EMBO/sd-character-level-ner huggingface dataset:
A-bound regions mark the highly interacting long-distance contacts in ESCs A.Figure 4A. Number of BAZ2A-bound regions in A and B compartments of ESC+serum. B. Hi-C contact matrices for a zoomed in region on chromosome 3 at 100-kb resolution showing the presence of strong long-distance contacts corresponding to regions in A compartment bound by BAZ2A in ESC+serum. Low panels. HiC contacts of a zoomed region at 50-kb resolution in ESC+serum, NPCs and CNs are shown. Data are from (Bonev et al., 2017). HiC contacts were visualized using Juicebox C. BAZ2A-bound regions mark A subcompartments that are characterized by the strongest far-cis contacts in ESC+serum. Boxplots indicating observed/expected contact values between BAZ2A-bound regions and loci not bound by BAZ2A at near-cis and far-cis contacts in all genome or in A compartments. Statistical significance (P-values) was calculated using the unpaired two-tailed t-test (****< 0.0001). D. Long-range contacts marked by BAZ2A binding are specific for ESCs. Boxplots indicating observed/expected contact values between BAZ2A-bound regions and loci not bound by BAZ2A at far-cis contacts in all genome or in A compartments of ESC+serum, NPC and CN. Statistical significance (P-values) was calculated using the unpaired two-tailed t-test (**** < 0.0001). Box plots depict the minimum and maximum values. The mean is represented by a horizontal line within the boxes. E. Number of BAZ2A-bound regions in A and B compartments in ESC+2i. F. BAZ2A-bound regions mark A subcompartments that are characterized by the strongest far-cis contacts in ESC+2i. Boxplots indicating observed/expected contact values between BAZ2A-bound regions and loci not bound by BAZ2A at near-cis and far-cis contacts in all genome or in A compartments of ESC+2i. Statistical significance (P-values) was calculated using the unpaired two-tailed t-test (** < 0.001, ****< 0.0001). Box plots depict the minimum and maximum values. The mean is represented by a horizontal line within the boxes. G. BAZ2A binding correlates the best with eigenvector values. Heat map showing Pearson correlation values of ChIPseq data sets with the eigenvector values from HiC data in ESC+serum and ESC+2i, respectively. H. Boxplots showing H3K4me1 and H3K27ac content of BAZ2A-bound regions and loci not bound by BAZ2A at far-cis contacts in ESC+2i. Data from regions with lowest 10% and top 10% contact strength are shown. Box plots depict the minimum and maximum values. The mean is represented by a horizontal line within the boxes.
Nucleolar RNA is enriched in specific intron‐encoded aluRNAsLeft: Scheme illustrating the origin of forward or reverse intronic aluRNA and processing into left arm aluRNAs (aluRNAL) and right arm aluRNAs (aluRNAR).Figure 2Left: Scheme illustrating the origin of forward or reverse intronic aluRNA and processing into left arm aluRNAs (aluRNAL) and right arm aluRNAs (aluRNAR). Right: Heatmaps of read density of forward and reverse intronic Alu RNA species present in the total RNA sample. Top: Heatmaps of read density of intronic Alu repeats in the nucleolar RNA fraction. For each aluRNA variant, the percentage over all transcribed intronic Alu repeats is indicated. Bottom: Ratio of nucleolar versus total RNA from higher (red) to lower (green) read density. View of nucleolus‐enriched Alu elements exemplified at two genomic loci. Mapping of normalized reads from total (red), nucleoplasmic (orange) and nucleolar (blue) RNA to intron #5 of DCP1A and intron #22 of UBE4B. Heatmaps of total RNA read density ratios between untreated (control) and α‐amanitin‐ or ML‐60218‐treated HeLa cells.
Figure 2(A) Overall identified O-glycoproteins in all HepG2 cell lines (631 glycoproteins) with comparison to the HepG2 O-glycoproteome presented in Schjoldager et al., 2012 (74 glycoproteins) and the human O-glycoproteome identified in 12 SCs presented in Steentoft et al., 2013 (662 glycoproteins). (B) Relative numbers of all identified sites, quantified sites and quantified single sites identified in this study. (C) Distribution of glycosites per protein identified in this study.
