EMBO/sd-character-level-ner huggingface dataset:
G Relative mTORC1 expression in pan-tumor samples segregated based on high (LDHAhi) (n=2917) and low LDHA (LDHAlo) (n=2914) expression. The p-value of paired Wilcoxon test following the adjustment multiple hypothesis correction (FDR). Data information: In the boxplots, the top and bottom horizontal lines represent the 75th and the 25th percentile respectively, and the middle horizontal line represents the median. The size of the box represents the interquartile range and the top and bottom whiskers represent the maximum and the minimum values respectively.
MT regrowth assay in centrinone-treated WT, pcnt KO and akap KO RPE-1 cells stained with antibodies to EB1 and giantin. At right, quantification of EB1 intensity at the Golgi membranes in WT and pcnt KO cells treated with centrinone, as a measure of MT-nucleation from the GA. Data were collected from two independent experiments and normalized to WT mean.
C Total counts of engrafted donor CD45.1+ cells in PB of mice from (B). Three independent experiment. Comparisons at day 18\24 were performed with an LME model, accounting for multiple experiments, followed by appropriate post-hoc analysis (see Appendix Supplementary Statistical Methods; ***p=0.0002 and ****p<0.0001 in all comparisons). CPA200 was not included in the analysis because n=4. Mean ± SEM.
C. Steady-state protein levels in three control and three FBXL4 patient-derived fibroblasts analyzed by western blotting.
B) Fluorescence polarisation assays against ubiquitin-TAMRA and ISG15-TAMRA using indicated SARS2 PLpro variants performed in technical triplicate and n=2 for each mutant, and compared to wild-type PLpro as shown in Appendix Fig S1A. Catalytic efficiencies were calculated as described in Methods.
HeLa cells were transfected with the indicated siRNAs (two control/non-targeting siRNAs and two PRMT1-specific siRNAs) and irradiated at 150 J/cm2 UVC or not irradiated. After 24 hours, cell cycle distribution was analyzed by flow cytometry using propidium iodide (PI) DNA staining for a representative experiment ((A), Appendix Fig. S8).
A. Injection schema for blood-retina barrier assay in adult Tspan12-/- mice with established retinal vascular defects.
C. Percent of host or donor CD8+ T cells in aged or naive hosts where each line represents all the CD8+ T the cells in blood from one mouse.
(P) Live/dead stain counts for non-adherent cells, recovered at 48 hpi from supernatants of SARS-CoV-2 infected Calu-3 cells, depleted for MAVS or RNA sensors, compared to siCtrl. Non-adherent cell counts were determined by acquisition by flowcytometry for a defined period of time. Mean +/-SEM, n=3. Total numbers are compared to siCtrl by unpaired t-test, *** (p<0.001).
(A) In vivo pull-down of HA-tagged SUMM2 by FLAG-tagged CRCK3.FLAG-tagged full-length or truncated CRCK3 and HA-tagged SUMM2 proteins were transiently expressed in N. bethamiana by agro-infiltration. FLAG-tagged CRCK3 proteins were immunoprecipitated with anti-FLAG conjugated beads (Sigma) and detected by Western blot using anti-FLAG antibody. HA-tagged SUMM2 protein co-immunoprecipitated with FLAG-tagged CRCK3 was detected by Western blot using anti-HA antibody. Similar results were obtained in two independent experiments.
C) Surface Stress radial kymograph of a representative time-lapse movie Phases are represented in different densities of brown. Stress values color scale is on the right.
, B. The SARS-CoV-2 spike protein S consists of the two subunits S1 and S2 and binds through S1 to ACE2 on host cells, which is required for syncytia formation (B). (B) For syncytia formation, the S protein expressed in an infected cell binds at the plasma membrane to ACE2 on another cell. This requires S priming by TMPRSS2 or ADAM10. The priming step may happen in trans while SARS-CoV-2 is still bound to ACE2 or after insertion of the S2 subunit - which contains the fusion peptide - into the membrane of the ACE2-expressing cell, which may be seen as a cleavage in cis. Inhibitors of TMPRSS2 (e.g. camostat) or ADAM10 reduce cell fusion and syncytia formation. The figure was created with BioRender.
