EMBO/sd-character-level-ner huggingface dataset:
Figure 1(A) Top: SERA6 architecture. Positions of the papain-like domain, catalytic triad residues, SUB1 cleavage sites and epitopes for the antibodies used are indicated. Positions of the inserted mTAP or myc3 epitope tags (between codons Asn886-Val887) introduced in SERA6-mTAP:loxP parasites are shown. Bottom: time-course analysis of egress by western blot, showing processing of SERA6-mTAP into multiple protein species (representative of 2 independent experiments). Schizonts were sampled at the indicated times following washout of C2-mediated arrest. Nomenclature for each protein species was based on apparent molecular mass or predicted composition. Note that the predicted mass of the p40 species is ~27 kDa, but its migration on SDS PAGE may be aberrant due to its acidic nature; the predicted pI of sequence between SUB1 site 1 and the central papain-like domain (Asp371-Lys605) is ~4.4 (see https://web.expasy.org/compute_pi/). (B) Coomassie-stained SDS PAGE gel showing proteins immunoprecipitated from schizonts expressing SERA6-mTAP, sampled at the indicated times following C2 washout or arrested with E64 (50 μM). Control, proteins immunoprecipitated from SERA6:loxP (Thomas et al, 2018) schizonts, which express non-epitope-tagged SERA6, sampled at 5 min following C2 washout. Arrowed, species analysed by mass spectrometric peptide mapping. Fragments A-H were identified as SERA6-mTAP-derived, while fragments I and J were derived from a different protein, PF3D7_1014100. (C) Schematic of the likely identities of the different SERA6-mTAP species identified from peptide mapping, relative to full length SERA6-mTAP and the papain-like domain (hatched). The predicted mass of each species based on its amino acid composition is indicated. Detection of a semi-tryptic SERA6-derived peptide (filled block arrowhead) unambiguously confirmed cleavage at SUB1 site 1 while the coverage allowed mapping of up to five residues upstream of SUB1 site 2 (fragment E). No evidence was found for processing at a third putative SUB1 cleavage site (Ruecker et al., 2012). The N-terminus of fragment H (assumed to correspond to PAP-mTAP) could not be precisely mapped; attempts to assign its N-terminus by Edman degradation were unsuccessful. Also see Figs EV1 and EV2.
Figure 1Blood samples were obtained from 42 pediatric cases of natural chikungunya (CHIKV) infections at an acute (d1-2) and a convalescent (d15-17) timepoint, relative to reported symptom onset. Samples were separated into whole blood, serum, and peripheral blood mononuclear cell (PBMC) aliquots for transcriptomic analysis, CHIKV viral titer assays, multiplex ELISA for cytokines, and mass cytometry (CyTOF). These data were combined with clinical data, including a severity score and a d15-17 anti-CHIKV antibody titer titer, to create a multiscale network of interactions during the observed course of CHIKV infection.
Asymptomatic COVID-19 patients exhibit more robust SARS-CoV-2 specific Th17 responses compared to symptomatic patients.Figure 3A. Expressions of genes associated with T cell functionality were compared between acute asymptomatic (n=30) and symptomatic (n=26) COVID-19 patients. Heatmap of DEGs with FDR adjusted P value < 0.05, scaled based on log2RPKM values, with blue and red colors indicating low and high expressions, respectively. B. Mass cytometry was performed on PBMCs obtained from acute symptomatic (n=37) and acute asymptomatic (n=19) COVID-19 patients and healthy controls (n=10). Naïve, TEMRA, Central memory (CM) and effector memory (EM) T cells were characterized based on CD45RA and CCR7 expressions. Data are presented as Mean ± SD *P<0.05; **P<0.01 (Mann-Whitney U test). C. SARS-CoV-2 specific CD4+ non T follicular helper (TFH) cells from symptomatic (n=5) and asymptomatic COVID-19 (n=5) patients were characterized with flow cytometry based on the expression of IL-17a, IFN-γ, TNF-⍺, IL-4 and IL-10 upon SARS-CoV-2 peptide stimulation. Representative plot overlay of unstimulated (black) on peptide stimulated (red) was performed on asymptomatic patient. Data are presented as Mean ± SD **P<0.01 (Mann-Whitney U test).
