EMBO/sd-character-level-ner huggingface dataset:
Figure 3(A) Crystal structures of ZIKV xrRNA1 (Left, PDB code: 5TPY) and MVEV xrRNA2 (Right, PDB code: 4PQV). (B) Two views of the overlay of ZIKV xrRNA1 and MVEV xrRNA2 crystal structures. (C, D) The scattering profiles (left), the PDDFs (middle) and the dimensionless Kratky plots (right) of individual xrRNA1s (C) and xrRNA2s (D) of DENV2 (cyan), ZIKV (red), WNV (blue) and MVEV (green). The insets in (C) and (D) show the Guinier regions of the respective scattering profiles with a linear fit line. The back-calculated scattering profile of ZIKV- xrRNA1 crystal structure (black, open circle) can be nicely fitted onto its experimental SAXS data (red line) in (C), while the back-calculated scattering profile of MVEV- xrRNA2 crystal structure (black, open circle) fits poorly to its experimental SAXS data (green line) in (D). The asymmetric PDDFs of xrRNA1 (C) and xrRNA2 (D), indicate slightly elongated molecules with asymmetric shapes. The dimensionless Kratky plots of xrRNA1 (C) and xrRNA2 (D) of DENV2, ZIKV, WNV and MVEV suggest all individual xrRNA1s and xrRNA2s are well-folded and of reduced flexibility in solution. (E, F) The ab initio shape envelopes of the individual xrRNA1s (E) and xrRNA2s (F) of DENV2 (cyan), ZIKV (red), WNV (blue) and MVEV (green) are fitted with the homologous atomic models built by ModeRNA.
Atg38 is required for the localization of PI3-kinase and Atg18 to the PAS.Figure 3.(A) Wild-type and atg38Δ cells expressing the indicated GFP fusion proteins and Ape1-mRFP were cultured at 30°C in the presence of rapamycin. After 60 min, cells were analyzed by fluorescence microscopy. Of the total pool of Ape1-mRFP signal, the percentage of Ape1-mRFP colocalizing with GFP was calculated. This percentage is shown as mean ± SD of two independent experiments, with the number of cells counted indicated at the bottom of the graph. (B) Examples of wild-type and atg38Δ cells expressing either Atg14-3 × GFP or Vps34-3 × GFP in A. Bar, 5 µm. (C) Cell extracts were prepared from the cells in A, then analyzed by immunoblotting with anti-GFP and anti-Pgk1 antibodies.
Figure 2A. Single-cell traces of hESCs before and after treatment with 100 ng/mL BMP4. Cells were imaged for 42 h prior to BMP4 treatment. Mean nuclear OCT4 levels were quantified every 5 minutes and individual cells were tracked from the cell division event that created the cell until its own division. B. 24 h after BMP4 treatment, cells were fixed, stained for expression of CDX2, and returned to the microscope for registration with the final time-lapse image. Mean nuclear OCT4-mCherry and CDX2 were quantified for each cell, and the resulting distribution was fit to a 2-component mixed Gaussian distribution representing self-renewing (OCT4+/CDX2-) and differentiated (OCT4-/CDX2+) cells hESCs that could be assigned to either distribution with >99% confidence (red and green dots, Pdiff < 0.01 or Pdiff > 0.99) were considered for pro-fate analysis. Cells that did not reach this threshold (gray dots) were not used to determine pro-fate. Because cells in the final frame were assigned to only self-renewing or differentiated categories, pro-mixed cells C. Distributions of OCT4 levels in pro-self-renewing, pro-mixed, and pro-differentiated cell populations. D. Distributions of OCT4 pulse frequencies in pro-pluripotent, pro-mixed, and pro-differentiated cell populations. To gain an unbiased look at preexisting determinants of cell fate in panels only cells who completed their entire cell cycle duration before BMP4 addition (t = 0) were included in the analysis. E. Distributions of cell cycle durations in pro-pluripotent, pro-mixed, and pro-differentiated cell populations. To gain an unbiased look at preexisting determinants of cell fate in panels , only cells who completed their entire cell cycle duration before BMP4 addition (t = 0) were included in the analysis. F. Decision surfaces for logistic regression classifier. Cells with self-renewing (red) or differentiated (green) pro-fates are plotted above the predicted class surfaces. (left) Slice of decision surface for pulse frequency vs. OCT4 levels at a cell cycle duration of 14 h. (right) Slice of decision surface for cell cycle duration vs. OCT4 level at a pulse frequency of 0.25 h-1. * P < 0.05, ** P < 0.0005; ns, not significant.