The human genome contains 1399 high-confidence MXEs.Figure 1A) Schematic representation of the various annotated and predicted exon types included in the MXE candidate list. For MXE validation at least three restraints must be fulfilled: the absence of an MXE-joining read (R1), except for those leading to frame shift, and the presence of two MXE-bridging SJ reads (R2 and R3). B) Prediction and validation of 1399 1SJ (855 3SJ) human MXEs. Top: Dataset of 6541 MXE candidates from annotated and predicted exons. Bottom left: MXE candidates for which splice junction data is currently missing hindering their annotation as MXE or other splice variant. Bottom right: Validation of the MXE candidates using over 15 billion RNA-seq reads. The outer circles represent the validation based on at least a single read for each of the validation criteria (1SJ), while the validation shown in the inner circles required at least three reads (3SJ). C) MXE saturation analysis. Whereas increasing amounts of RNA-seq reads should lead to the confirmation of further MXE candidates, more RNA-seq reads might also result in the rejection of previously validated MXEs. The green curves show the number of validated MXEs in relation to the percentage of total RNA-seq reads used for validation. The orange curves indicate the number of initially 'validated MXEs' that were rejected with increasing amounts of reads. Grey dashed lines indicate the point of saturation, which is defined as the point where a two-fold increase in reads leads to rejection of less than 1% of the validated MXEs. Of note, whereas the rejection of validated MXEs saturates with 20% of the data the amount of novel MXE validations is still rapidly increasing. D) Distribution validated MXEs in two-exon and multi-exon clusters. E) Size and distribution of multi-cluster MXEs. F) The CUX1 gene (cut-like homeobox 1) contains 2 interleaved clusters of MXEs (clusters 1 and 2) and 2 standard clusters each with two MXEs (clusters 3 and 4). The exon 3 and exon 4 variants each are orthologous exons. The exon 4 variants are mutually exclusive (cluster 2). Exon 3a is a differentially included exon and only spliced together with exon 4a. The exons 3b, 3c, 3d and 3e are part of a cluster of four MXEs (cluster 1) and are only spliced together with exon 4b (Appendix Fig. S16 and S17). Novel exons are labelled with an asterisk.
Figure 5A) Decision tree to classify AD vs non-AD participants based on the protein levels of a core 26 protein set. Splits are indicated by black triangles. A tree with a minimum depths of six can correctly classify the participants by AD status. B) Receiver operating characteristic (ROC) curve for the model based on XGBoost. The diagonal line indicates random performance. Black line represents the mean performance of the model when trained on six stratified train - test splits (K-fold). The grey areas represent the standard deviation of ROC values. C) Confusion matrix indicating model performance when predicted on the test split of the cross-validation. Overall accuracy is 0.85.
