EMBO/sd-character-level-ner huggingface dataset:
a) NOX2 activity in Slamf1+/+ and Slamf1−/− BALB/c peritoneal macrophages stimulated for 0–100 min with E.figf2(a) NOX2 activity in Slamf1+/+ and Slamf1-/- BALB/c peritoneal macrophages stimulated for 0-100 min with E. coli F18, S. aureus or PMA, assessed with lucigenin. Data are representative of five independent experiments (mean ± s.d.). (b) Phagosomal pH of Slamf1+/+, Slamf1-/- and gp91phox-/- B6 primary macrophages loaded for 0-200 min with pHrodo-coated E. coli or S. aureus, analyzed by flow cytometry. Data are representative of three independent experiments (mean ± s.d.). (c) NOX2 activity in primary macrophages in response to LPS, purified OmpC, peptidoglycan (PGN) or PMA, assessed with lucigenin. TLR4-KO, TLR4-deficient (strain del/Jtht; C3H). Data are representative of five independent experiments (mean ± s.d.).
Vasculotide delays the dissemination of spontaneous lung metastases from orthotopic breast cancer (LM2-4luc) xenografts without having effect on primary tumors.Figure 5A-C Vasculotide (VT) did not alter the growth kinetics of orthotopic LM2-4luc tumors; n = 8 mice per group. Sunitinib (SU) treatment significantly lowered primary tumor burden as measured by volume (B: one-way ANOVA, P < 0.01) or bioluminescent activity (C: one-way ANOVA, P < 0.001). D VT treatment did not alter the mean thickness of the viable peripheral regions of primary tumors, as quantified by H&amp;amp;E histological staining. E VT treatment did not alter the mean thickness of the CAIX+ hypoxic edge between the viable periphery and necrotic core of a primary tumor; IHC control and sample images taken at 5× are shown. F Ex vivo bioluminescent imaging suggests that VT treatment lowered the proportion of mice with metastatic involvement in the lungs, without the corresponding effect on lymphatic metastases. Representative bioluminescent vs. photographic images of dissected lungs in a 24-well plate are shown. G Human LM2-4luc cells can be identified on histological sections of mouse lung tissue by IHC staining of human vimentin, a cytoskeletal protein. Size distribution analysis show VT monotherapy reducing the occurence of bigger metastatic nodules.
Figure 7A) Lineages of animals of the indicated genotypes were grown at 25°C or 20°C for the indicated generations. Each generation six animals were isolated in order to maintain the lineage and to determine numbers of progeny. Data are represented as +/- s.e.m. N=3 for generation 8 or 9, N=6 for all other generations. B) Volcano plot showing log2 fold change of the number of endogenous siRNAs targeting each C. elegans gene in zsp-1(gg682) vs WT animals on x-axis and log10 (adjusted p-value) of changes on the y-axis. Dots in red indicate siRNAs pools targeting this gene were significantly different in zsp-1(gg682) and WT animals. Dashed lines indicate log2(fold change)=-1 or 1. Adjusted P-value was calculated using Benjamini Hochberg method in DEseq2. C) Representation factor indicates the number of genes overlapping divided by the expected number of genes overlapping in two datasets, with a value of one (dashed line) indicating random overlap. A representation factor >1 indicates more overlap between datasets than expected by chance. Data show a comparison of lists of genes differentially targeted by endo siRNAs in zsp-1(gg682) animals relative to WT animals against lists of genes known to be targeted by CSR-1 (Claycomb et al. 2009), HRDE-1 (Buckley et al. 2012), WAGO-4 (Xu et al. 2018), or all WAGO-class endo siRNAs (Gu et al. 2009). D) Venn diagram of the overlap between genes that are both regulated both ZSP-1 and targeted by WAGO-4 endo siRNAs and lists of genes targeted by either WAGO (Gu et al. 2009) or CSR-1-class endo siRNAs (Claycomb et al. 2009). E) Fluorescent micrographs showing a piRNA sensor transgene (mex-5p::gfp::his-58::21UR-1::tbb-2 3'UTR) (Bagijn et al. 2012) expressing GFP in the -1 to -3 oocytes of adult C. elegans of the indicated genotypes. Scale bar, 20μm. F) oma-1(zu405) is a temperature sensitive (ts) lethal (embryonic arrest at temperatures > 20°C) allele of oma-1 (R. Lin 2003). oma-1 RNAi suppresses