EMBO/sd-character-level-ner huggingface dataset:
Exogeneous ROS can restore mucin granule accumulation defects.Figure 7Exogeneous ROS can restore mucingranule accumulation defects. (A) Quantification of fluorescent DCF normalized to controls (n=3 independent experiments/group; 10-30 spheroids quantified/culture). *P0.05 as determined by ANOVA with Tukey's multiple post‐test comparisons (B) Images of control and Atg5VC colonic epithelial spheroids labelled with TRITC‐UEA (red) after treatment with either vehicle or 300 μM H2O2. Bars=20 μm. (C) Quantification of average mucin area/goblet cell (n=6 samples/group; 150 cells were quantified/sample). ***P0.001, ****P.0001 as determined by ANOVA with Tukey's multiple post‐test comparisons. (D) Relative expression of Muc2 mRNA in differentiated spheroids from control and Atg5VC mice treated with either vehicle or 300 μM H2O2 as measured by qRT-PCR. No statistically significant differences were found in either group as determined by ANOVA. (E, G) Images of p22 mut or Dynasore‐treated colonic epithelial spheroids (mucin is labelled with TRITC‐UEA; red) after treatment with either vehicle or 300 μM H2O2. Bars=20 μm. (F, H) Quantification of average mucin area/goblet cell (n=6 sample/group; 30-60 cells were quantified/sample) corresponding to experiments performed using p22 mut (E) and Dynasore‐treated wild‐type spheroids (G) Atg5VC and p22 mut spheroids were treated with 300 μM H2O2 plus or minus 100 μM BAPTA‐AM (I) or PMA/ionomycin (J). (I, J) Quantification of average mucin area/goblet cell (n=5 sample/group; 150 cells were quantified/sample). Error bars indicate s.e.m. ****P0.0001 as determined by the Student's t‐test (I) or ANOVA with Tukey's multiple post‐test comparisons (J).
Figure 3A) Representative MEA trace (left) showing the effect of increasing concentrations of LUF7346 on FP contour, measured in LQT2N996I-CMs. Average data (right) showing the effect of LUF7346 on QT interval duration relative to baseline in CMs derived from all the hiPSC lines used in this study. * = p<0.05 vs baseline. Colour in the heatmap defines the magnitude of QT shortening (blue) after treatment with increasing concentrations of LUF7346. N: 7-14. B) Representative AP from LQT2corr-, LQT2N996I-, LQT1corr-, and LQT1R190Q-CMs paced at 1 Hz, under baseline conditions (black) and after application of increasing concentrations of LUF7346 (colour-code is shown). C) Effect of LUF7346 on APD90, APD70, and APD50, measured in LQT2corr- (blue), LQT2N996I- (green), LQT1corr- (purple), LQT1R190Q- (dark blue) paced at 1 Hz; * = p<0.05 vs baseline. The colour of the asterisk indicates comparisons and respective statistical significance. ° = p<0.05 vs the genetically-matched corrected control. # = p<0.05 LQT1corr vs LQT2corr. N: 12-15. D) Effect of LUF7346 on APA and Ediast, measured in LQT2corr- (blue), LQT2N996I- (green), LQT1corr- (purple), and LQT1R190Q-CMs (dark blue) paced at 1 Hz. N: 12-15.
