EMBO/sd-character-level-ner huggingface dataset:
Aurora B inhibition affects DNA-dependent actin clearance during anaphase(A) Stills from time-lapse sequence of representative Hela cells expressing Lifeact-GFP and H2B-mCherry, exiting mitosis under different conditions i.Figure 5(A) Stills from time-lapse sequence of representative Hela cells expressing Lifeact-GFP and H2B-mCherry, exiting mitosis under different conditions i.e. Control siRNA, knock-down of centralspindlin protein RACGAP1, in the presence and absence of Aurora B kinase inhibitor (2µM ZM). Scale bar - 10 µm (B) High magnification view of boxed regions in (A) pseudo-colored showing failure to reduce actin during anaphase in cells treated with Aurora B inhibitor and even greater effect with Aurora B inhibitor upon RACGAP1 depletion. Scale bar - 1 µm. (C) Quantification of actin intensity at the poles of cells at 6 mins post anaphase onset, before furrow formation under different conditions as in (A). Actin fails to be cleared from the poles upon Aurora B kinase inhibition during mitotic exit and is stronger when both centralspindlin and Aurora B kinase activity are affected, while it is cleared upon RACGAP1 depletion. Data represented as mean±SD for treatments - siControl- 0.98±0.05, siRACGAP1 - 0.99±0.04, ZM - 1±0.039, siRACGAP1+ZM - 1±0.037. Ordinary one-way ANOVA with multiple comparison- siControl vs siRACGAP1 - n.s., p=0.896, siControl vs ZM - **p=0.0018, siControl vs siRACGAP1+ZM - ****p<0.0001 and ZM vs siRACGAP1+ZM - p=0.1528. (D) Schematic representation for quantifying actin intensity profile along the cortex as in E (E) Quantification of average actin intensity profile of siControl, siRACGAP1, ZM and siRACGAP1+ZM treated cells shown in (A). While actin levels reduced from the polar regions in both siControl and siRACGAP1 treated cells seen by levels lower than 1 (dashed red line) in segments 2-8, they failed to be cleared from ZM and siRACGAP1+ZM treatment conditions. Date represented as mean±SD . (F) Stills of representative Hela cells expressing Lifeact-GFP and H2B-mCherry captured at early anaphase under different conditions i.e. Control siRNA, knock-down of RACGAP1, in the presence and absence of Aurora B kinase inhibitor (2µM ZM). The masks below were generated for each cell and the shortest distance between the centroid of DNA (+) and the masked cortical actomyosin network (*) was quantified in (G) with Matlab. Scale bar - 10 µm (G) Quantification of shortest distance between centroid of DNA and cortical actomyosin network. While chromatin is able to reach close to the cortex in both siControl and siRACGAP1 treated cells, the distance of chromatin from cortex increases upon ZM treatment and is further enhanced in the double treatment condition. Data represented as mean±SD. siControl - 4.7±0.87, siRACGAP1 - 4.4±0.64, ZM - 5.47±0.87, siRACGAP1+ZM - 6.35±1.14 microns. Ordinary one-way ANOVA with multiple comparison, siControl vs ZM, ***p=0.002, siControl vs siRACGAP1, p=0.2823, siControl vs siRACGAP1+ZM, ****p<0.0001, siRACGAP1 vs siRACGAP1+ZM, ****p <0.0001, ZM vs siRACGAP1, ****p<0.0001, ZM vs siRACGAP1+ZM, ***p=0.0004 .