oma-1(zu405ts) lethality. Animals were exposed to oma-1 RNAi and the number of embryos that hatched was scored at 20°C. N=3. Error bars are +/- s.d. Animals harboring mutations in genes specifically required for the inheritance of RNAi would respond normally to direct exposure to oma-1 dsRNA but fail to inherit this silencing (Wan et al. 2018). zsp-1 mutants fail to respond immediately to direct exposure to oma-1 dsRNA, indicating that ZSP-1 is not specific to inheritance and, therefore, is generally required for RNAi (see also Fig. EV5B-D). G) RNAs of 16-30nts in length were sequenced from +/- oma-1 RNAi treated animals of the indicated genotypes. One replicate of the experiment is shown here and another replicate, which shows similar results, is shown in Fig. EV5E. Y axis shows # small RNA reads per million sequenced reads. X axis is the oma-1 gene.
Characterization of proteins that aggregate in heat shock.Figure 2A Definition of aggregating proteins. Solubility is measured as the log2-transformed ratio of protein intensities in the soluble fraction between heat shocked and mock shocked samples. Proteins with significant reduction in solubility [Benjamini-Hochberg adjusted p-value < 0.05 and solubility < log2(2/3)] are then considered as aggregators. Dotted horizontal line shows the cut-off at solubility of log2(2/3). Comparisons of physicochemical properties between soluble proteins and aggregators. Hydrophobicity (gravy scores) (B) and isoelectric points (C) are shown as combined violin- and boxplots (p-values are for non-parametric Wilcoxon test). Difference in median amino acid composition between soluble proteins and aggregators is compared to hydrophobicity (gravy score) for each amino acid (D). In D, Pearson coefficient (r) with p-value is shown for the correlation analysis. Comparison of structural features between soluble proteins and aggregators. Fraction of protein sequence predicted to contain intrinsically disordered regions (E), molecular weight (F) P-values in E-G are for non-parametric Wilcoxon test. Comparison of structural features between soluble proteins and aggregators. fraction of protein sequence predicted to contain secondary structure elements (alpha helix, beta sheet or coil) (G) are compared. P-values in E-G are for non-parametric Wilcoxon test. H Protein complex members in soluble proteins and aggregators. Pie charts show the fraction of proteins annotated to be a member of a protein complex. The number of proteins in each segment is indicated. P-value is for Fisher's exact test. I Protein complexes involving aggregators. Heatmaps show the absolute difference in solubility change after heat shock between each complex member. Protein complexes with at least 75% of its members quantified and containing at least 60% of its members as aggregators are shown. 'DNA repair complex' = 'DNA repair complex NEIL2-PNK-Pol(beta)-LigIII(alpha)-XRCC1'.
Figure 7Distribution of peptides containing sulfhydrated cysteine residues relative to their H/L ratios as determined by the BTA analysis of cell extracts isolated from C1 and P1 patient cells (indicated in A) and C2 and P4 fibroblasts (B). Values of H/L ratios are plotted against the number of identified peptides. The gray line marks the H/L ratio >2, consisting of cysteine-containing peptides in proteins from patient cell lines that exhibited higher reactivity with H2S compared with the respective control. Indicated on the X axes are representative sensitive cysteine peptides, which are common between the ones found in the RedoxDB database, and by the BTA assay.
Figure 5(A-B) Supernatants from the panel of infected (A) Vero E6-TMPRSS2 and (B) Vero E6 cells were harvested when CPE was observed and the viral titer (FFU/mL measured by focus forming assay) was determined for the final time point of each sample that successfully isolated. Samples are colored according to the day CPE was observed and the sample was harvested. Total E RNA CT of the original clinical sample used for viral isolation is plotted on the X axis. Data shown is from the subset of samples that successfully isolated in each cell line. Limit of detection for infectious titer is 400 FFU/mL.
