EMBO/sd-character-level-ner huggingface dataset:
Upstream factor identification through enhancer analysis using ChIL-seq.Figure 2(A) Tissue specificity of identified enhancers by H3K27ac ChIL-seq. The odds ratios of hits in the reference tissue-specific enhancer list identified by bulk-tissue ChIP-seq data(Shen et al, 2012) are shown. Odds is defined in the Materials and Methods. The cells enclosed by black squares indicate the maximal odds ratios (i.e., maximal specificity) for each row. (B) The IGV tracks of H3K27ac ChIL-seq at identified tissue-specific enhancers of Rxra, Gnat3, Eps8, and a house-keeping gene of Actb loci are shown with the replicates. (C) Specific motif enrichment analysis was conducted using chromVAR(Schep et al, 2017). Hierarchical clustering of deviation-Z scores of three replicates of each tissue is shown. (D) Super-enhancer identification. Tissue-specific enhancers are identified, so that they are listed more than twice (twice: blue, all: red) in the top 5% in all enhancer candidates and are not in the SEs of other tissues. Grey shades indicate the top 5% of tag count in enhancer candidates. (E) Immunofluorescent images of mouse liver sections. Tissues were stained with anti-Hnf4ɑ antibody and visualized by a fluorescent-labeled anti-mouse ChIL probe. DNA was counterstained with Hoechst 33342. Scale bar: 200 µm (top), 10 µm (bottom). (F) Hnf4ɑ binds to the SE at Alb gene loci. (G) Gene set enrichment analysis of Hnf4ɑ-bound genes (top), and their rate of Hnf4ɑ-bound genes in sliding windows of 100 genes (bottom).
Figure 3(A) Electron microscopy images of healthy, crushed, and 4-AP treated crushed sciatic nerve at 21 days post-injury. Scale bar at 3000x = 5m; scale bar at 8000x = 2m) (B) Comparison of the axonal area of randomly chosen individual axons of vehicle treated and 4-AP treated mice (n=4 for each experimental group; 40 axons analyzed per mouse). 4-AP treated sciatic nerve showed statistically greater axonal area compared to the vehicle-treated group (p<0.05; ANOVA; restricted-maximum-likelihood). Moreover, 4-AP treated mice had a greater proportion of axons with areas greater than the mean value for uninjured mice (shown in the green line), with inset figure displaying all axons with values above this mean. This experiment represents a single group of mice of one of three replicates on NCV recovery.
Figure 8Postulated organization of the amphipathic H3 helix in the post-fusion trimer. Left panel: proposed arrangement of the H3 helix in our post-fusion trimer model, lying flat on the fused membrane (in light orange) and projecting the TM segments (not drawn) away from the trimer axis. Second panel: potential conformation of the sE448r trimer, with the H3 helices trimerizing via their hydrophobic surfaces. Third panel: the sE-linker construct. As the linker is not long enough (only 8 residues), the H3 helix cannot trimerize as in the full-stem sE448r trimer. It would be exposed on the sides of the trimer, potentially leading to aggregation. Right-hand panel: the sE-linker* did not lead to aggregation, and at high concentrations it crystallized in a form that had the tip disordered. Trimerization of the three H3 helices in the trimer may have caused this effect, as they must displace the trimer tips in order to oligomerize. This is a further measure of the highly dynamic breathing behavior of the E trimers, as the sequence required to complete the stem zipper was absent from this construct. Schematic of the viral surface in the mature particle, with E and M in side view, color coded as in Fig 1. The breathing behavior within and between E dimers, transiently exposing the fusion loop and regions normally buried at the dimer-dimer interface, is indicated by curved lines. A vertical arrow points to the hinge between domains I and II, a region that supports E protein breathing. ii). Dissociation of E dimers in the acidic milieu of an endosome, exposing the FLs and allowing their insertion into the endosomal membrane. iii). Upon fusion loop insertion, the E proteins form "extended intermediate" trimers bridging the two membranes. Curved blue arrows indicate the subsequent re-location of domain III, in blue, to the sides of the trimer to initiate the "hairpin" conformation of the individual subunits of the post-fusion trimer. iv). Upon relocation of domain III, the upstream part of the stem becomes anchored to the the