Nine tissues/cells from the F1 hybrid of the two mouse inbred strains C57BL/6J and SPRET/EiJ were subject to the 3'mRNA-seq. Sequencing reads were mapped to the reference genomes of the parental strains respectively and classified as allele-specific reads or common reads. Only reads uniquely mapped in PASs annotated in the PolyA_DB were used for PAS usage calculation. Total PAS usage (calculated by the sum of allelic and common reads) in different tissues was utilized in within-tissue comparisons and analysis of APA patterns across tissues. The allelic reads were utilized to study allele-specific APA patterns and cis-regulatory divergence.
A. On the left, schematic of chr. VIIL variants, either with 2 or with 20 kb between a DSB and a telomere, used to study the effect of DNA damage checkpoint activation on cell viability in panels. Triangles represent HO sites, dashed lines - telomeric sequences, TG81 - 81 bp of (TG1-3) n and grey boxes - genes with the grey arrows above showing promoters. The diagram on the right outlines the DNA damage response (DDR) activation as the reaction of the two different stains on DSB induction by the addition of galactose (GAL).
The RNAi efficiency of Pten.1524 and Pten.932 shRNAmir mediated by miR-AB vs. miR-E was tested in NIH-3T3 cells in the context of <30% transduction efficiency (GFP as reporter). Representative western blots of Pten protein expression of the FACS-sorted GFP+ cells from two biological replicates were shown. β-actin served as loading control.
H 3D surface projection of two adjacent OHC from a mouse injected with AAV.Syne4. In the left cell myosin VIIa and DAPI signals were removed to only show FLAG staining.
Hierarchical clustering on the 20% of most correlated 297 genes with PC1 showed 2 major clusters that includes young HO-1+/+ LT-HSCs in one and old LT-HSCs with young HO-1-/- LT-HSCs in second.
(A, B) Mature S. aureus biofilms were allowed to develop for 24 h in polystyrene plates and then treated for the indicated time intervals with cell suspensions, or culture supernatants of the CV2 or the CV2-DispB strains (A) or the WT or the WT-DispB strain (B). Biofilm presence was assessed by crystal violet staining and included a negative staining control (i.e., crystal violet without biofilm). The results are expressed as mean ± s.d. of OD 595 nm absorbance values obtained from three different biological replicates (n=3) and statistically compared by one-sided ANOVA followed by the post-hoc Fisher's PLSD test: *P ≤ 0.05; **P ≤ 0.005; ***P ≤ 0.0005; ****P ≤ 0.00005
(B) Cytoplasmic and mitochondrial iron content was measured for Huh7 cells with DFP treatment (0, 0.01, 0.1, 0.5 or 1.0 mM as indicated) for 24 h. (n=5, biological replicates) The central horizontal bar and the error bards indicate mean ± SD. *: P<0.05, **: P<0.01.
f-i) Induction of autophagy in Bax/Bak−/− MEFs by etoposide. (f) Electron micrograph (×14,800) of Bax/Bak−/− MEF treated with etoposide (20 μM) for 18 h. Representative features of Bax/Bak−/− MEFs that have received no treatment (NT; that is, are healthy) are also shown (NT: ×14,000).
MST measurements of the heterocomplex (preformed using 50 nM HMGB1 and 50 μM CXCL12) in the presence of increasing concentrations of DFL. Decreasing MST signal (from ∼950 to ∼930 a.u.) upon DFL titration indicates that CXCL12 has been displaced from HMGB1. n=3; error bars represent avg ± sd.
D The mRNA levels of WT and the variant FOXL2 in control (left) and miR-1236-/- KGN cells (right) after transfecting siRNAs against AGO mRNAs were determined by allele-specific real-time RT-PCR. The data (mean ± SEM) are from three independent experiments, performed in triplicate.
Representative images (E) and nose-to-anus length (F) of male mice at 6 weeks of age (n=6-10 per group).
We selected six resources and five species for additional experiments. Resources are shown in relation to glycolysis and the citric acid cycle. Species are shown with a phylogenic tree with connections in the correct order but not to scale.
