EMBO/sd-character-level-ner huggingface dataset:
Paused promoters have high levels of paused Pol II throughout the embryo, even in tissues where the genes are not expressed; they show high promoter accessibility in all tissues, low gene expression noise when active, but also high background expression when inactive. TATA promoters mediate highly tissue-restricted expression and only show Pol II occupancy when active; they have lower chromatin accessibility and background expression but show higher expression noise when active.
d, Representation of the contribution of the plasma membrane to autophagosome precursor formation. Cholera toxin is internalized by clathrin-dependent and clathrin-independent endocytosis. Clathrin knockdown significantly decreases cholera toxin uptake. Clathrin-coated vesicles (CCV) budding immediately from the plasma membrane (EEA1-negative) are precursors to early endosomes (EE) (EEA1-positive). Previous studies showed that delivery of fully formed autophagosomes to lysosomes requires fusion of such autophagosomes with early or late endosomes or multivesicular bodies (LE/MVB) to form amphisomes, which are Atg16L1-negative, LC3-positive and positive for endosomal markers (red arrows). We show here that inhibition of clathrin-dependent internalization inhibits the formation of early Atg16L1-positive precursors that mature to form phagophores and, later, autophagosomes (blue arrows). These Atg16L1 vesicles were positive for other early autophagosomal markers (Atg5 and Atg12) but not for early endosomal markers (EEA1). AP2 is a clathrin adaptor at the plasma membrane, whereas AP1 localizes to the trans-Golgi network and endosomes. Uncropped images of blots are shown in Supplementary Information, Fig. S9.
B, PTPN3 (L232R) inhibits TGF-β-induced reporter gene activity. HepG2 cells were transfected with expression plasmids for PTPN3, PTPN3 mutants and CAGA-luc reporter, and Renilla-luc reporter. Cells were harvested for luciferase assay after 8 h of TGF-β (2 ng/mL) treatment. Data are shown as mean ± SEM; n = 3. Statistical analysis was performed using two-tailed Student's t-test. *P<0.05, **P<0.01.
Tabular summary of ATP hydrolysis by CtUba4 WT and mutants in the presence of CtUrm1. The amount of released PPi was measured using a fluorometric pyrophosphate assay kit. After PPi hydrolysis, Pi was also detected by a Malachite Green kit. Changes in enzymatic activity are indicated as a color gradient from blue to red, normalized to CtUba4 WT. PPi: pyrophosphate; Pi: inorganic phosphate; SD: standard deviation; n≥3
(f) NADH-driven H2O2 production was determined using Amplex Red. Results are shown from five hearts analysed in duplicate.
(g) Amino acid sequence comparison of TREM2-TMH wild-type and K186A with Notch1 and APP. α-helical secondary structure elements in the free form of each protein are labeled by grey boxes. Positively charged residues are marked by blue and polar residues close to the cleavage site (indicated by the comma mark) by yellow boxes. Red boxes indicate the unfolded sequence stretches in the bound state preceding the cleavage site. The secondary structure of the substrate helices (or derived from the structural model for TREM2-TMH) in complex with γ-secretase is shown above.
A. Cells from mock-organoids display a healthy nucleus labeled by DAPI (blue) (i). SARS-CoV-2-positive cells (green) display condensed (middle panel, ii) and fragmented DNA (bottom panel, iii, arrows). At least 75 cells from two (n=2) independent batches of organoids were examined. Figures display scale bars.
(A) Am80 selectively activates RARα and RARβ to alter expression of RA-target genes, leading to modest myeloid expansion and effective neutrophil differentiation. Up arrow, increased expression of cell cycle inhibitor p21Cip/Kip; down arrow, decreased expression of p21Cip/Kip.
(B) The mitochondrial potential was detected by JC-1 staining. Data are presented as means ± S.E.M. (paired t-test, n = 3, biological replicates per group, ** p < 0.01).
B COS‐1 cells were incubated in medium containing Alexa 488‐Tfn for 40 min, washed, fixed, and stained with recombinant 2xPH. Magnified image is shown in the right panel. Scale bar, 10 μm.