Figure 2A. m6A peak distribution of BZLF1 in different EBV infection stages was analyzed based on MeRIP-seq data. The data presented from the top down are the EBV acute infected (24 hpi) NPEC1-Bmi1 sphere-like cells (NPEC-sp), and HK1 cells; latently infected CNE2EBV cells; and CNE2EBV cells with reactivated EBV induced using NaB alone or TPA/NaB together. For acute EBV infection, RNA was harvested at 24 hpi. TPA (30 ng/ml) and NaB (2 mM) were used alone or together to treat the cells for 24 h to reactivate EBV. The PA-m6A-seq peak is indicated by the green box, and the potential m6A sites on the indicated sequence are shown in red. The region indicated by the dotted box was PCR-amplified to construct the wild-type BZLF11-262nt plasmid. B. The BZLF1 m6A levels were measured using MeRIP-qPCR in CNE2EBV, Raji and B95-8 cells with latent EBV infection or lytic reactivation. TPA (30 ng/ml) and NaB (2 mM) were used to treat the cells for 24 h to reactivate EBV. The cellular RNA was harvested for MeRIP-qPCR assays. The fold enrichment was determined by calculating the 2-ΔCt of the MeRIP sample relative to the input sample. The mean value of the results in mock-treated cells was normalized to 1. C. The relative product abundance of SELECT at the predicted m6A sites in the latently infected or lytic reactivated CNE2EBV cells. TPA (30 ng/ml) and NaB (2 mM) were used to treat the CNE2EBV cells for 24 h to reactivate EBV. The mean value of results in mock-treated cells was normalized to 1. D. The validation of the BZLF1 m6A peaks by MeRIP-qPCR in PDXs, NPC1 and NPN. One nontumor control biopsy (NPN), one NPC sample (NPC1), and two PDX samples (PDX-NPC04L and PDX-NPC03W) were used to perform the MeRIP-qPCR assays. The fold enrichment was determined by calculating the 2-ΔCt of the MeRIP sample relative to the input sample. E. Construction of the WT-BZLF1 and mut-BZLF1 plasmids. The WT-BZLF1 plasmid was constructed by inserting the BZLF11-262nt fragment (1-262 nt of the BZLF1 mRNA) into pcDNA3.1+ plasmid. The mut-BZLF1 plasmid was constructed by introducing A>T mutations at the putative m6A sites of BZLF11-262nt. F. The IgG or anti-m6A antibody enrichment of the WT-BZLF1 and mut-BZLF1 mRNAs was measured by RIP-qPCR in the CNE2EBV cell lines. Blank indicates CNE2EBV cells without plasmids transfection. The fold enrichment was determined by calculating the 2-ΔCt of the RIP sample relative to the input sample. The mean value of the m6A enrichment level on WT-BZLF1 mRNA was normalized to 1.
Models predict the sequence of gene activation during resetting to naïve pluripotency.Figure 2(A) Model predictions of the number of regulation steps required for permanent activation of each network component. Light blue regions indicate where only some concrete networks predict that the given gene has permanently activated (B) Heatmap of average gene expression normalised to β-actin over an EpiSC resetting time course in 2i+LIF. Each row is coloured according to the unique minimum and maximum for that gene. The values shown are average expression of four independent experiments (C) Gene expression for Stat3, Klf2, Esrrb and Tfcp2l1 during EpiSC resetting relative to established mouse ESCs. β-actin serves as an internal control. Mean +/- SEM, n=4 independent experiments. * = p<0.05 Student"s t (D) Left: Local network topology for Tfcp2l1 and Esrrb
Figure 3A-L Double immunolabelling of CETN3 (green) and SYCP3 (red) on squashed WT mouse spermatocytes at (A) leptotene, (B) zygotene, (C) pachytene, (D) metaphase I, (E) telophase I, (F) early interkinesis (F), late interkinesis (G), prophase II (H), metaphase II (I), telophase II (J), round spermatid (K), and elongated spermatid (L). Amplifications at 300% magnification for the centrosomic CETN3 signals are presented for each image. Chromatin has been stained with DAPI (blue). Scale bar in L represents 10 μm.