Figure 2(a) Kymograph of a single DNA-bound Scc1Halo-TMR-TEVcohesin complex after 750 mM NaCl buffer wash. (b) Distribution of background subtracted EMCCD counts of Scc1Halo-TMR-TEVcohesin complexes immobilized on the coverslip. Peaks at ~ 4 x 103 and ~ 8 x 103 EMCCD counts correspond to single fluorophores and a small fraction of double fluorophores respectively. n = 290 regions with fluorescent molecules, 228 regions with background. (c) Photobleaching kinetics of a single DNA-bound Scc1Halo-TMR-TEVcohesin complex. (d) Diffusion coefficients of Smc1/3 wild type or K38A ATP binding deficient 'KA' forms of Scc1Halo-TMR-TEV-cohesin in the presence or absence of ATP. Diffusion coefficients were calculated based on the linear fit of the average mean square displacement of ≥ 13 freely diffusing molecules per condition over a 1 second time period.
Cyclin D1b selectively rescues Ccnd1−/− phenotypes.Figure 2Neonates from 2 independent litters of Ccnd1+/KI × Ccnd1+/KI crosses were sacrificed at birth and genotyped. Mice were organized by genotype and total size measured.Mass of neonates was calculated at birth and average mass quantified (n = at least 4 mice per group).Mice were weighed weekly, and growth is plotted as the average mass of each genotype/week (left: male, right: female, n > 5 per gender and genotype). Mice were held by the tails and analyzed for the leg clasping phenotype. Representative images of Ccnd1+/+ and Ccnd1KI/KI age-matched mice are shown (n > 10 for each group). Top: Individual whole-cell lysates were generated from the eyes of mice of each genotype and analyzed for expression of +/+ cyclin D1 (cyclin D1a) and KI/KI cyclin D1b via immunoblot using antisera specific to each isoform. Bottom: Representative H&amp;amp;E staining of retinal tissue from Ccnd1+/+ and Ccnd1KI/KI mice showing individual layers of the retina. The ganglion cell layer and inner plexiform layer (G + IPL), inner nuclear layer (INL), and outer nuclear layer (ONL) were quantified (right). Images were taken at 400× magnification. Comparison of the phenotypes described in the Ccnd1−/− mouse with those observed in the Ccnd1KI/KI mouse, previously described (Sicinski et al, 1995).
Figure 3(A, B) Colonogenic assays were performed by adding increasing concentrations of pemetrexed to control wild-type (WT) 293A cells and HMCES knockout (HMCES KO) cells (A) and quantification of results were shown (B). Increasing numbers of cells were seeded with increasing concentrations of the compound to obtain optimal drug-dose response curve. Seeding density with respect to control is indicated below each image. The images shown are representative of three biological repeats, each performed in duplicate. (B) Data are presented as mean±s.e.m. and student's t-test was performed for statistical analysis (****, p<0.0001). (C) Western blot showing APEX2 levels upon 293A cells infected with viruses encoding scrambled or two independent APEX2 shRNAs. (D) Colony forming assay was performed to indicate colony forming efficiency in 293A and HMCES KO cells upon infection with viruses encoding scrambled or APEX2 shRNAs. (E) Sensitivity assay was conducted with WT and HMCES KO cells upon treatment with AP Endonuclease inhibitor III (APEi). Results are presented from two independent experiments and data are presented as mean±s.e.m. (F) Colony forming assay was performed with WT and HMCES KO cells upon either pemetrexed treatment alone or in combination with APEi. Data are presented as mean±s.e.m (n=2 each with technical repeats). (G, H) Western blotting revealed levels of POLH (G) and REV1 (H) in two independent clones upon selection following transfection with indicated shRNA. (I) Colony formation assay was conducted to reveal proliferation of WT, POLH KD, REV1 KD, HMCES KO, HMCES KO + POLH KD and HMCES KO +REV1 KD cells. (J) Representative images showing proliferation of respective shRNA resistant reconstitution of POLH in 293A and HMCES KO cells stably expressing POLH shRNA. Experiment was repeated two independent times in duplicates. (K) Mutagenesis frequency analysis was conducted in 293T scrambled and 293T HMCES shRNA cells using sup F shuttle vector-based assay system. Experiments were repeated three independent times and data are presented as mean±s.e.m. Student's t-test was used to calculate statistical significance (**, p<0.01).