(A) Western blot analysis of DIV 5 cortical neurons transfected with siMecp2 or siControl (siCtl) and treated with 1 µM FK506 or vehicle for 1 h, and quantification of pS421 HTT (n=9 per group). Data are presented as means ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001, Mann-Whitney test.
(B) Percentage of RNA (1 μg) remaining after a 15´ digestion with increasing doses of RNase A in the presence or absence of (PR)20 (5 μM) (n=3). Data represent mean values ± SEM.
(e) STUB1 knockdown induces cellular senescence. The siRNA transfection was performed as in c and repeated 4 days later, followed by incubation for 3 days and SA-β-Gal staining. Percentages of SA-β-Gal-positive cells were from triplicate experiments (mean±s.d.). **P0.001 (Student's t-test).
(H) Summary of region on CSAG2 required for SIRT1 interaction. HEK293FT cells stably expressing HA-SIRT1 were transfected with indicated CSAG2 constructs for 48 h before IP with anti-HA followed by SDS-PAGE and immunoblotting for anti-Myc.
(E) Representative picture and quantification of cardiomyocyte cell division in fetal cardiomyocytes 48 hours after application of the adenoviruses as indicated and immunostained for aurora B (Auro B), troponin I and DAPI. Scale bar: 50µm.
J The average length of procentrioles was quantified for wild‐type and Ccno‐deficient MCCs of E17 tracheae (49 procentrioles in n = 3 independent samples for wild‐types and 47 procentrioles in n = 3 independent samples for Ccno‐deficient MCCs).K The diameter of deuterosomes (41 deuterosomes in n = 3 independent samples for wild‐types; 38 deuterosomes in n = 5 independent samples for Ccno‐deficient MCCs) was measured in wild‐type and Ccno‐deficient MCCs of E17 tracheae.
3D reconstruction of representative images of ASM (A) or ceramide (B) staining in HeLa cells infected with Shigella WTor mock-treated, analyzed at 3 hpi. ASM, ceramide, Hoechst and Shigella staining were surface converted by voxel distance
C. Reconstruction of SEM serial sections of a complete rounded up concavin(-) sporozoite; plasma membrane in blue and IMC in yellow. Scale bar 1µm. In total 6 parasites were reconstructed from two different preparations.
Schematic of single-cell dual-fluorescence miRNA sensor assay. The control sensor plasmid consists of GFP and dsRed expressed under a separate promoter (top panel). The miR16 sensor includes two miR-16-5p binding sites at the 3'UTR of dsRed. Binding of endogenous miR-16-5p to the dsRed mRNA would inhibit the translation of dsRed but not GFP (bottom panel).
E Histogram showing isoproterenol-induced heart hypertrophy in mdx NaCl (n=5) mice, surviving mdx (n=9), mdx/CD38-/- NaCl (n=9) and mdx/CD38-/- (n=9) mice, expressed as heart weight/body weight ratio (HW/BW). Data information: one value/mouse. After normality and variance comparison tests, significance was assessed using Kruskal-Wallis followed by Mann-Whitney tests Values are expressed as means ± SEM. Significance: *p<0.05, **p<0.01, ***p<0.001.
(E) PTEN protein modulation by miR-10b. Upper panel: PTEN protein levels in SKBR-3, and MCF-7 cells were analyzed by Western blot. GAPDH was used as a loading control with cells cultured as in (B). Lower panel: Densitometric analysis of Western blot assays performed with three biological replicates.
A hypothetical composite structure of Nrxn1 bound to both Nxph1 and Nlgn1 is shown. The Nxph1-LNS2SS2A+ complex structure was superimposed on the near full-length crystal structure of Nrxn1α (3R05) containing LNS2-LNS6 domains (Chen et al, 2011). Note that there are no coordinates for the Nrxn1α LNS1 or EGFA domains; structures shown here are models based on the EGFC and LNS6 domains of Nrxn1α (3R05). Nlgn1 from the crystal structure of Nrxn1β-Nlgn1 complex (3BIW) (Araç et al, 2007) is modeled by superimposition of that complex with the structure of Nrxn1α. Stalk regions, trans-membrane domains, and intracellular regions for Nlgn1 and Nrxn1α were modeled based on mouse protein sequences using PyMOL. For Nrxn1, the cysteine-loop was modeled based on the reported cysteine-loop sequence (Sterky et al, 2017), and glycosylation of the stalk region was modeled based on predicted O-linked glycosylation sites (Steentoft et al, 2013). Note the opposite effects on binding affinities of inserts at SS2 and SS4 for the Nrxn1 LNS2-Nxph1 and Nrxn1β-Nlgn1 complexes, respectively.