Figure 2(A) 0.25 μg of purified long mouse DNGR-1 ECD was diluted into 10 mM Tris buffer of the indicated pH (left panel) or into 10 mM Tris pH 7.4 buffer supplemented with increasing amounts of NaCl (1 - 250 mM; right panel), and reduction sensitivity of DNGR-1 was assessed by reducing SDS-PAGE and Western blot. (B) 0.25 μg of purified DNGR-1 long mouse ECD was diluted into buffers of different pH and ionic strength, and its reduction sensitivity was assessed by reducing SDS-PAGE and Western blot. The intensity of bands corresponding to dimer and monomer was determined densitometrically, and the ratio was plotted as a function of buffer ionic strength and pH. (C) 0.25 μg of purified long mouse DNGR-1 ECD was diluted into PBS or 10 mM MES pH 6.1 buffer, and its reduction sensitivity was tested under mildly (Laemmli buffer) or strongly denaturing (8 M urea) conditions by reducing SDS-PAGE and Western blot. (D) 0.25 μg of purified long mouse DNGR-1 ECD was diluted into PBS or 10 mM MES pH 6.1 buffers, and its reduction sensitivity after different lengths of heat-denaturation in Laemmli buffer was tested by reducing SDS-PAGE and Western blot. HRP-conjugated anti-FLAG antibody was used for detection of all proteins. Numbers on the side of blots indicate positions of molecular weight markers. Reduction-resistant dimers are indicated with an * and reduction-sensitive protein with a +.
Impact of the BH3‐only protein Bad on autophagy.Figure 7(A, B) Interactions between Bcl‐XL, Beclin‐1 and Bad in conditions of autophagy induction. Cells were either treated by nutrient depletion (A) or addition of 1 μM rapamycin, 1 mM lithium chloride, 100 μM L‐690,330 or 50 μM carbamazepine (B), followed by immunprecipitation of Bcl‐XL (as in Figure 2C) and revealing the immunoblots by antibodies specific for Bcl‐XL, Beclin‐1 or Bad. (C) Interaction between endogenous Bad and Bcl‐2. HeLa cells were treated with ABT737 (1 μM) or nutrient‐depleted, followed by immunoprecipitation of Bcl‐2 and immunodetection of Bad. (D, E) Impact of Bad depletion on autophagy. Cells were transfected with LC3‐GFP together with siRNAs specific for Bad, siRNAs specific for emerin or the Beclin‐1‐associated PI3 kinase Vps34. Forty‐eight hours later, when the siRNAs had down‐regulated the proteins of interest (D), cells were subjected by nutrient depletion (NF) or addition of rapamycin (E) and the frequency of cells exhibiting LC3‐GFP aggregation as a marker of autophagy was assessed after 18 h (means±s.d., *P0.05, n=3 separate experiments). (F) Effect of the Bad knock‐out on autophagy. Wild Type (WT) or Bad−/− MEF were transfected with LC3‐GFP and then subjected to nutrient depletion (NF) and/or treatment with ABT737 (18h; means±s.d., n=3). The asterisks denotes a significant (P0.05) effect of Bad deficiency. (G) Effect of the Bad knockout on the Beclin‐1/Bcl‐2 interaction. WT or Bad−/− MEF were subjected to the indicated treatments, followed by immunoprecipitation of Bcl‐2 and immunobloting of Beclin‐1. (H) Overexpression of Bad triggers autophagy. Cells were transfected with LC3‐GFP alone (Control), together with vector‐only or with a vector encoding Bad, and subjected 48 h later to nutrient depletion (NF). Cells were cultured in the continuous presence or absence of the pan‐caspase inhibitor Z‐VAD‐fmk (50 μM) for 16 h, followed by assessment of autophagy as in (E) (means±s.d., n=3 separate experiments, *P0.01). Immunoblots are representative of at least three independent experiments.
