EMBO/sd-character-level-ner huggingface dataset:
Figure 1Treatment of HCT116 cells with siRNA targeting Ku70 (A-I) or control followed by western blot analysis (A) The basal levels of p53 normalized to GAPDH were arbitrarily set at 1.0 and the fold change is shown in (ii) (n=4). Treatment of HCT116 cells with siRNA targeting Ku70 or control (A-I) combined or not with a treatment with doxorubicin (Doxo) (positive control for phosphorylation of ATM and p53) (B) followed by western blot analysis. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), ASF (C) (positive control for y-H2AX activation [29]) or control (A-I) followed by western blot analysis. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), or control (A-I) combined or not with a treatment with wortmannin (WTN) (D) followed by western blot analysis. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), or control (A-I) combined or not with a treatment with NU-7441 (E,F) followed by western blot analysis. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), XRCC4 (F) or control (A-I) combined or not with a treatment with NU-7441 (E,F), followed by western blot analysis. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), or control (A-I) followed by RT-qPCR analysis (G). In RT-qPCR analysis (G), mRNA levels were standardized against HPRT mRNA (n=3). The two panels of Fig. 1C are partion of the same gel. Statistical analysis by unpaired t-test (*P<0.05; **P<0.01). All error bars reflect SEM. Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), or control (A-I) combined or not with a treatment with MG132 (H) followed by western blot analysis (A-F, H-I). Treatment of HCT116 cells with siRNA targeting Ku70 (A-I), or control (A-I) combined or not with a treatment with cycloheximide (CHX) (I) followed by western blot analysis (A-F, H-I).
Figure 5In vivo effect of EV20 (10 mg/kg, n=4), untreated control (PBS, n=4) and EV20/MMAF (3.3 mg/kg, n=5; 10 mg/kg, n=4) on tumor growth in mice injected with BTRH cells. Mice were treated when tumors reached a volume of 500 mm3. Data are represented as mean + SEM. In vivo effect of single-dose EV20 (10 mg/kg, n=3) untreated control (PBS, n=4), single-dose EV20/MMAF (10 mg/kg, n=4) or weekly trastuzumab (30 mg/kg, n=4) on tumor growth in mice injected with BTRH#10 cells. Data are represented as mean + SEM. In vivo effect of single-dose EV20 (10 mg/kg, n=4), untreated control (PBS, n=4), single-dose EV20/MMAF (10 mg/kg, n=4) or weekly trastuzumab (30 mg/kg, n=4) on tumor growth in mice injected with HCC1954 cells. Data are represented as mean + SEM. Analysis of pHistone H3, HER3, pHER3 and cleaved caspase-3 in tumors of mice injected with BTRH and treated (T) or not (C) with EV20/MMAF (3.3 mg/kg for 24 hours or two weeks). Numbers below the blots show the quantitation of the signal of each band, referred to that of a control untreated tumor. Long-term monitoring of mice injected with BTRH cells and treated with EV20/MMAF (3.3 mg/kg, n=5; 10 mg/kg, n=4). Where indicated, mice were treated with EV20/MMAF. Data were represented as mean + SD of the different mice groups.
Figure 3(A) gag mRNA expression (left) and relative viability (right) of HIV-1 infected Jurkat-TAg cells transfected with non targeting siRNA (NC) or siRNAs targeting the HMOX-1 gene. Cells were infected 24hr post-transfection and assayed for mRNA expression (by qPCR) or viability (by MTT assay) 48hr post-infection. Data (mean±SEM; n=3 technical replicates) were normalized using the NC control and analyzed by unpaired t-test. P<0.05; **P<0.01. (B RNA-Seq analysis of different time points of primary CD4+ T-cells infected in vitro with HIV-1 or mock infected. The gene set used for the analyses is GO iron ion import (12 genes, GO:0097286). Data in (B) were analyzed by Fisher test. Boxplots depict median and 25-75 percentiles while whiskers extend from the hinge to the highest or lowest value that is within 1.5 * IQR (inter-quartile range) of the hinge. Data beyond the end of the whiskers are outliers and plotted as points. Dots below boxplots illustrate, for each time point, the pathway enrichment analysis of genes up-regulated in infected vs matched mock-infected controls. Dots are color-coded based on the enrichment q-values and their size indicates the fraction of DEGs in the pathway; grey dots are not statistically significant. (C) Heatmaps