Figure 7(A) The Mog1E65K Ran-binding mutant complements the slow growth of mog1ΔsetΔ mutants. Ten-fold serial dilutions of wild-type (WT), mog1Δ, set1Δ, and mog1ΔsetΔ strains transformed with wild-type MOG1 (pMOG1) or with the Ran-binding mutant (pMOG1E65K) as indicated were spotted on SC-LEU and incubated for 2 days at the indicated temperatures. (B) The Mog1E65K Ran-binding mutant complements the low H3K4me3 and H2Bub1 levels of mog1Δ. The expression levels of H3K4me3, total H3, H2Bub1, total H2B and PGK1 in whole cell lysates from wild type (WT) or from mog1Δ cells transformed with an empty plasmid (+pEmpty), wild type MOG1 (pMOG1) or a Ran-binding mutant (pMOG1E65K) were analyzed by western blotting using the corresponding antibodies. (C) Gsp2 is not involved in H2Bub1 regulation. The expression levels of total H2B and of H2Bub1 were analyzed in whole cell lysates from wild type (WT) and gsp2Δ strains by western blotting using anti-total H2B and anti-H2Bub1 antibodies.
Figure 6A. Representative confocal images from EGFP-GSK3α FRAP experiments. Upper left panel ('Baseline') shows EGFP-GSK3α expression in a dendrite and dendritic spine of organotypic slices cultured from rat hippocampus. The spine was bleached ('Bleach'; yellow dashed circle) and its fluorescence partially recovered 90 s later ('Recovery'). Single dendritic spines were imaged once per second for 10 seconds prior to, and for 90 seconds following, spine photobleaching. Such single FRAP experiments were performed on different spines during basal conditions ('basal'; upper panels), immediately after cLTD induction with 20 µM NMDA for 5 minutes ('cLTD'; middle panels), and immediately following 10 minutes of washout ('washout'; lower panels). Scale bars = 1 µm. B. Time course of fluorescence recovery during FRAP experiments described in A. White circles ('basal') show the average spine/dendrite fluorescence of FRAP on EGFP-GSK3α-expressing spines in slices prior to cLTD induction. Pink circles ('cLTD') show fluorescence in spines in which cLTD had been induced using 20 µM NMDA. Green circles ('washout') represent the average of spine fluorescence after NMDA washout for 10 minutes. Recovery trajectories for the three conditions are fitted with two-phase exponential curves (red lines). C, D. As A, B but for spines expressing EGFP-GSK3β. Scale bars = 1 µm. E. Quantification of last 10 seconds of fluorescence recovery (90-100 seconds) in B and D. Bars represent mean ± SEM. Statistical significance was calculated according to the Mann-Whitney U test. N = 12, 11 and 11 spines for EGFP-GSK3α baseline, cLTD and washout groups and 20, 11 and 9 spines for EGFP-GSK3β baseline, cLTD and washout groups, respectively. F. Spine/dendrite ratios showing enrichment of EGFP-GSK3α or EGFP-GSK3β in 'baseline', 'cLTD' or 'recovery' conditions. Bars represent mean ± SEM. Statistical significance was calculated according to the Mann-Whitney U test. G. Representative western blots from whole extracts of organotypic slices incubated in ACSF (untreated; 'baseline'), treated with 20 µM NMDA for 5 minutes ('cLTD') or treated with 20 µM NMDA for 5 minutes and allowed to recover in plain ACSF for 10 minutes ('washout'). Blots were probed for pSer21/9 GSK3α/β, total GSK3α/β, pSer845 GluA1, and total GluA1. H. Quantification of experiments as those shown in G. Error bars represent SEM and statistical significance was calculated according to the Mann-Whitney U test. N = 4 independent experiments for all conditions. Asterisks (**) denote P < 0.01.
