EMBO/sd-character-level-ner huggingface dataset:
Representative images (above) and quantification (bottom) of TRF1 telomeric foci in ERK1/2 RNA interfered p53-/- MEFs. Scale bars, 5μm. Data are representative of n=2 independent experiments.
Decreased Drp1S616 phosphorylation in PINK1-null mouse substantial nigra tissues. Lysates of PINK1-null (PINK1KO) and parkin-null (parkinKO) mouse substantial nigra tissues (e), and their matched wildtype controls (WT) were immuno-detected with antibodies against either phospho (Ser616)-Drp1 (pS616), phospho (Ser637)-Drp1 (pS637) or Drp1. β-actin was detected as a loading control. Quantitation of pS616/Drp1 for each experiment is shown Two independent PINK1KO and parkinKO substantial nigra from two separate PINK1KO and parkinKO mice (1, 2) were shown. Student's test. *p<0.05, ***p<0.001, ns: no significance. Data was presented as mean ± SEM of three independent experiments.
(e-g) Effect of inhibition of macroautophagy by treatment with 3-methyladenine (3-MA) on the lysosomal content of KFERQ-PS-CFP2 in cells maintained in serum-free media for 16 h. (e) Immunoblot for the indicated proteins of the same fractions as described in (b). The amount of KFERQ-PS-CFP2 associated to lysosomes (f) and the changes in the intracellular content of KFERQ-PS-CFP2 were calculated as in (c, d). Values are mean+s.e. of three different experiments and differences are significant (*) for P=0.032 (t-test).
E, In vitro ubiquitination of TRAF2 in the presence of UbcH7, Ub, and PknG or its mutants. The reaction products were immunoblotted using antibodies against TRAF2 or TAK1.
knockdown JAK2 by siRNA (I, J) abolished hTau-induced STAT1 phosphorylation at Tyr701 in total extracts I) and the nuclear fraction J) (n=3). Data information: Data were presented as mean ± SD, *, p<0.05 vs Ctrl, eGFP or WT.
B-F: GFP-MOSPD2 (green) expressing cells were transfected wit Flag-STARD3 (C and labelled using anti-Flag (B-E; magenta) and anti-Lamp1 (B-D; red as markers of late endosomes/lysosome Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm
Relationship between the extent of EMX2 upregulation induced by EZH2i and the DNA methylation level at the EMX2 promoter in GBM cell lines. Values for EMX2 upregulation are the fold-change in mRNA expression induced upon treatment with 10μM EZH2i (see: panel G), and are expressed relative to a DMSO control. Values represent mean ± SEM from three technical replicates. The SEM is indicated to show reliability of the RT-PCR values, due to the low endogenous levels of EMX2. Methylation values are averages of DNA methylation at CpGs across the promoter of EMX2 in GBM cell lines. All data sourced from the CCLE. The significance of the anti-correlation between mRNA levels and DNA methylation levels across cell lines is indicated (Spearman rank correlation).
Western blot showing STN1 knockdown. shLUC (siRNA targeting luciferase) was used as the control.
(d) GFP-tagged empty vector or GFP-BRLF1- or GFP-BRLF1 L578A-expressing plasmids were transfected into A549 cells, after which the cells were then infected with HSV-1, or transfected into HNE-1-EBV-ΔBRLF1 cells. The cells were then collected and lysed. Immunoprecipitation with GST-affinity beads was subsequently performed, and then whole-cell lysates and immunoprecipitated complexes were analyzed as indicated.
A. Mitochondria were visualized by mitochondrially targeted mCherry (red) in WT cells expressing Q97-GFP for 4 h. Fluorescence micrographs are maximum intensity projections of z stacks subjected to deconvolution. Bar, 5 µm.
Demographic representation of HADA labeling. Bacteria were sorted according to their cell length and oriented with the old pole at the bottom of the demograph by PdhS-mCherry fluorescence signal. NFI: normalized fluorescence intensity. n = 393 bacteria (3 biological replicates).