Figure 7Western blot showing total TDP-43 levels, quantification in B, ratio paired-t-test. Total cell lysates were fractionated into soluble and insoluble fractions and analysed by Western blot, representative Western blots of soluble and insoluble fraction without C and with D HDAC6 inhibitor treatment. Quantification of soluble full-length TDP-43 levels, ratio paired-t-test and E quantification of insoluble full-length TDP-43 levels, ratio paired-t-test. Western blots of insoluble fraction with (right) and without (left) HDAC6 inhibitor treatment. Quantification of insoluble c-terminal fragment of 35kDa (CTF-35; H) and insoluble c-terminal fragment of 25kDa (CTF-25; I), ratio paired-t-test.
Figure 1Assessment of mtDNA damage by quantitative PCR. HEK293 cells infected with control (empty vector) or shATG5 for 5 days. Cells were then incubated with DMSO or 4 mM 3-NPA for 2 h and either immediately harvested or washed with fresh medium and incubated for another 1 h. Cells with or without washout were used for the extraction of total DNA. All DNA samples were used for amplification of 8.9 kb mtDNA fragment using quantitative PCR and were normalized to amplification of a 221 bp mtDNA fragment. PCR products were quantitated by PicoGreen staining using Micro Plate Reader. Data are presented as mean ± SD (n = 3 independent experiments), statistical significance was assessed by two-tailed student's t-test, N.S., not significant, *p < 0.05, **p < 0.01. The data in (A) were further calculated for the frequency of mtDNA damage. The equation was seen in "Materials and Methods". Data are presented as mean ± SD (n = 3 independent experiments), statistical significance was assessed by a two-tailed student's t-test, N.S., not significant, **p < 0.01. Representative images show 8-oxo-dG staining. Control or shATG5 HeLa cells were treated with DMSO, 200 µM H2O2, or 4 mM 3-NPA for 2 h, and then either fixed immediately or washed with fresh medium and incubated for another 1 h. Cells were immunostained with DAPI and anti-8-oxo-dG antibody and analyzed by confocal microscopy. Scale bar, 10 µm. Quantification of the relative 8-oxo-dG fluorescence intensity in (C). Data are presented as mean ± SD (n = 3 independent experiments, 20 cells per experiment), statistical significance was assessed by a two-tailed student's t-test, N.S., not significant, *p < 0.05, **p < 0.01. Control or shATG5 HEK293 cells stably expressing mito-Keima. Control cells were treated with DMSO, 200 µM H2O2, or 4 mM 3-NPA for 2 h, and shATG5 cells were treated with 200 µM H2O2 as a negative control. Cells were then imaged with 458 nm (measuring mitochondria with a neutral pH) and 561 nm (measuring mitochondria with an acidic pH) laser excitation for mito-Keima by confocal microscopy. Right panels show the pixel intensity of red (mitochondria within lysosomes) and green (mitochondria in the cytoplasm) from a line. Scale bar, 10 µm. Quantification of the relative ratio of red to green fluorescence intensity (561 nm/458 nm) of the cells described in (E). Data are presented as mean ± SD (n = 3 independent experiments, 20 cells per experiment), statistical significance was assessed by a two-way ANOVA, N.S., not significant, **p < 0.01. HeLa cells expressing GFP-LC3 were treated with DMSO, 200 µM H2O2, or 4 mM 3-NPA for 2 h. Cells were stained with anti-Tom20 and anti-DNA antibodies, then analyzed by confocal microscopy. The white arrows were indicated the LC3 punta colocalizing or contacting with mtDNA/Tom20. Scale bar, 10 µm. Quantification of the LC3 punta colocalizing or contacting with mtDNA/Tom20 in a cell. n = 30 cells from 3 coverslips, Data are presented as mean ± SD (n = 30), statistical significance was assessed by a one-way ANOVA with Tukey's multiple comparisons test, ***p < 0.001.
