EMBO/sd-character-level-ner huggingface dataset:
Electron micrographs showing the immunogold localization of α-amylase in attached cotyledons cells (A–E), detached cotyledons cells (F), or control for immunogold labeling (G).Figure 3.Electron micrographs showing the immunogold localization of α-amylase in attached cotyledons cells (A-E), detached cotyledons cells (F), or control for immunogold labeling (G). (A) Anti-α-amylase antibody immunogold-stained PSV, but not SG. (B and C) Golgi complex and PSVs were both immunogold-labeled with anti- α-amylase antibody. (D) Immunogold localization of α-amylase (15-nm particles) and SH-EP (10-nm particles). α-Amylase and SH-EP were localized in PSVs and KVs, respectively. (E) Gold particles from anti-α-amylase antibody were detected in LVs as well as PSVs. (F) PSVs in detached cotyledons were immunogold labeled with anti- α-amylase antibody. (G) Immunogold staining of detached cotyledon cells without first antibody (anti-α-amylase antibody). No gold particles were observed in the cell. CW, cell wall; G, Golgi complex; KV, KDEL-tailed cysteine proteinase-accumulating vesicle; LV, lytic vacuole; Mt, mitochondrion; PSV, protein storage vacuole; SG, starch granule. Bars, 200 nm.
Transcription in the miR-205 miRNA locus from selected species.Figure 3UCSC browser snapshots show tracks from small and long RNA-seq analyses of mouse , bovine and porcine oocytes Tracks were constructed as described in material and methods. The y-scale for small RNAs depicts counts per million (CPM) of mapped 19-32 nt reads. The y-scale for long RNA-seq depicts CPM of mapped fragments. Grey rectangles and dashed lines represent exon-intron structure and transcript splicing inferred from RNA-seq data. Despite the splice acceptor of the terminal exon appears conserved, murine RNA-seq data do not support splicing of the primary miR-205 transcript. Instead, the mmu-miR-205 locus produces a detectable antisense spliced maternal lncRNA while the precursor of mmu-miR-205 is undetectable. SA, splice acceptor for the last exon of lncRNA originating from the locus.
Figure 5(A) Representative western blots showing the level of SQR, TST, ETHE1 and SUOX proteins in two controls (C1, C2) and two patients fibroblasts (P3, P4). (B-E) Relative levels of proteins normalized to Vinculin.
Regulation of apoptosis in MCL‐1 conditional knockout mice.Figure 6(A) Cortices from the indicated genotypes and ages were analysed by western blot for the presence of active caspase‐3, p62 and MCL‐1. Primary cortical neurons treated for 12 h with camptothecin were used as a positive control for active caspase‐3. (B) Active BAX (6A7) and active caspase‐3 were also quantified by staining the cortex of 1‐ to 14‐day‐old MCL‐1Δ/Δ mice. Representative images from at least three sections from each animal were counted. The total number of cells was determined by counting the Hoechst‐positive nuclei. Data are expressed as percent of positive cells for at least three animals per genotype±s.d. *P0.05. (C) Active BAX is present in a subset of LC3‐positive cells. Sections from P7 and P14 MCL‐1Δ/Δ brains were stained for LC3 (Green) and active BAX (6A7; Red). Confocal images were taken using × 63 objective. Arrows point to BAX‐positive, LC3‐negative cells. Representative images are shown. Scale bars=50 μm. Primary cortical neurons of the indicated genotypes were infected with a lentivirus expressing Cre at the time of plating and analysed for the presence of the caspase‐cleaved fragment of the caspase‐3 substrate CAS5 days later. Alternatively (E), the neurons were analysed for cytochrome c release by immunofluorescence. Data are expressed as percent of positive cells for at least three embryos per genotype±s.d.
