EMBO/sd-character-level-ner huggingface dataset:
(B and C) Fresh weights of calli after incubation on CIM (B) or CIM + SIM (21+ 21) (C). Box plots: centerlines show the medians; box limits indicate the 25th and 75th percentiles; whiskers extend to the minimum and maximum. Results were obtained from 3 independent experiments with at least 20 calli per condition, asteriscs mark statistical significance to WT according to the T -test (** P<0.001; *** P<0.0001).
Blood glucose levels of mice intravenously injected with either IgM isotype ctrl (n=6), anti-insulin PR-IgM (d85, n=5), anti-insulin primary IgM (d7, n=4) after Insulin-specific pulldown. Mean ± SD, statistical significance was calculated by using repeated measure ANOVA test showing comparison of red and black line, ***P < 0.001, ****P 0.0001.
(H) HDAC activity and its inhibition by panobinostat in B cells from CLL patients (red) in comparison to healthy donors (grey). Error bars indicate standard deviation measured in four biological replicates.
Treatment of HeLa cells with DFP (1mM for 24 h) decreases levels of CHIP (n=8, biological replicates; ***p<0.001; paired t-test).   Treatment of HeLa cells with DFP (1mM for 24 h) leads to decreased levels of CHIP mRNA (n=3, biological replicates; *p<0.05; unpaired t-test).
C The scatter plot depicted the colon length of Ccdc50+/+ and Ccdc50-/- mice treated with DSS (Ccdc50+/+ mice, n = 8 mice; Ccdc50-/- mice, n = 7 mice). Data information: Data are representative of at least five biological replicates and shown as the mean with SEM *P <0.05, ***P < 0.001; two-tailed unpaired Student's t-test.
(E) Morphologies of cells isolated from mouse lesions and analysed by SEM. Scale bar: 2 µm.
On the right, Western Blot (WB) analyses of the indicated protein expression in FaDu cells as in A. Actin was used as loading control. Right graph reports the quantification of protein expression normalized on Actin loading control. Data represent the mean (±SD) of three independent experiments. Unpaired t test was used to verify the statistical significance.
A, Western blot of DNA damage signalling in BRPF3 depleted cells. siRNA transfected U-2-OS cells were treated with HU for 2 h or 24 h. One representative experiment out of two biological experiments is shown.
B. The cumulative distribution of the proportion of constricting cells for the ftsA* mutant and its parent were plotted against cell age. We measured the degree of constriction of all cells using Oufti. For each strain, we had two independently acquired image sets (n = 496 and 1,168 cells for the WT replicates and n = 692 and 1,506 cells for the ftsA* replicates). The distributions of the degree of constriction were not significantly different among the four datasets at a threshold p-value of 0.01 using a Kruskal-Wallis multi-comparison test. Moreover, Bonferroni-corrected post-hoc pairwise tests did not allow a distinction between WT and ftsA* samples.
Representative CD20 immunoprecipitation using anti-pSer+pThr in (c) C2C12 cells treated with IGF1 for two hours (2 h) or overnight (ON) as indicated. Densitometric analysis of data are expressed as the ratio of CD20/vinculin or pSer+pThr/CD20 and are shown normalized to vinculin in arbitrary units in the lower panels shown Two-way ANOVA. *p<0.05; **p<0.01; ****p<0.0001. Each experiment was performed in triplicate wells. All values are expressed as the mean ± SEM.
(A) NF-κB luciferase assay in HEK293T wildtype (WT) and NOD1/2 knockout (KO) NF-κB reporter cells upon various stimulations.
Cells were infected with HCV for 72 or 96h (C). Some infected cells were treated with inhibitors of replication (DAA) the last 24 or 48h of infection (C). All HCV infections were done with a moi of 0.3. Levels of EGOT (A-E), or genome equivalents (GE) of HCV RNA per μg of total RNA (C-D) were calculated by qRT-PCR in RNA isolated from the control, treated or infected HuH7 cells. GAPDH expression was also evaluated in all cases and used as a reference to calculate the relative levels of each transcript. The experiment was performed twice in triplicate (n=6). Each value shows the average and error bars indicate standard deviations. Asterisks mark significant differences (ns, non-significant; * denotes p<0.05, ** p<0.01) obtained with a two-tailed non-parametric Mann-Whitney U-test.