A. Various neurotransmitter genes as identified in 3E. Genes are marked (color-code) to represent the neurotransmitter receptor they encode for: Glycine (Red), Glutamate (Purple), GABA (Yellow) and Serotonin (Blue).
E. Heatmap of selected genes differentially expressed in 3C-CD34+α6+ and 3C-CD34¯α6+ cells. Note upregulation of multiple key HFSC lineage identity transcription factors and bulge markers (triangles) in 3C-CD34+α6+ cells, and upregulation of genes implicated in lineage progression in 3C-CD34¯α6+ cells (circles).
F BRCA2-deficient (-BRCA2) human DLD1 cells were treated with 2 µM pyridostatin (PDS) for 24 hours. Next, pyridostatin was removed and cells were released in a medium containing 250 nM NU-7441 (NU). Whole-cell extracts were prepared at the indicated timepoints after release and immunoblotted as shown. SMC1 was used as a loading control. KAP1 phosphorylation site is indicated in red.
iBMDMs stably expressing GFP-tagged PLK4 and parental iBMDMs were left untreated or stimulated with LPS for 4h. Anti-GFP immunoprecipitates were prepared for immunoblot analysis for the interaction between GFP-PLK4 and endogenous NEK7.
C57BL6xFVB mice were injected with G-CSF, CoPP or solvent controls (NaCl, DMSO) daily for 5 days. Samples were collected 6 h after the last injection. Cytokine and growth factor concentrations in plasma measured by Luminex assay. CoPP induces a group of cytokines (red box) which are not induced by G-CSF.
D, E Percentage of donor CD4+ and CD8+ cells gated within CD3+CD45.1.2+ cells in PB of CPA treated (D) or NC mice (E) from (B). Mean ± SEM.
D Model of substrate binding in the presence of GSIs showing substrate accumulation at exosites provided by NCT, PEN-2 and the PS1 NTF.
(C) 293A cells were transfected with plasmids encoding Flag-tagged NEMO and HA-tagged full-length M45, truncated M45, or an unrelated MCMV control protein (m142), respectively. Lysates were subjected to immunoprecipitation (IP) with an anti-HA antibody. Immunoprecipitates and the whole cell lysates (WCL) were analyzed by immunoblotting (IB) with the indicated antibodies.
B. Irf8-VENUS expression on A1 and A2 progenitor cells isolated from the E 9.0 yolk sac of Irf8-Venus PAC reporter mice. Representative blots obtained from 17 individual embryos are shown. Mean fluorescence intensity (MFI) of Irf8-VENUS signal in A1 and A2 progenitor cells. Each symbol represents one mouse. Data are from one representative experiment out of two and represented as mean ± s.e.m.
B. Similar to (A), but the experiments were performed at three time points, 45 min, 60 min or 75 min in anaphase extracts. Quantification of phosphorylation levels of T532 are shown. The intensities of pT532 signals in WT at each time point (45 min, 60 min or 75 min) were arbitrarily set to 1.0. The level of T532-phosphorylation was normalised to pan-Apc1. Error bars, SEM from three independent experiments.
Sequence alignment of miR-183 (G) and miR-96 (H) and their target sites in the 3'UTR of ATGL from various species.
C) Histoblasts Elongation Ratio decreases over time for all (black - the whole nest), central (blue) and peripheral histoblasts (red). D) Color coded snapshots of cell elongation at different time points. Color scale on the right. Scale bar = 40 µm. Data information: In (D), the top panel represents the central histoblasts and the bottom panel the peripheral. We employed an arbitrary annotation and consider peripheral those cells present in the 2 outermost rows of cells all throughout the perimeter of the nest at the earliest recorded time point.