(F) The membrane fraction of the pancreas from the mice (A) was measured for Nox activity using luminometric assays. Data information: Data from 3 experiments are presented as the mean ± SD. One-way ANOVA test was used for statistical analysis of differences between groups, and P (*) < 0.05; P (**) < 0.01 and P (***) < 0.001 are considered statistically significant.
A. Representative blots of ATG13 (left) and VPS34 (right) phosphorylation levels in wt and KO-OFD1 cells transiently expressing GFP-ATG13 and FLAG-VPS34 in FM.
A Telomere restriction fragment analysis of genomic DNA from wt cells and cells carrying two tiTELs (CAF110) grown for 24 h in EMM medium containing THI and TSA as indicated. DNA was digested with HindIII and hybridized first with a nmt1 probe and successively with a telomeric probe. Marker molecular weights are on the left in kilobases. Numbers at the bottom indicate gel lanes.
C, D Hypocotyl lengths in seedlings of different genotypes. Seedlings were grown under continuous white light at 20 °C for 7 days or at 20 °C for 4 days followed by 3 days at 28 °C. Representative seedlings are shown in (C). Numbers in (D) indicate the ratio of hypocotyl lengths (28 °C/20 °C). Different letters above each box indicate statistically significant differences (ANOVA and Tukey's HSD; P < 0.05; n > 12). In the box plots, the thick lines indicate median values, the lower and upper ends of the boxes represent the 25th and 75th percentiles, and the ends of the whiskers are set at 1.5 times the interquartile range.
B. CYP26C1protein representation and luciferase assay. Variants found in CYP26C1 are indicated in green. Cignal-RARE system luciferase assays to test the impact of CYP26C1 variants on its RA degradation activity were performed in U2OS cells treated with 250 nM all-trans retinoic acid (ATRA) for 24 hours (n = 4). Experiments were performed in triplicates.pIRES2-EGFP empty vector was used as control. The residue C459 represents the Iron binding residue (Q6VOL0, UniProtKB) and was mutated to Ala and used as a positive control. TM, Transmembrane helix; P450, cytochrome p450 domain. RLU, Relative Light Units.
E Effects of ML-SI1 treatment on LFS-induced LTD in CA1 from 2-3 weeks old mice. Data information: Slopes of fEPSPs were normalized to the average values recorded during the first 10 min baseline Insets show representative traces of evoked fEPSPs before and 40 min after LFS. Scale bar 0.5 mV/10 ms.
MLE-12 cells were incubated with WT PR/8 + AM-EVs at 4 °C (to permit surface binding but not entry) for 1 h, washed, and processed for (B) RNA isolation (B) Data represent mean M gene transcripts relative to the means of the corresponding untreated condition, each normalized to β actin from 3 independent experiments.
Effect of Bumetanide on LARP6 binding to Col1a1, Col1a2, and Col3a1 mRNA in cultured MRC-5 cells. MRC-5 cells were treated with TGF-β1 and Bumetanide (2 μM) for 24 h. The cells were ultraviolet-cross-linked and RNA-bound proteins were immunoprecipitated with LARP6 antibody followed by RNA extraction for qRT-PCR. *P < 0.05 vs Vehicle, #P < 0.05 vs Control (two-way ANOVA); n = 6 for all groups.
Upper flow cytometry histograms of Ki67 expression in CD4 T cells from the representative subjects as indicated on the right, after stimulation with A549-SC3 cells. Vertical dotted line indicates the cut-off value of positive vs negative Ki67 expression. The percentage of Ki67-expressing CD4 T cells is shown within the histograms. Below, same data represented as a scatter plot from a sample of G1 and G2 donors as indicated, with relevant statistical comparisons with the test Mann-Whitney.
(F) LN229 cells treated with DMSO or thapsigargin (TG) were lysed and subjected to immunoprecipitation with a Merlin antibody or IgG. The immunoprecipitated products were subjected to western blotting. The number of conjugated ubiquitin (Ub) moieties was marked. Tetra-ubiquitinated Merlin is indicated by an arrow.
Scatterplot of mRNA expression and ribosome profiling data from control and YTHDF1 knockdown cells. The upregulated (red) and downregulated (blue) genes in translational efficiency (TE) are highlighted.