of the standardized expression of the GO iron ion import pathway genes in HIV-1 infected and mock infected samples over time. Expression levels were standardized [(mean gene expression - SD)/SD] for each gene in each time point. The TFRC (TfR1) gene, which was considered for further analysis, is highlighted. (D) TfR1 expression in HIV-1 infected or mock infected CD4+ T-cells as visualized by confocal microscopy. Fluorescent values shown were calculated as described in (McCloy et al, 2014). Data were analyzed by one-way ANOVA followed by Sidak´s post-test to compare corresponding mock infected vs infected pairs. Black dash indicates median, n= number of cells. Scale bar = 2μm. (E) Western blot (upper panel) and relative expression (lower panel) of FTH-1 protein in HIV-1 infected vs mock infected CD4+ T-cells over time. Western blot quantification was performed with Fiji-Image J (Schindelin et al, 2012), normalized to the housekeeping protein beta-actin and expressed as Log2 fold change in HIV-1 infected vs mock infected cells (mean±SEM; n=3 biological replicates). P<0.05; (F) Percentage of intracellular p24+ gag CD4+ T-cells infected with HIV-1 and left untreated or treated for 48hr with the iron donor Fe-NTA at 150μM concentration (n=3; mean ± range of three biological replicates). (G) Percentage of GFP+ J-Lat cells (clones 9.2 and 15.4) left untreated or treated for 48hr with TPA (10μM), the iron donor FeNT (150μM) or a combination of the two. Data are expressed as mean±SEM of four technical replicates for each cell line and were analyzed by one-way ANOVA followed by Tukey´s post-test. *P<0.05; ***P<0.001; ****P<0.0001.
a) IB3-1 cells cultured with 250 μM cystamine or transfected with either 50 nM human TG2 siRNA or scrambled oligonucleotides.figf3(a) IB3-1 cells cultured with 250 μM cystamine or transfected with either 50 nM human TG2 siRNA or scrambled oligonucleotides. IP, immunoprecipitation; IB, immunoblot. (b) IB3-1 cells cultured with the proteasome inhibitor MG132 (50 μM). (c, d) IB3-1 cells transfected with either human TG2 siRNA or scrambled oligonucleotides followed by incubation with MG132. (c) Cells immunostained with anti-beclin 1 (green) and anti-HDAC-6 (red) antibodies. Yellow indicates co-localization. N, nucleus. Scale bar, 10 μm. (c, d) IB3-1 cells transfected with either humanTG2 siRNA or scrambled oligonucleotides followed by incubation with MG132. (d) FRET analysis of beclin 1-Alexa 546 (Al546) fluorescence after HDAC6-Cy5 photobleaching. F/F0 indicates the post-bleaching/pre-bleaching fluorescence ratio of Al546/Cy5. (e) IB3-1 cells were transfected with either human TG2 siRNA or scrambled oligonucleotides. Immunoblot of TG2, Ambra-1, hVps15, hVps34, Atg14L and beclin 1 in IB3-1 cells grown in starvation. Uncropped images of blots are shown in Supplementary Information, Fig. S10. (f) IB3-1 and C38 cells were transfected with either 1 μg of pLPCX-WT-TG or pLPCX-Cys277Ser-TG or the empty vector. Immunoblot of beclin 1. Uncropped images of blots are shown in Supplementary Information, Fig. S10. (g, h) IB3-1 cells were treated with cystamine or 10 mM EUK-134 or transfected with either 50 nM humanPIASy siRNA or scrambled oligonucleotides. (g) Percentage of cells containing more than five GFP-LC3 punctate dots. Means ± s.d. for three independent experiments. Asterisk, P 0.001 versus control; ANOVA. (g, h) IB3-1 cells were treated with cystamine or 10 mM EUK-134 or transfected with either 50 nM humanPIASy siRNA or scrambled oligonucleotides. (h) Left: electron micrographs of IB3-1 cells showing areas enriched in autophagosomes (arrows) on PIASy gene silencing, EUK-134 or cystamine. Scale bar, 100 nm. Right: number of autophagosomes per cell counted in 20 cells per experiment. Means ± s.d. for three independent experiments. Asterisk, P 0.001 versus control; ANOVA. (i) IB3-1 cells treated with cystamine. Immunoblot of p62. (j) IB3-1 cells treated with cystamine or EUK-134 or transfected with human PIASy siRNA or scrambled oligonucleotides. Immunoblot of beclin 1 (left) and PIASy (right). Uncropped images of blots are shown in Supplementary Information, Fig. S10. The experiments were repeated three times. β-actin was used as loading control for all blots. Densitometric analysis of blots and quantitative measurement of co-localizations are shown in Supplementary Information, Fig. S9.