Fig.Figure 1Schematic representation of the TCF/LEF protein structure. Below, the guideRNAs (gRNAs) used for each specific gene are annotated. Grey boxes indicate the DNA-binding HMG box Western blots for detection of TCF7, LEF1, TCF7L1, TCF7L2 and β-catenin (B") proteins in wild-type (WT) and knockout (K.O.) TCF-reporter TOP-flash assay performed on the indicated cell lines treated with 10 μM CHIR, for 24 hrs. The control FOP-flash reporter is not activated upon CHIR administration. The data represent the mean ± s.e.m. of averages of nine independent experiments (N = 9). dBcat, β-catenin knockout cells; d4TCF, TCF/LEF quadruple knockout cells Transfection of individual TCF-expressing plasmids could restore the ability of d4TCF cells to respond to Wnt3 as measured in a TOP-flash assa Quantitative RT-PCR analyses of Axin2 transcripts, performed on the different cell lines treated with 10 μM CHIR or DMSO (control) for 24 hrs. Error bars show that standard deviation obtained from 3 independent experiments Transfection of individual TCF-expressing plasmids could restore the ability of d4TCF cells to respond t CHI as measured by Axin2 mRNA abundance
Figure 2A. Representative electron micrographs of synapses in the hippocampal CA1 region in wt and NPC1nmf164 mice (black arrows indicate post-synaptic densities, d-dendrite, sv-synaptic vesicles, m-mitochondria, MLB-multilamellar bodies). B. Mean ± SEM synapse density (n=3 mice, 3 month-old, unpaired Student t test, *p = 0.0377). C. Mean ± SEM synaptic vesicle density (3,088 vesicles from 101 wt and 121 NPC1nmf164 synapses from 3 different mice (3 month-old) per group, unpaired Student t test, ***p < 0.0001). D. Synaptic vesicle diameter (528 wt and 507 NPC1nmf164 vesicles from 3 different mice (3 month-old) per group). E. Mean ± SEM post-synaptic density length (101 wt and 121 NPC1nmf164 post-synaptic densities from 3 different mice (3 month-old) per group, unpaired Student t test, ***p < 0.0001). F. Mean ± SEM post-synaptic density thickness (n=3 mice, 3 month-old, unpaired Student t test, ***p < 0.0001). G. Mean ± SEM cholesterol levels in synaptosomes (n=4 mice, 3 month-old, unpaired Student t test, *p = 0.0476) from wt and NPC1nmf164 mice expressed as percentage of wt mice.
Lineage B clade-specific determinants for human ACE2 usagea, Schematic overview of clade 1, 2 and 3.Figure 4:Schematic overview of clade 1, 2 and 3. Highlighted in yellow are the 14 residues that contact human ACE2. Deletions in loops 1 and 2 are indicated for clades 2 and 3. Structure of human ACE2 and the SARS-S RBD (PDB: 2AJF), with loops highlighted in gray. VSV pseudotypes were generated with the indicated RBD and used to infect BHKs transfected with either human ACE2 or empty vector. Shown are data for 3 replicates. Westernblot of producer cell lysates and concentrated pseudotyped particles.
Figure 3YAP driven aberrant sphere formation of MCF10A cell lines is ablated by AIB1 knockdown, but further enhanced by ANCO1 knockdown. Indicated cell lines were plated on Matrigel and allowed to form mammospheres for 7 days. 10x and 40x magnifications shown, Scale bar = 200μm. Quantification of aberrant spheres per field displayed in panel (A), n= number of spheres. Quantification of size (arbitrary units) of aberrant spheres per field displayed in panel (A), relative to control YAP S94A shGFP spheres. n= number of spheres. MCFDCIS xenograft tumors have increased expression of AIB1-YAP, EMT, and 1q21.3 genes during the invasive transition. Schematic of experiment shown. Gene set enrichment analysis of RNA-seq data generated from MCFDCIS xenografts at indicated time points. Invasive lesions have significantly less ANCO1 than encapsulated DCIS lesions. Representative images of H&E, p63, and ANCO1 stained MCFDCIS xenografts as DCIS and invasive lesions. Yellow circles indicate DCIS lesions and black arrows indicate invasive lesions. 20x magnification; Scale bar = 100μm. Quantification of ANCO1 positivity in DCIS and Invasive lesions shown in panel (E).
Figure 8Working model of autophagy regulation by IO. Acute IO leads to inhibition of mTORC1 leading to autophagy induction. Prolonged IO prevents ALR-mediated production of new lysosomes through AKT-TSC-Rheb-mTORC1-UVRAG signaling defects. The lack of free lysosomes contributes to autophagy inhibition and insulin resistance in skeletal muscle.