D. miRNA-target duplex with miRNA nucleotides numbered from 5' end (Ago2 removed for clarity). Dashed line indicates disordered nucleotides in bridge region.
D, H1299 cells transfected with a Myc-tagged USP7 or a Myc-tagged USP7 together with a Flag-tagged p53 were subjected to IF analysis (upper). A549 cells transfected with a p53-specific siRNA or a control siRNA (siCont.) were examined by IF analysis, and the RNAi efficiency is shown (lower). Representative images are shown (magnification, ×40).
(e) Shown is the time course (left panel) and averaged 10 min secretion (columns) of the lysosomal marker acid phosphatase in the saliva collected from wild-type (black) and Trpml1-/- mice (red) stimulated with 1 mg/kg pilocarpine and 0.6 mg/kg isoproterenol. The mean±s.e.m is from 4 mice of each line.
(A) Absolute numbers of NK cells in one of the 4 draining lymph nodes in mice treated or not with the anti-CD62L antibody to inhibit NK cel ingress in the lymph node from blood. Where indicated mice were challenged with LPS.
(A) Immunoblots of murine wt non-tumorous and tumor tissue lysates were probed for CASP2, RAIDD, and p53. HSP90 and PonceauS protein staining served as loading controls.
A. Histograms of crossovers (blue) per F2 individual in the indicated populations, with the Poisson expectation plotted in red. Mean values are indicated by the black dotted lines. The genome average SNPs/kb for each cross are printed above the plots.
Expression of CBFs targets (E) in Col-0, atann1, and atann1 AtANN1 (#10) plants. Twelve-day-old plants grown on MS medium at 22°C were placed at 4°C for 24 h (E), and total RNA was extracted and subjected to qRT-PCR analysis. Relative transcript levels in untreated Col-0 plants were set to 1.
Genome-wide log2-ratio plots of chromosome 6 of two primary tumour biopsies and their corresponding non-regressed tumour showing no amplification in the primary tumour (upper panels) and a small amplification in the non-regressed tumours (bottom panels, yellow arrow).
E. Quantification of CD4+ (top panel) and CD8+ T cells (bottom panel) in lung circulation (Cir) or parenchyma (Par) 60 d.p.i.
Immunoblot analyses of TDG, TDG-SUMO in whole cell extracts of mESCs TDG and TDG-SUMO before and after H2O2 treatment (10 mM, 15 min).
Drp1 oligomerization by chemical crosslinking in wild-type CNs with or without rIFN-β for 2 h. B. Quantification of A.
a, Percentage of S. Typhimurium coated by the indicated galectins. HeLa cells stably expressing YFP-tagged galectins were treated with the indicated siRNAs. YFP-positive bacteria were counted by microscopy at 1 h p.i. siRNAs are further characterized in Supplementary Fig. 2.
K. The ratio of FACL4 immunogold particles on the MAM and mitochondrion. n = 80 MAMs (one MAM means one mitochondrial ER tethering site in the corresponding mitochondria).
A. CryoEM structure of EAAT1WT trimer with tranDs and scaDs depicted in magenta and teal, respectively. Non-protein extra density corresponding to lipid/detergent molecules is depicted in grey.
shCTRL- and shNAV3-MDA-MB-231 cells were plated in invasion chambers, and 18 h later, invaded cells were stained. For 3D growth, cells were seeded in Matrigel and photos were taken 5 days later (scale bars, 200 μm and 50 μm).Histograms (means ± SD) of the results presented in (A) (from three experiments).
B Body weight measurements were performed for indicated time points in ERp57WT (n=50), ERp57Nes+/- (n=32) and ERp57Nes-/- (n=19) mice.
B. Mixed neuroglial, neuron-enriched, and glial cortical cultures were assayed for the presence of LAG3 where no band could be found. Activated T cells display band at expected size.