8-week-old female C57Bl/6 mice were infected intranasally with 150 pfu of influenza A/Scotland/20/74 (H3N2) virus (IAV) and treated or not simultaneously with 4 mg of succinate (Suc; by the intranasal route). Some mice were euthanized at 4 days post-infection and lungs as well as BAL fluids were collected to determine: (a) the viral load by a plaque forming unit assay; Data information: Statistical analysis was performed using the Mann-Whitney test (a (*P < 0.05).
A) TLA analysis for ABCB1 contacts in RPE-1 parental and Taxol resistant clone sg6C9 covering the whole genome. Green arrow in sg6C9 shows a de novo interaction found between ABCB1 and a region in chromosome 6. B) TLA analysis for RPE-1 Parental and sg6C9. Zoom in in the region of chr6 with de novo interaction for sg6C9. Image modified from IGV viewer.
F In vitro ubiquitin discharge assay of UbcH7~Ub by PknG or its isopeptidase activity-dead mutant (PknG I87A). The reaction products were immunoblotted as in B.
(C) Western blot analysis of LC3-II and tubulin levels in MEF and SKNSH cells were treated as in (A) in complete media or glucose-free media in the presence of 100 nM YM-201636, 200 nM Wm, and 400 nM BAF for 4 hr.
(A) Upper panel: schematic representation of the 17-beta-dehydrogenase domain shared between the HSD17B1 and HSD17B7 enzymes with positions of the NADP and steroid binding sites and the active site (Breton et al., 1996). Two conserved charged residues in these domains, Tyrosine (Y) 193 and Lysine (K) 197, were changed into Glycine (G). Lower panel: closeup views of active site of HSD17B7 showing the NADP and steroid binding pocket and position of the Y193G and K197G amino acid substitutions.
(A) Wild-type (WT) K562 cells and indicated deletion isolates were treated with 100nM Bafilomycin A1 (BafA1) or DMSO for 18h and analyzed by flow cytometry for red:green ratio of tf-NBR1. Median values for each sample are identified by a black line within each violin. The red dotted line across all samples corresponds to the red:green ratio in wild-type cells. The red solid line across all samples corresponds to the ratio observed under maximally inhibited conditions (ATG9AKO cells). (n > 10,000 cells).
F) Overall survival in patients with high (top 50%) PTGES expression in relation to high (top 50%) and low (bottom 50%) NK cell gene expression signature.
(M-O) Area covered by diffusing CD22 with the nanocluster parameters or the diffusion coefficient of Cd22R130E. Data are from seven independent simulation runs.(M) Computer simulation of diffusing CD22 with the nanocluster parameters of Cd22R130E cells.(N) Computer simulation of diffusing CD22 with the median diffusion coefficient of Cd22R130E.(O) Computer simulation of diffusing CD22 nanoclusters with the real diffusion coefficient and nanocluster parameters for CD22 observed in wild-type and Cd22R130E cells. The graph shows the area covered.Data are from seven independent simulation runs.*P <0.01, **P <0.001, ***P <0.0001 (Student's t-test).
K. Quantification of data in (J) for S phase (EdU-positive) cells (red bars, mean).
(B) The percentage of each analysed SNARE observed in DRM fractions was quantified from blots (ImageJ densitometry analysis) and displayed as relative amount versus WT.
C Cells were stained for LAMP1 and mTOR. Note the increase in lysosomes and lysosomal dissociation of mTOR in UBE2QL1 depleted cells. Scale bar: 20 µm.
K. Inset showing the cavity formed through the centre of the putative Elp456 density. Black scale bars in this figure correspond to 50 Å.
(F) Validation of the binding specificity for the derived WW domains from the Hippo WW domain-containing proteins. HEK293T cells were transfected with the indicated SFB-tagged constructs and subjected to the pulldown assay.
(B) Immunohistochemical quantifications of fibrinogen depositions in tumour-bearing mouse lung after an injection of B220+CD11c+NK1.1+cells derived from tumour-bearing- FX+/− or FX−/− mouse liver. Sorted B220+CD11c+NK1.1+cells were prepared from three mice per each genotype, and were i.v. injected. Shown are averages (N = 6/group, 4 sections/mouse) with SEM and Welch"s t-test
K Re-introduction of MOAP-1 and MOAP-1-L120E, but not the p62 binding defective MOAP-1-M3, reduces Nrf2 transcriptional activity in the MOAP-1 KO cells. MOAP-1 KO LO2 cells were transfected with Myc-tagged MOAP-1, MOAP-1-L120E or MOAP-1-M3 and ARE-Nrf2-firely and renilla luciferase reporter constructs for overnight and subjected to luciferase assay.