Figure 6(A and B) Quantitative analysis of immunofluorescence microscopy images for colocalization of M.tb with Lamp-1 in WT (A) and Mavs -/- (B) BMMs infected with M.tb for 24 hr and then treated for an additional 24 hr with moxifloxacin (MFX) and/or EVs from M.tb-infected BMMs (EVs+MFX) in the presence of recombinant mouse IFN-ɣ. Mock, no EVs or moxifloxacin treatment; EVs_Control, EVs from uninfected BMMs. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). LC3 (C), colocalization with M.tb were analyzed. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). NOX2 (D) colocalization with M.tb were analyzed. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). p47phox (E) colocalization with M.tb were analyzed. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). Mavs -/- BMMs were used and colocalization of M.tb with LC3 (F) were quantified. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). Mavs -/- BMMs were used and colocalization of M.tb with NOX2 (G) were quantified. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). Mavs -/- BMMs were used and colocalization of M.tb with p47phox (H) were quantified. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed). (I and J) M.tb CFU analysis in WT (I) and Mavs -/- (J) BMMs infected with M.tb for 24 hr followed by treatment with EVs, moxifloxacin or combination in the presence of IFN-ɣ for 24 and 72 hr. Data shown are representative of three independent infections. The results are the mean ± SD of three independent experiments. n.s., not significant; * p < 0.05, ** p < 0.01 and *** p < 0.001 by Student's t-test (two tailed).
Figure 5A. Unsupervised hierarchical clustering of proteins significantly differentially abundant across cell types, with columns showing three biological replicates (n=3) of four cell types and rows significant proteins. Significance was calculated by one-way ANOVA followed by Benjamini-Hochberg correction for multiple hypothesis testing (FDR < 0.05). Frames and numbers indicate four clusters of proteins highly enriched in each cell type. B. GOBP map specific for hHEP (proteins in cluster 1). Each circle represents an individual biological process term significantly enriched, with the number of proteins and FDR-corrected p-value indicated by size and degree of transparency. Significance was calculated by one-way ANOVA followed by Benjamini-Hochberg correction for multiple hypothesis testing (FDR < 0.01). The leading term given by the most proteins associated within a group is indicated. C. GOBP map specific for hKC (proteins in cluster 2). Significance was same as Panel (B). D. Comparative analysis of GOBP enrichment for hHSC and hLSEC from Cluster 3 and Cluster 4 from Panel (A) A, with blue terms specific for hHSC and pink specific for hLSEC and gray terms shared between them. Significance was same as Panel (B).
Altered PI(4,5)P2 levels modulate metastatic migration and invasive capacity.Figure 4(A,B) LM2 cells transfected with siRNA targeting PTPRN2 (A), PLCβ1 (B) or a control siRNA were transfected with Lyn11-FRB and INPP5E-FKBP, treated with either DMSO or 100 nm rapamycin and subjected to the migration assay. N = 5 inserts/group. (C,D) MDA-MB-231 cells overexpressing PTPRN2 (C), PLCβ1 (D) or control vector were treated with carrier alone or carrier incubated with PI(4,5) P2 for one hour and then immediately subjected to the migration assay. N = 5 inserts/group. (C,D) MDA-MB-231 cells overexpressing PTPRN2 (C), PLCβ1 (D) or control vector were treated with carrier alone or carrier incubated with PI(4,5) P2 for one hour and then immediately subjected to the migration assay. N = 5 inserts/group. (E) Kaplan-Meier curve representing distal metastasis-free survival of a cohort of breast cancer patients (N = 1609) as a function of their primary tumor's mean PIP5K1A, PIP5KB, and PIP5KC expression levels (Data from KMPlot (Gyorffy et al., 2010)). Patients' primary tumors' combined PIP5K expression levels were classified as low (blue) or high (red) expression. (F,G) Migration (F) and Matrigel invasion (G) of LM2 cells transduced with a retroviral vector overexpressing PIP5K1A or control vector. Data normalized to control values. N = 5 inserts/group.