ACE2 levels increase upon telomere shortening in human cells and in lungs of G3 Terc-/- mice.Figure 2A RT-qPCR detection of ACE2 mRNA expression levels in early passages (PD 34-37) and late passages (PD 61-64) human normal fibroblasts (BJ) (n = 3-4 replicates per group). B ddPCR detection of ACE2 mRNA expression levels in early passages (PD 22.37) and late passages (PD 140) human bronchial epithelial cells (HBEC) (n = 6 replicates per group). C Representative images and quantitative analyses of ACE2 immunohistochemical staining in lungs of age-matched wild type and G3 Terc-/- mice (n = 3-4 mice per group). Scale bar, 200 µm. D Double-marker immunofluorescence and quantitative analyses of ACE2 intensity level in type II pneumocytic pro-SP-C-positive and CD31-positive endothelia in lungs of age-matched wild type and G3 Terc-/- mice (n = 3-4 mice per group). Scale bar, 100 µm. a.u. = arbitrary units.
Figure 8(A) Identical starting number of cells from the MEF lines were serially passaged following identical schedules. The differences in cell number were determined after 3 and 5 passages. The results are average of 3 experiments (mean +/- SD). (B) Identical number of cells from the MEF lines were plated at low density and cultured for 14 days. Colony formation efficiency was determined by crystal violate staining. (C) Survival curves of p53-/-;MDMXSG/SG and p53-/-;MDMXwt/wt mice. Thymic lymphomas were the major cause of mortality in both genotypes. (D) A model of Wnt signaling regulation by MDMX. In the absence of stress, MDMX inhibits CK1α, increases basal expression of Wnt/β-catenin target genes and stimulates proliferation. Interaction with mutant p53 stimulates MDMX-CK1α binding and promotes proliferation. DNA damage promotes MDMX degradation and inhibits MDMX-CK1α binding, reducing Wnt signaling.
Figure 1 A Western blot analysis of LC3 in SH-SY5Y cells after IU1 treatment (100 μM, 24h). Graphs represent mean± SEM. N= 8  B-D Electron microscope images were taken to quantify the number of autophagosomes and autolysosome present in control (DMSO) or IU1 treated cells. At least 55 cells were analysed from 3 different experiments. Graphs represent mean± SEM  E Western blot analysis of mitochondrial marker proteins ATP5a, HSP60 and normalised by actin after 24h of DMSO (C) or IU1 (100 μM) treatment. N=3. Graphs represent mean± SEM  F-I Confocal images of cells transfected with mito-YFP and treated with DMSO (F, control) or IU1 (G) for 24h. Fragmentation index (H) and total mitochondrial volume / cell were measured and represented in (I), mean ±SEM. At least 35 cells were analysed from 3 different experimental conditions. Scale bar 10 μm  J-L Mito-YFP transfected cells were treated with IU1 (100 μM, 24h), fixed and immuno labelled for LC3. Number of LC3 dots / cell (K) and LC3 dot overlapping mitochondria (L) were counted. At least 35 cells from 3 independent experiments were analysed. Scale bar: 10 μm  M-N Cells were transfected with mito-keima and treated with DMSO or IU1 for 24h and subjected to FACS analysis, counting 10,000 cells per experiment. (M) represents the shift of the signal intensity in IU1 treated cells when excited at 560 nm (false colour). (N) represents the average intensity of the mito-keima protein when excited at 560 nm and the ratio between 560: 405 nm. Graphs represent mean ± SEM from 3 different sets of experiments