(b) Δ133p53α is ubiquitinated at the C-terminal lysine residues. MRC-5 fibroblasts expressing wild-type FLAG-Δ133p53α in the absence of STUB1 knockdown were used in IP with anti-FLAG antibody, and the resulting immunoprecipitates were analysed using mass spectrometry. Representative MS/MS spectrum of an [M+3H]3+ peptide from FLAG-Δ133p53α shows sites of ubiquitin modification. The peptide sequence for the spectrum is shown, with the sites of modification marked as asterisks: amino-acid positions 248 and 249, which correspond to positions 381 and 382 in full-length p53. The same peptide was also observed in the unmodified state.
(C) Three-dimensional side view of the A-C linker from the lumen of the centriole.
A. Inhibition of p38 or JNK does not block osmotic stress-induced YAP nuclear localization. HEK293A cells were pretreated with 2 μM p38 inhibitor (SB203580) or 20 μM JNK inhibitor (SP600125) before treatment with 0.2 M sorbitol for 1 hr in the absence of serum. Cell lysates from identically treated samples were examined for phosphorylation of p38 substrate MK-2 and JNK substrate c-Jun.
E. Correlation between H-DNA forming sequences and all genes (white bar), and genes with DRIP peaks in wild type (blue) and primpol cells (red). Data information: P values calculated with Mann-Whitney U test. In violin plots, bar = median; box = interquartile range (IQR); whiskers = upper and lower inner fences (1st / 3rd quartile + 1.5*IQR).
(A) Immunoblot of brain tubulin with or without carboxypeptidase A (CPA). Note CPA specifically removes the α-tubulin C-terminal tyrosine without affecting other PTMs.
(b) Bif-1+/+ and Bif-1−/− MEFs were transfected with GFP-LC3, cultured in LC3 or complete medium for 3 h, then analysed by fluorescent microscopy. The number of GFP-LC3 dots per GFP-positive cell was counted (mean ± s.d.; n = 35).
(C) Representative images of SOD1G93A/ChAT::tdTomato or WTChAT::tdTomato neuromuscular junctions at P90. White: denotes BTX, Red: denotes direct ChAT, Green: denotes CRMP4, Yellow: denotes Z projection of 3D Imaris co-localization of CRMP4 and ChAT. Scale bar: 10 µm. (D) Quantification of CRMP4 positive NMJs in gastrocnemius muscles from 3 WT or 3 SOD1G93A P90 mice. Total of 44 NMJ's in WT condition and 60 NMJ's in SOD1G93A condition. Student's t-test, n=3, Data presented as mean ±SE, *p = 0.0157. (E) Quantification of the percent of partially denervated NMJ's in the presence or absence of CRMP4 immunostaining in 3 different SOD1G93A mice. We counted 24 NMJ's in SOD1G93A CRMP4 negatives and 67 NMJ's in SOD1G93A CRMP4 positives. Student's t-test, n=3, Data presented as mean ±SE, *p = 0.0352.
B Volcano plot analysis derived from FPKM values showing DE genes (p<0.05, Student's t test) in sorted MitoTneg and MitoTpos CD3+ cells (from 4 different donors of PBMCs).
(d) Miz1-binding sites contain a conserved sequence element. Sequences (±50 bp) around all 261 Miz1-binding sites observed in NPCs were analysed by MEME-ChIP ('Motif Analysis of Large DNA Data sets').
(C) (upper panel) LC-MS/MS analysis was performed to measure 15N-glutamine-labeled purine synthesis intermediates in BGC823 cells transfected with or without the WT-UHMK1 or UHMK1-K54R constructs. (lower panel) LC-MS/MS was used to analyze metabolites labeled with 13C-glycine in BGC823 cells transfected with or without the WT-UHMK1 or UHMK1-K54R constructs(three biological replicates). Data information: *P< 0.05, **P< 0.01, ***P< 0.001, # marked no significance.
B Representative in vivo and ex vivo images of vehicle and DEBIO-0932 treated mice 5 weeks (experimental endpoint) after intracardiac inoculation of H2030-BrM cells.
E Colocalization of Hwa-Flag (red) and ZNRF3-HA (green) or ZNRF3ΔRING-HA (green) in Hela cells is indicated by yellow dots. Baf-A1, 50 nM, 12 hrs. Magnification: 100 ×; scale bar: 10 μm.