(e) Representative confocal images of muscle 4synapses stained for both DVGLUT (green) and FasII (red) in wandering larvae of wild-type (C155-Gal4/+), Atg1Gap-GFP (C155-Gal4/+; UAS-Gap-GFP/+; UAS-Atg16B/+), and Atg1NTAP-Rae1 (C155-Gal4/+; UAS-NTAP-Rae1/+; UAS-Atg1/+). Scale bar represents 10 μm. (f) Quantification of the number of boutons at muscle 4synapses in wild-type, Rae1 expression (C155-Gal4/+; UAS-NTAP-Rae1/+), Atg1Gap-GFP and Atg1NTAP-Rae1 wandering larvae (segment A2-4, n = 28, 27, 33 and 34, respectively). Error bars denote s.e.m. *P 0.001, **P 0.05.
Schematic of biotin-5caC DNA substrate pulldown experiment. 0.5 µM TDGwt or TDGcat (TDG(N140A)) incubated with biotinylated 5caC substrate (G•5caC DNA, 1 µM) and homoduplex competitor DNA (G•C DNA, 10 µM) as indicated and equimolar amounts of XRCC1-SUMO (>50% SUMOylated), POLβ and LIGIII.
B. RIP analysis using EZH2 antibody for the co-transfection of Flag-EZH2 and NDIME exons 1-3 (P = 0.0177) or exon 4 (P = 0.4335) in HEK293T extracts compared with IgG.
Representative polysome profiles from CPEB2-WT and -KO BAT. RT-qPCR analysis of the polysomal distribution of Ucp1L mRNA with RNA isolated from each fraction
(F) Kidney ILC2s isolated from C57BL/6 mice were cultured with Medium only or Medium with IL-33 and IL-2C for 30 minutes. The expression of phosphorylated STAT5 was examined in kidney ILC2s by flow cytometry. p-STAT5 (red lines) and isotype controls (gray-filled areas) are shown. Data represent the mean ± SEM of evaluations of MFI from each group (n=6 per group) and an unpaired t-test was performed; ***P<0.001. MFI, mean fluorescence intensity.
(l) NRK cell lines stably expressing LAMP1-Cherry;clathrin-GFP were transfected with nonspecific- or PIP5K1A-RNAi and starved for 8 h. Scale bar, 5 μm. The middle panels show enlarged regions of interest from the upper panels. The distance between clathrin dots on each reformation tubule in the upper panels is shown in diagrammatic form in the lower panels. Black line: reformation tubules; red dot: clathrin. (m) The distance between two clathrin dots on reformation tubules (RTs) in cells from l was accessed in a blind fashion. Twenty-five cells were counted. The average distance between two clathrin dots is indicated above each column. Uncropped images of blots are shown in Supplementary Fig. S6.
(e) Thirty-nine LC3 family sequences including isoforms A-C were aligned with ClustalW and displayed using WebLogo, with symbol heights corresponding to relative amino acid frequency. Asterisks denote residues predicted to contact CL in the favoured (magenta) and alternative conformations (cyan).
Chromosome maps of differentially expressed genes in cay1Δ cells shown as fold enrichment over wt. Red dots are overexpressed genes and Tf2 retrotransposons, blue dots are overexpressed LTR sequences.
(C) CD59-KO (CRISPR CD59) or control (CRISPR Control) CD4 T cells infected with HIV WT (left) or HIVΔVpu (right) (strain NLAD8) were cultured with the indicated antibody, and NHS or HIHS for 24h. The % of CDC was calculated as the % of disappearance of Gag+ cells after culture with NHS compared to HIHS. Each dot represents a single donor of CD4 T cells. n= at least 6 donors of CD4 T cells.
(a) Representative immunohistochemical staining of LC3 (green) along with NeuN staining (red) in WT and H3R−/− mice (scale bar, 25 μm).
G: degradation rate of Aβ1-42 by purified 6-His tagged PITRM1WT and PITRM1R183Q proteins. Quantification of these experiments is displayed below the blots. Experiments were performed in duplicate.
qPCR analyses of Fgf21 expression in 9-week-old livers (n=3-6 per group, 3 independent repeats). Ribosomal L32, loading control for qPCR analysis.