/CDK4 phosphorylates and stabilizes ATOH1 proteinA, B.Figure 3A, B. Cycloheximide (CHX) assay. (A) N2a cells transfected with Atoh1-GST, Ccnd1 and Cdk4 plasmids 2 days prior were harvested after 0, 4 and 8 hr treatments of 50ug/mL CHX and lysates were subjected to immunoblotting with anti-Atoh1 or β-actin as a housekeeping control. (B) Quantification of relative levels of ATOH1 to the β-ACTIN. N2a transfected with only Atoh1-GST is referred as WT and N2a transfected with Atoh1-GST, Ccnd1 and Cdk4 plasmids is referred as 4D. Same experiments were performed three times. Error bars are representing as Mean ± SEM. C. N2a cells transfected with Atoh1-GST plasmids were treated with DMSO(Control), Palbociclib (Palb) or Palb and 10 uM MG132 (MG132) for 6hrs before harvest. Lysates of N2a were subjected to immunoblotting with anti-Atoh1 or β-actin. Quantification of relative ATOH1 to the β-ACTIN (right panel). Tukey-Kramer test was performed between Control, Palb, MG132. Bars and error bars are representing as Mean ± SEM from three trials of each condition. ***p < 0.001, *p < 0.05, N.S.: Not Significant. Three slots of each experimental condition represent the three biological replicants of each condition. D. in vitro phosphorylation assay of ATOH1. GST-tagged full length ATOH1 (FL), N-terminus ATOH1 including the bHLH domain (N) and C-terminus of ATOH1 downstream of the bHLH domain (C) were purified from N2a cells using glutathione-beads. ATOH1 proteins were incubated with either Casaine kinase 1α (CK1α) or CCND1 and CDK4 in the presence of [γ-32P]-ATP. Phosphorylation signals detected with FL, N and C by the phosphorylation of CK1α (lanes 1, 2, 3) are indicated by white arrowheads. Phosphorylation signals were also detected with FL and C by the phosphorylation of CCND1-CDK4 (lanes 5, 6, 7). Phosphorylation signals of N with CCND1-CDK4 were not detected. Strong CK1α self-phosphorylation signals were observed (lane 4). E. Phosphorylation sites of ATOH1-GST purified from N2a cells were determined by LC-MS/MS. 5 phosphorylation sites (Y78, S82, S309, S311, S328) were identified. F. In vitro kinase assay with either wild type ATOH1 (WT) or non-phosphorylated form of the Serine-to-Alanine 309 (S309A). WT or S309A were incubated with CCND1 and CDK4 (4D) in the presence of [γ-32P]-ATP. While WT with CCND1 and CDK4 gave rise to strong signals (lanes 1,2 white arrowhead), S309A showed no obvious signal (dashed arrowhead indicated the predicted size of ATOH1). G. Atoh1 expression plasmids were transfected into N2a cells with Ccnd1 and Cdk4 expression plasmids (4D). N2a cells were treated with 10 nM, 100 nM, 1000 nM Palb for 6 hrs before harvest. Lysates from N2a cells were subjected to immunoblotting with anti-Atoh1, p-S309-Ab and β-actin. Quantification of relative ATOH1 to the β-ACTIN (right panel). Tukey-Kramer test was performed between 0, 10, 100, 1000nM. Bars and error bars are representing as Mean ± SEM from three trials of each condition. **p < 0.01, *p < 0.05, N.S.: Not Significant.
Figure 1(A) Representative images of COS7 cells expressing GFP-COP1 (green) co-transfected with empty vector or the indicated TRIB1-FLAG construct (red). Cells were stained with anti-FLAG (Sigma) and anti-mouse Alexa Fluor 568. Nuclei were counterstained with DAPI (blue). Scale bar, 10 µm. See also Appendix Fig S1B for western blots showing the expression levels of the constructs used. (B) Quantification of the average ratio of nuclear/cytoplasmic fluorescence of GFP-COP1 in (A). Mean values ± S.E.M. are shown for three independent experiments where 50 individual cells per experiment were analyzed. Significance relative to GFP-COP1 with empty vector was calculated using the Student t test (****p < 0.00001). (C) Representative images of COS7 cells showing GFP-COP1 (green) and TRIB1-FLAG (red) colocalizing in punctate nuclear structures. Insets show enlarged images of these nuclear puncta. Cells were stained with anti-FLAG and anti-mouse Alexa Fluor 568. Nuclei were counterstained with DAPI (blue). Scale bar, 10 µm. (D) Representative images of COS7 cells showing that GFP-COP1 (green) and TRIB1-HA (red) are diffuse in the nucleoplasm when coexpressed with C/EBPα (cyan). Cells were stained with anti-HA (Santa Cruz), anti-C/EBPα (Cell Signaling), anti-mouse Alexa Fluor 405, and anti-rabbit Alexa Fluor 680. Scale bar, 10 µm. See also Fig EV2C for images of cells expressing each construct alone or in pairs.