C) By mutually exclusive inclusion into transcripts, MXEs of a cluster are supposed to encode the same region of a protein structure. If the respective regions of the protein structures are embedded within secondary structural elements (the ends of the exon-encoded peptides are part of α-helices and/or β-strands), it is highly unlikely that the translation of a transcript will result in a folded protein in case the respective exon is missing (skipped exon). If the MXEs have highly similar sequences and do not encode repeat regions, it seems unlikely that either could be present in tandem or absent at all in a folded protein. Here, we have combined protein structure features (colours) with splicing regulation information (symbols). Accordingly, 87% of the MXE encoded protein regions are embedded in secondary structural elements (orange and green symbols), and most of the remaining MXEs can only be spliced mutually exclusive because splicing as differentially included exons would lead to frame shifts (blue circles). As examples, we labelled many MXE clusters distinguishing annotated MXEs (purple letters), known exons that we validated as MXEs (orange letters), and clusters containing novel exons (dark-grey letters).
Heatmap of k-means clustering derived from mean-centered expression values (log2 scale) of DEGs selected from 6000 DEGs (Fig EV4A) obtained from RNA-seq (Fig 2). See also Dataset EV1.
(C) Representative confocal laser-scanning microscopy pictures of biofilm development of bacteria cultured for 4 d in minimal medium in chambered cover slides under light or dark conditions. The scale bar represents 30 µm.
Gli1 mRNA expression levels upon SAG exposure in MEF cells (n=7 WT and n=9 null samples) (F; SAG 100 nM) and cortical neurons (n= 15 WT and n=12 null samples) (G; SAG 400 nM). The graphs show the mean ± S.E. of the fold change derived from at least three independent experiments (mean ± S.E., **p<0.01, by two-way ANOVA, followed by Bonferroni post hoc test).
F, Electron microscopy pictures from 150 INS-1 cells transfected as described in G.
(A) U2OS cells were left untreated, exposed to nutrient starvation for 6 h or to metabolic stress for 24 h, with or without treatment with AO1 (NAC (2 mM) and L-ascorbic acid (2 mM)) for 24 h. The percentage of cells with GFP-LC3 puncta was calculated, and data are shown as the mean and standard deviation from three independent experiments.
Immunohistochemistry of dystrophin-positive fibres in body-wide muscles from mdx mice treated with repeated doses of PMO-M (n=4) or PMO alone (n=3) (scale bar: 100 μm) (*p<0.05, **p<0.001; two-tailed t test).
(f, g) Immunoblots (IB) of polyubiquitin (f) in starved C38 and IB3-1 cells. Uncropped images of blots are shown in Supplementary Information, Fig. S10.
Expression of SBP enzymes PHGDH and PSPH in the METABRIC dataset. The expression of a proliferation related gene set and ATF4 are also shown. Samples were stratified by Pam50 intrinsic subtypes and ER status. Brown-Forsythe and Welch ANOVA test with unpaired t-test with Welch correction were applied. *p<0.05, **p< 0.01, ***p<0.001, ****p<0.0001.
(d) HCT116 cells were treated with AGK2 for 24 h at different doses, as indicated. The cytosolic lysate was extracted for co-immunoprecipitation with anti-FoxO1, followed by probing with anti-acetylated lysine.
miR-AB shRNAmir cloning strategy. A shRNAmir sequence obtained from SplashRNA or shERWOOD platform are converted to two oligos (75 bp and 67 bp respectively) by Microsoft Excel-based miR-AB Oligo Tool. These two oligos are synthesized, annealed and ligated to BamHI/ApaI-cut miR-AB viral vector for transfection into 293T cells to produce miR-AB virus.
E) Graph comparing the median distance of REC8 foci with computationally predicted even and random distributions. Median distances between foci subdivided in three categories (11-14, 15-18 and 19-22 REC8 foci per chromosome). Purple asterisk indicates the median distance of all REC8 foci.
(h,i) Immunoprecipitation (IP) and immunoblot (IB) analysis of HEK293 cells transfected with Flag-ΔN85-ATG16L1 (h) or Myc-FL-ATG16L1 (i), together with HA-Nod1, HA-Nod2 or HA-tagged enhanced GFP (EGFP-HA). Ig, immunoglobulin (loading control). Results are representative of one of at least three independent experiments (a-g) or one of three independent experiments (h,i).