signalling and ILK-controlled lymphatic vascular growth.Figure 9A In quiescent LECs, VEGFR3 and β1 integrin are physically separated. ILK directly or indirectly interacts with β1 integrin and connects it to the F-actin cytoskeleton via intracellular proteins, such as α-parvin, a component of the IPP complex. B Upon mechanical stretch, the complex of β1 integrin and ILK (along with the entire IPP complex) transiently disrupts. C This releases β1 integrin, resulting in its interaction with VEGFR3, and thus in increased VEGFR3 tyrosine phosphorylation ("P" in yellow circle). As a consequence, LEC proliferation and lymphatic vascular growth are induced. D The absence of ILK results in permanent interaction between VEGFR3 and β1 integrin, leading to upregulated VEGFR3 tyrosine phosphorylation ("P" in yellow circle), LEC proliferation, and non-physiologic lymph
Figure 2(A) Levels of BRCA1 promoter hypermethylation, levels BRCA1 mRNA and the presence of BRCA1 nuclear foci by immunofluorescence are shown T127 and T162 were used as positive controls for hypermethylated BRCA1 promoter. Error bars indicate SEM from independent tumors (n ≥ 2). Dashed line indicates mean of BRCA1 mRNA levels in normal breast. PARPi-response is shown in the summary underneath: white box: PD; black box: PR/CR. Alterations in HRR-related genes in PDX are also indicated. (B) Western blot of PALB2 detected in U2OS cells and PDXs. Three biological replicates of PDX093 are shown; PDX302 is used as PALB2 wild type PDX control. (C) YFP-PALB2 recruitment to laser-induced DSBs is impaired in HeLa cells expressing PALB2 p.M296Nfs (n=4, unpaired t-test at 16min). Gene-targeting efficiency using Cas9/mClover-LMNA1 homologous recombination assay of (D) siRNA PALB2 cells (n=4, one-way ANOVA) or (E) cells with no PALB2 depletion complemented with wild type and p.M296Nfs siRNA-resistant constructs (n=7, one-way ANOVA). Western blots of PALB2 wild type and PALB2 p.M296Nfs for each condition are shown. Error bars indicate SEM from independent experiments.
PER2::LUC stability oscillates in CRY-deficient cells(A) Actual PER2::LUC levels (dark symbols (3-hour moving average, n=3 ±SEM, 4 outliers removed)) as assayed in acute luciferase assays on cell lysates from cells harvested every hour over 48 hours, compared with parallel longitudinal co-recordings from cells in the presence of 0.Figure 4(A) Actual PER2::LUC levels (dark symbols (3-hour moving average, n=3 ±SEM, 4 outliers removed)) as assayed in acute luciferase assays on cell lysates from cells harvested every hour over 48 hours, compared with parallel longitudinal co-recordings from cells in the presence of 0.1 mM luciferin (light lines (n=6, mean ±SEM)). (B) PER2::LUC recording of asynchronous WT and CKO cells pulsed with proteasome inhibitor MG132 (10 µM, applied at the arrow) (n=3, mean ±SEM). (C) Quantification of relative PER2::LUC induction upon proteasome inhibition, n=3, mean (solid) ±SEM (dashed). (D) Phase-dependent PER2::LUC half-life was determined by inhibiting translation at different circadian phases and fitting the resulting data with a one-phase exponential decay curve (n=3, mean ±SEM). Left image depicts the timing of cycloheximide (CHX, 10 µM) pulses (labelled I (PER2 levels going up) and II (PER2 levels going down)), plotted on PER2::LUC bioluminescence traces of control cells (dark colours). A representative trace of CHX-treated cells at time point I is shown in light colours. Right image shows quantifications, p-values were calculated by unpaired t test. (E) A simple model incorporating mRNA, protein translation and PER2::LUC stability that were measured experimentally (inputs) shows that the observed oscillating stability of PER2 is sufficient to generate rhythmic PER2::LUC expression (output).