Figure 1(A) Normalized Hi-C contact maps showing contact frequencies between 10-kb bins across the Caulobacter genome during exponential growth (Le et al, 2013) and following nutrient starvation. Axes indicate genome position of each bin. (B-C) Hi-C contact maps for intra-chromosomal arm interactions (dashed box in panel A) rotated 45o clockwise. Images were resized in each dimension after rotation, but otherwise unprocessed relative to panel A. CID boundaries are marked with vertical purple lines and numbered. Directional preference plots were used to assign domain boundaries (see Materials and Methods and Appendix Fig S1B-C). Left- and rightward preferences are shown as green and red bars, respectively. Abrupt transitions from left- to rightward preference correspond to chromosomal interaction domain boundaries. Plots of gene expression 'rpkpm * transcript length' for each 10-kb bin along the chromosome are shown beneath the directional preference plots. Genomic locations of long, highly-expressed genes or operons (see Dataset EV2) are indicated by orange vertical lines. (D-G) Comparison of normalized Hi-C contact maps for regions of the Caulobacterchromosome during exponential growth and following starvation. The black dashed line shows the genomic location of (D) an rRNA locus, (E) a ribosomal protein gene cluster, (F) an ATP synthase gene cluster and (G) starvation-induced genes CCNA03325, CCNA03327, CCNA03328 and CCNA03337.
AGO6 incorporates 21–22nt siRNAsAGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.Figure 2AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.A A representative microRNA, miR158B. The location of miR158B from this locus is shown in orange. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.B The SimpleHat2 non‐autonomous TE targeted by Pol IV‐RdDM.C The RDR6‐RdDM target locus TAS3a. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.D The Athila6A consensus element in the wt Col epigenome.E The same Athila6A consensus element as in (D) upon transcriptional activation in the ddm1 epigenome. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.F The total small RNAs produced in the wt Col TE‐silenced epigenome.G The total small RNAs in ddm1 mutants. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.H Bisulfite sequencing of different regions of the Athila6A element. Error bars represent Wilson score interval 95% confidence limits. Differences in the methylation status were analyzed with a two‐tailed unpaired Student's t‐test. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.I A second biological replicate of the FLAG‐AGO6‐IP and no‐antigen control followed by deep sequencing shown for the Athila6A consensus element in ddm1 mutants. The transcriptional start sites of the LTR TSS‐ and IR‐driven transcripts are shown. AGO6 relative enrichment of small RNAs (solid lines) and total small RNAs (dashed lines) are shown for each small RNA size class (21, 22 and 24nt) at four genomic loci.J Analysis of the first base of AGO6‐enriched siRNAs from the wt Col TE‐silenced epigenome (left) and the ddm1 TE‐transcriptionally active epigenome (right).
Schematic model of GlaB mechanisms of actionSeveral tumors harbor a number of genetic/epigenetic changes that activate Hh signaling to induce the Gli1 transcriptional activity.Figure 9Several tumors harbor a number of genetic/epigenetic changes that activate Hh signaling to induce the Gli1 transcriptional activity. These mechanisms may be targeted by specific inhibitors (i.e. against Smo, BRD4, kinases, HDAC1/2). Gli1 suppression may be achieved by the common route of the interaction of Gli1 to DNA that is mediated by specific residues such as K350 localized in Gli1 ZF4. GlaB achieves the most profitable interactions on a groove sculpted by K340 residue on the surface of ZF4 as well as a direct binding to K350. In this way, GlaB would mask critical DNA docking sites and impair the Gli1/DNA interaction, thus blunting the expression of Gli1 target genes and repressing cellgrowth consequently.
Figure 1(A) Number of genes identified by MAJIQ that display 20% or more variation in splicing between wild-type and B52 depleted cells, in each dataset. We identified 108 shared alternative splicing events, corresponding to 105 genes. A comprehensive list of the 108 events is given in Appendix Table S1. (B) High confidence protein-protein complexes identified by MIST (Molecular Interaction Search Tool) within the 105 genes identified in (A). Interactions between proteins can be direct or indirect within the complexes. (C) GO term enrichment analysis of the 105 genes identified in a, performed by PANTHER with Bonferroni correction for multiple testing. (D) Schematic representation of yki locus and its two isoforms: Yki2 (includes exon 3 and contains 2 WW domains) and Yki1 (skips exon 3 and contains 1 WW domain). (E and F) RT-PCR and western blot showing the effect of RNAi-mediated B52 depletion in S2R+ cells (E) and in larval wing discs (F).
