL-ENL. (C-E) RNA-seq log2-fold change of key self-renewal genes (C), RT-qPCR validation of Hox/Meis1 expression represented as mean ± SD of 3 independent experiments. (C-E) RNA-seq log2-fold change of key self-renewal genesTwo-tailed t-test was performed (D) (C-E) RNA-seq log2-fold change of key self-renewal genes GSEA showing "Hoxa9_dn.v1_up" (Faber, Krivtsov et al., 2009) for the β-catenin deleted LSK-MLL-ENL to β-catenin deleted GMP-MLL-ENL comparison (E). (F) Colony numbers in serial replating assay of indicated MLL-ENL transduced cells. Data are represented as mean ± SD. (G) Kaplan-Meier survival curve of mice transplanted with Hoxa9-/- (n=18) or WT LSK-MLL-ENL (n=13) transformed cells (solid lines) and secondary recipient mice (n=5 for Hoxa9-/- and n=4 for wt, dotted lines) as indicated. Comparisons between Hoxa9 WT and Hoxa9-/- were not significantly different (ns). See also Appendix figure S3.
Figure 8Sororin is indicated in green, SYCP3 in red, and their colocalization in yellow. The kinetochores, and the microtubules and centrioles, are indicated in blue and light blue, respectively. Nuclei and condensed chromosomes, as well as spermatocyte cytoplasms, appear in grey and white, respectively. To simplify, the scheme only represents one autosomal bivalent and the sex pair XY. Chromosomes are telocentric, and the metaphase I autosomal and sex bivalents show a single interstitial and distal chiasma, respectively. Amplified details of the centromeric areas of an autosome in metaphase I and metaphase II, and of a segregating chromatid in anaphase II, are also shown.
Figure 1(A) SA-β-gal staining of U2-OS cells, harvested at indicated times post irradiation (IR); h, hours; D, days; UT, untreated. n = 3. Scale bar: 50 μm. Bar Graph: Significance was determined by Brown Forsythe ANOVA with Dunnett's 3T test, using n = 3 biological replicates. SD shown. *** p < 0.001, n.s. = not significant. (B) Nuclear fractions from U2-OS cells were harvested at indicated time points post IR (20 Gy) as in (A). Top panel: NF-κB DNA binding was analyzed by EMSA. Lower panels: Western blot of indicated NF-κB subunits and PARP1 as loading control. The two phases of nuclear translocation (C) Gene expression analysis of two NF-κB phases following damage. RNA-Seq analysis (n = 5 biological replicates per group) of U2-OS cells, either untreated or analyzed 1.5 hours or 7 days after irradiation (20 Gy), as indicated. The heatmap shows significantly regulated genes (log2 value > 0.5 and p value < 0.05). Black arrow points to NFKBIA. Significance determined by ANOVA with Bonferroni correction for multiple testing. See also Dataset EV1. (D) Female mice 12-16 weeks old (n = 6 mice per condition (with three technical repeats per mouse), were irradiated (5 Gy) and sacrificed after 1 hour (first phase), 7 days (second phase) or left untreated. Skin or renal tissue RNA was analyzed by RT-qPCR, shown as a mean +/- SD. Significance was confirmed by ANOVA with Tukey multiple comparisons test. SD shown: * = p < 0.05, ** = p < 0.01. (E) In situ hybridization using an anti-sense IκBα/Nfkbia riboprobe on longitudinal skin sections from male and female mice (5-8 weeks of age) treated as in (D) for the time indicated. Representative sections shown from n = 6 mice per condition. Bu, bulge region (dashed lines), HS, hair shaft, IFE, interfollicular epidermis. Scale bar: 50 μm. (F) Quantitation of (E) from n = 6 mice per condition and 3-4 sections per mouse. Bulge regions positive for Nfkbia were counted and presented as percentage of total. One way ANOVA analysis with Tukey multiple comparisons test was performed. SD shown. ** = p < 0.01. (G) Skin sections as in (E) analyzed by immunofluorescence with IκBα (red) or IL-6 antibody (green) and nuclear DAPI staining (blue). Dotted lines delineate hair follicles, Bu, bulge region, HS, hair shaft, IFE, interfollicular epidermis, SG, sebaceous gland. Scale bar: 50 μm. (H) Quantitation of (G) from n = 6 mice per condition and 3-4 sections per mouse. Bulge regions positive for IκBα or IL-6 were counted and presented as percentage of total. One way ANOVA analysis with Tukey multiple comparisons test was performed. SD shown. * = p < 0.05, *** = p < 0.001.