J. A model delineating relationships between Rabl2 and IFT machinery. Rabl2-GTP enters cilia by binding to the Ift74-Ift81 heterodimer of IFT-B. After reaching the ciliary tip through anterograde IFT, it undergoes retrograde IFT towards the ciliary base. When its GTP is hydrolyzed into GDP during its IFT, the resultant Rabl2-GDP dissociates from IFT-B. The trafficking of IFT machinery continues in the absence of Rabl2.
Relative quantification of Coasy, Tfrc and Drd1 mRNA from GP by number of days post-cessation of treatment with 5μg of 4'-phosphopantetheine per g of body weight. n= 11 (WT-veh-D0), 5 (KO-veh-D0), 4 (WT-pPanSH-D0), 5 (KO-pPanSH-D0), 5 (KO-pPanSH-D1), 5 (KO-pPanSH-D2), 5 (KO-pPanSH-D3), 5 (WT-pPanSH-D7), 4 (KO-pPanSH-D7),
C, Olaparib and ETC-159 synergize to prevent the growth of HPAF-II xenografts in mice. NSG mice with established HPAF-II subcutaneous xenografts were randomized into four groups. Mice were gavaged daily with ETC-159 (10 mg/kg), Olaparib (50 mg/kg) or a combination of ETC-159 (10 mg/kg) and Olaparib (50 mg/kg). Treatment was initiated after HPAF-II tumors were established. (C) Tumor volumes were measured starting from day 0 and during the course of treatment as shown. Data points represent the mean ± SD. n = 7-8 tumors/group. p-values indicate significant difference compared to the vehicle group.
(A, B) Endogenous CAMK2B redistribution to ER membrane structures labeled by mCherry-FAM134B and BAP31 upon Tg treatment. Scale bars, 10 μm. The scale bars in the magnification boxes are 2 μm. The colocalization was analyzed by Pearson's correlation coefficient (PCC) in (B). For Ctrl, 16 cells were counted(n=16); for Tg treatment, 18 cells were counted (n=18). ***P<0.001, one-way ANOVA, error bars indicate SEM.
E Representative sections of brains from vehicle and DEBIO-0932 treated mice in (B-D). The dotted lines surround the metastases (GFP+). Representative field of view of metastasis stained with GFP and cleaved caspase 3. Scale bars: slices,1mm; cleaved caspase 3, 50 µm. F Quantification of established metastases found in vehicle and DEBIO-0932 treated brains from panel (E). Values are shown in box-and-whisker plots where every dot represents a different brain and the line in the box corresponds to the median. The boxes go from the upper to the lower quartiles and the whiskers go from the minimum to the maximum value (vehicle: n=10 brains; DEBIO-0932: n=14 brains). P value was calculated using two-tailed t-test. G Quantification of number of cleaved caspase 3 (CC3+) in cancer cells found in vehicle and DEBIO-0932 treated brains from panel (E). Values are shown in box-and-whisker plots where every dot is a metastatic lesion and the line in the box corresponds to the median. The boxes go from the upper to the lower quartiles and the whiskers go from the minimum to the maximum value (n=8 metastatic lesions from 4 brains per condition). P value was calculated using two-tailed t-test.
g, Representative flow-cytometry plots of Ki-67 and PD-1 among splenic CD8 Tem of 45-wk-old mice.
F. Gene ontology analyses for up- and down-regulated genes. Panel F summarizes top significant up- and down- regulated biological processes corresponding to each microRNA and comparison among them. Over-expression of miR-181a-5p and miR-148a-3p led to down-regulation of genes related to cognition and synaptic functions, while downregulated genes due to increased expression of miR-146a-5p represent ncRNA processing, defense response and protein folding mechanisms. The up-regulated genes due to overexpression of these microRNAs represent several processes including extracellular matrix, endoplasmic reticulum stress.