MVP-dependent escape from autophagy leads to increased Listeria survival.Figure 6A. InlK and ActA expression in Listeria strains used for survival assays. Total lysates of L. monocytogenes EGD-(pADc-GFP), EGD-(pADc-inlK), ΔactA-(pADc-GFP) and ΔactA-(pADc-inlK) grown in BHI were immunoblotted using anti-ActA and anti-InlK antibodies. B. Intracellular survival of EGD-(pADc-GFP), EGD-(pADc-inlK), ΔactA-(pADc-GFP) and ΔactA-(pADc-inlK) in RAW 267.4 macrophages. Statistical analyses were performed on the results of 3 independent experiments using the Student t test. P values of <0.05 were considered statistically different and are labeled here as *. C. Intracellular survival of ΔactA-(pADc-GFP) and ΔactA-(pADc-inlK) in MVP-GFP transfected Jeg3 cells. Statistical analyses were performed on the results of 3 independent experiments using the Student's t test. P values of <0.05 were considered statistically different and are labeled here as *. D. Intracellular survival of ΔactA-(pADc-GFP) and ΔactA-(pADc-inlK) in MVP-GFP transfected HeLa cells. Statistical analyses were performed on the results of 3 independent experiments using the Student's t test. P values of <0.05 were considered statistically different and are labeled here as *. E. MVP levels in RAW 267.4 macrophages treated with MVP-siRNA. Western blot is representative from 3 independent experiments. F. Intracellular survival of ΔactA-(pADc-GFP) and ΔactA-(pADc-inlK) in MVP knock-down RAW 267.4 macrophages. Statistical analyses were performed on the results of 3 independent experiments using the Student t test. P values of <0.05 were considered statistically different and are labeled here as *.
Figure 4a, Summary of the markers distinguishing each of the four stromal subpopulations identified in this study. b, FACS validation in matched patient tissue. Stromal cells are negatively gated for EPCAM (epithelial), CD45 (immune) and CD31 (endothelium) and positively selected for PDGFRβ. Subsequent markers PDGFRα and CD146 (MCAM) are used to distinguish CAFs and PVL cells, respectively. Expression of FAPHIGH, FAPLOW, CD36+ and CD36- are further used to define myofibroblast-like CAFs, inflammatory-like CAFs, immature-PVL cells and differentiated-PVL cells, respectively. c-d, Immunofluorescence of cultured human CAFs (c) and PVL cells (d) from passage 8, staining for CD34 (CAFs), ⍺-SMA (myCAFs and PVL cells), CD146 (PVL cells) and CD36 (imPVL cells). e-f, Quantitative analysis of collagen abundance (e) and orientation (f) using second harmonic generation (SHG) from cellular derived matrices from stromal subsets and representative images multiphoton SHG images (n = 3 biological replicates). All SHG intensity values within each replicate were normalised to the SHG intensity of the myCAFs. Error bars represent standard deviation. Statistical significance for collagen abundance (e) was determined using unpaired two-tailed Student's t test with equal standard deviation with p‐values denoted by asterisks: *p < 0.05, **p < 0.01 and ***p < 0.001. After normalization of the orientation peak distributions (f), statistical significant was determined using a Kruskal-Wallis test with Dunn's post-hoc multiple comparisons test (p value < 0.0001). , Immunohistochemical staining of PDGFRβ, ⍺-SMA, CD34 and CD146 in serial sections cut 4 μm apart from matched cases; Patient-2 (g) Images were aligned using FIJI. Tumour (T) regions are annotated by the solid yellow line. Co-localisation of CD34 and CD146 was used to distinguish blood vessels (BV), where their differential staining was used to identify CAFs and PVL cells. g, MyCAFs were found to be localised at the invasive stromal interface (insert A), whilst iCAFs were located at distal regions (insert B) with a high abundance of tumour infiltrating lymphocytes (TILs). Immunohistochemical staining of PDGFRβ, ⍺-SMA, CD34 and CD146 in serial sections cut 4 μm apart from matched cases; Patient-4 (h). Images were aligned using FIJI. Tumour (T) regions are annotated by the solid yellow line. Co-localisation of CD34 and CD146 was used to distinguish blood vessels (BV), where their differential staining was used to identify CAFs and PVL cells. h, Case with a high abundance of perivascular-like (PVL) cells in regions surrounded by blood vessels. i, Validation of disseminated PVL cells from blood vessels using co-immunofluorescence of CD31 (red), CD146 (green) and DAPI (blue). Representative images from Patient-4 is shown.