Figure 6Western blots of supernatants and cell lysates from THP-1 cells engineered by CRISPR/Cas9 to induce USP7 and/or USP47 KOs. Deficiency of both USP7 and USP47 was induced by doxycycline (Doxy) treatment (1 µg/mL; 3 days), as indicated. All cells were differentiated with PMA and either unprimed or LPS-primed (1 μg/mL, 4 hrs.) and treated with nigericin (10 μM, 45 mins.), as indicated. Bands in the figure represent: Pro-IL-1β (31 kDa); mature IL-1β (mIL-1β, 17 kDa); pro-caspase-1 (pro-Casp-1; 45 kDa); mature caspase-1 (mCasp-1; 20 kDa); USP7 and USP47. β-actin is shown as a loading control. Blots are representative of at least 3 independent experiments. IL-1β release measured from doxycycline treated (1 µg/mL; 3 days) PMA-differentiated and LPS-primed (1 μg/mL, 4 hrs.), nigericin activated (10 μM, 45 mins.) THP-1 cells, which contain the indicated sgRNA. Bars represent the mean percentage of IL-1β release relative to their respective non-doxycycline treated control cells, ± S.D. LDH-release measured from cells as treated in B. Bars represent the mean percentage of LDH release relative to their respective non-doxycycline treated control cells, ± S.D.
Figure 1(A) Domain organization of TRIM25 and TRIM32 including fragments used in this study. (B) SEC-MALLS traces of different TRIM25 and (C) TRIM32 constructs. The proteins are colour coded and the domain architecture is reported next to the respective MALLS curve. The constructs were analysed over different concentration ranges and the data are reported for: T25R, 4 mg/mL; T25RB1, 3.2 mg/mL; T25RB1B2, 3.6 mg/mL; T25RBCC, 0.5 mg/mL; T32R, 3 mg/mL; T32RB, 4 mg/mL; T32RBCC, 4 mg/mL.
Figure 1HCT116 and U87 cells were treated with 20 µM LXR623 for 24h. Transcriptome and gene set enrichment analysis was performed. Shown are enrichment plots. NES: normalized enrichment score. HCT116 cells were treated with 20 µM LXR623 for 24h. Transcriptome and gene set enrichment analysis was performed. Shown are enrichment plots. NES: normalized enrichment score. U87 GBM cells were treated with 20 µM LXR623 or 50 µM Simvastatin (Sim) for 48h. Thereafter, lysates were collected and analyzed for total cholesterol levels. Shown are means and SD (n = 3). Shown is a volcano plot of patients from the TCGA (database) with high vs. low levels of ABCA1. A close-up of the most significantly down-regulated genes is presented (mRNAs related to the electron transport chain and mitochondrial metabolism). The x-axis highlights the fold changes (log2 scale) (down-regulated genes, negative values), whereas the y-axis indicates the level of statistical significance (shown in -log10 format). GBM12 GBM cells (short term patient-derived xenograft), U87 GBM cells, HCT116 colonic carcinoma cells and A375 melanoma cells were treated with the indicated concentrations of ABT263, LXR623 or the combination for 72h. Thereafter, cellular viability was analyzed and statistical analysis was performed. , CI (Combination Index) value indicates whether the drug combination is additive (CI value = 1.0), synergistic (CI value < 1.0) or antagonistic. Normalized isobolograms are presented for the combination treatments in the various solid tumor cell lines.
Schematic of a mitochondria-controlled TMX-ROS-NFAT1 signaling axis in melanoma (Left panel) In TMXHIGH cells, the undisturbed ER-mitochondria contacts allow Ca2+ and lipid transfer and thus optimal mitochondrial function.Figure 8In TMXHIGH cells, the undisturbed ER-mitochondria contacts allow Ca2+ and lipid transfer and thus optimal mitochondrial function. Here, mitochondria and NOX4 generate no or low levels of ROS which can be eliminated by mitochondrial/cellular antioxidants. ORAI channel-governed SOCE activates NFAT1 translocation by its dephosphorylation through calcineurin and initiates the expression of target genes promoting melanoma proliferation and migration. NFAT1 controls the gene expression of mitochondrial proteins, thus suggesting a regulatory mitochondria-NFAT1 feedback loop.