Figure 5EGFP-Rab7a wild-type (wt), Q79L (constitutively active mutant), and T22N (dominant-negative mutant) were transiently expressed in Vero cells. The cells were then infected with SARS-CoV-2 at an MOI of ~0.3. Using flow cytometry, cell populations with levels of EGFP-Rab7a expression varying by roughly one-log increments were selected, and the proportion of infected cells within each population was quantified at 8 hpi. Data were normalized to that of the cell population with the lowest EGFP-Rab7a intensity. Unpaired t-test with Welch's correction was applied. *, p < 0.05; **, p < 0.01. n = 5-6 biological replicates. Vero and A549* cells were pretreated with colcemid (B) or MG-132 (C) at the indicated concentrations and subsequently infected with SARS-CoV-2 (MOIs ~0.3 and 0.2, respectively) in the continuous presence of inhibitors. Infection was analyzed by flow cytometry, and data were normalized to samples where inhibitors had been omitted. n = 3-4 biological replicates. Unpaired t-test with Welch's correction was applied. *, p < 0.05; **, p < 0.01; ****, p < 0.0001. SARS-CoV-2 particles were bound to A549* and Vero cells (MOIs of 0.2 and 0.3, respectively) on ice for 90 min and then switched rapidly to 37°C to allow infectious penetration. MG-132 (3.7 µM) was added to cells at the indicated times to block further late endosomal maturation. Infection was analyzed by flow cytometry, and data were normalized to that of control samples that had not been treated with MG-132. n = 2. As in panel B but using a MOI of 0.4 and Caco-2 cells instead of Vero cells. n = 6 biological replicates. As in panel C but using a MOI of 0.3 and 0.4 and Calu-3 and Caco-2 cells. n = 2-8 biological replicates. The timing of the MG-132-sensitive step during SARS-CoV-2 infectious entry into Calu-3 and Caco-2 cells was assayed as detailed in D but using 60 µM MG-132 for Caco-2 cells and MOIs of 0.5 and 0.6, respectively. n = 2. A549* cells expressing or lacking TMPRSS2 were pretreated with MG-132 at the indicated concentrations and subsequently infected with SARS-CoV-2 (MOI ~0.2) in the continuous presence of the drug. The proportion of infected cells was quantified by flow cytometry as described in Fig. 1D, and data were normalized to that from control samples for which MG-132 had been omitted. n = 6 biological replicates.
Comparison of APEX2:RAB21-, 5-, 4- and 7-generated proteomes.Figure 2(A) VENN diagram highlighting the number of interactors with each RAB and their overlap. (B) Reactome analysis of all APEX2:RAB21, APEX2:RAB5, APEX2:RAB4 and APEX2:RAB7 interactors/neighbors. Green, blue and beige shadings along with RAB names refer to previously identified interactions between RABs and the depicted proteins. (C) ProHits-viz generated dotblots of RAB effectors, GEFs, GAPs and sorting complexes. Green, blue and beige shadings along with RAB names refer to previously identified interactions between RABs and the depicted proteins.
A Noc‐Spo0J hybrid can recruit DNA to the membrane in E.Figure 7A, B Effects of Spo0J and Noc30‐Spo0J overproduction on cell division and nucleoid morphology in E. coli. Cells of strains DWA271 (PA1/04/03‐spo0J) (A) and 272 (PA1/04/03‐noc‐spo0J) (B) were grown in LB in the presence of 1 mM IPTG, to induce the expression of either Spo0J (A) or the Noc‐Spo0J hybrid (B), and were examined 1 h post‐induction. Cell membranes and DNA were stained with FM5-95 and DAPI, respectively. Scale bar, 5 μm.
Figure 7A - Histograms show flow cytometry analysis of exponentially growing cells of E. coli strain MC4100 relA-, and its mutant derivatives hfq-F42A (hfq-prox), hfq-Y25D (hfq-dist), hfq-R16S_R17A_R19A_K47E (hfq-rim), ∆hfq, harboring the translational fusion plasmid pDgcM::GFP in combination with pOmrA, pOmrB, or a control vector. Data was obtained on a MACSQuant VYB instrument. B - Northern blot analysis of total RNA isolated from the same cultures at the same time point as used for flow cytometry, probed for OmrA (strains containing pOmrA) or OmrB (strains containing pOmrB). 5S rRNA was used as loading control.
Figure 5Pulldown of biotinylated RNA probes of (A) repetitive GGACU sequences incubated with 4 pmol His-NT-Fmr1 and increasing concentrations of GST-Ythdf The indicated P-values were determined by unpaired, two-tailed Student's t-test. Pulldown of biotinylated RNA probes of (B) the respective quantification plotted as the median of the m6A/A signal intensity +/- SEM of three replicates. The indicated P-values were determined by unpaired, two-tailed Student's t-test. Pulldown of biotinylated RNA probes of (A) repetitive GGACU sequences incubated with 4 pmol His-NT-Fmr1 and increasing concentrations of GST-Ythdf and (C) AAACU sequences RNA probes incubated with either 32 pmol recombinant GST-Ythdf or/and 6xHis-NT-Fmr1. Pulled down proteins were analyzed by immunoblotting using α-Ythdf and α-Fmr1 antibodies. Pulldown of biotinylated RNA probes of (A) repetitive GGACU sequences incubated with 4 pmol His-NT-Fmr1 and increasing concentrations of GST-Ythdf and (D) EMSA assay using purified GST-Ythdf and His-NT-Fmr1 proteins and RNA probes in the AAACU sequence context. (E) Intersection of Fmr1 RNA immunoprecipitation target genes for wildtype and Ythdf KO brains, displaying a considerable number of genes gained and lost with Ythdf loss. (F) Intersection of genes lost (i.e., bound in wild type but not in Ythdf KO) with Ythdf target genes determined by a TRIBE assay. P-value calculated with hypergeometric test.