Figure 5A, B, Primary cortical neurons transfected with cyt-Fluc-EGFP (A) or nuc-Fluc-EGFP (B) (green), in combination with nuc-mHTT (upper rows) or cyt-mHTT (lower rows) (magenta). Cells were fixed at DIV 3+2 and stained for the neuronal marker MAP2 (grey). Nuclei were labeled with DAPI (blue). Arrowheads point to Fluc-EGFP foci. Dashed lines mark the nuclei. Schemes on the right summarize the cellular distribution of the respective Fluc-EGFP (green) and mHTT (magenta) proteins. Corresponding cultures transfected with control HTT are shown in Appendix Fig. S3A. C, Quantification of Fluc-EGFP foci formation. N=3 independent experiments. Two-way ANOVA with Tukey's multiple comparisons test. ANOVA: HTT, ****p<0.0001; Fluc, n.s.; Interaction HTT x Fluc, n.s. Significant pairwise comparisons within the respective Fluc-EGFP group are indicated above the bars. D, Quantification of the subcellular localization of the respective Fluc-EGFP protein in the presence of the indicated mHTT constructs. N=3 independent experiments. Three-way ANOVA with Tukey's multiple comparisons test. ANOVA: Localization, *p=0.019; HTT, n.s.; Fluc, n.s.; Localization x HTT, **p=0.0012; Localization x Fluc, ****p<0.0001; HTT x Fluc, n.s.; Localization x HTT x Fluc, ***p=0.0009. E, Quantification of Fluc-EGFP foci localization in the presence of cyt-mHTT. N=3 independent experiments. Two-way ANOVA with Bonferroni's multiple comparisons test. ANOVA: Localization, ****p<0.0001; Fluc, n.s.; Localization x Fluc, *p=0.01. Dark-green, light-green and striped asterisks in D and E indicate significant comparisons of the fractions of cells with Fluc-EGFP localized in the cytoplasm, in the nucleus, or in both compartments, respectively.
Figure 2A. Western blot showing abundance of BRCA1 and BARD1 protein in Brca1Δ2/Δ2;Trp53bp1-/- mouseB cells. B. Analysis of chromosome aberrations in mice with targeted deletion of Brca1 exon 2 and Trp53bp1. Left, metaphase spreads were prepared from B cells treated with olaparib. Arrows show examples of chromosome aberrations. Right, quantification of chromosome breaks (CSB), chromatid-type breaks (CTD), radial chromosomes, and other abnormalities in mouseB cells. Error bars indicate SD, N=3 C. RAD51 foci in B cells after 10 Gy IR exposure. Chart shows mean percentage of cells with >5 foci, N=3. Error bars indicate SD, statistical analysis by two-tailed Student t test. D. Percentage of cells showing >5 RAD51 foci after 4hrs treatment with 10 μM olaparib (OLA) or 10 μM camptothecin (CPT). N=2. E. Average number of γ-H2AX foci per cell in cells that were not treated (NT) or exposed to 2 Gy ionizing radiation. N=2. F. Sister chromatid exchanges in cells that were not treated (NT) or exposed to 2 μM olaparib (OLA) for 16 hrs. Black arrows in images show SCEs in olaparib-treated cells. Scale bar: 5 μm. N=2.
Figure 2Genetic interactions between condensin and (A) subunits of the SAGA complex. Fivefold serial dilutions of fission yeast strains were spotted onto complete medium. Genetic interactions between condensin and (B) catalytically inactive Gcn5. Fivefold serial dilutions of fission yeast strains were spotted onto complete medium. Genetic interactions between condensin and (C) multiples HATs. Fivefold serial dilutions of fission yeast strains were spotted onto complete medium. Genetic interactions between condensin and (D) Gcn5 and Mst2. Fivefold serial dilutions of fission yeast strains were spotted onto complete medium.
Formation of autophagosomes in TAΔDHPR.Figure 4(A) Ultrathin sections were imaged by electron microscopy, M, mitochondria; C, collagen fibres; T, T‐tubule. Arrows show double‐membrane vesicules called auphagosomes. (B) Transversal cryo‐sections were stained with anti‐LC3b (red), anti‐dystrophin (green) antibodies, nuclei with Dapi (blue) and imaged by confocal microscopy. Upper panel: TACtrl (Ctrl) and lower panel: TAΔDHPR (ΔDHPR). Bars represent 20 μm. (C) Myofibres isolated from control (Ctrl) or 6 months post‐injected FDB (ΔDHPR) muscles were processed for immuno‐fluorescent labelling for P62 (green) and LC3b (red) and imaged by confocal microscopy. Scale bars, 10 μm.