Figure 1A, Rb1 protein (RB) was expressed in uterine luminal epithelium and stroma during early pregnancy in mice. Scale bar = 100μm; arrowhead, embryo. B, Phosphorylated RB (pRB) protein, the inactive form of RB, was expressed in the luminal epithelium on days 2 and 3 of pregnancy. Scale bar = 100μm. C, RB was efficiently deleted in the uterus of mice with uterine deletion of Rb1 (Rb1d/d mice) on day 4 of pregnancy. Scale bar = 100μm. D, RB expression was normal in the ovary of Rb1d/d mice on day 4 of pregnancy. Scale bar = 100μm. E, The number of new-born pups was decreased in Rb1d/d dams (n=37 different dams) compared to their littermate controls (Rb1f/f mice, n=20 different dams) (mean ± SEM, Student's t test).
DHR3 is a specific dS6K interacter.Figure 1Induction of UAS constructs and EPs by the ap promoter directed Gal4 expression within the dorsal compartment of the wing imaginal disc: (A) ap-Gal4 control; (B, E, H, J, K) UAS-dS6K; (C, F, I) UAS-S6K1dE/D3E; (D, E, F) DHR3-EP; (G, H, I, K) UAS-DHR3-RNAi and (J) unidirectional DHR3-EP. The bending down of the wing indicates a slight overgrowth of the dorsal compartment, whereas a bending up reveals a slight growth deficit of this compartment.
Figure 3Representative images of adult females wings in combinations of Dl∆β1-2 with loss of function Delta alleles. In combinations with Dlrev10 (A), Dl∆Exon2 (B), or Df(3R)DlFx3 (C), vein thickening is strongly enhanced (right panels) compared to heterozygous mutants alone (left panels). Vertical square brackets indicate the regions used for vein thickness quantification. Quantification of wing-vein thickness in females of the indicated genotypes. Representative images of adult female wings demonstrate that Dl∆β1-2 rescues the wing notching phenotype, caused by a Notch loss of function allele (N55e11). Horizontal square bracket indicates the L5 vein "delta" at the intersection with the wing margin analyzed in EV2E. Quantification of wing notching in females of the indicated genotypes, Dl∆β1-2 rescues notching in a similar manner to Dlrev10.
Experimental outline of the cell surface labeling and preparation for mass spectrometryImmunostaining of HeLa cells probed for FITC‐labeled streptavidin and DNA and the merge streptavidin (red) and DNA (blue).Figure 1A Immunostaining of HeLa cells probed for FITC‐labeled streptavidin and DNA and the merge streptavidin (red) and DNA (blue). Cell surface was labeled using amine biotinylation. Scale bar, 10 μm. B Western blot analysis of cell surface preparation. The surface‐biotinylated cells were lysed using a detergent containing lysis buffer, incubated with and then eluted from streptavidin beads (i.e., cell sur. fraction). Cell surface fraction and the whole‐cell lysate were blotted against a cell surface protein, EGFR, and a cytoplasmic protein, actin. C Workflow of the cell surface proteome. Cell surface labeling and isolation of cell surface proteins in SILAC‐labeled interphase versus mitotic HeLa cells are shown. D Schematic of all identified proteins (blue circle) and cell surface proteins (pink circle). Cell surface/plasma membrane proteins were identified based their GO annotations; their distribution is shown in a pie chart (top).
FOXD3 recruits SUV39H1 to repeat elements.Figure 4(A) GFP immunoprecipitation analysis using GFP-SUV39H1 and GFP-SUV39H2 expressing Suv39h dn mESCs. The top panel represents immunoblot using GFP antibody and the bottom panel represents immunoblot using FOXD3 antibody. The experiment was repeated 3 times using biological replicates. (B) Volcano plot depicting significantly upregulated and downregulated repeats in Suv39h dn ES cells compared to wild type ES cells determined by RNA-Seq analysis. MSR (GSAT_MM), MERVL-int, L1MdA and MT2_Mm are labelled. (C) Metadata profile showing enrichment of SUV39H1 and H2 over the MERVL consensus sequence including 1kb sequences flanking the start (D) ChIP-qPCR analysis of endogenous SUV39H1 enrichment over MERVL, MSR and L1MdA in control and Foxd3 KO cells. Actb promoter is used as negative control. Data is represented as average percentage input from 3 biological replicates. Error bar indicates SEM (*: p<0.05 paired t-test). (E) ChIP-qPCR analysis revealing H3K9me3 enrichment