Figure 5A Epo-induced signaling upon AKT or MEK inhibitor treatment. Growth-factor deprived cells were pretreated for half an hour with AKT VIII or U0126, respectively, and subsequently stimulated with 5 U/ml Epo. PDI served as loading control. B Model calibration with integrated pAKT, ppERK and pS6 data upon inhibitor treatment and 5 U/ml Epo stimulation of mCFU-E, 32D-EpoR and BaF3-EpoR cells. Area under curve from time-resolved, quantitative immunoblotting data was calculated and is represented by filled circles. Error bars represent standard deviation estimated by an error model. Solid lines represent model trajectories. C Analysis of cell-cycle indicator genes upon AKT VIII and U0126 treatment. Growth-factor deprived cells were pretreated for half an hour indicated doses of a single inhibitor, followed by stimulation with 5 U/ml Epo for 0 h and 3 h. The expression of cyclinD2, cyclinG2 and p27 was measured by quantitative RT-PCR and normalized to the Rpl32 gene. Genes were selected based on microarray analysis. Experimental data is shown as fold change to unstimulated cells with mean ± standard deviation, N=3. Welch Modified Two-Sample t-Test, n.s. not significant, * p<0.05, ** p<0.01, *** p<0.005.
Fig.Figure 3A. Hypergeometric overlap analysis comparing genes deregulated in CamkIIδc TG mice to genes differentially expressed in the hippocampal CA1 region of Kat2A, Kmt2A and Kmt2b knock out mice. Fisher's hypergeometric test, Benjamini-Hochberg (BH) correction. B. Experimental scheme for ChIP-seq analysis. NeuN+ neuronal nuclei were FACS sorted and used to perform chromatin immunoprecipitation (ChIP) for H3K4me3 C. Pie chart showing the distribution of H3K4me3 peaks in the neurons of the hippocampal CA1 region from CamkIIδc TG mice. D. MA plot showing the number of significantly altered neuronal H3K4me3 peaks when comparing CamkIIδc TG and control mice. E. NGS plot showing H3K4me3 peaks at the TSS of genes down-regulated in the hippocampus of CamkIIδc TG mice. Inset shows statistical analysis, (unpaired t-test, two-tailed, ***P < 0.001). Bars and error bars indicate mean ± SEM. H3K4me3 ChIP analysis was performed on 4 replicates/group.
Time- and concentration-dependent stimulation of autophagy by rapamycin.Figure 1A-B) Western-blot analysis for GAPDH and LC3 of protein lysates obtained from HeLa cells treated with DMSO or 1 µM rapamycin (Rapa) for the indicated time periods (A) (n = 7) or for 5 h with the indicated concentrations (B) (n = 6). One hour before harvesting, 100 nM bafilomycin A1 was added. Upper panels: representative Western blots; lower panels: quantification of the LC3-II/GAPDH ratio. C) GFP-LC3-punctae quantification in HeLa cells treated for 5 h with different concentrations of rapamycin. Left: representative pictures. The scale bar represents 10 µm. Concentrations are mentioned in the right lower corner. Right: Quantification of the number of punctae per cell (n = 3). * p<0.05, repeated measurements ANOVA.