hnRNP L confers a spectrum of protection from NMD A qPCR of selected transcripts found by RNA-Seq to decrease in abundance in the absence of hnRNP L.Figure 3A qPCR of selected transcripts found by RNA-Seq to decrease in abundance in the absence of hnRNP L. Values represent the mean and standard deviation of 3 independent biological replicates. Significance of hnRNP L KD and UPF1 KD were compared to NT, and the double knockdown was compared to hnRNP L KD. For clarity, only a single star is used to indicate significance; in all cases P ≤ 0.025 in two-tailed Student's t-tests. B Half-lives of selected transcripts under conditions of non-targeting or hnRNP L knockdown. Measurements were calculated based on proportional recovery of transcript labeled with 5-EU in 1 hour versus total transcript. Significance was determined using two-tailed Student's t-test: *P ≤ 0.05, **P ≤ 0.01, ****P ≤ 0.0001. Values represent mean and standard deviation of 3 independent replicates. C The stability of β globin reporters with the CSRP1 3'UTR, along with several variants in which putative hnRNP L motifs in the indicated intervals were mutated, were evaluated in HeLa Tet-off cells. The schematic indicates areas of 3' UTR CA-enrichment in light blue and includes the sequence of the first 200 nucleotides of the CSRP1 3'UTR, where putative hnRNP L binding elements are in red typeface. The northern blot is a representative example from 3 independent replicates. Mean values plotted with SD at each timepoint were used to quantify decay (right panel). Significance is measured compared to the WT reporter using the sum-of-squares F test. D The stability of the WT and L mut 1-200 CSRP1 reporters in HeLa Tet-off cells treated with non-targeting RNAi or siRNA against UPF1 was evaluated. The northern blot is a representative example from at least 3 biological replicates. Decay quantification and statistics were performed as in (C), with significance measured compared to the non-targeting condition.
Figure 1 A. RT-PCR of CNPase (left) and MBP (right) mRNAs expression in MO3.13 precursors under control conditions and following 10-200 μM D-Asp exposure for 3 days. Graphs show quantification of ratio of CNPase, and MBP to L19 Data information: The values represent the means ± S.E.M. Level of significance was determined by using: in A, left panel, one-way ANOVA P<0.0001 followed by Tukey's post hoc test, *P< 0.05 versus control (n=3); A, right panel, one-way ANOVA P<0.0001 followed by Tukey's post hoc test, *P<0.05 versus control (n=3  B. Western blotting (left) and densitometric analysis (right) of MBP expression in absence or in presence of 10-200 μΜ D-Asp exposure for 5 days Data information: The values represent the means ± S.E.M. Level of significance was determined by using ); B, one-way ANOVA P=0.0001 followed by Bonferroni post hoc test, *P<0.05 versus control (n=3  C. Cell growth analysis of human MO3.13 oligodendrocytes in absence or in presence of 200 μM D-Asp for 1-5 days. The density of M03.13 oligodendrocytes was daily recorded through trypan blue dye exclusion. Mean of daily measurements were recorded. The data of each experimental group were normalized to the density of cells plated at day 0 and expressed as percentage of ctrlday0 Data information: The values represent the means ± S.E.M. Level of significance was determined by using C, for each day, two-tailed Student's t test, *P<0.05 versus control at 3 days (n=5)  D. FACS-based cell cycle distribution analysis after PI incorporation of MO3.13 oligodendrocytes in absence or in presence of 200 μM D-Asp for 3 days. Representative FACS plots of biological replicates are shown (n=5 independent experimental sessions)  E. FACS-based cell cycle distribution analysis after PI incorporation of rat primary OPC in absence or in presence of 100 μM D-Asp for 1 day. Representative FACS plots of biological replicates are shown (n=3 independent experimental sessions)  