Primary ovarian tumour cells were characterised for AXL and OPCML levels by (I) Western blottin PAX8 was used to identify ovarian cancer cells
MEFs were stimulated with IFNγ for 24 h (100 U/ml). IFNγ-stimulated MEFs were infected with Pru, PruΔgra15 or RH for 3 h and subsequently fixed, permeabilized and stained for (C) total ubiquitin, For analysis, at least 100 vacuoles were scored. All experiments were performed 3 times. On the right-hand side, a representative fluorescent image is shown for the Toxoplasma Pru strain, which expresses GFP. DNA was stained with Hoechst 33258. Scale bar is 10 µm. The yellow box inside each representative image is shown as an inset picture with magnification. All experiments were performed 3 independent times.
J Co-IP assays of SCR with SEU and SDG4 in Arabidopsis. Proteins extracted from SDG4-myc and SDG4-myc SCR-GFP plants were immunoprecipitated using anti-GFP antibody, and immunoblotted using anti-myc and anti-SEU antibodies.
d, Percentage of cilia that remained intact, underwent ectocytosis during live-cell imaging from (a) and (c). Three 3 biological replicates, n>100 cilia per sample and repetition. Data include mean ±s.d. and P values are calculated by two-tailed unpaired student t-test.
C Level of histone gene mRNAs was determined by qRT-PCR. Data are the mean of four independent biological repeats and error bars are ±SEM. Two-tailed unpaired t-tests showed no significant differences (P > 0.05) between wild type and abo1Δ cells.
(F) Schematic diagram of tumor resection in inguinal lymph node pre-removed or sham control 4T1 tumor-bearing mice.
I Rhizosphere acidification of MdCAX3-suppressed in leaves Md/Mx. Bromocresol purple was used as a pH indicator for visualization. Scale bars: 1 cm.
(B) Schematic representation of the areas containing GnRH migrating cells during embryogenesis at E14.5 and representative immunofluorescence showing migrating GnRH neurons at the level of the nose, the olfactory bulb (OB) and the ventral forebrain (VFB) in control and mutant embryos. Scale bar: 50μm. VNO, vomeronasal organ; OE, olfactory epithelium; OB, olfactory bulbs; VFB, ventral forebrain.
F Representative kymographs depicting EB1 comet movement patterns in the proximal dendrites of ddaC neurons at 96 h AEL. Horizontal arrow indicates the direction towards the somas, and vertical arrow indicates the time. (B) RNAi knockdown of klp10A in msps810/mspsP18 mutants restored the retrograde movement pattern of EB1 comets. MT depolymerization or severing induced by Colchicine treatment (D) or overexpression of Kat-60 (F) led to mixed MT orientation in ddaC neurons.
Viability of single and double mutants of RNAPII and rad52Δ, mre11Δ and cdc44‐8. Tenfold serial dilutions of double mutants obtained by genetic crosses were tested for growth in plain SC medium or supplemented with 10 mM HU or 0.005% MMS.
A. Transmission electron micrographs of wild type sporozoites with highlighted secretory organelles in magenta, IMC in yellow, microtubules in green and plasma membrane in blue. Scale bar 500 nm.
(F) Cellular fractions were prepared from HEK293 cells that transiently expressed mutant COA7HIS and were treated with MG132. The samples were analyzed by reducing SDS-PAGE and Western blot.
D Bile acid levels in the liver of mice in (B). n=8 in mice on RD and n=6 in mice on HFD. *P<0.05, **P<0.01 relative to GFP by Student's t-test.
B. KT299 (MATa ssk2/22Δ hkr1Δ msb2Δ) was exposed to the indicated concentrations of α-factor for 15 min in the presence or absence of 0.8 M NaCl, and Hog1 phosphorylation was determined using the Phos-tag band-shift assay. Data information: (B) Representative results from three independent experiments.