Altered fuel preference in glycolytic muscle of 65-day-old asymptomatic SOD1G86R mice.Figure 4Relative mRNA levels of Pdk4 and Pdk2 evaluated by qPCR at the indicated ages in tibialis anterior of WT and SOD1G86R mice. Graphs represent mean fold change ± from age-matched WT. Pdk4: **P-values versus WT: 0.014 at 65 days and 0.018 at 105 days; Pdk2: ***P-value versus WT < 0.0001 at 105 days. n = 7 and 8 for WT and SOD1G86R, respectively, at 65 days; n = 5/genotype at 105 days, two-way ANOVA followed by Fisher's LSD post hoc test. Relative mRNA levels of genes involved in lipid handling evaluated by qPCR at the indicated ages in tibialis anterior of WT and SOD1G86R mice. Graphs represent mean fold change ± SEM from age-matched WT. P-values versus WT: Lpl **P = 0.0009, Cd36 *P = 0.051, Acsf2 ***P < 0.0001 at 65 days, **P = 0.0059 at 105 days and Cpt-1β **P = 0.01 at 65 days and *P = 0.02 at 105 days (n = 6 and 7 for WT and SOD1G86R, respectively, at 65 days; n = 10 and 8 for WT and SOD1G86R, respectively, at 105 days, two-way ANOVA followed by Fisher's LSD post hoc test). Relative levels of (left) ATP and (right) NADH and NAD+ measured in total tibialis anterior homogenates of WT and SOD1G86R mice. Left: Graphs represent mean fold change ± SEM in ATP from age-matched WT. **P = 0.002 (n = 9 and 12 for WT and SOD1G86R, respectively, at 65 days; n = 10/genotype at 105 days, two-way ANOVA followed by Fisher's LSD post hoc test). Right: The amounts of NADH and NAD+ relative to protein content in whole tibialis anterior homogenates are represented as mean ± SEM. *P = 0.027 and **P = 0.001 (n = 5, Student's t-test).
Targeting caspase-1/caspase-11 or IL1R1 rescues loss of visual function in aged VEGF-Ahyper mice.Figure 10A. Graphs show means of a- and b-waves of dark-adapted and light-adapted ERG responses (ERG intensities in μV) for both the left and the right eyes from 21-months old VEGF-Ahypermice, VEGF-Ahyper/CASP1-/-/CASP11-/-mice, CASP1-/-/CASP11-/-mice, VEGF-Ahyper/IL1R1-/-mice, IL1R1-/-mice, VEGF-Ahyper/IL18-/-mice, and IL18-/-mice. None of these mice had cataracts. Graphs show mean ± SEM. B. Representative ERG tracings are shown for VEGF-Ahyper mice and for VEGF-Ahyper/CASP1-/-/CASP11-/- mice. X-axes show milliseconds (ms). Y-axes show ERG intensities (in μV). C. For each experimental mouse group means for a- and b-waves of ERGs were calculated and p-values are shown for left and right eyes using a two-tailed unpaired Student's t-test.P-values of the VEGF-Ahypermouse group are compared to those VEGF-Ahypermice lacking either CASP1/CASP11, IL1R1 or IL18, as well as to the KO mice (CASP1-/-/CASP11-/-mice, IL1R1-/-mice, or IL18-/-mice) without the VEGF-Ahyper allele. In addition, p-values are shown comparing values between to KO mouse groups and those that have the VEGF-Ahyper allele. Mouse numbers for each experimental mouse group is indicated as well.
physically associates with miR-200s A.Figure 3A. The prediction for miR-200s binding sites in the LINC00115 transcript sequence. B and C. MS2-RIP followed by qRT-PCR analysis of endogenous microRNAs association with LINC00115. 12XMS2 empty vector, LINC00115-12XMS2, or LINC00115-mut- 12XMS2 with Flag-tagged MS2 were co-transfected into 1123 GSCs. Data information data are representative of three independent experiments. Error bars, ± SD. , ***, p < 0.001, by two-tailed t-test. D. Schematics of the predicted binding sites of miR-200s on LINC00115 and LINC00115 mutants. E. RNA pull-down and qRT-PCR assays of LINC00115 binding with miR-200s. GSC1123 cell lysates were incubated with biotin-labeled LINC00115. Data information data are representative of three independent experiments. Error bars, ± SD. p < 0.05, **, p < 0.01, by One-way ANOVA. F. Luciferase activity analysis of 1123 GSCs co-transfected with miR-200s and luciferase reporters containing LINC00115 WT, mutant, or empty vector (EV). Data information: data are representative of three independent experiments. Error bars, ± SD. p < 0.05, **, p < 0.01, by One-way ANOVA.