(A, B). H3K9me3 association with the Xist promoter (P1 and P2) in two independent Rif1+/+ and Rif1-/- cell lines, analysed by ChIP-pPCR in mESCs (A) and 2d EBs (B). Myf5 serves as a positive control region, β actin negative. Average ±standard deviation of two independent experiments.
E. Detection of HA-tagged Sufu binding to a Myc-tagged Fbxl17 construct encompassing the residues 318 to 701 (corresponding to the F-box motif and C-terminus region) assessed by in vitro binding assay. Both proteins were synthetized in vitro using a T7-coupled reticulocyte lysate system.
(H) TMR-dextran-based assessment of vascular permeability in vivo, comparing heart homogenates from control and Ntras-silenced mice. Data normalized to organ and body weight (n = 11-12 mice per group). Experimental outline on the left. Data information: data are represented as mean ± S.E.M. n.s.: non-significant, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. H) two-tailed unpaired t-test
Ultrastructural analysis depicting in the nucleolus (arrows for nucleolar interstice) (C), condensed chromatin blocks (arrows) (D) and nuclear speckles or interchromatin granule clusters (arrows for dense cords) Nu: nucleolus; G: granular component; D: dense fibrillar component; FC: fibrillar center; C: condensed chromatin.
(B) Structures of S. cerevisiae Hrr251-394 K38R bound to CK1-7 (Chijiwa et al, 1989; Xu et al, 1996) and C. glabrata Hrr251-403 K38R bound to ADP, with domains colored as in (A). Bound PO4-/SO4- ions are shown as spheres. See Figure EV2A for additional C. glabrataHrr25 structures, and Figure EV3A for SO4- ion electron density.
(C) Flag-tagged Raptor was transfected with HA-tagged ArfGAP1 or HA-tagged RFP1 in wildtype (WT) or RagA/B knockout (KO) HEK293A cells for 24 hours. Cell lysates were immunoprecipitated (IP) with anti-Flag beads. IP or WCL samples were probed for HA-tagged ArfGAP1 or HA-tagged RFP1, Flag-tagged Raptor, RagA, RagB, and Vinculin the loading control.
(D) Mitochondria recruitment experiment with wild-type PI4KB and c10orf76. Left: AKAP1-FRB-CFP is localized to the mitochondria before (top) and 5 minutes after rapamycin (100 nM) treatment (bottom). Middle: mRFP-FKBP12-PI4KB at high expression level is located in the cytosol before rapamycin (top) and translocates to the mitochondria after rapamycin induction (bottom). Right: eGFP-c10orf76 at a high expression level is located in the cytosol before rapamycin (top) and translocates to the mitochondria after rapamycin induction (bottom).
A H4 Cas9 cells stably expressing N-terminally (Myc-TMEM41B) or C-terminally Myc-tagged TMEM41B (TMEM41B-Myc) were probed by calnexin (CANX) and Myc tag immunostaining and imaged with an automated CV7000 confocal microscope. Scale bar: 20 μm
(C) Endonuclease activities of the WT and mutant TmMR proteins towards a bubble DNA molecule in the absence (left panel) or presence of ATP (right panel). See Appendix Fig S4C and S4D for the quantified values.
B, Cells stably expressing NPC1-mCherry were treated with control or OCRL siRNAs for 2 days, or transfected with GFP-ORP2 or -ORP2-mHHK (PI(4,5) P2 binding-deficient mutant) for 1 day, stained with PI(4,5) P2 antibody and imaged by confocal microscopy. For FAK inhibition, GFP-ORP2 transfected cells were treated with 10 μM PF228 for 4 h. Dashed lines indicate cell edges. Arrowheads indicate colocalization of PI(4,5) P2 and NPC1. C, Quantification of B. Values from Ctrl siRNA and Vector control were pooled and are shown as Ctrl siRNA/Vector ctrl. Mean ± SD, n = 20-43 cells pooled from 2 independent experiments. Student's t-test.
TNBC cells were pretreated with specific antagonist against Smad3 (SIS3, 3μM) for 1h and then cultured with or without TGF-β. The indicated proteins were determined by immunoblotting analysis.