Figure 5(A). Heatmap showing log2-transformed fold changes of Reno1 depleted cells compared to their respective controls. All genes which were significantly differentially expressed (P<0.05) in any of the treatments are shown. (B) Boxplots indicating the median, quartiles, and 5th and 95th percentiles of changes in expression levels of Reno1m/m cells compared to WT cells, for genes with significant (P<0.05) change in cells where Reno1 was perturbed with the indicated method. Number of genes in each group is indicated in the x axis. ** P<0.01 (two-sided Wilcoxon rank sum test). (C) Mean expression levels of genes in genes which were significantly (P<0.05) dysregulated by at least two out of three Reno1 perturbations at day 2 in fig. 5A (n= 168 down-regulated and 94 up-regulated), during an eight time point neuronal differentiation and maturation of mouse ES cells: Neurons1 corresponds to day in vitro (DIV) 1; Neurons2 - DIV7; Neurons3 - DIV16; Neurons4 - DIV21; and Neurons5 - DIV28 (Hubbard et al., 2013). (D) Expression levels of Reno1 following Dox addition in WT and Reno1m/m cells. Mean ± SEM is shown for three independent experiments, * P<0.05 (unpaired two sample t-test). (E) Cell count of WT and Reno1m/m cells following four days of differentiation, with or without Dox addition. Cells were harvested from one well of a 6-well plate, and counted using Orflo MOXI Z Mini Automated Cell Counter. Mean ± SEM is shown for three independent experiments. * P<0.05 (unpaired two sample t-test). (F-G) Boxplots indicating the median, quartiles, and 5th and 95th percentiles of changes in expression levels of WT (F) or Reno1m/m (G) cell treated with Dox for the first two days of differentiation compared to untreated cells, in genes which were significantly (P<0.05) downregulated (n=252) or upregulated (n=223) by at least two out of three Reno1 perturbations at day 2 in fig. 5A, compared to all genes (n=16,511). ** P<0.01 (two-sided Wilcoxon rank sum test).
Figure 1Retrotransposon transcription in WT (Hu303), fft2Δ (Hu1955), fft3Δ (Hu1867), and fft2Δfft3Δ (Hu2000). A) Transcription in fft2Δ relative to WT over chromosome 1. The top and bottom panels represent fold change in tiling array signal from the forward and reverse strands, respectively. Coordinates are shown in the middle panel; genes more than 2-fold upregulated are marked: Tf2 retrotransposons (black); antisense to Tf2 elements (gray); coding genes (green); noncoding RNAs (blue) Retrotransposon transcription in WT (Hu303), fft2Δ (Hu1955), fft3Δ (Hu1867), and fft2Δfft3Δ (Hu2000). B) Northern blot for the Tf2 ORF; rRNA staining of the same membrane by methylene blue is shown below. Two different exposures of the molecular size marker are shown (left). Retrotransposon transcription in WT (Hu303), fft2Δ(Hu1955), fft3Δ (Hu1867), and fft2Δfft3Δ (Hu2000). C) qPCR scheme. Amplicon 1 spans the end of the LTR and into the ORF. Amplicon 2 does not include the LTR but covers the translation start site. Amplicon 3 is entirely within the ORF. D) Bar graph representing RNA levels for the amplicons depicted in (C), relative to a control locus (SPAC1F8.07c) and to WT levels, as measured by qPCR. Error bars represent standard deviation of duplicate reverse transcriptions of biological triplicates. Retrotransposon transcription in WT (Hu303), fft2Δ (Hu1955), fft3Δ (Hu1867), and fft2Δfft3Δ (Hu2000). E) 5'RACE products run on a 1.5% agarose gel. Retrotransposon transcription in WT (Hu303), fft2Δ (Hu1955), fft3Δ (Hu1867), and fft2Δfft3Δ (Hu2000). F) The sequence of a Tf2 retrotransposon transcript from the 'R' sequence of the upstream LTR to the start codon of the coding region. The long and short RACE products (E) were sequenced and the transcript start sites are highlighted (yellow). Orange text, R. Light blue, U5. Dark blue, self-primer and primer binding site. Green, start codon.
A proposed model for UBTD1 controlling cancer cell aggressiveness through YAP signaling and ubiquitylation At high cell density, UBTD1 is highly expressed and is associated with β-catenin at the cell-cell adhesion complex.Figure 7At high cell density, UBTD1 is highly expressed and is associated with β-catenin at the cell-cell adhesion complex. Cytoplasmic UBTD1 is associated with the YAP degradation complex composed of UbcH5, β-TrCP and β-catenin and promotes YAP ubiquitylation for proteasomal degradation (Left). At low cell density, or during cancer progression, UBTD1 is weakly expressed. Low UBTD1 expression induces RhoA activation, increases cell traction forces and activates YAP signaling via ROCK2 fostering cancer cell aggressiveness (Right).