F. Confocal microscopic images displaying the coexpression of MAG or MBP with Phalloidin-594 (a-b and c-d, respectively) in rat primary OPC cultured in absence or in presence of 100 μM D-Asp for 4 days. (e), Single representative MBP++ cell. Scale bars: 50 μm in a-d; 10 μm in e. (f-g); Quantitative analysis of Alexa-594-phalloidin-positive oligodendrocytes (f) or MAG+ and MBP++ oligodendrocytes (g) scored in control and D-Asp exposed cells. Alexa-594-phalloidin-positive oligodendrocytes were scored in 4 categories, according to their morphological complexity. In each category, data were normalized on the total number of oligodendrocytes Data information: The values represent the means ± S.E.M. Level of significance was determined by usin F, for each category two-tailed Student's t test, *P<0.05 versus control (n=3)
Figure 7HeLa cells with or without transient expression of H2B-GFP or EGFP-TOP2A were exposed to nocodazole for 3 hr. Mitotic cells were cytospun onto coverslips and subjected to immunofluorescence staining with DAPI and antibodies for GFP, Sgo1 and ACA. Example images are shown (A). The immunofluorescence intensity ratio of centromeric Sgo1/ACA was determined on 400 chromosomes in 20 cells (B). The means and S.D.s from three independent experiments with the Sgo1/ACA ratio determined on around total 1200 chromosomes in 60 cells are shown (C). Data information: Means and error bars representing S.D. are shown unpaired t test). Scale bars, 10 μm. HeLa cells with or without transient expression of H2B-GFP or EGFP-TOP2A were exposed to nocodazole for 3 hr. Mitotic cells were cytospun onto coverslips and Lysates of asynchronous cells were immunoblotted (D). * represents non-specific bands. U2OS-LacO cells transiently expressing Myc-LacI-Bub1, with or without EGFP-TOP2A (970-1531), were subjected to immunofluorescence staining with DAPI and antibodies for H2ApT120 and Sgo1. Example images are shown (E). Scale bars, 10 μm. U2OS-LacO cells transiently expressing Myc-LacI-Bub1, with or without EGFP-TOP2A (970-1531), were subjected to immunofluorescence staining with DAPI and antibodies for H2ApT120 and Sgo1. The relative enrichment of EGFP-TOP2A (F) and Sgo1 (G) at the LacO repeats was quantified in 120 cells. Data information: Means and error bars representing S.D. are shown unpaired t test). U2OS-LacO cells transiently expressing Myc-LacI-Bub1, with or without EGFP-TOP2A (970-1531) Cell lysates were immunoblotted (H). U2OS-LacO cells transiently expressing EGFP-LacI-Bub1-K together with VSV-Sgo1 or a control vector were subjected to immunofluorescence staining with DAPI and antibodies for TOP2A and VSV-tag. Example images are shown (I). The relative enrichment of TOP2A (J) and VSV-Sgo1 (K) at the LacO repeats was quantified in 50 cells. Data information: Means and error bars representing S.D. are shown unpaired t test). Scale bars, 10 μm. U2OS-LacO cells transiently expressing EGFP-LacI-Bub1-K together with VSV-Sgo1 or a control vector Cell lysates were immunoblotted (L). * represents non-specific bands.
Figure 5A Induction of Treg following PBMC treatment with either native or proteinase K-treated CLys or CSN (10µg/ml) for 72h. B Mass spectrometry analysis of CVF, CLys and CSN and identification of distinct and shared proteins. From the shared proteins in CLys and CSN, relative detection scores of GDH and IDH were presented. C PBMCs were treated with GDH inhibitor bithionol (BT) (20µM) or IDH inhibitor IDH-305 (50nM) alone or in combination before incubation with CLys or CSN. Percentage of Treg was determined and PGE2 and IL-10 levels were evaluated. D Treg induction from PBMC by CLys, CSN and TsGDH (5µg/ml) and levels of PGE2 and IL-10 in culture supernatants. E Treg induction from monocyte-naïve CD4+-TsGDH culture and levels of PGE2 and IL-10 in monocyte-CD4+-TsGDH and monocyte-TsGDH culture supernatants. F Treg induction from PBMC by CLys, CSN and rhGDH (10µg/ml).