Figure 5A Mutation of either S947 alone or all 12 serine and threonine residues in the SET domain reduced O-GlcNAc modification level of ATX1ΔN. B Site mutation of S947 or all 12 serine and threonine residues in the ATX1 SET domain inhibited the activation of ATX1 by SEC; moreover, mutation of five conserved amino acids in the SEC functional domain inhibited SEC activity. C Quantitative analysis of HKMT activity of ATX1ΔN, ATX1ΔN-12m, and ATX1ΔN-m and the effect of SECΔN and ΔSEC5m in activating those proteins. Experiments were repeated three times. Error bars are SD. His-ΔSEC5m and His-SECΔN were used as the control.
Fig.Figure 3(A) Fluorescence micrographs of uninfected Caco-2 cells or Caco-2 cells infected with the indicated V. parahaemolyticus strains. Cells were stained with DAPI (blue) and anti-BrdU antibody (green). (B) Fluorescence micrographs of Caco-2 cells transfected with siRNA targeting EBP2 (siEBP2) or control siRNA (siNC) and subsequently infected with vopZ'ΔvscN1. Cells were stained with anti-BrdU antibody (green) and DAPI (blue). (C) The percentage of BrdU-positive cells (green) among the total cells (green + blue) was analyzed for 100 cells in three experiments. Error bars represent mean ± standard deviation (n=3 biologically independent experiments). One-way analysis of variance (ANOVA) was used for statistical analysis. *P<0.05 (when compared to the uninfected Caco-2 cells). (D) The percentage of BrdU+ cells was analyzed as above for cells treated with siNC or siEBP2. Error bars represent mean ± standard deviation (n=3 biologically independent experiments). Two-way ANOVA was used for statistical analysis. *P<0.05 (when compared to siNC-transfected cells that are infected with vopZ'ΔvscN1). (E) Immunoblot of EBP2 and Actin in Caco-2 cells transfected with siEBP2 or siNC.
Effect of Dyn A/B on induced epileptiform activity in human hippocampal slices.Figure 4: depicts representative traces under conditions of increased KCl. B: Same as A together with Dyn A/B and C: same as A under co-application of Dyn A/B with 5'-GNTI. D: Burst events and E: interictal spikes were recorded from area CA1. Treatment with Dyn A/B markedly reduced the amplitude and number of bursts G: but not of interictal spikes. Each color represents the last 5 minutes of each application for one slice of 4 individual patients. Data sets for each patient represent mean ± SEM for amplitude or absolute number of events.
Figure 2Treatment of HeLa cells with DFP (1mM for 24 h) decreases levels of CHIP (n=8, biological replicates; ***p<0.001; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) leads to decreased levels of CHIP mRNA (n=3, biological replicates; *p<0.05; unpaired t-test).   Treatment of DFP (1mM for 24 h)-treated HeLa cells with 50μM MG132 for the final 6 h does not recover reduced levels of CHIP induced by DFP. Histogram shows relative levels of cellular CHIP upon four different conditions (n=3, biological replicates; ****p<0.0001; *** p<0.001; repeated measures analysis of variance followed by Tukey's multiple comparisons test). CHIP knockdown increases levels of SENP3 but not SENP5 in HeLa cells. Nsi or CHIPi (Nsi, non-specific siRNA; CHIPi, CHIP siRNA; concentration, 20 nM) was transfected into HeLa cells for 48 h. Lysate samples were blotted as indicated.   CHIP knockdown prevents the DFP-induced increase in SENP3 levels. Nsi, SENP3i or CHIPi (Nsi, non-specific siRNA; SENP3i, SENP3 siRNA; CHIPi, CHIP siRNA; siRNA concentration, 20nM) was transfected into HeLa cells for 48 h. Two days post-transfection the cells were treated with DFP for a further 24 h.