Figure 3A, B. Identification of mitochondrial ribosomal proteins and pseudouridine synthase interacting proteins by sucrose gradient centrifugation in control cells (A) Individual fractions were separated by SDS-PAGE and immunoblotted with the indicated antibodies. The migration of the mt-SSU (28S), the mt-LSU (39S) and the mitochondrial monosome (55S) are shown. A, B. Identification of mitochondrial ribosomal proteins and pseudouridine synthase interacting proteins by sucrose gradient centrifugation in cells treated with siRNA (B). Individual fractions were separated by SDS-PAGE and immunoblotted with the indicated antibodies. The migration of the mt-SSU (28S), the mt-LSU (39S) and the mitochondrial monosome (55S) are shown. C. Quantification of the levels of the mt-SSU (28S), the mt-LSU (39S) and the mitochondrial monosome (55S) normalized to the SDHA levels. The graph represents the relative abundance of individual subunits in cells treated with specified siRNA versus controls. The bars represent the mean ± SEM. D. 2D-immunoblot analysis (BN-PAGE/SDS-PAGE) of pseudouridine synthase interacting proteins. The migration (sizes in kDa) of known protein complexes in the first dimension is indicated on the top of the blot.
Figure 5 Figure 5. cDC1 are required for the development of parasite-specific IFN+ TNF+ Th1 responses during PbA blood stage malaria (A) Experimental protocol to analyze T cell responses in Karma mice at day 6 post-infection with PbA pRBC. Mice were injected with DT prior to infection and repeatedly every 60 h thereafter. (B) Gating strategy to assess efficiency of cDC1 depletion. (C) Percentage (mean +/- SEM) of CD8α+ out of Lin- B220- CD11c+ MHC IIhi cells. *, P = 0.0043 by Mann-Whitney test. (D) FACS plots showing the proportion of CD11a+ CD49d+ CD4 T cells that produce IFN and TNF in response to MutuDC loaded with RBC, PbA pRBC or ETRAMP10.2 peptide. (E) Proportion of CD11a+ CD49d+ CD4 T cells that co-produce IFN and TNF (mean +/- SEM) in response to MutuDC loaded with the indicated antigen, isolated from Karma mice treated (open circles) or not (black circles) with DT. Asterisks show significant differences between no DT and + DT group by multiple unpaired t-tests without assuming consistent SD. RBC, P = 0.18 ; GAPDH.1, P = 0.019 ; EF1a, 0.039 ; ETRAMP, P = 0.00054 ; OVA, P = 0.25 ; pRBC, P = 0.015. (F) Absolute numbers of IFN+ TNF+ CD11a+ CD49d+CD4+ T cells responding to the indicated stimuli (mean +/- SEM). GAPDH.1, P = 0.0047 ; EF1a, 0.00039 ; ETRAMP, P = 0.00022 ; pRBC, P = 0.00019 by multiple unpaired t-tests without assuming consistent SD. (G) Blood parasitemia (mean +/- SEM) at day 4 and 6 pi in Karma mice treated (open circles) or not (black circles) with DT. Day 4, P = 0.022 ; Day 6, P = 0.77 by multiple unpaired t-tests without assuming consistent SD. C,E,F,G: No DT group, N = 5 mice ; + DT group, N = 6 mice. Representative of 3 independent experiments. (H) Absolute numbers of CD4+ T cells producing IFN, TNF or IL-10, either individually or in combination, following restimulation with MutuDC+PbA pRBC (mean +/- SEM). These cells are called cytokine+ in the next panels. *, P = 0.0023 by unpaired t-test without assuming consistent SD. (I) Multifunctionality analysis (IFN, TNF, IL-10) of cytokine+ T cells responding to MutuDC loaded with PbA pRBC. (J) Proportion of IL-10+ IFN- TNF- among cytokine+ T cells, in response to MutuDC loaded with the indicated antigen (mean +/- SEM). GAPDH.1, P = 0.32 ; EF1a, 0.16 ; ETRAMP, P = 8.5x10-5 ; OVA, P = 0.11 ; pRBC, P = 0.0015 by multiple unpaired t-tests without assuming consistent SD. H,I,J: N = 3 mice per group.