Figure 3A Bar graph showing take rates in % of PDXs from the treatment naïve patient ILCs. In parentheses number of injected glands (n) with growth over total n of injected glands. Red dotted line shows average take rate. B Principal component analysis plot of global gene expression profiles of ILC and non-ILC ER+ HER2- (NST) PDX cells purified by FACS-sorting based on GFP expression, n=3. C Barplot showing Reactome pathway enrichment analysis of differentially upregulated genes. D CNET plot showing network associations between genes and pathways in the top 5 terms with increased expresion in ILC. E Barplot of top 10 Reactome enriched pathways upregulated in ILC predicted by ISMARA. F Barplot of top 10 up-regulated MSigDB C2.CP curated canonical pathways collection in ILC PDXs predicted by ISMARA. G Representative photomicrographs of picrosirius red stained histological sections of non-ILC (left) and ILC (right) PDXs. Scale bars 100 μm. H GSEA plots showing gene sets that are differentially regulated between ILC and non-ILC FACS sorted intraductal PDXs, related to luminal progenitor features (left) and increased expression of basal and mesenchymal versus luminal genes (middle and right). Significance scores (P-values) of GSEAs are computed by nonparametric permutation tests, a procedure that yields only estimates of the P-values.
Dcp-1 regulates autophagic flux in vitro.Figure 2.(A) Drosophila S2 cells stably expressing GFP-RFP-Atg8a showed an increase in the percentage of cells containing two or more autolysosomes after starvation, which was blocked after Bafilomycin A1 (BafA1) treatment. At least 50 cells were manually quantitated in three independent experiments (n = 3). Statistical significance was determined using one-way ANOVA with a Dunnett post test (**, P < 0.01). (B) Representative images of S2-GFP-RFP-Atg8a cells after the indicated treatments. (C) S2-GFP-RFP-Atg8a cells were treated with dsRNAs and subjected to fed or starvation conditions as indicated. At least 50 cells were manually quantitated in three independent experiments (n = 3). Statistical significance was determined using one-way ANOVA with a Dunnett post test (**, P < 0.01; ***, P < 0.001; ****, P < 0.0001). (D) Representative images of S2-GFP-RFP-Atg8a cells after the indicated RNAi treatments. Error bars represent the means ± SD. Bars, 10 µm.
Figure 3Volcano plot adjusted p values for differential protein expression to averaged normalised SILAC ratios of two replicates. Red (higher expression in HNSCC) and blue (higher expression in SQCLC) dots indicate significantly regulated proteins (p < 0.05). Two-class comparison of genetic dependencies from a publically available genome-scale CRISPR-Cas9 screen of HNSCC (12 cell line) versus SQCLC (10 cell lines) identified a subset of differentially expressed proteins that were also differential dependencies in these tumor types. The x-axis represents the effect size of the mean difference of dependency scores in HNSCC compared to SQCLC cell lines. Positive effect size indicates a greater mean dependency in HNSCC; negative effect size indicates a greater mean dependency in SQCLC. The y-axis represents the statistical significance of differential enrichment calculated as -log10(p-value) from a two-sided t-test. The p-values used for this plot are uncorrected for multiple hypothesis testing. Highlighted in green are genetic dependencies that were also identified as differentially expressed proteins in our study. Immunohistochemical analysis of the expression of HMGCS-1 (p=0.0014),FTL (p=0.0001) (C),LGALS7 (p=0.0001) and FAM83H (p=0.0001)(D) in an independent cohort of 212 SQCLC and 343 HNSCC cases. Scale bar indicates 100µm. Shown are mean values and standard deviation. Statistical significance was assessed using Wilcoxon-Mann-Whitney test. Pathway-enrichment analysis for proteins differentially expressed in HNSCC and SQCLC.