(a) Study overview and data types. Data modalities are shown in different rows (D = number of features) and samples in columns, with missing samples shown using grey bars
d, Average normalized responses of EGFP-positive TRPML1Va-transfected cells (typically n = 20-40 cells) to 1 or 10 mM Fe2+ (pH 4.6).
(I-J) RAB21 knockout reduces endosomal F-actin levels. (I) Immunofluorescence of endogenous F-actin using Alexa-488 conjugated phalloidin and EEA1 in RAB21-depleted cells. Boxed region is magnified and single channel images are depicted. (J) Per cell Pearson Correlation between internal F-actin and EEA1; SEM, n=3 independent experiments. Data information scale bars represent 10µm or 2.5µm in enlarged views. Statistical tests used: Mann-Whitney tests. All error bars are SEM. Number of cells presented on each graph represents the total number of cells analyzed for all repeats.
LBPA intensity in a mixed population of cells expressing HRH3-GFP. After transfection with HRH3-GFP, uncloned stably expressing cells (C) were labelled with anti-LBPA antibodies and analysed by automated microscopy (C). Unbiased quantification (D) shows the inverse correlation between HRH3-GFP expression (high expressing cells in green) and the endosome integrated intensity of LBPA staining (high LBPA labelling in red). (n=2 independent experiments with 500,000 cells analysed automatically, error bars = SD).
Fat body nuclei increase in size during development. Graph shows mean ± SD for each day during development of larvae. DO, day-old.
(F) Gr-1 immunostaining of spleens isolated from WT and FAKΔmyeloid mice 5 days post-infection with S. typhimurium strain SL1344. Panels b and d are enlarged sections indicated by black boxes in panels a and c. Gr-1-positive cells appear red-brown.
(D) The postnatal day of hair coat recovery by 25%, 50% 75% and 100% in shaved Sirt7f/f and Sirt7f/f;K15-Cre mice. n = 6 mice per genotype. Box-and-whisker plots: mid-line, median; box, 25th and 75th percentiles; whiskers, minimum and maximum.
A The senescence phenotypes of five-week-old vector control, proSAG12::CDF4 and proSAG12::CDF4 & 35S::CAT2 plants, from old to young. Scale bars indicate 1.5 cm.
D CRISPR-Cas9 system mediated NAT10 gene editing. The genome targeting for knockdown NAT10 was performed as described in Experimental Procedures. The HCT116 NAT10 knockdown cell lines were harvested and western blot was performed on the cell lysates for the indicated proteins.
An NMD activating mRNP composition will result in the activation of NMD in the presence of UPF3B (left part) or when UPF3B was depleted (middle part). In the absence of UPF3A and UPF3B, NMD is inactive (right part). In normal cells, UPF3B can engage in multiple, potentially transient interactions (depicted by blue sphere), e.g. with the terminating ribosome via eRF3a, with UPF2 or with the EJC (left). When UPF3B is missing, UPF3A can at least partially compensate for it (depicted by pink sphere, middle). UPF3A interacts with UPF2 and possibly also with eRF3a. The interaction of UPF3A with the EJC is weaker than that of UPF3B, therefore UPF3A cannot establish a stable bridge between UPF2 and the EJC (middle).
D. Southern blot analysis of lysates obtained 72 h post-transfection of HepG2 cells with the same plasmids as described in panel A.
HeLa cells expressing GFP-centrin1 arrested at prometaphase or forced mitotic exit (Ro3306 5') and stained with phalloidin. Scale bar- 5µm and for zoom-2µm.
C. Relative levels of metabolites in glycolysis and the TCA cycle.n=9 biological repeats; upper whiskers, maximum-the third quartile; lower whiskers, the first quartile-minimum; boxes, the third quartile-the first quartile; the central line indicates the median; p-values were calculated using two-sample t-test.
Percentage of iNKT cells in total live cells in FTOC with or without BTP2 (1uM) addition at the start of culture (day 0);
I-J. Axin2 mRNA levels in the above 50 GBM specimens were examined through RNAscope technology. The staining was scored as 1-12 according to the intensity and percentage. The significance of the correlations was determined by Pearson's correlation test.