Miro1 ubiquitination and degradation is required for mitochondrial clearance through mitophagy.Figure 3(A) Representative images at 3 hours of FCCP treatment (10 μM) of YFPParkin (green) expressing MiroDKO cells co-transfected with the specified myc-tagged versions of Miro1 (cyan). Tom20 (red) was used to reveal the mitochondrial network. A detail of YFPParkin translocation onto the mitochondrial network is also shown for each condition. (B and C) Quantification of the fraction of cells showing advanced stages of the mitophagic process (cells showing mitochondrial aggregation due to advanced Parkin translocation or the complete translocation of Parkin onto all the mitochondrial network) at 1 hour (B) or 3 hours (C) after FCCP treatment (data collected from at least 3 independent experiments: WT n=7, MiroDKO n=8, +mycMiro1WT=7, mycMiro15R=3, mycMiro1allR=3; mycMiro2=3; one way ANOVA with Dunnett post test). Error bars represent s.e.m. Significance: *p<0.05, **p<0.01 and ***p<0.001 D) Western blots and quantification of Miro1 degradation in MiroDKO cells co-expressing YFPParkin and the indicated myc-tagged Miro1 constructs and treated with FCCP (10 μM) for the indicated times (n=3 different experiments; 2-way-ANOVA with Dunnett post test; comparisons between Miro1WT and Miro1allR are represented with yellow asteriscs, comparisons between Miro1WT and Miro15R are represented in green and comparisons between Miro15R and Miro1allR are represented in black). E). Representative images at 24 hours of FCCP treatment (10 μM) of YFPParkin (green) expressing MiroDKO cells co-transfected with the specified myc-tagged versions of Miro1 (cyan). Tom20 (red) was used to reveal the mitochondrial network. Expression of both ubiquitin mutants (Miro15R and Miro1allR - arrowheads) protects from the mitophagic clearance of mitochondria (arrows) after damage. (F) Quantification of the fraction of cells showing the complete loss of Tom20 signal of mitochondria after 24 hours of FCCP treatment (data collected from at least 3 independent experiments: MiroDKO n=8, mycMiro1WT=7, mycMiro15R=3, mycMiro1allR=3; one way ANOVA with Dunnett post test). Error bars represent s.e.m. Significance: *p<0.05, **p<0.01 and ***p<0.001
Mutation of genes encoding RpoE (HI0628) and MclA (HI0629) has similar effects on biofilm formation and antibiotic tolerance in H.Figure 5Mutation of genes encoding RpoE (HI0628) and MclA (HI0629) has similar effects on biofilm formation and antibiotic tolerance in H. influenzaeBiofilms of different H. influenzae strains were developed after 24 h in μ-well chambers in sBHI medium and were then treated with the antibiotic azithromycin as specified.(i) H. influenzae wild-type; (ii) rpoE (HI0628); (iii) mclA (HI0629); (iv) H. influenzae wild-type treated with 150 μg/ml azithromycin; (v) rpoE (HI0628) treated with 150 μg/ml azithromycin; (vi) mclA (HI0629) treated with 150 μg/ml azithromycin. For these experiments, H. influenzae was visualized with SYTO9 (green strain), as described in Materials and Methods. Scale bars = 20 μm. Images shown are representative of 12 images from five independent experiments.The biofilm biomass after treatments was quantified using COMSTAT. Data are presented as the average of five replicates, with error bars representing the standard deviation of the data. Statistical significance by two-tailed Student's t-test is indicated: **P < 0.01.