Selected features of ssc-miR-205.Figure 4(A) ssc-miR-205 turnover. Fully-grown transcriptionally quiescent oocytes were cultured for indicated periods of time and selected miRNAs were quantified by RT-qPCR in four (ssc-let-7a), five (ssc-miR-22) or six (ssc-mir10b and ssc-miR-205) independent experiments. Error bars = SD. (B) Comparison of mean relative changes of miRNA level at 20 h of culture ssc-miR-205 in oocytes is less than ssc-miR-10b, ssc-miR-22, or ssc-let-7a. Error bars = SD. Asterisks indicate statistical significance (p-value) of one-tailed paired t-test, (*p < 0.05, **p < 0.01). (C) Treatment by sodium periodate followed by qPCR does not support 2'-OH modification of either ssc-miR-205 or bta-miR-10b miRNA. Shown are relative levels of the sodium periodate-treated samples to non-treated samples, which were set to one. The experiment was performed three times. Error bars = SD. Efficiency of periodate treatment was confirmed using non-methylated and methylated miR-221 RNA oligonucleotides (D) 3′ tailing of small RNAs in murine, bovine, and porcine oocytes Shown are percentages of miRNAs tailed at 3′ end with mono- and oligonucleotides.
Figure 4A Down-regulating MICAL2PV promotes mitochondrial infiltration into the cortical cytoskeleton. Control and shM2 H1299 cells were subjected to immunofluorescence staining with anti-Tom20 antibody (green) and staining using phalloidin (red). Scale bar: 25 μm. Data information: nuclei were stained with Hoechst 33342 (blue). B Down-regulating MICAL2PV enhances mitochondrial infiltration. Mitochondrial infiltration index was quantified as the ratio of the radial distance from the nucleus to the furthest mitochondria and the radial distance from the nucleus to the plasma membrane. Fifty cells were quantified for each group. Data information: data were analyzed using one-way ANOVA with Bonferroni post-test and shown as mean ± SEM; * P < 0.05, *** P < 0.001. Three independent experiments were conducted. C Down-regulating MICAL2PV enhances the mitochondrial localization of Miro1, Miro2 and KIF5B. Mitochondria were purified from control and shM2 H1299 cells and subjected to Western blotting. The purity of the mitochondrial fraction was confirmed by detection of mitochondrial ATP5B and the absence of the cytoplasmic β-tubulin protein. D The band intensity of target proteins in the mitochondrial fraction was determined [gels shown in (C)] and expressed relative to the Control. Data information: data were analyzed using one-way ANOVA with Bonferroni post-test and shown as mean ± SEM; * P < 0.05, *** P < 0.001. Three independent experiments were conducted.
Fig.Figure 4Hepatic CREBH protein levels in the WT and L-HRD1 KO mice. (n=6 for each group). Hepatic CREBH mRNA (B) levels in the WT and L-HRD1 KO mice. (n=6 for each group). Data information: The data are representative of three independent experiments (mean ± s.d.). (C) Western Blot analysis CREBH protein stability after HRD1 over-expression. Data information: The data are representative of three independent experiments (mean ± s.d.). *: P<0.05. **: P<0.01 by unpaired student's t test. (D) Western Blot analysis of hepatic CREBH and HRD1 protein levels in WT and HRD1 LKO mice. The mice were fasted 16 hours and refed for another 4 hours. (E) Western Blot analysis interaction of CREBH and HRD1 after immunoprecipitates of Flag-agarose in transfected HEK293T. (F) Endogenous interaction between CREBH and HRD1 in liver. (G) Western Blot analysis ubiquitination of CREBH after immunoprecipitates of Flag-agarose in transfected HEK293T. (H) Hepatic CREBH ubiquitination level in the WT and L-HRD1 KO mice (I) Relative protein levels of CREBH, HRD1 and ubiquitination of CREBH. Data information: The data are representative of three independent experiments (mean ± s.d.). *: P<0.05. **: P<0.01 by unpaired student's t test. (J) Primary hepatocytes were isolated from WT and HRD1 LKO mice and Western Blot analysis CREBH protein stability. (K) Full-length and C-terminal of HRD1 were generated. (L) Interactions of full-length and C-terminal of HRD1 with CREBH were measured by Co-IP. Data information: The data are representative of three independent experiments (mean ± s.d.). *: P<0.05. **: P<0.01 by unpaired student's t test.