B) Tubb1 is expressed in the developing mouse thyroid at E13.5, E15.5, and E17.5 and in the adult mouse thyroid: quantitative PCR results normalized to one of three thyroid tissues at E13.5 value and peptidylprolyl isomerase A. Experiments with at least three tissues per stage. Statistical comparisons versus E13.5 (black), **p<0.01 using ANOVA test
(m) Diagram of the three-chamber social interaction test and mouse movement trace. HET mice did not show preference or stay longer in the stranger1 or stranger2 chambers than the WT mice did in the two test phases. (n) Statistical analysis of the percentage of time spent in each chamber with one empty chamber and another chamber containing stranger1; n for WT=16 adult mice, n for HET=13 adult mice. (o) Comparison between the time that WT and HET mice spent in stranger1 chamber. n for WT=16 adult mice, n for HET=13 adult mice. (p) Time spent in each chamber when one contains familiar mouse stranger1, as well as new mouse stranger2. n for WT=16 adult mice, n for HET=13 adult mice. (q) Comparison between the time of WT and HET mice staying in the stranger2 chamber. n for WT=16 adult mice, n for HET=13 adult mice. Data information: Error bars represent the means ± S.E.M. Two tailed unpaired T-tests were used to analyze the data, n.s. (no significant difference), p<0.05(*), p<0.01(**), p<0.001(***).
Western blot analysis of nephrin expression in immortalized podocytes cultured in normal (5 mM) or high glucose (HG; 30 mM) or high mannitol (Mann; 5 mM glucose plus 25 mM mannitol) for 24, 48 and 72 hours. *P < 0.05 versus normal controls for the indicated time points (Wilcoxon two-sample test; n = 3).
A. Time-lapse of actin filaments constantly exposed to nGr-Tpm1.8 (yellow), WT mCh-cofilin-1 (blue) and S3D-cofilin-1, and to MICAL1 from time t=100s onwards. Here we mixed WT with an excess of S3D-cofilin-1 to mimic cellular conditions in which most of cofilin-1 is inhibited by phosphorylation. While Tpm prevents cofilin from binding to non-oxidized filaments, MICAL1 rapidly oxidizes actin, allowing cofilin to bind and sever filaments.
Representative OXPHOS (E) proteins. One month of edaravone treatment in both wt and LowOXPHOS mice and rewired the upregulation of ETF-A, ETF-B and SDHA proteins to the levels of wt (E). Two samples per condition are shown. Each sample contains extracts from 3 mice. Tubulin is presented as a loading control. Quantifications in lateral histograms (wt, n= 6; LowOXPHOS, n= 6; wt + edaravone, n= 6; LowOXPHOS + edaravone, n= 6).
A/B. Hybrid Rab7/Arl8b compartment. A. Representative confocal images of fixed HeLa cells expressing Arl8b-GFP (green), immunolabeled against endogenous Rab7 (eRab7, magenta). Zoom insets (3x) highlight select regions of colocalization (white), white arrowheads point to vesicles positive for both GTPases. B. Colocalization (Mander's overlap) of endogenous Rab7 with Arl8b-GFP versus free GFP (EV), n EV=4, n Arl8b=9 images (3≥ cells per image) analysed from 2 independent experiments. Significance: 2-tailed student's t-test, * p<0.05.
Schematic drawing of the experimental design to investigate RNAPII transcription speed in vivo. In brief, Arabidopsis seedlings of NRPB2WT-FLAG Col-0 and NRPB2Y732F-FLAG Col-0 were grown on MS media for 12 days and then were transferred to MS liquid media for 2 days. Flagellin peptides (flagellin 22) were added into media and treated samples were collected in a 0 minute (no treatment), 2 minutes and 4 minutes time course. The nascent RNA was isolated and used for reverse transcription and qPCR analyses to reveal RNAPII accumulation at different region in genes. See Methods for technical details.
K Blood glucose and quantification of the AUC during a 120-min pyruvate tolerance test (PTT, 2.0 g/kg pyruvate, i.p.) in HFD-fed WT (n = 6) and NOD2−/− mice (n = 6), *P = 0.0005.
A The expression level of two GA-degradation and two GA-biosynthesis enzymes in expanding leaf tissue (third leaf at 15 DAS) of wild-type plants 20 min, 40 min, 1 h, 2 h, 4 h, 8 h, 12 h, 16 h, and 24 h after mannitol treatment. The fold changes (FC) were calculated relative to control conditions.Data information: In A, data are presented as mean ± SEM, n = 4 independent experiments, *= FDR<0.05 (unpaired two-sided Student's t-test)
Measurements of swim path length confirms the training performance deficit of cDKO mice compared to LM controls. During the reversal phase, the performance of AAV-APPsα injected cDKOs is significantly improved at day 5 (genotype F(2,46)=16.70, p<0.0001, genotype x day F(8,184)=5.788, p<0.0001; *p<0.05, p<0.01).