Fig.Figure 7A. Q25-GFP and Q97-GFP were expressed in wild type cells for 4.5 h. Cell extracts were subjected to mass spectrometry-based proteomics. The levels of Mia40 and other mitochondrial proteins (mito., yellow) and carrier proteins (orange) are visualized in a volcano plot (p-values were derived using a moderated t-test). The position of Rnq1 is indicated in blue. B. The open reading frames of the indicated proteins were cloned downstream of a GAL promoter in plasmids and transformed into cells harboring the Q97-GFP expression plasmid. Cells harboring an empty vector (ev) were used for control. Cells were shifted to galactose medium overnight before they were dropped onto glucose or galactose plates. If cells are directly dropped from lactate medium onto galactose plates, only individual cells (indicated by red protein names) escaped the polyQ toxicity (see Fig. EV4C). C. Wild type and GAL-Mia40 cells were transformed with plasmids that express Yellow Fluorescent Protein (YFP) under the control of a minimal promoter containing the Hsf1-driven heat shock element (HSE). Cells were grown at 30 or 37°C before fluorescence was measured. Mean values and standard deviations of three replicates are shown. D. Cells expressing the indicated carrier proteins from GAL promoters were dropped onto galactose plates. Cell growth was quantified by densitometry from one representative plate. OM refers to carrier proteins that were expressed as fusions with an N-terminal outer membrane anchor. See Fig. EV4D, E for scans of the respective plates. E. The formation of protein aggregates was detected with a PROTEOSTAT protein aggregation assay. The expression of the most abundant mitochondrial carrier protein, Pet9, resulted in a strongly increased fluorescence in wild type cells but not in GAL-Mia40 cells. Mean values and standard deviations of three replicates are shown. F. Using the SPELL data collection of transcription data sets, we calculated the co-expression of yeast genes with genes relevant for proteostasis (chaperones, proteasome, and others; see Materials and Methods for details). Genes for mitochondrial import components are highlighted.
a) Nox2-specific ROS production was assessed using the Nox2 redox biosensor p47-roGFP redox biosensor Cat: catalase, PEG-Cat: pegylated catalase.f1(a) Nox2-specific ROS production was assessed using the Nox2 redox biosensor p47-roGFP redox biosensor Cat: catalase, PEG-Cat: pegylated catalase. ( b) Measurement of intracellular glutathione redox potential with Grx1-roGFP2. (c) Analysis of Rac1 and (d) Src. . (e) Immunoblot of precipitated p47phox probed with an anti-phosphoserine or anti-p47phox antibody. (f) Nox2-specific intracellular ROS production was measured using p47-roGFP redox biosensor. g) Extracellular ROS production was assessed using Amplex-red dye. (h) Plasma membrane calcium influx was measured by analysing the Fura-2 fluorescence quench rate upon addition of extracellular Mn2+. (i) Intracellular RNS generation was measured using DAF-FM. Bars represent average±s.e.m. from n=15 individual fibres for each condition in (a,b,f,g,i ,j). Markers of autophagy were analysed in isolated fibres (incubated with or without PP2) from FDBs. (k) Autophagosome formation was analysed using fluorescence microscopy (scale bar=100 μm) and illustrated LC3 localization and autophagosome formation. (l) Confocal microscopy detected p62-LC3 localization in single fibres from FDBs (scale bar=140 and 50 μm for white box areas). All immunoblots were performed with isolated proteins from FDBs and probed with antibodies as indicated. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was detected as a loading control. Representative images are shown. Bars represent average ±s.e.m. from n=3 independent biological experiments. Statistical differences between groups were determined using analysis of variance (ANOVA) with Tukey's post hoc test. *P0.05 and **P0.01.
Figure 1(A) LC-MS/MS extracted ion chromatograms of 28 modified ribonucleosides analyzed in parasite tRNA. Signal intensities of the modifications are scaled to that for m1A, the ribonucleoside with the highest MS signal (B) Comparison of P. falciparum tRNA modifications (red) to those in all three kingdoms of life (Grosjean & Benne, 1998, Jackman & Alfonzo, 2013). Modifications in the box are those found in eukaryotic cellular organelles and the assignment of ms2t6A to both eukarya and bacteria is supported by published work (Arragain, Handelman et al., 2010). The figure has been adapted from Grosjean and Benne, 1998; and Jackman and Alfonzo, 2013 (C) Comparison of tRNA modification profiles among P. falciparum, S. cerevisiae and H. sapiens. tRNA wobble modifications are highlighted in red, with wobble-specific modifications bolded and underlined. The assignments of tRNA modifications to H. sapiens and S. cerevisiae are based on published work (Cantara et al., 2011, Chan et al., 2010, Jühling, Mörl et al., 2009, Kellner, Neumann et al., 2014, Machnicka et al., 2013, Phizicky & Hopper, 2010, Su et al., 2014)