Figure 3Migration and invasion of TNBC cells transfected with LINC00665 ORF and control lentiviruses for 48h. Right panel is the quantification of TNBC cells that migrated through the membrane without the Matrigel (upper) and invaded through Matrigel-coated membrane (lower). Migration and invasion of wild-type and CIP2A-BP knockout TNBC cells and CIP2A-BP re-expressed Hs578TKO and MDA-MB-231KO cells. Right panel is the quantification of cells that migrated through the membrane without the Matrigel (upper) and invaded through Matrigel-coated membrane (lower). Wound-healing assay on Hs578T and MDA-MB-231 cells transfected with LINC00665 ORF expressing and control lentiviruses. The wound edges were photographed at the indicated time points after wounding. Right panel is the quantification of the relative wound healing area. Wild-type and CIP2A-BP knockout TNBC cells and CIP2A-BP re-expressed Hs578TKO and MDA-MB-231KO were seeded on cell culture inserts for wound-healing assay. The wound edges were photographed at the indicated time points after wounding. Right panel is the quantification of the relative wound healing area. Macroscopic and histological analysis of the lungs of nude mice injected with the indicated Hs578T (G) and MDA-MB-231 cells (H) (1.0×105-106 cells/mouse) via tail vein; lung metastatic nodules were visualized 8 weeks post-transplantation (5 mice per group). Right panel is the quantification of pulmonary metastases.
a,b).f7(a,b). Representative fluorescent images of HeLa cells co-tansfected with GFP-tagged Beclin 1 wild type, MutM L178A/L192A or MutStab E189L/A217L/E224L/A255L (green) and AsRed-Atg14L (red) (a) or FLAG-UVRAG (red) (b); Co-localization of AsRed-Atg14L with GFP-tagged Beclin 1 wild type or MutM is shown in discrete puncta. (c-e) Representative fluorescence images of HeLa cells co-transfected with Myc-Beclin 1 wild type, MutM L178A/L192A or MutStab E189L/A217L/E224L/A255L (blue), AsRed-Atg14L (red) and GFP-tagged Atg12 (c). The scale bar is 20 μM. (c-e) Representative fluorescence images of HeLa cells co-transfected with Myc-Beclin 1 wild type, MutM L178A/L192A or MutStab E189L/A217L/E224L/A255L (blue), AsRed-Atg14L (red) and GFP-tagged LC3 (d) and DFCP (e) (green). The scale bar is 20 μM. (f). A proposed model for the dynamic transition among homodimer (antiparallel) of Beclin 1, and the heterodimers of Beclin 1-Atg14L and Beclin 1-UVRAG. Multiple homo- and hetero-oligomeric forms of Beclin 1 protein exist intracellularly, and the nature of metastability in Beclin 1 CCD interface facilitates the transition of Beclin 1 homodimer to Beclin 1-Atg14L/UVRAG heterodimer that are compatible with activated autophagy-lysosomal degradation. The star shapes indicate the imperfect dimer interface. The thicker arrows pointing towards Beclin 1-UVRAG complex indicate its stronger affinity.
Human S100A9 expression restricts ex vivo MMLV replication and inhibits MMLV RTase activity in vitro(A) MDTF cells were transfected with an empty vector (pCDNA3.Figure 8MDTF cells were transfected with an empty vector (pCDNA3.1) or a vector expressing myc-tagged human S100A9 whose expression was analysed by Western-blotting with anti-myc 48h post-transfection. MDTF cells were then challenged with MMLV-YFP vectors (MOI 1) for 48h. Flow cytometry dot-plots from a representative experiment are shown. Human S100A9 expression in CHO cells was analysed by immunoblotting or flow cytometry as in Figure 7. E2C- and S100A9-expressing CHO cells were challenged with indicated MOI of MMLV-YFP vectors for 24h or 48h. The percentages of YFP+ CHO infected cells analysed by flow cytometry and data were represented on graphs.