Adipocyte hypertrophy is visible in the absence of ATMs. Representative images of hematoxylin and eosin staining (H&E) (E) and corresponding adipocyte diameters (F) of lean and obese (HFD 16 weeks) eWAT from WT or CD169-DTR mice treated for 7 or 12 days with DT. Scale bar, 50 μm. Each group comprised 3-5 mice. Statistical significance was determined using one-way ANOVA test. **P < 0.01; ****P < 0.0001; ns: no significant difference. The box and whisker plots show the median value and 10-90 percentiles of adipocyte diameters.
(B) Box plots showing Log2 FC ERS/Control in mouse liver for ERS UP genes 8h and 34h after tunicamycin injection in WT or ATF6 KO mice (3 mice per experimental condition). Box plots are composed of a box from the 25th to the 75th percentile with the median as a line and min to max as whiskers. One-sample t-test with BH correction for multiple testing was used to determine if the mean Log2 FC ERS/Control is statistically different from 0, *P < 0.05. NS, not significant. (C) Similar analyses to panel (B) for ERS DOWN and LIVER-ID genes (3 mice per experimental condition). Box plots are composed of a box from the 25th to the 75th percentile with the median as a line and min to max as whiskers. One-sample t-test with BH correction for multiple testing was used to determine if the mean Log2 FC ERS/Control is statistically different from 0, *P < 0.05.
Proliferation as quantified by number of BrdU+ cells per unit in WT and Atf3−/−vehicle±HDT. Each data point = mean of counts from 40-50 gastric glands from an individual mouse, n=3-4 mice per treatment group. Significance by unpaired two-tailed t-test. Error bars denote standard deviation.
E) Forelimb grip strength was measured in 15-week-old WT and Dup18-30 mice. WT, n = 9; Dup18-30 n = 12.
C HeLa cells stably expressing GFP, GFP-FXR1 wild type (WT) and GFP-FXR1 mutated in the sequence recognized by FXR1 siRNA-1 (GFP-FXR1-MUT-siRNA1) were treated with the indicated siRNAs, synchronized by double thymidine block, released for 24 hours and analysed by immunofluorescence microscopy (C). The percentage of cells with irregular nuclei was quantified, 1000 cells were analysed (mean ±SD, **P < 0.01, ***P < 0.001; N = 3).
D. Suppression of retinal progenitor division in vivo (grey bars) and ex vivo (black bars) by Rb∆K11 and p16 but not by wildtype pRb. Shown are quantification of BrdU+;GFP+ cells relative to all GFP+ cells in samples transfected with the indicated vectors (bars represent mean ±SD; ** denotes p<0.01; *** p<0.001 by one-way ANOVA and Tukey's multiple comparison analysis, n=3 biological replicates).
(B) UBE2D1~UbAtto discharge assays with TRIM25 wild-type RING, the fused RING constructs and different mutants important for dimerization or the interaction with ubiquitin. Time point zero for the T25-R Linker and T25-R Linker V72R samples was taken before the addition of E3 as discharge is very fast. The loss of UBE2D1~UbAtto is plotted as the average of experimental duplicates (± s.d.).
C Quantified fluorescence of UbG76V-GFP normalized to mRFP from lysates of transgenic nematodes at L4+48 hours that express these reporters in the hypodermis. Animals were exposed to bacteria containing the indicated RNAi clone or an empty RNAi vector as a control. *P<0.05, ANOVA with Dunnetts posthoc comparison to the empty RNAi vector control. N=3 trials. Error bars indicate SEM.