T.Figure 8A, B, C HBMEC were challenged with ΔHx (WT), MIC1 ko, MIC3 ko, MIC1-3 ko T. gondii at MOIs that yielded similar percentages of infected cells (A). Cell lysates were obtained and used to examine total EGFR and phospho-tyrosine 1068 EGFR (B) or total Akt and phospho-Ser 473 Akt (C) by immunoblot. HBMEC were incubated with 10 nM of recombinant MIC4, MIC3, M2AP or MIC6 for 15 minutes followed by examination of total EGFR and phospho-tyrosine 1068 EGFR (D) or total Akt and phospho-Ser473 (E) by immunoblot. Densitometry data represent means ± SEM of 4 experiments. Band densities from infected cells or cells treated with MICs were compared to bands from uninfected or untreated cells (Un), which were given an arbitrary number of 1. Results shown are representative of 4 independent experiments.
a, Pancreatic sections stained with Atg7, LC3 and p62 showing Atg7−/− next to Atg7+/+ regions after Pdx-Cre-mediated recombination.f1a, Pancreatic sections stained with Atg7, LC3 and p62 showing Atg7−/− next to Atg7+/+ regions after Pdx-Cre-mediated recombination. Right-side images are cropped and magnified regions (boxed) from left-side images. The magnification is × 2.5. b, Haematoxylin and eosin (HE), p53 and caspase-3 staining of exocrine tissue in Atg7+/+ and Atg7−/− pancreases. c, Quantification (median, s.e.m.) of p53 and caspase-3 activation in Atg7-deficient acinar tissue. d, Kaplan-Meier analysis comparing overall survival of mice either wild type (blue) or Atg7−/− (red) in the pancreas. Mantel-Cox test was used for statistics. 'n/a', 14/16 mice were killed >500 days when completely healthy. e, Biochemical analysis of endocrine function in moribund Atg7−/− or age-matched healthy control mice. Detailed information about the mice is tabulated. A Mann-Whitney U-test was used for statistics. Scale bars: 50 µm. **P 0.01 and ***P 0.001.
Pacs1 deletion suppresses abnormal lymphocyte accumulation in models of lymphoproliferation (A) Spleen size and FACS analysis of abnormally expanded B220+CD23+CD21+/low FOB cells in Pacs1+/−;Bcl2TG and Pacs1−/−;Bcl2TG mice.Figure 8(A) Spleen size and FACS analysis of abnormally expanded B220+CD23+CD21+/low FOB cells in Pacs1+/−;Bcl2TG and Pacs1−/−;Bcl2TG mice. (B) Number of circulating B cells in the blood and FOB cells in the spleen of Pacs1+/−;Bcl2TG and Pacs1−/−;Bcl2TG mice. Horizontal bars indicate mean value. Two-tailed Mann-Whitney test, *P < 0.05, **P < 0.01. (C) B cells were purified from the spleens of Pacs1+/−;Bcl2TG and Pacs1−/−;Bcl2TG mice (CD45.2), labelled with CTFR and CTV proliferation dyes, respectively, and transplanted into unirradiated CD45.1 recipients. Donor B cells were measured in the spleen of recipient mice 7 days after B cell transfer based on CD45.2 expression and proliferation dye fluorescence. (D, E) The fraction of proliferating (D) and recovered (E) donor cells from the experiment in (C). Symbols represent individual recipient mice and data are from two independent adoptive transfer experiments. Two-tailed unpaired t test, **P < 0.01, ***P < 0.001. (F) Fraction of apoptotic B cells in the adoptively transferred B cell populations in the experiment in (C). Symbols represent individual recipient mice and data are from one adoptive transfer experiment. Two-tailed unpaired t test, ***P < 0.001. (G) Splenocytes from Pacs1+/−;Bcl2TG and Pacs1−/−;Bcl2TG mice were stained with cell surface antibodies to identify FOB cells and treated with 100 µM H2O2 for 35 minutes. Cells were then labelled with TMRE to monitor MMP. TMRE fluorescence was measured by FACS analysis. Data are presented as mean ± SD of 3-4 replicates, unpaired t test, ns = not significant, ***P < 0.001. Results are from one experiment. (H) Lymph node size and flow cytometry of lymphoproliferative CD3+B220+ cells in Pacs1+/+;Faslpr/lpr­ and Pacs1−/−;Faslpr/lpr mice. (I) Enumeration of CD3+B220+ cells in the peripheral blood and lymph nodes of Faslpr/lpr dependent on Pacs1 expression. Horizontal bars indicate median value. Two-tailed Mann-Whitney test, **P < 0.01.