Figure 7. Schematic representations of mechanisms that may underlie brain maldevelopment in ASD. In control brains (top panel) both smaller and bigger brains develop and function normally. At the cellular level, there are no significant alterations of cell division phases and the number of cells produced is within normal variation. At the molecular level, there are no genetic alterations and changes in gene expression are within normal variation with no significant alterations of network structure and function. In ASD brains (bottom panel), brain development is abnormal and smaller and bigger brains represent two different anatomical and functional outcomes. In smaller ASD brains, the cellular characteristics are currently less clear compared to bigger brains. We hypothesize that G1/S phase transitions may be longer and/or checkpoints may stall/delay the timing of cell divisions leading to a reduced number of cells. Alternatively, there are an increased number of apoptotic cells. These cellular phenotypes may be related to genetic mutations (red lightning bolt) of Hc-genes that, for instance, regulate chromatin modification as to the case of CHD8 (blue). The mutation leads to altered regulation (red arrows) of downstream transcription factors or regulatory elements (yellow, cyan, green diamond shapes) that in turn regulate the expression of brain-relevant genes (purple square). Mutated CHD8 can also alter directly the expression brain-relevant genes, as to the case of CCNE2. Gene expression and networks functions are altered, but closer to normal brains. In bigger brains, cellular and molecular phenotypes are more pronounced compared to smaller ASD brains. Cellular evidence suggests that the increased number of neurons may be due to the shortening of G1/S phases. At the molecular level, this may be related to mutations and changes in gene expression that lead to a reorganization of networks controlling neuroprogenitor cell divisions. In addition to the downstream effects of Hc mutations (i.e. CHD8, red arrows), pronounced gene expression changes cause a substantial reorganization of the network with the activation of new regulatory genes (i.e. TOP2A, TYMS, triangle shapes) and new interactions (green arrows), thus leading to altered or different network functions.
B. Deconvolved fluorescence images of SINC-containing nuclei in vps4Δ and vps4Δpom152Δ cells expressing Chm7-GFP and Nup170-mCherry (green, red and merged images are shown). Scale bar is 1 μm.
E. Fluorescence imaging of GL261 intra-striatal grafts at 1 and 2 h after an intra-peritoneal injection of BODIPY FL prazosin, the green-fluorescent derivative of prazosin. Scale bar = 2 mm.
Heatmap showing the normalized expression (z score) of DEGs. Red indicates up-regulated, whereas blue indicates down-regulated genes.
A. Confocal images from the mito-QC-expressing fibroblast lines. Scale bar 25 μm. B. Quantification of the mitolysosomes/cell area (μm2) using the mito-QC counter plugin for ImageJ (Montava-Garriga et al, 2020). At least 3 technical replicates for each line and n>15 cells analyzed for each technical replicate. Data is presented as 25 to 75 percentile box with the indicated median and whiskers representing 5-95 percentile interval. Non-parametric Mann-Whitney test; ***, p<0.001.
C) Schematic diagram showing alteration of the Wnt signalling gradient upon Lats1/2 dKO in the intestine, with crypt expansion being accompanied by an elongation/flattening of the Wnt signalling gradient, whose elevated level gradually declines over time.
Analysis of mouse cortex tissue expressing mCherry in axonal projections after stereotaxic viral injections in the thalamus and post-hoc immunostained for ChgA as DCV marker. Representative confocal image and zoom of cortex tissue containing labeled axonal projections. Measurements of ChgA puncta (black) were performed within axonal projections (mask, yellow). Zoomed areas are indicated in top, right image (M). Scale bars: 10 µm (M), 5 µm (N
I Representative Western blot analysis of pAkt, Akt, pERK1/2 and ERK1/2 from MVF cell extracts. MVF were treated with vehicle or insulin for 3 h. Quantitative analysis of pERK1/2 / ERK1/2 (J) ratio in PI+MVF treated with vehicle or insulin for 3 h. Data are expressed as relative intensities (n = 3 each). Mean ± SD. Unpaired Student's t-test was used for statistical analysis. *P < 0.05 vs. vehicle.