D Protein degradation in indicated yeast strains monitored by pulse-chase experiments for the Hrd1 model substrate PrA*-3xHA (left) and by CHX decay assays for the Doa10 model substrate Deg1-eGFP2 (right). Values for each time point are reported as means ± standard deviation (n = 4 for PrA*-3xHA and n = 3 for Deg1-eGFP2).
C, D FRAP analysis of dendritic lysosomal movement in Raptor KD neurons. C Representative images of a neuron at 0 and 4 min after photobleaching. D Quantification of the lysosomes in C undergoing retrograde (red) or anterograde (cyan) transport (N = 7 and 8 neurons for shSc and shRaptor, respectively, from 3 independent experiments). Data information: Data with error bars are represented as means ± SEM. Statistical significance was assessed by Mann-Whitney U test ***P < 0.001 as compared to shSc; n.s., not significant.
(a) Subcellular localization of ceramide, generated by wild-type CerS1 induction (+ tet), was visualized in mitochondria by colocalization of ceramides and mitochondria with anti-ceramide and anti-Tom20, respectively, using confocal microscopy.
A. Schematic of the predicted domain architecture of Chm7 and truncations. Green circle is a Chm7-specific insertion (relative to Vps25; see Appendix Fig S1). MIM is microtubule interacting motif. Red stripes denote putative autoinhibitory helix. Numbers are amino acid residues.
MEFs were immortalized with the adenoviral oncogene E1A.12S (E1A MEF) and treated with 0.4 µg/ml Doxo for 17 hours. Apoptosis (Annexin V) was quantified by flow cytometry. +/+ and -/-: n=3; EE/EE: n=6. Western blots show expression of E1A and β-actin as loading control.
c) SAFB phosphopeptide abundance changes during SARS-CoV-2 infection. Vero cell phosphopeptide data from(Bouhaddou et al, 2020). Data previously acquired from Vero cells infected with SARS-CoV-2 (Bouhaddou et al, 2020). S604 phosphosite that temporally coincides with SAFB thermal destabilisation (Potel et al, 2020). Shown above the line plot are the thermostability and abundance profiles of SAFB in SARS-CoV-2 infected Caco-2 cells as described in 1A (n=3). Asterisk denotes significant regulation (see methods).
The promoter of sigA and mec was fused to luciferase, and transformed cells were either left untreated or exposed to CHP stress for 6 h and luciferase activity was measured (mean ±SD; n=6)
(D) HeLa cells were transfected with the indicated myc-SNX18 plasmids for 6 h and starved for an additional 2 h before harvest. Equal parts of cytosol (C) and membrane (M) fractions were analyzed by Western blotting. The graph shows the average membrane-bound myc-SNX18 ± SEM (error bars), n = 3.
(C) Duolink proximity ligation assay between Cdc25C and CLU in PC3 cells. Confocal microscopy was used to detect the interaction (red dots). DNA was counterstained with DAPI (blue). PC3 cells transfected with siCLU were used as a negative control. Scale bar represent 10 µm.
(E Immunohistochemistry of dystrophin-positive fibres in body-wide muscles from mdx mice treated with PMO at 12.5 mg/kg/week for 3 weeks mixed with MOTS-c (500 μg) (PMO-M) intravenously (scale bar: 100 μm) (n=3; *p<0.05). PMO represents PMO in saline.
C. The percentage of cells with Q97-GFP aggregates were quantified in control (mock transfected) and MIA40-expressing cells. Data are displayed as mean ± standard deviations and were analyzed by two-tailed Student's t-test, n = 6.; * P ≤ 0.05.
(F) Codon-specific ribosome occupancies for 21 nt RPFs from WT and eEF3d cells (two biological replicates). CGA and AGG codons are shown in red and purple, respectively. Variance for each dataset is indicated.
B TRAF6 mRNA expression by human Tregs and non-Treg CD4+ T cell. Human Tregs (CD3+/CD4+/CD8-/CD25HIGH/CD127low/CD39+) and non-Treg CD4+ T cells (CD3+/CD4+/CD8-/CD25-) were obtained from the peripheral blood of healthy donors by FACS after Ficoll-Paque PLUS gradient centrifugation and magnetic bead enrichment of CD4+ T cells. TRAF6 mRNA was measured by qRT-PCR.