AD risk factors cause loss of gene-expression control.Figure 7Figure 7: AD risk factors cause loss of gene-expression control.Our central hypothesis is that the various AD risk factors eventually lead to aberrant gene-expression and loss of transcriptional control. This is based on the assumption that proper gene-expression is a core feature of cellular homeostasis and that changes occurring at various compartments of the cell (e.g. at the synapse) will eventually signal to the nucleus and cause gene-expression changes. In the case of FMN2 we provide for example evidence that chronically low levels of FMN2 disturb synaptic actin dynamics and thereby affect ERK1/2-dependent gene-expression programs. Thus, even if an AD risk factor may play no direct role in gene-expression it may eventually contribute to a loss of transcriptional plasticity. We therefore suggest that targeting pathological gene-expression could be a suitable therapeutic approach, especially for multifactorial diseases such as AD, where it is nearly impossible to determine all of the genetic and environmental factors that eventually contribute to clinical phenotypes.
Figure 3(A-D) Grip power (A), body weight (B), rotarod performance (C) (Two-way ANOVA with Tukey's test) and survival rate (D) (Kaplan-Meier analysis and log-rank test) of AR-97Q mice treated with or without RG108 (DMSO, n =20; 0.5 mg/dL, n = 20; 2.0 mg/dL, n = 20). All parameters were significantly improved in AR-97Q mice treated with 2.0 mg/dL RG108 compared with those treated with DMSO: *P < 0.05 (grip); *P < 0.05 (body weight); *P < 0.05 (rotarod); and *P < 0.05 (survival). (E) Representative photograph of a 13-week-old AR-97Q mouse treated with DMSO (left) and an age-matched AR-97Q mouse treated with 2.0 mg/dL RG108 (right). AR-97Q mouse medicated with RG108 maintained muscle volume and body size compared with DMSO-treated AR-97Q mouse of same age. The white compact masses (arrow) of mice head are the cement covering the surgical site. (F,G) Immunoblots (F) and relative intensities (G) of the Dnmt1-immunoreactive bands of spinal cords from AR-97Q mice treated with or without RG108 (n = 3 per group). (H,I) Dnmt1 immunohistochemistry in spinal cord sections from AR-97Q mice treated with or without RG108 (n = 3 per group).
Figure 2Representative confocal microscope image of L6 cells stably expressing tandem fluorescent-eGFP-mCherry LC3B treated with iron (FeSO4, 250 µM) for the indicated time points (4 h, 8 h, 24 h). Quantification of mean LC3B puncta per cell from (A). Experiments were repeated three times, and one representative experiment is presented here. Representative western blot images and quantification of p62 to actin in L6 cells after iron treatment (FeSO4, 250 µM) at multiple time points. Representative western blot images and quantification of LC3B-II to GAPDH in L6 cells after 4h iron treatment stimulated with chloroquine (CQ, 30 µM). * Representative western blot images and quantification of LC3B-II to GAPDH in L6 cells after 24 h iron treatment stimulated with CQ 30 µM. Representative confocal microscope z-stack image of mCherry-LC3B L6 cells after iron treatment (250 µM, 24 h). Quantification of LC3B puncta size from (F). Experiments were performed three times, and one representative experiment is presented here. Quantification of autophagosomes motility from live cell microscopy (Supplementary Movie 1) from mCherry-LC3B L6 cells under basal, iron (250 µM, 24 h) and starvation (amino acid free) condition. Experiments were performed three times, and one representative experiment is presented here. *P < 0.05 (unpaired Student's t-test compared to Iron 24h). Representative TEM images of autophagosome and autolysosomes in L6 cells after iron treatment (250 µM, 24 h). Representative confocal microscope images of eGFP-mCherry-LC3B L6 cells with Lysotracker DeepRed after iron treatment (250 µM, 24 h).