COP1Lys422Ala shows a delayed flowering phenotype under long days and suggests that CO - CRY2 may compete for the COP1 VP-binding site.Figure 5A. Representative image of wild-type (Ws), cop1-5/Pro35S:YFP-COP1, cop1-5/Pro35S:YFP-COP1Lys422Ala, cop1-5/Pro35S:YFP-COP1Tyr441Ala and cop1-5/Pro35S:YFP-COP1Trp467Ala transgenic lines grown for 39 days in long day conditions. B. Representative image of individual wild-type (Ws), co-11, cop1-5/Pro35S:YFP-COP1 and cop1-5/Pro35S:YFP-COP1Lys422Ala plants grown for 39 days in long days conditions. C. Quantification of flowering time. Means and SD are shown (n = 14). D. Number of rosette and cauline leaves at flowering. Means and SD are shown (n = 14). E. Immunoblot analysis of CRY2 and actin (loading control) protein levels in wild-type (Col), cry2-1, wild-type (Ws), cop1-5/Pro35S:YFP-COP1 and cop1-5/Pro35S:YFP-COP1Lys422Ala seedlings grown for 4 days in darkness. F. ITC assays between the CO VP-peptide (left) and CRY2 VP-peptide (right) versus the COP1 WD40 and the COP1Lys422Ala WD40 domains, with a table summarizing their corresponding affinities. The following concentrations were typically used (titrant into cell): CO - COP1 (2240 μM in 195 μM); CO - COP1Lys422Ala (1200 μM in 120 μM); CRY2 - COP1 (750 μM in 70 μM); CRY2 - COP1Lys422Ala (3000 μM in 175 μM). The stoichiometries of binding (N) for CO - COP1 = 1.03 ± 0.00; CO - COP1Lys422Ala = 0.89 ± 0.03; CRY2 - COP1 = 1.04 ± 0.20; CRY2 - COP1Lys422Ala = 1 (fixed) (All measurements ± standard deviation). G, H. Binding kinetics of the (G) full-length activated CRY2 and (H) CRY2NC80 versus the COP1 WD40 domain obtained by GCI experiments. Sensorgrams of protein injected are shown in red, with their respective heterogenous ligand binding model fits in black. The following amounts were typically used: ligand - COP1 (2000 pg/mm2); analyte - CRY2 (highest concentration 14 μM), CRY2NC80 (highest concentration 450 μM). ka = association rate constant, kd = dissociation rate constant, Kd = dissociation constant.
Sept2 localizes to mitochondrial constrictions.Figure 3(A) Immunofluorescence microscopy analysis of endogenous Sept2 (green) and mitochondria (Tom20, red) in HeLa cells. The insets show a twofold enlargement with arrowheads highlighting the presence of small Sept2 structures on mitochondria. Scale bar 10 µm. (B) 3D rendering of confocal microscopy sections showing Sept2 at mitochondrial constrictions (white arrowheads) and constricted mitochondria that are not marked by Sept2 (yellow arrowhead) Scale bar 2 µm. (C) CoDiM super-resolution imaging of endogenous Sept2. Arrowheads highlight the presence of Sept2 at mitochondrial constrictions. Insets show a threefold enlargement Scale bar 10 μm. (D) Still images from a live cell imaging experiment showing Sept2 (green) accumulation during two mitochondrial fission events (mitotracker orange, red), highlighted by yellow (first fission) or green arrowheads (second fission). Scale bar 2 µm.