Figure 6(A) Standard replication assay on undamaged (UD), 8-oxoguanine (8oxoG), and thymine glycol (Tg) templates in the absence of TLS components. (B) Pulse chase reaction on UD and 8oxoG templates. Where present, Pol δcat (10 nM) was added directly after addition of the chase. (C) Pulse chase reactions on the 8oxoG template in the absence and presence of 10 nM Pol δ analysed on a denaturing gel.
A network perspective on adaptive responses.Figure 4A Left: A consolidated overview of PLSR-derived variable importance in the projection (VIP) scores mapped onto a simple schematic of immediate-early signaling, indicating the fraction of 10 studied cell lines in which early (1-10 h) or late (24-48 h) up- or down-regulation of each of the cell state markers and phospho-proteins is predictive of response to RAF/MEK inhibitors (72-h viability). Up-regulation or down-regulation of signals with VIP score >1 is shown. Right: Correlation between up- and down-regulation of selected pathways in response to RAF/MEK inhibition across the 10 studied cell lines. Correlations between signaling changes in response to RAF/MEK inhibition were evaluated based on pairwise Spearman's correlation between VIP scores for selected phospho-proteins at 24 and 48 h post-treatment across the 10 studied cell lines. P-values <0.05 were considered significant. Time-dose-response plots indicating changes in RPPA measurements for p27 versus p-histone H3 for two selected cell lines (C32 and WM115) after exposure to PLX4720. Mean values of four biological replicates are shown. Protein levels represent log2 fold change of each signal (at a specific dose and time) relative to a DMSO-treated control. B-D Time-dose-response plots indicating changes in RPPA measurements for pERK(Thr202/Tyr204) versus pAKT(Ser473) for two selected cell lines (C32 and WM115) after exposure to PLX4720. Mean values of four biological replicates are shown. Protein levels represent log2 fold change of each signal (at a specific dose and time) relative to a DMSO-treated control. Time-dose-response plots indicating changes in RPPA measurements for pS6(Ser235/236) versus total c-Jun levels for two selected cell lines (C32 and WM115) after exposure to PLX4720. Mean values of four biological replicates are shown. Protein levels represent log2 fold change of each signal (at a specific dose and time) relative to a DMSO-treated control.
Fungi were harvested from the stomachs of C57BL/6 (a–c), IL-17F-deficient (d–f) or IL-17A-deficient (g–i) mice infected with GFP–Candida for 2 days, and stained using PE-labeled antibody to IL-17A.f5Fungi were harvested from the stomachs of C57BL/6 (a-c), IL-17F-deficient (d-f) or IL-17A-deficient (g-i) mice infected with GFP-Candida for 2 days, and stained using PE-labeled antibody to IL-17A. Cells were green in (a,d and g), red upon staining with anti-IL17A-PE (b,e and h) and double stained in the merged images (c,f and i). Scale bars, 10 μm. Images are from one representative experiment out of two. (j-l) SEM images showing fungal germination in the stomachs of C57BL/6 mice (j), filamentation in IL-17F-deficient mice (k) and neither one in IL-17A-deficient (l) mice intragastrically infected 2 days before. Scale bars, 10 μm. (a-l) Images are from one representative experiment out of two. (m-o) C57BL/6 mice were infected intragastrically with C. albicans MKY 378 strain (m) or the crh11Δ (n) and crh12Δ (o) mutants, and were monitored for fungal growth (log10 colony-forming units (CFU) at 2 (open circle) or 8 (black circle) days post infection). Data are mean±s.d. (n=3). The actual P-values are indicated (two-tailed t-test) for 2 and 8 days (m) and for 2 days (o). Fungal cells were untreated (none) or exposed to 10 ng ml−1 IL-17A for 4 h before injection.