(E) Top: Subcellular fractionation showing amount of RAD51 in DLD1 stable clones expressing BRCA2-T207A (T207A) in chromatin fraction. When indicated, cells were irradiated (2h treatment) and/or transfected with a plasmid expressing RNase H1 (RH) 48h prior fixation. α-tubulin and Histone H3 signals are shown as markers of cytoplasmic and chromatin fraction respectively. Bottom: quantification of RAD51 levels detected by Western blot with a RAD51 specific antibody in chromatin fraction relative to RAD51 levels in WCE. Results are presented as the fold change compared to the untreated sample. The data represents the mean ± SD of three independent experiments. Statistical significance of the difference was calculated with unpaired t test and the p-values show the significant difference.
E The odorant habituation results shown as the ratio of odor sniffs/water of the miR-200b/a KD mice were significantly reduced compared to those of the NC mice. These odorants included male urine (1:50), female urine (1:50), isopentyl acetate (50 μM), citral (50 μM), 2-heptanone (50 μM) and propyl propionate (50 μM) (n=6 mice each group; data represent the mean±SEM; *p<0.05, **p<0.01, ***p<0.001; Student's t-test).
Survival curve of Trf1+/+ K-Ras+/G12V p53−/− and Trf1−/− K-Ras+/G12V p53−/− mice.
(I) Immunostained images of adult mouse heart tissue. These were stained with a phosphorylated AMPK antibody (I, upper), a phosphorylated CLIP-170 antibody (I, lower) and a plakoglobin antibody (middle). The representative images from more than 5 analyzed tissues were shown.
B) Cells transiently transfected with RISC Free control siRNA, or siRNA duplexes directed against TRAPPC8 or RAB1A and B were stained for the cis-Golgi markers GM130 or RAB1B and analysed by confocal microscopy. Scale bars = 20 μm in main panels, 10 μm in inset panels. Bar graph: > 100 cells per siRNA were scored for fragmented or normal juxtanuclear Golgi stacks from three independent experiments. Error bars ± s.e.m., * p<0.05, ** p<0.01, one way ANOVA with Sidak's multiple comparison test.
F PARP cleavage in HCT116 Bax/Bak DKO cells expressing similar levels of mitochondrial Bax or BaxTBak analyzed by Western blot without apoptotic stimuli. Actin is used as loading control. n = 3.
Wild type, mrc1∆ and rad9∆ cells were synchronized in G1 with α-factor prior to release with pronase in the presence of 0.033% MMS. Cells were collected at the indicated times and Rad53 phosphorylation was monitored with an in situ phosphorylation assay - ISA (F).
D, Western blot detection of phosphorylated eIF2α (Ser51) and total eIF2α levels in cell lysates of control and mutant MDA-MB-231 cells. GAPDH and Tubulin serve as loading control for phosphorylated eIF2α and total eIF2α blots respectively. Numbers indicated below the blots represent the band intensities of target proteins normalized to the loading controls, with the value in the control line taken as baseline of 1.
(A) In a mixed population, flies reach the end of a "pre-mortality plateau phase" (PMP) at an age of four weeks, where 90% of the flies are still alive. We define this age as their "midlife" point. At this stage, the rate of the population decline accelerates from 1% (plateau) per day in the previous 10 days to 3.5% per day in the following 10 days. Median survival=39 days, N=260.
(C) Extracellular acidification rate (ECAR) was measured in TFAM knockdown and WT SNU-368 cells with serial addition of glucose, oligomycin and 2-DG (n = 3 independent experiments). The panel shows the calculated glycolytic parameters, including glycolysis and maximal glycolysis. Glycolysis was calculated as maximum rate measurement before Oligomycin injection minus last rate measurement before glucose injection. While glycolytic capacity was calculated as maximum rate measurement after Oligomycin injection minus last rate measurement before glucose injection. Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test for , C *P < 0.05; **P < 0.01.
(C) Mass spectrometry identification of SMCR8 phosphorylation sites by in vitro TBK1 kinase assay of HIS-tagged C9ORF72:SMCR8:WDR41 complex purified from baculovirus infected insect cells.
A Top: Sequence alignment of human H2A.B and H2A. Residues not conserved in H2A.B and H2A are colored in red. Secondary structures are indicated above. H2A.B N-terminal domain (NTD) and the docking domain (DD) are indicated. H2A.B residues subject to mutagenesis analyses are indicated as boxes. Bottom: Schematic view of mutants used for MNase analysis. The H2A and H2A.B sequences are indicated by blue and gray, respectively.