activitiesA.Figure 5A. Similar experiment as described in (Figure 4A) with naïve MCF7 and thermotolerant MCF7aTT cells as indicated. means +/- SEM, n = 3, * p<0.05, **p<0.01 (calculated with the two-stage linear step-up procedure of Benjamini, Krieger and Yekutieli). B. Same experiment as in (A) with thermotolerant MCF7aTT cells incubated with the HSP70 inhibitor VER-155008 (50 μM, 1h) or MG132 (20 μM, 1h). The signal obtained for ribosomal protein RPS19 and RPL7 are shown for reference, documenting residual polysomes despite the flat UV trace. Right panels are quantifications of K48-ub (means +/- SEM, n = 2), black vertical lines indicate splicing of lanes that were run on different gels due to limitations in lane capacity. The bar graphs represent the P/M ratios (means +/- SD, n = 3, numbers above bars indicate p values calculated with the two-stage linear step-up procedure of Benjamini, Krieger and Yekutieli). C. Sucrose density gradient fractions described in (B) were assayed for partitioning of HSPA1 and HSPH1 by immunoblotting. HSPA1 data were quantified in a line graph (means +/- SEM, n = 4). D D. Cell lysate was spun through a 30% sucrose cushion to pellet polysomes. Nascent proteins were released from polysomes by dissociating 80S ribosomes with EDTA (see Appendix Figures S3). Soluble and insoluble fractions were prepared by centrifugation and assayed for the level of K48-linked polyubiquitin. The levels of K48-linked polyubiquitin were quantified (means +/- SD, n = 3). Immunoblots within black squares were run on the same gel but irrelevant lanes were removed as indicated by white lines.
The apical α-βH Spectrin cytoskeleton binds to and co-localises with Expanded protein and acts genetically upstream or at the level of Expanded to control wing growth.Figure 3A Co-IP of V5-tagged Expanded with the FLAG-tagged N-terminal region of Karst/βH-Spectrin, but not other Karst truncation constructs. B, C Expanded co-localises with Karst at the apical membrane of wing imaginal disc cells (apical view, B; cross section, C). D Control nubbin.Gal4 expressing wing.E Overexpression of Ex activates Hippo signalling to produce a small wing.F RNAi knock-down of α-Spectrin results in an overgrown wing.G Overexpression of Ex blocks the effect of α-Spectrin RNAi.H RNAi knock-down of βH-spectrin/karst results in an overgrown wing.I Overexpression of Ex blocks the effect of βH-spectrin/karst RNAi.
Figure 8A, B. Representative images and analysis of AIF accumulation in the nuclei of HME control (vec) and OAS single-knockout cells (KO), and HME control (vec) and OAS double- and triple-knockout cells (DKOs and TKO), 24 h after treatment with 1000 μM H2O2. Data information: Nuclei were visualized by DAPI staining. Arrowheads: AIF translocated to nuclei. Untreated control cells are shown on the left. Right panels, AIF-positive nuclei were counted from pictures taken from three independent fields in two independent experiments. Means ±SD are plotted. HME: * p<0.005; ** p<0.001 compared to H2O2-treated control (vec) cells; C. Representative images and analysis of AIF accumulation in the nuclei of PC3 DKO2/3 and PC3 DKO2/3+1 cells, 24 h after treatment with 100, 200 or 400 μM H2O2. Data information: Nuclei were visualized by DAPI staining. Arrowheads: AIF translocated to nuclei. Untreated control cells are shown on the left. Right panels, AIF-positive nuclei were counted from pictures taken from three independent fields in two independent experiments. Means ±SD are plotted. PC3: *** p<0.0001 compared to untreated cells of each genotype as assessed by Student's t-test. Scale bars, 10μm.
Gene expression profiles between primary and orthotopic xenograft MPNSTs are similar.Figure 6Heat map showing the correlations between expression levels of genes in the molecular signature of MPNSTs. Hierarchical clustering of tumors and xenografts groups all primary tumors with their derived orthoxenografts. Moreover, sporadic tumors and NF1-related tumors form two different clusters. PCA of genes in the molecular signature of MPNSTs. All primary-xenograft pairs cluster together. The first component distinguishes between sporadic and NF1-related tumors; the second component differentiates primary tumors (and derived orthoxenografts) from the cell line (and the derived orthoxenograft).