(E) Hierarchical clustering of the expression profiles of genes differentially expressed both, between p38γ/δLyzs-KO and MCD-diet-fed Lyzs-Cre and between Lyzs-Cre treated with anti-Ly6G and MCD-diet-fed Lyzs-Cremice. Genes up-regulated in both comparisons (red cluster) were mainly regulated by PPARα, SREBF1 and SREBF2, while the enriched upstream regulators of genes down-regulated in both conditions versus the control group (blue cluster) were IL1β, TNFα and PDGFB β.
C Gene Ontology (GO) analysis of significantly enriched proteins utilizing PANTHER classification system.
(h) Quantification of the number of GFP-positive or GFP-LC3-positive bacteria. p62−/−/p62 (wild type, ΔLIR, ΔUBA, ΔLIR-ΔUBA or K7AD69A) cells were infected with ΔactA2 bacteria and stained with anti-bacteria and anti-Myc antibodies (Supplementary Information, Fig. S4e). Data are mean ± s.e.m., at least 500 LC3 were counted in each experiment (n = 3 experiments). P 0.001 versus p62-positive LC3 in p62−/−/p62 wild-type cells, P 0.001 versus GFP-p62-positive bacteria in p62−/−/ubiquitinwild-type cells. Ub, ubiquitin. WT, wild type. A full scan of g is shown in Supplementary Information, Fig. S7.
(e) Isolated acini before and after stimulation with 10 nM CCK for 30 min were stained for the lipid ceramide. The figure shows example images before stimulation and the averaged fluorescence intensity.
E In vitro methylation assays. Purified fusion protein of GST-tagged LSD1 WT or the four methylated arginine mutants (R-A), or four methylated arginine mutants in combination (4RA), were incubated with GST-CARM1 in the presence of 3H-SAM. Methylated proteins were detected via autoradiography and total amounts of proteins were visualized by Coomassie Blue staining (arrows: the position of LSD1; arrowheads: the position of CARM1).
Live imaging of a T cell using SIM shows the inter-foci connections (arrows in the inset) as predicted in the simulations; and their loss concomitant with loss of actin foci, as the T cell polarizes to initiate motility (compare insets between 0 and 120 sec). '0 sec' refers to the beginning of the observation of the cell, after it has attached to the substrate, spread and maintained synapse for 12 min. Scale bar, 5µm in main images and 1.5μm in the magnified insets.
(E) qRT-PCR analysis with total RNA isolated from HT-29 cells transfected with control siRNA or IRGM siRNA or doubly transfected with IRGM siRNA and IRF3 siRNA as indicated.
(c) H1299 cells were transfected with Flag-FoxO1(WT), SIRT2(WT) or SIRT2(168A). At 24 h after transfection, the cell lysate was extracted and immunoblotted with anti-Flag, anti-SIRT2, anti-LC3, anti-p62 or anti-β-actin..
D. Histological studies of testis sections from wild-type mice or mice with MLS deletion at different time points after birth. Scale bars: 50 m.
(B) Acetylated cohesinHalo488 was treated as in (A), or treated with CDK1 or Aurora B rather than Plk1. Sororin intensity on DNA was measured after immunofluorescence microscopy with anti-Sororin antibody (n > 100, *p < 0.0001, two-tailed Mann-Whitney test).
C Changes in pHi evoked by repetitive release (arrows) of cGMP from caged cGMP. The first UV flash was delivered at t = 0.
(f) H1299 cells were transfected with a control plasmid, Flag-FoxO1(WT), Flag-FoxO1(K262R), Flag-FoxO1(K265R), Flag-FoxO1(K274R) or Flag-FoxO1(KR). At 24h after transfection in the presence or absence of E64 and pepstatin A and immunoblotted, the cell lysate was extracted and immunoblotted with anti-Flag, anti-LC3, anti-p62 or anti-β-actin.
U2OS cells were transfected with the indicated siRNA oligonucleotides in a setup similar to the primary screen, but with 10 Gy of IR and in the presence of ATM and ATR inhibitors. Mitotic entry was analyzed by MPM2 and PI staining by flow cytometry. For normalization see materials and methods. Error bars represent the SEM of three independent experiments.