WOX5 positively regulates PLT3 expression.Figure 1A, Schematic representation of the Arabidopsis root meristem. The QC cells (red) maintain the surrounding stem cells (initials) outlined in black together building the root stem cell niche (SCN). The different cell types are colour coded. QC = quiescent center (red); CSC = columella stem cells (yellow); CC = columella cells (green); LRC = lateral root cap (light purple); ep = epidermis (purple); c = cortex (light turquoise); en = endodermis (dark turquoise); bright turquoise = cortex/endodermis initials; dark purple = epidermis/lateral root cap initals; dark orange = stele initials; stele = light orange; grey dots = starch granules. SCN = stem cell niche B, C, Representative images of pPLT3::erCFP (cyan) expressing and PI-stained (red) Arabidopsis roots in Col or wox5 background, respectively. D, Mean fluorescence intensities of the pPLT3::erCFP roots summarized in box and scatter plots. The mean fluorescence intensity of the CFP signal in Col roots was to set to 100 %. D, Box = 25-75 % of percentile, whisker = 1.5 interquartile range, − = median, □ = mean value, = minimum/maximum. The data was statistically analyzed by one-way ANOVA and Holm-Sidak post-hoc multiple comparisons test. Asterisks indicate statistically significant differences (α = 0.01). Number of analysed roots (n) (biological replicates) is indicated for each genotype and results from two technical replicates. B, C, Scale bars represent 10 µm. PI = propidium iodide CFP = cyan fluorescent protein. E, F, Representative images of pPLT3::PLT3-YFP (yellow) expressing and FM4-64-stained (red) Arabidopsis roots in Col or wox5 mutant background, respectively. g, Mean fluorescence intensities of the pPLT3::PLT3-YFP expressing roots summarized in box and scatter plots. The mean fluorescence intensity of the YFP signal in Col roots was to set to 100%. G, Box = 25-75 % of percentile, whisker = 1.5 interquartile range, − = median, □ = mean value, = minimum/maximum. The data was statistically analyzed by one-way ANOVA and Holm-Sidak post-hoc multiple comparisons test. Asterisks indicate statistically significant differences (α = 0.01). Number of analysed roots (n) (biological replicates) is indicated for each genotype and results from two technical replicates. , E, F, Scale bars represent 10 µm. YFP = yellow fluorescent protein
Figure 2Cells were fixed and the viral bait fused with the V5-tag was visualized by immunofluorescence staining using Alexa Fluor-488 labeled anti-V5 immunostaining (yellow), actin with Alexa Fluor594-labeled phalloidin (magenta), and nuclei were stained with DAPI (cyan) (Scale bar: 10 μm).
Figure 6KGN cells were transfected with 200 nM of scrambled control or FOXL2-specific siRNAs for 24 h. Then, KGN cells were further transfected with 20 nM of control miRNA, miR-1236, control anti-miRNA, or anti-miR-1236. The proportion of annexin-V-positive apoptotic cells (A and B) were analyzed by flow cytometry. The data are presented as the mean ± SEM of three independent experiments. Different letters denote statistically significant differences (p < 0.0001; Student-Newman-Keuls test). KGN cells were transfected with 200 nM of scrambled control or FOXL2-specific siRNAs for 24 h. Then, KGN cells were further transfected with 20 nM of control miRNA, miR-1236, control anti-miRNA, or anti-miR-1236. The population at S phase (C and D) were analyzed by flow cytometry. The data are presented as the mean ± SEM of three independent experiments. Different letters denote statistically significant differences (p < 0.0001; Student-Newman-Keuls test). Efficient silencing of FOXL2 using specific siRNAs was confirmed by western blotting. E, F KGN cells were transfected with control siRNA or siFOXL2 for 24 h. Then, KGN cells were further transfected with control miRNA or miR-1236 (E) or with control anti-miRNA or anti-miR-1236 (F) for 48 h, and Transwell-migration assays were performed. The migrated cells were imaged under a bright-field microscope (100 × magnification, scale bar = 100 µm). The results are from three independent experiments and represent fold-changes in the average number of cells/field (mean ± SEM). Different letters denote statistically significant differences (p < 0.01; Student-Newman-Keuls test). G, H COV434 cells were transfected with miR-control or miR-1236 for 48 h, after which cell viabilities (G) and migration abilities (H) were measured. The data are presented as the mean ± SEM of three independent experiments. Immunoblots showing no change in FOXL2 protein are presented in the top panel, and images of migrated cells are presented in the right panel. The migrated cells were imaged under a bright-field microscope (100 × magnification, scale bar = 100 µm). I-K The properties of miR-1236-/+ and miR-1236-/- KGN cells versus control KGN cells were assessed by measuring cell viability (I), cell proliferation (J), and cell migration (K). The data are presented as the mean ± SEM of three independent experiments. The migrated cells were imaged under a bright-field microscope (100 × magnification; top of Fig. 6K). Scale bar = 100 µm. L No difference in the cell-migration activities of control and two independent miR-1236-/- (#1 and #2) COV434 cell lines. The migrated cells were imaged under a bright-field microscope (100× magnification, scale bar = 100 µm; top) and the results (bottom) represent fold-changes in the average number of cells/field. The data are presented as the mean ± SEM of three independent experiments.