over MERVL, MSR and L1MdA in control and Foxd3 KO cells. Actb promoter is used as negative control. Data is represented as average percentage input from 3 biological replicates. Error bar indicates SEM (**: p<.005, *: p<0.05 paired t-test). (F) Metadata analysis of H3K9me3 enrichment over MSR and MERVL consensus sequences in WT mESCs (blue) and Suv39h dn mESCs (green). Relative enrichment is plotted along the repeat start (RS) and repeat end (RE) including ± 1 kb flanking sequences. Y axis represents reads per genome coverage (RPGC). (G) Immunofluorescence analysis of control and Foxd3i GFP-SUV39H1 expressing Suv39h dn mESCs stained with DAPI, FOXD3 and H3K9me3. The percentage of counted cells (~150 cells per experiment) exhibiting fluorescence signal is indicated. Scale bar=2μm. n= 3 biological replicates. (H) FOXD3 represses MERVL and MSR in mouse ES cells: FOXD3 (green) binds to MERVL and MSR and recruits SUV39H1 (yellow) to the repeats. SUV39H1 facilitates repression of the repeats by establishing H3K9me3 (orange hexagons). Foxd3 depletion disrupts the recruitment of SUV39H1 and leads to a reduction in H3K9me3 over MERVL and MSR. This in addition to an increase in H3K4me3 (green hexagons) on the region results in the activation of MSR and MERVL and a concomitant increase in 2CLC specific genes, shifting the balance between mESCs (Green and purple) and 2CLCs (Brown and blue). This is accompanied by a perturbation of the classical condensed heterochromatic foci in the nucleus. Foxd3 thus helps maintain ES cells in a pluripotent state by regulating MERVL and MSR.
Figure 5Confocal maximum projections of otoferlin-immunolabeled organs of Corti from p15 Wt and 1/3-DKO mice illustrated with an intensity-coded look-up table where brighter colors indicate higher intensity. Scale bars: 5 µm. Otoferlin levels were reduced by approx. 28.4 ± 3.5 % in 1/3-DKO IHCs compared to Wt IHCs (relative intensities normalized to Wt; Wt n = 31 images (229 IHCs)/ N = 5 organs of Corti (3 animals); 1/3-DKO n = 20 images (196 IHCs)/ N = 5 organs of Corti (4 animals); unpaired student's t-test; ***p < 0.001; box plots illustrate the median with the interquartile range, whiskers indicate 10-90 % of data points, the squares present the respective mean value). Normalized otoferlin intensity line profiles through single IHCs of the respective genotypes at a longitudinal central plane through the nucleus from apical (0.0) to basal (1.0) revealed similar distribution patterns of otoferlin in IHCs of Wt and 1/3-DKO and an overall reduction of otoferlin levels throughout the entire cell in 1/3-DKOs. Traces indicate the mean pixel intensity ± SEM; Wt n = 123/ N = 3; 1/3-DKO n = 124/ N = 3. Interaction of otoferlin and endophilin-A1 detected by co-IP in HeLa cells co-expressing GFP-otoferlin and RFP-endophilin-A1. Otoferlin-GFP was immunoprecipitated (IP) by GFP-Trap beads, and blots were probed with a KO-validated anti-endophilin-A1 antibody. Exogenously overexpressed GFP-otoferlin was immunoprecipitated using GFP-Trap beads and incubated with purified endophilin-A1 (pA1). IP was then followed by immunoblotting with an anti-endophilin-A1 antibody.
Fever incidence within 24 hours of study vaccines.Figure 2a) Kaplan-Meier to first fever (≥38°C) episode within 24 hours of receiving vaccines administered at 4 months of age; control n=88 and 4CMenB n=89. b) Principal component (PC) analysis of RNA-sequencing data (14837 genes, n= 253) from all study time points. The lines connect participant's pre- and post-vaccination samples. The ellipses are the two-dimensional (PC1 and PC2) 95% confidence intervals for each study time point. A contribution plot (top right) displays the genes contributing most to PC1/PC2, i.e. 0.15 implies 0.15% contribution of that variable to the principal components displayed. c) Plotted are the CIBERSORTx neutrophil fractions from whole blood RNA-sequencing data, with median and interquartile range. P-values were determined from a two sample Wilcoxon rank sum test. The number of individuals in each group is display in the x-axis. d) Spearman's rank correlation between neutrophil counts measure by full blood counts (n= 142) and those estimated by CIBERSORTx, using the LM22 signature matrix.