Figure 1A Schematic representation of the S. pombe telomeric transcriptome. Telomeric repeats are shown in black, subtelomeric sequences in grey. Oligonucleotides used for RT-PCR and for 3' end RACE are indicated. B Growth curves of the indicated strains shown as population doublings (pds) on the y axis and hours in liquid culture on the x axis. Before inoculation in liquid YES medium, single cells underwent approximately 25 pds to produce visible colonies on solid medium. For each strain, averages and SDs (error bars) of three biological replicates are shown. Arrows indicate times of harvesting and corresponding pds for experiments shown in (C) and (D). C Telo-PCR analysis of telomere length in wt (ter1+ and est1+) and two independent clones (a and b) deleted for ter1+ or est1+ grown for ~ 25+10 (~ 35), ~ 25+14 (~ 39) and ~ 25+18 (~ 43) pds in YES medium. PCRs were performed with the subtelomeric oligonucleotide oF5, located 620 bp upstream of the first telomeric repeat, and the oligo(dG)-containing oligonucleotide odG18. Marker (m) molecular weights are on the left in kilobases. tels: telomere-containing Telo-PCR products. D qRT-PCR analysis of G-rich TERRA, polyA+ TERRA and ARRET/αARRET levels in cells as in (B). Values are normalized to ACT1 mRNA and expressed as fold increase over control wt strains (ter1+ and est1+) grown in parallel. Bars and error bars are averages and SD from 4 (ter1Δ clones) or 3 (est1Δ clones) independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.001 (relative to wt; two-tailed Student's t test). E ChIP experiments with anti-RNA polymerase II (RPII) antibodies and extracts from ter1+ and ter1Δ cells grown for ~ 43 pds. Quantitative PCRs were performed using oligonucleotides oF1 + oR1 (subtel) or oligonucleotides amplifying a region from the actively transcribed act1+ locus. Values are expressed as fold increase over ter1+. Bars and error bars are averages and SD from three independent experiments. * P < 0.05 (relative to ter1+; two-tailed Student's t test). The western blot to the right shows no difference in RNPII levels between ter1+ and ter1Δ cells used in ChIP experiments. Ponceau Sstaining serves as loading control. F 3' RACE analysis of polyA+ TERRA in ter1Δ cells grown for ~ 43 pds. The sequence at the top is the genomic reference sequence (gen) taken from the pNSU70 plasmid (nucleotide positions are indicated). The sequences below derive from all positive clones isolated. Sequences boxed in blue, green and yellow correspond to the oligonucleotide used for RACE (o3'RACE), subtelomeric DNA and telomeric repeats, respectively. Genomic adenines corresponding to putative RNA cleavage sites (CS) are highlighted. G Western blot analysis of cellular fractions from ter1+ and ter1Δ cells grown for ~ 43 pds. Total proteins (tot) and equivalents of soluble (sol) and insoluble (ins) proteins were loaded. Actin serves as a control for soluble proteins, H3K9ac for chromatin-bound proteins. H qRT-PCR analysis of total TERRA and polyA+ TERRA in cellular fractions as in (G). Values are expressed after normalization to total cellular extracts. Bars and error bars are averages and SD from 3 independent experiments.
Figure 3Representative images of day 21 BALO and tdTomato+ rMC stained with LipidTOX (green). Scale bar represents 100 μm. Fluorescence images of αSMA (green) and neutral lipids (LipidTOX red) staining in WT‑derived rMC and BALO (dotted lines indicate single BALO or insert) at day 21 of culture. Scale bars represent 50 μm (left) and 25 μm (right). Representative flow cytometric dot plots and histograms of PDGFRα expression in rMC (EpCAM-CD45-CD31-Sca-1+) isolated from the lung homogenate of PdgfraGFP reporter mice. Representative images of BALO formation at day 8, 15 and 21 of co-culture. WT BASC were co‑cultivated with sorted rMC (total PDGFRα+ population) or rMC expressing either low or high levels of PDGFRα‑GFP. Scale bars represent 100 μm. tSNE plots and violin plots depicting selected genes representing MYO (Pdgfrα, Tagln, Acta2, Eln, and Axin2) (top panels) and LIF associated-genes (Fgf10, Apoe, Serpina3n, Gsn, and Gas6) (bottom panels). Each violin plot shows the frequency distribution of the mean transcript level (log2). C3 (MYO, orange) and C4 (LIF, green) refer to the scRNA-Seq experiment in Fig. 1G. Fluorescence image of αSMA+PDGFRαhigh MYO (yellow arrows) and LipidTOX+PDGFRαlow LIF (red arrows) from sorted PDGFRα-GFP rMC after 21 days of BALO culture. Scale bars represent 50 μm (left) and 25 μm (right). Fluorescence image containing PDGFRαhigh MYO from sorted PDGFRα-GFP rMC within BALO derived from tdTomato+ mice at day 21 of culture. Scale bars represent 100 μm (left) and 50 μm (right).