Figure 5A, Fixed cells labelled with phalloidin shows filamentous actin accumulation on the edges of the pillars and along the top and bottom surfaces. Cross-sections along the direction of migration (yellow) and perpendicular to the direction of migration (blue, 1-4) are shown below. B, A confocal slice on the surface of the nucleus indicates that GFP-nesprin-2 co-localizes with actin cables at the surface of the nucleus. Dashed white lines represent the outlines of the constriction. C, Spinning disk confocal microscopy on live cells reveals the elongation state of nesprins at the nuclear envelope. Dashed white lines represent the outlines of the constriction. The distance between mCh-LAC and GFP-nesprin-2 was measured at the back, at the constriction, at the front near the constriction, and at the center of the front. (N = 17 cells, over 3 biological replicates. The alignment of the two channels was corrected using at least 28 fluorescent beads.) D, Two-photon laser ablation reveals that ablation at the front of the nucleus results in a rearward movement of the nucleus, but not ablation at the back. The nuclei of DN-KASH, which show a decrease of endogenous nesprin at the nuclear envelope show a dampened rearward movement upon ablation at the front of nuclei. Bars represent the average value. Error bars represent the standard error on the mean. Back: N= 9, Front: N= 30, Cell Front: N = 16, and DNKASH: N=31, 3 biological replicates.
Figure 6(A) (Top) Illustration of the reporter constructs. The black and red lines indicate H2AA3-SL and H2AA3-SLD PCR products, respectively. The dashed line indicates the FISH probe. (Bottom) FISH to detect the distribution of the corresponding mRNA at 2 hr after injection of H2AA3-SL and H2AA3-SLD PCR products. C and N indicate cytoplasmic and nuclear FISH signals, respectively. C/N ratios were determined for 20 cells in each experiment. Data information: Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, ***P< 0.001. (B) (Top) Illustration of the reporter constructs. The dashed line indicates the FISH probe. FISH to detect mRNA distribution at 24 hr after transfection of the H1C-SL-Rb (cH1C) and H1C-SLD-Rb (pH1C) constructs. C/N ratios were quantified Data information Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, ***P< 0.001. (Top) Illustration of the reporter constructs. PCR products of cG-SL and cG-SLD were used instead. cG, β-globin cDNA. Data information: Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, ***P< 0.001. H1C-SL-Rb or H1C-SLD-Rb construct, together with VSV M were co-transfected into HeLa cells, followed by IPs with IgG or the ALYREF antibody at 24 hr post-transfection. The immunoprecipitates were subjected to western analysis to detect ALYREF (D) H1C-SL-Rb or H1C-SLD-Rb construct, together with VSV M were co-transfected into HeLa cells, followed by IPs with IgG or the ALYREF antibody at 24 hr post-transfection. RT-qPCRs to detect RNAs (E). The bars in the graph shows the relative ALYREF IP efficiency, with that for the H1C-SL-Rb mRNA set as "1". Data information: Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, ***P< 0.001. (F) H1C-SLD-Rb construct with wild-type (H1C-SLDWT-Rb) or mutant (H1C-SLDmut-Rb) U7-binding sequence, together with VSV M, was co-transfected into HeLa cells, followed by IPs with IgG or the ALYREF antibody at 24 hr post-transfection. The immunoprecipitates were subjected to western analysis (left panel) and RT-qPCRs (right panel). The bars in the right graph show the relative ALYREF IP efficiency, with that for the H1C-SLDWT-Rb mRNA set as "1". Data information: Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, ***P< 0.001.