PTX3 activity is inhibited by thrombospondin 1 through direct interaction.Figure 5A) PTX3 binds TSP1 and TSP-2 but not TSP-4. Different amounts of human recombinant PTX3 were incubated in microplate wells coated with purified human TSP1 or recombinant TSP-2 and TSP-4. Binding is reported as Absorbance at 450 nm (Mean±SD). Data are from one experiment out of three performed B) N-terminal PTX3 binds TSP1. Binding of PTX3 N-term domain was performed on TSP1 immobilized on plastic wells. Data are reported as Absorbance at 450 nm (Mean±SD) and are representative of one out of two experiments performed C) Schematic representation of TSP1 monomer and the TSP1 proteolytic fragments (N-term, C-term) and recombinant domains (P123-1, E123-1, E123CaG-1, Ca-1) used in the study D) PTX3 binds the C-term proteolytic fragment of TSP1. TSP1 and its fragments C-term, N-term, and Ca-1 (type III repeats) were immobilized in plastic wells (5µg/ml) and binding of PTX3 is reported as Absorbance at 450 nm (Mean±SD). Data refers to one out of two experiments performed with similar results E) PTX3 and its N-terminal domain bind TSP1 C-terminal globular domain. 50 nM of P123-1 (type I 'properdin' repeats), E123-1 (Type II EGF repeats), E123CaG-1 (type II repeats plus type III repeats and globular C terminus) and TSP1 were immobilized in plastic well. Binding with PTX3 or N-terminal domain (both at 220 nM) was analyzed. Data are reported as Absorbance at 450 nm (Mean±SD) and refers to one out of two experiments performed with similar results F) Representative images of 14DIV control and PTX3-treated cultures stained for the presynaptic marker bassoon (blue) and the microtubule protein tubulin (red). Scale bar: 10μm. G) Quantification of synaptic density (Bsn/μm) in the different experimental conditions (Ctr=0.385±0.029, PTX3=0.407±0.032, TSP1=0.721±0.039, PTX3+TSP1=0.707±0.037, E123=0.764±0.047, PTX3+E123=0.719±0.041. Number of fields examined: 86, 80, 75, 72, 46, 55 respectively; Kruskall-Wallis test, p<0.0001 followed by Dunn"s test as indicated in figure; at least 3 independent experiments, data are presented as mean values H) Representative images showing 14DIV neurons stained for surface AMPAR (GluA, green), the presynaptic protein Bassoon (blue) and tubulin (red) in the different tested conditions. Arrowheads point to postsynaptic GluA clusters. Scale bar: 5µm. I) Quantification of the synaptic surface GluA density (number of sGluA puncta/μm) in the different tested conditions showing that GluA density increases in neuronal cultures treated with PTX3 or E123+PTX3 but not in cultures treated with the full length TSP1+PTX3 (Ctr=1.000±0.055; PTX3=1.367±0.074; TSP1=1.04±0.061; PTX3+TSP1=0.968±0.076; E123=0.971±0.063; PTX3+E123=1.613±0.077. Number of dendrites examined: 103, 80, 88, 57, 51, 62 respectively; Kruskall-Wallis test, p<0.0001 followed by Dunn's test as indicated in figure; at least 3 independent experiments, data are presented as normalized mean values±SEM).