Figure 7A. Overall survival (OS) curves in Pancreatic Adenocarcinoma patients (PDAC) (n=172) from The Cancer Genome Atlas (TCGA), showed that combined expression of AXL, GAS6 and ENAH (hMENA), is a prognostic signature in PDAC. B. Overall survival (OS) curves in Lung Squamous cancer patients (LUSC) (n=501) from The Cancer Genome Atlas (TCGA), showed that combined expression of AXL, GAS6 and ENAH (hMENA), is a prognostic signature in LUSC.
Figure 5A. Fold change (FC) and significance for transcripts in Kdm6apKO vs. control pancreas. Genes significant at q<0.05 are shown as colored dots above the horizontal line. B. GSEA showing that genes specific to differentiated acinar cells were downregulated in KDM6A-deficient pancreas, but not genes specific to islets or duct cells. Upregulated genes were enriched in genes specific to mesenchymal cells. Lineage-enriched genes were obtained from (Muraro et al., 2016). C. Most enriched functional annotations in genes differentially expressed in Kdm6apKO. D. GSEA showed increased expression of indicated oncogenic pathway genes in Kdm6apKO pancreas. E. KDM6A functions as a transcriptional activator of direct target genes in pancreatic cells. Left: Downregulated, but not upregulated, genes in Kdm6apKO pancreas were enriched for KDM6A binding. P-values and odds ratios (O.R.) were calculated by Fisher's exact test. Right: Venn diagrams showing overlap of down and upregulated genes with KDM6A binding. F. KDM6A-bound enhancers and promoters showed increased H3K27me3 and decreased H3K27ac in Kdm6apKO pancreas. G. Genes that were downregulated in Kdm6apKO pancreas (KDM6A-dependent genes) showed greatest changes in histone marks. Box plots show fold-changes of H3K27me3 (left) and H3K27ac (right) signals in Kdm6apKO compared to control. Signals were analyzed in KDM6A bound regions in promoters and enhancers of genes that were KDM6A dependent (grey; n=420 regions) and independent (white; n=8035 regions). The signals are average values from ChIP-seq experiments in two biological replicates. The horizontal central line marks the median. Box limits indicate the first and third quartiles and whiskers extend to highest and lowest data points within 1.5 x IQR outside box limits. P-values were determined by two-tailed Mann-Whitney U test. H. Changes in KDM6A, H3K27me3 and H3K27ac ChIP-seq profiles in the Ppp4r4-Serpina10 locus in Kdm6apKO pancreas. I. qPCR of H3K27me3, H3K27ac and H3K4me1 changes in promoter and enhancer regions highlighted in (H) (R1, R2, R3), and Serpin10 mRNA. Error bars show +/-SD and P-values are from two-tailed Student's t test, n=3. J. KDM6A-bound genes that showed decreased expression in Kdm6apKO pancreas (functional KDM6A binding sites) showed simultaneous gain of H3K27me3 and loss and of H3K27ac.
Figure 4(A) qRT-PCR in TGFß treated MCF10A cells after siRNA mediated knockdown of ZEB1, FOSL1 or YAP compared to control knockdown cells. Knockdown efficiency of all three factors is shown on the left. The right panel shows their effects on 11 ZEB1/YAP/AP-1 activated target genes. (B) Overlap of ZEB1, YAP and JUN peaks at the promoter of CYR61, CTGF and ANKRD1 and in a known enhancer region of the DOCK9 gene. (C) ZEB1-only peak at the promoter of LLGL2, a well-known ZEB1 repressed target. (D) Luciferase reporter assay in MDA-MB-231 cells comparing control and transient siRNA-mediated knockdown of ZEB1. Reporter constructs of regulatory regions of genes For reporter assays, firefly luciferase activity was normalized to co-transfected renilla luciferase. (E) Luciferase reporter assays with the ANKRD1 promoter and DOCK9 enhancer constructs in MCF7 cells upon overexpression of ZEB1, YAP and JUN/FOSL1 (AP-1) or different combinations of the factors. n = 5 (ANKRD1) and n = 3 (DOCK9); shown is mean ± s.e.m.; *P ≤ 0.05; ratio paired t-test. For reporter assays, firefly luciferase activity was normalized to co-transfected renilla luciferase.