Limited impact of potential palmitoylation for shape maintenance A.Figure 5A. Cysteine 7 is predicted to be palmitoylated and was changed into alanine in the concavinC7A-GFP mutant. B. Expression and localization of concavinC7A-GFP in blood stage parasites. Scale bar 5 µm. C. Oocysts number in infected mosquitoes. Data points represent individual midguts observed between d12-17 post infection of 3 independent cage feeds. Shown is the mean ± SEM. P-value calculated using the Mann Whitney test. D. Quantification of concavinC7A-GFP cell shapes at the indicated days from midgut (MG) or salivary gland (SG) derived sporozoites. Numbers above bars indicate investigated sporozoites. E. Average speed of salivary gland sporozoites. Data points represent individual sporozoites from 3 independent biological replicates. Shown is the mean ± SEM. P-values are calculated using the Kruskal Wallis test followed by the Dunns multiple comparison test. F. Localization of concavinC7A-GFP in normal and deformed sporozoites (arrowheads point to deformations). Scale bar: 5 µm.
Figure 5(A) Lamin B2 pA-DamID z-scores of a representative locus in HAP-1 cells sorted for G1, mid-S and G2 phases. Arrows point to regions that lose NL interaction from G1 to G2. The red star indicates the end of the chromosome. Average of three biological replicates. difference in pA-DamID z-scores between G2 and G1 sorted pools as function of distance to either telomere or centromere for HAP-1 (n=3), HCT116 (n=2) and K562 cells (n=2). (C-E) LAD size distribution in basepairs (C), density of active genes (D) and Repli-seq scores (E) for stable and dynamic LADs. Repli-seq data are not available for HAP-1 cells. (F) Overlapping fractions of stable and dynamic LADs and iLADs with Hi-C subcompartments in HAP-1 and K562 cells, as imputed by SNIPER A permutation test with 1000 permutations (R-package regioneR was used to test for a significant enrichment of B1 subcompartments in decreasing LADs, resulting in P-values of 1e-3 and 1e-3 for HAP-1 and K562, respectively.
Figure 3A. RNA expression ratio (D2-to-D0) of candidates selected from day 2 microarray data. B. Timeline followed for proliferation examination of neonatal CMs in vitro. C. Ki67, α-MHC, and DAPI staining and quantification of CMs treated with control (Ctrl), or combinations of a FoxM1 up-regulation adenovirus (F), Id1 up-regulation adenovirus (I), Hmgb2 up-regulation adenovirus (H) or a small molecule Jnk3 inhibitor (Ji). D. Ki67, α-MHC, and DAPI staining and quantification of CMs treated with Ctrl or combination of FI with Jnk3-shRNAs in place of the small molecule (Ji). The arrow head indicates Ki67+/α-MHC+ proliferated CMs. E. Morphology and quantification of FIJs-treated CMs undergoing cytokinesis by H3P and Aurora kinase B (AURKB) staining. F. Timeline followed for adult reprogrammable mice administered the control or doxycycline in vivo. G. RNA expression ratio of Oct4, FoxM1, Id1, or Jnk3 in doxycycline-injected versus control adult CMs isolated from reprogrammable mice. H. Immunofluorescence of H3P and cTnI and quantification of the heart tissue sections from control or doxycycline injected mice.
In vivo activity curves of representative E.Figure 7Steady state levels of different metabolites obtained from metabolomics were used to derive activity curves for (A) Adenylate Kinase (AK) (B) Uridylate Kinase (PyrH) (C) Adenylosuccinate Lyase (PurB) and (D) GAR Transformylase (PurN). AK, PyrH and PurB appear to follow hyperbolic MM kinetics in vivo, while PurN shows Hill-like dependence with cooperativity. In panel (A), the red data points were acquired during growth in amino acid supplemented M9 medium in the absence of external AMP and are derived from WT and various stabilizing and destabilizing point mutations of AK (Adkar et al., 2019) as well as DHFR mutants used in the present study, while black points were derived exclusively from cultures of AK mutants supplemented with 1mM external AMP (Adkar et al., 2019). Data points for all other panels (B-D) are derived from mutant DHFR strains in the present study. Since SAICAR, AICAR, GAR and FGAR levels were undetectable in WT, all data points in (C-D) are relative to those observed in I91L+W133V mutant following 4 hours of growth at 40°C. Data points in all panels represent metabolite levels for individual biological replicates without averaging.