Figure 1A Summary of disease history in 77 SCA8 families in our cohort. B No correlation between length of combined repeats and age of onset in SCA8 patients, n=85, p=0.9847 or in the subset of SCA8 patients with CCG•CGG interruptions (int.), n=26, p=0.2096, linear regression analyses. Open circles indicate SCA8 patients with pure CAG repeat expansions. Red squares show average expansion size for individuals with two expanded alleles, individual allele repeat lengths: 137/177,110/320, 104/130, 96/109. Red triangles show average expansion size for individuals with two expanded alleles and CCG•CGG interruptions: 84/114, 92/100. Grey triangles indicate individuals with CCG•CGG interruptions. C Allele length distribution of affected (n=111) and asymptomatic (n=88) expansion carriers; box: 25th to 75th percentile; whiskers: minimum to maximum value; p=0.0672, Mann-Whitney test. D-H SCA8 family pedigrees: squares represent males, circles represent females, diamonds mask gender. Filled symbols represent affected individuals, symbols with dot represent asymptomatic expansion carriers, open symbols represent individuals with non-expanded alleles, diagonal line indicates a deceased individual. Combined repeat number, age (y - years) at onset (Onset) or age still asymptomatic, and interruption status (Pure or Int. [CCG•CGG interrupted]) are noted below the symbols. (G) Abbreviated pedigree I SCA8 allele configurations in the CAG direction as determined by sequencing. Family or individual and affected status indicated on left: Sporadic 1 - figure 1D indicated by #, Family 1 - figure 1G, Family 2 - figure 1H; A - affected, AS - Asymptomatic; CGG interruptions represented by black boxes. Sequences used in figure 5C are indicated by *. See Appendix Table S1 for allele configurations. J Age of onset correlates with the number of CCG•CGG interruptions, n=24, p=0.0016, linear regression analyses. Grey triangles indicate individuals with CCG•CGG interruptions. Red triangles indicate the average expansion size for individuals with two expanded alleles and CCG•CGG interruptions, individual allele repeat lengths are: 84/114, 92/100. Data information: (I, J) Individuals identified as having CCG•CGG interruptions by restriction digest are not included.
Figure 6(A) Schematic representation of wild type and mutant COA7. (B) Cellular protein extracts were isolated from HEK293 cells that were transfected with a plasmid that encoded wildtype or mutant COA7. The samples were analyzed by reducing SDS-PAGE and Western blot. (C) Cellular fractions were prepared from HEK293 cells that were transfected with a plasmid that encoded wildtype or mutant COA7. The fractions were analyzed by reducing SDS-PAGE and Western blot. T, total; C, cytosol; M, mitochondria. (D) HEK293 cells that transiently expressed wildtype or mutant COA7HIS were solubilized, and the affinity purification of COA7HIS was performed. The samples were analyzed by reducing and non-reducing SDS-PAGE and Western blot. Load: 3 %. Eluate: 100 %. (E) Equal amounts of wildtype and mutant radiolabeled [35S]COA7 precursors were imported into mitochondria isolated from HEK293 cells. The samples were analyzed by reducing SDS-PAGE and autoradiography. The results of three biological replicates were analyzed, quantified, and normalized to wildtype COA7 at 45 min. The data are expressed as a mean ± SEM (n=3). IAA, iodoacetamide.
Figure 7Boxplots (plotted as in Fig 3B) of gene expression of ELP3, ALKBH8 and CTU2 upon treatment with 17β-E2 and/or ICI-182780 in MCF7 cells (extracted from (Wardell et al, 2012; Data Ref: McDonnell et al, 2012)). Wilcoxon-Mann-Whitney tests (two-sided) were used to assess differences between conditions (n = 10) and a global Bonferroni adjustment was applied on p-values. Immunoblotting of MCF7 cell extracts. MCF-7 cells were grown in phenol red-free RPMI media supplemented with 5% charcoal-stripped serum for 24 hours and then treated with 1 nM E2 and/or 100 nM ICI-182780 for 24 hours. Graphs represent mean ± SD of n=5 experiments. Data were analyzed using two-sided paired Student's t tests. ChIP-seq reveals ER binding sites proximal to the ELP3 gene. Each track represents a distinct ChIP-seq dataset. Immunoblotting of MCF-7 cell extract for indicated proteins following deletion of ELP3 gene by CRISPR-Cas9 technology. Quantification of mcm5S2-U, mcm5U and cm5U levels in ELP3-null MCF-7 cells and in MCF-7 cells treated with 100 nM ICI-182780 for 72 hours versus their respective controls by liquid chromatography-mass spectrometry (LC-MS) analysis. The MCF-7 cells were grown in phenol red-free RPMI media supplemented with 5% charcoal-stripped serum for 24 hours prior to ICI-182780 treatment. Graphs represent mean ± SD of n=4. Data were analyzed using two-sided paired Student's t tests. Vh, Vehicle; ICI, ICI-182780.