Figure 2Two gRNAs flanking the Dl Exon 2 were used to replace the exon with a modified version where 5 amino-acids in the β1-2 loop were removed. Red lettering highlights the genomic sequence of the β1-2 loop. Adult wings from Dl∆β1-2 flies. No defects are detected in wings from Dl∆β1-2/+ (B), Homozygous Dl∆β1-2/ Dl∆β1-2 have extra vein tissue near L5 and uneven L2 veins (arrowheads; C). Microchaetae are arranged in rows on the thorax of control (yw) flies; these become disordered and more dense in Dl∆β1-2/Dl∆β1-2. White rectangle indicates area scored for E Number of microchaetes per central area (white rectangle in D) in the indicated genotypes.
NKp46 antibody-coated NPs do not induce spontaneous non-specific NK cell activation or cytokine secretion.Figure 5(A) YTS KIR2DL1 cells were incubated with SHP-1 and Cbls siRNA-loaded NPs (red curve) or empty NPs (purple curve). After 48 hours, cells were loaded with Fluo-3-AM and analyzed for intracellular calcium levels for 20 min at 37°C. YTS KIR2DL1 cells that were treated with PBS served as a negative control (green curve). NK cells that were mixed with 721.221 Cw7 target cells served as a positive control (blue curve). Calcium levels were analyzed by spectrofluorometry. (n=3) (B) Freshly isolated PBMCs were treated with SHP-1 and Cbls siRNA-loaded NPs, empty NPs, PBS as negative control or PHA as a positive control. Following incubation, supernatant from each sample was collected and the levels of human cytokines TNF-α, IL-6, IL-10 and IFN-γ were determined using ELISA. The data represent four independent experiments (n=4). P values were calculated by One way ANOVA following Tukey's post hoc analysis, and are indicated by asterisks. *P≤0.008. Data are shown as mean ± SEM.
Figure 6Cartoon representation of the OTU fold (vOTU, PDB 3PHW), with the active site and S1 site indicated. The S1 site is composed of a common region (CR, red) surrounded by three variable regions (VR, blue) that are responsible for Ub binding. Comparison of structural adaptations in the VR1 arm region of the S1 site. VR1 has been observed to contribute to Ub binding as either a short α-helical segment (left), and extended α-helical region (center), or a β-hairpin (right). Examples of OTUs that follow each arrangement are provided to the right. Comparison of structural adaptations in the VR2 central β-sheet edge of the S1 site. VR2 has been observed to contribute to Ub binding in its most common arrangement (left), with additional or fewer β-strands (center), or altered with additional substructure (right). Examples of OTUs that follow each arrangement are provided to the right. Comparison of structural adaptations in the VR3 loop extending from the central β-sheet. This VR3 loop has been observed as short and not utilized in Ub binding (left), expanded and participating in unstructured interactions with Ub (center), or expanded with a β-hairpin motif that binds Ub (right). Examples of OTUs that follow each arrangement are provided to the right.
Characterization of Dnmt2−/− MSCs.Figure 3Flow cytometry analysis of bone marrow primary mesenchymal cells. Quantification of fibroblast colony-forming units (CFU-F). Quantification of alkaline phosphatase colony-forming units (CFU-ALP). Dnmt2 expression after 3 days of mesenchymal cell differentiation. Alizarin Red staining, indicating enhanced osteoblast differentiation of primary stromal cells from Dnmt2−/− mice. Scale bar: 50 μm. Quantification was performed with three biological replicates. qPCR analysis of the osteoblast markers Osterix and Osteocalcin. Oil Red O/hematoxylin staining, showing reduced adipocyte differentiation of primary stromal cells from Dnmt2−/− mice. Scale bar: 50 μm. Quantification was performed with three biological replicates. qPCR analysis of the adipocyte markers CEBp alpha and Glut4.