Figure 9Figure 9. The self-association deficient SPOP mutant mutBTB has a substrate degradation defect in vivo. UAS transgenes were expressed under the control of an epithelial driver C765-Gal4 in Drosophila melanogaster. (A) UAS vector served as a control and yielded a normal wing. Longitudinal veins are denoted by numbers 1-5. Expression of (B) SPOP WT resulted in a modest Hh loss-of function phenotype with fusion of LV3 and LV4 (arrow). (C) mutBTB acts in a dominant-negative manner, as evidenced by a modest Hh gain-of function phenotype with expansion of the LV3-LV4 intervein space. Expression of (D) mutBACK and (E) mutBTB-BACK does not induce a wing patterning defect. ~50 animals each were analyzed from two independent crosses.
Figure 3A-C. Analysis of TCR nanoclustering by EM in OT-II WT and CCR5-/- naïve cells (A; n = 6 cells/genotype; WT: 3427, CCR5-/-: 3528 particles), and IL-2- (B; WT, n = 8 cells, 15419 particles; CCR5-/-, n = 6 cells, 5410 particles) or IL-15-expanded lymphoblasts (C; WT, n = 8 cells, 27518 particles; CCR5-/-, n = 7 cells, 22696 particles). A representative small field image at the top of each panel shows gold particle distribution in the cell surface replicas of anti-CD3ε-labeled cells; at bottom, quantification (mean ± SEM) of gold particles in clusters of indicated size in WT (gray bars) and CCR5-/- cells (red). Insets show the distribution of clusters of one, two, three, four, or more than four particles, and statistical analysis. D, E. Posterior distribution in naïve (D) and IL-2-expanded lymphoblasts (E) of the clustering parameter b for WT (gray) and CCR5-/- cells (red); randomly generated distributions of receptors are shown in blue. The mean value of the b parameter is indicated for each condition. The probability of a chance distribution similar to that determined in cells is nearly 0% by the ROPE. F. Comparison of TCR oligomer size using BN-PAGE and anti-CD3ζ immunoblotting in day 10, IL-2-expanded WT and CCR5-/- OT-II lymphoblasts lysed in buffer containing digitonin or Brij-96. The marker protein is ferritin (f1, 440 and f2, 880 kDa forms). The ratio of TCR nanoclusters to monomeric TCR in each lysis condition was quantified by densitometry (right; n = 5). G. Top, representative small field EM images showing gold particle distribution in the cell surface replicas of CD4+ T cells isolated from OVA/OVA-immunized WT and CCR5-/- mice. Bottom, quantification (mean ± SEM) of gold particles in clusters of the indicated size (WT, gray bars; n = 5 cells, 14680 particles; CCR5-/-, red; n = 7 cells, 15374 particles). Insets show the distribution between clusters of one, two, three, four or more than four particles, and statistical analysis.
K6 and K9 Are Primary Sites for HTTex1p In Vitro SUMOylation.Figure 1(A) Schematic illustration of the SUMOylation pathway. SUMO is expressed as a precursor protein (SUMO-X) that is processed by SUMO-specific proteases (SENPs) to expose a C-terminal diglycine motif (-GG). Enzymatic reactions are similar to ubiquitination and include activation by the SUMO E1-activating enzyme (SAE1/UBA2), to the SUMO E2-conjugating enzyme (Ubc9), and transfer of SUMO to target lysines with or without the assistance of the SUMO E3-ligating enzymes. (B) Time course of SUMO-1 modification of His-tagged HTTex1p (25Q)-purified proteins (WT, K6R, K9R, K6,9R, K6,9,15R, and S13,16D) was performed in vitro. SUMOylation was visualized using anti-His antibody. WT HTTex1p is SUMOylated within 16 min, K6R or K9R mutations delay SUMOylation, and combined mutations (K6,9R or K6,9,15R) greatly reduce SUMOylation. Mutations that mimic phosphorylation (S13,16D) alter kinetics of SUMO-1 modification with SUMO modification observed beginning by 4 min.