(B) Htt-Q97-induced toxicity is increased in cells silenced for HOIP. SH-SY5Y cells transiently transfected with Htt-Q25 or Htt-Q97 and either control siRNA (CO) or HOIP siRNA were analyzed by immunocytochemistry using an antibody against phospho-c-Jun. For rescue experiments, HOIP was co-expressed 24 h after silencing. Quantification of Htt-expressing cells with nuclear phospho-c-Jun is based on Student's t test from 3 independent experiments. Data are displayed as mean ± SD unless otherwise stated. *p ≤ 0.05, ***p ≤ 0.001.
H. Relative luciferase activity in HeLa transfected with plasmid containing WT BTAF1 3'UTR (white) or BTAF1 3'UTR on which the miR-132 binding site is mutated (grey) downstream of renilla luciferase, in the presence of miR-132-3p or miR-212-3p mimics. Error bars indicate SEM from eight replicate treatments. Data information: Significance in (H) determined by unpaired t-test. * p<0.05, ** p<0.01.
A, B HeLa cells stably expressing a wild-type β-globin reporter (A) or a β-globin NS39 NMD reporter (B) were mock-depleted or depleted of UPF2, GNL2, SEC13, RAB27A, RAB27B or both RAB27A and RAB27B. The level of the β-globin mRNA was monitored by quantitative RT-PCR relative to two reference genes (POLR2J and ACTB). The values shown are the average fold-change (mean ± SEM) from four independent experiments relative to mock-depleted cells (control). Statistical analysis was performed using the Mann-Whitney U-test for non-parametric distributions. *P < 0.05. The level of depletion of NMD factors is shown in Supplementary Fig S3C.
D, WB showing mutCC Trnp1 and ∆1-16 Trnp1 protein turnover in P19 cells 24 h after transfection in the presence of cycloheximide (CHX) for the time indicated below in hours (h). GFP (half live:24h) was used as a loading control.
Primary astrocytes from OTUB1fl/fl and GFAP-Cre OTUB1fl/fl mice were treated with IFN-γ (10 ng/ml) for indicated times and analyzed by western blot with indicated antibodies.
F. Western blot analysis of in vitro ubiquitinated HBP1 by the hGID 5-subunit complex in the presence of the different ARMC8 isoforms (α or β) (n=2).
b, Immortalized MEFs of the indicated genotypes were deprived of growth factors, glucose, amino acids or glucose and amino acids for 1 h and re-stimulated with glucose and/or amino acids for 10 min. Whole-cell lysates were immunoblotted for the indicated proteins.
C. Antibacterial activity assay of wild type and mutants of 12D1, 1D1108 and 15955 was carried out in a ratio of 30:1 against E. coli harboring the plasmid pRL662. The target E. coli cells were serially-diluted and grown overnight on gentamicin-containing LB agar prior to photographing. Each competition was done at least four times and reproduced in three independent experiments.
Western blot protein analysis of Gata6-WT and Gata6-KOmye pMɸ stimulated with 100 ng.ml-1 LPS for 6 h. Data shown are representative of at least 3 independent experiments. Western blot was performed on whole cell lysates.
25HC inhibits membrane fusion mediated by SARS-CoV-2 S (F) 293FT or Vero cells were treated with EtOH or 25HC (5 µM) for 16h prior to transfection with pLVX plasmids encoding SARS-CoV-2 S (F) in the absence of EtOH or 25HC. At 24h post-transfection, syncytium formation was visualized by fluorescence microscopy. White arrows indicate syncytia. Quantification of membrane fusion was performed by calculating the percentages of nuclei involved in syncytia formation from all nuclei in GFP positive cells.
B - High magnification of rosettes in Rx-Dicer mutants at E12.5 (Type 1), E14.5 (Type 2) and E17.5 (Type 3), immunostained for PH3 and Tuj1. Dashed line indicates ventricular surface; arrowheads indicate basal mitoses; arrows indicate a stream of apical mitoses connecting the lumen of the rosette with the lumen of the telencephalic ventricle.
Macrophages were treated for 24 h with CL097, ssRNA40, ssRNA41 or rapamycin (Rapa). (A) Immunoblot of co-immunoprecipitation of cell lysates with BECN1 specific antibody using antibody to PIK3C3 and BECN1 with antibody to β-actin as loading control.