EMBO/sd-character-level-ner huggingface dataset:
(d-f). Drp1S616 phosphorylation is required for mitochondrial fission induced by PINK1. Drp1KO HeLa cells were co-transfected Δ110-PINK1-GFP-FKBP /FRB-MTS with either a plasmid encoding Drp1WT or a Drp1 dephosphorylation mimic mutant Drp1S616A. Cells were treated with 250 nM rapalog for 2 hours to activate PINK1 kinase, followed by immunodetecting mitochondria (TOM20, blue), exogenous Drp1 (myc, red), Δ110-PINK1-GFP-FKBP /FRB-MTS (PINK1-FKBP, green). Cells treated with solvent (DMSO) were used as a treatment control. Representative images were shown, scale bar=25 μm (d). Higher magnification images are also included (d, Zoom, scale bar=5 μm). Mitochondrial morphology in different transfections was quantified (e, f). Student's test. **p<0.01, ***p<0.001, ns: no significance. Data was presented as mean ± SEM of three independent experiments. For each condition, >100 cells were analyzed.
(e) Fresh weight of 15 d old plants with and without SA187 under NHS, HS and LAT temperature regimes. Data information: fresh weight, measuremts after recovery at 22°C for 4 days. All treatments are compared with direct 44°C heat stressed plants. Asterisks indicate a statistical difference based on the Mann-Whitney test (* P < 0.05 )
Representative confocal z-stack images of LV-infected dendritic segments in the DG granule neurons from control and susceptible mice with different treatment. White arrows indicate stubby spines, yellow arrows indicate mushroom spines, and blue arrows indicate thin spines.
LPS-primed and Nigericin-stimulated iBMDMs stably expressing a control shRNA or 3 different PLK4-specific shRNAs (F).
A Targeting strategy used for creating conditionally phosphorylated proteins. Upon integration of the targeting cassette and expression of the gene of interest (GOI), the targeted protein is C‐terminally tagged with an HA epitope. After Cre recombinase‐mediated excision, Clb2m‐HA is fused to the targeted protein.
C. Box plots showing pair-wise sequential-stage comparisons of fold change in accessibility (ATAC-seq) and corresponding changes in gene expression (RNA-seq dataset) (Owens et al, 2016). The data represents two biological replicates. The central band within the boxplot represents the median (50th percentile), the box represents the range between the first and third quartile (25th-75th percentile), and the whiskers show 1.5 x the interquartile range (IQR).
(E) Fusion activity. Vacuoles were isolated from BJ cells carrying Nyv1-S9-EGFP and DKY cells deleted for Nyv1 and carrying Vam3-S9-mCitrine or from wild-type cells and incubated in standard fusion reactions in the presence or absence of chlorpromazine (CPZ, 150 µM) and anti-Vam3 (3 µg). Content mixing was assayed via alkaline phosphatase activity. Means and s.d. are shown for three independent experiments.
Evaluation of M1 markers in macrophages by real-time PCR. Fold change of TNFA, CD80, CXCL9, and CXCL10 mRNA in IL10, MSO- and glufosinate (10 and 20µM)- stimulated IL10 macrophages (n=3). Evaluation of M2 markers in macrophages by real-time PCR. Fold change of MRC1, MSR1, mRNA in IL10, MSO- and glufosinate (10 and 20µM)- stimulated IL10 macrophages (n=3).
H) Quantification of long-range peroxisomal trafficking events (visualised with pxGFP) in HeLa cells transfected with scrambled, Miro1 and Pex14 siRNA, by blind scoring of a five-minute movie (n=18 cells over three independent experiments). Data information: For H) is a one-way ANOVA with Newman-Keuls post-hoc test. ** denotes p < 0.01. For H), no statistical difference between conditions was observed, unless stated. Data are represented as mean ± SEM.
(G) RT-qPCR of U2-OS cells irradiated (20 Gy) either 1.5h or 7 days prior to harvest. CHIR denotes treatment with GSK3 inhibitor CHIR-99021 (10nM overnight). DMSO overnight treatment was used as control. Quantitation was performed and statistical significance obtained from n = 3 replicates, using ANOVA with Tukey multiple comparisons test. SD shown. * = p < 0.05, ** = p < 0.01, *** = p < 0.001.
Hep1-6 cells were infected with pCDNA3.1-DOCK5 (DOCK5 +) or pCDNA3.1 (DOCK5-) for 48 h. - mice were incubated in control (GlcN-) or GlcN (GlcN+) medium for indicated times, followed with (Ins+) or without (Ins-) 100 nmol/L insulin stimulation for 20 min. Total and phosphorylated InsR, IRS-1Ser1101 and Akt in Hep1-6 cells.
Representative flow cytometric analysis of BAL from Csf2ra-/- recipients 7 weeks after neonatal i.n. transfer of pMΦ or fetal Mo from CD45.1+ E14.5 fetal livers, 7-week old un-transferred Csf2ra-/-, WT (CD45.2) mice. Pre-gated viable CD45+ singlets.
Kaplan-Meier survival curves of wild-type animals fed bacteria transformed with either empty-vector (con) or with vector expressing dsRNA agains isp-1 (G) and left untreated or treated with 600, 1200 or 2400 J/m2 UVB
B. The same experiment as described in (A) but with MCF7 cells exposed to MG132 (20 μM, 1 h) or puromycin (100 μg/ml, 10 min). Means +/- SEM, n = 2. C. Lysate from MCF7 cells exposed to MG132 (20 μM, 1 h) or puromycin (100 μg/ml, 10 min) as indicated were subjected to immunoprecipitation with anti-puromycin antibodies, followed by assaying for co-precipitation of K48-ubiquitylated proteins. Actin is shown as a reference.
Figure 5. The effect of cell-shape change during growth on the stability of Min protein patterns. A. Cytosolic fluorescence during growth of a cell from a small elliptical form into a large rectangular shape. Numbers in red indicate time in hours. Illustrations show the positions and orientations of the cell in the first and last time frames. Green and blue lines indicate the maximum and minimum Feret diameters, respectively. B. sfGFP-MinD patterns during the growth of the cell shown in A. Illustrations indicate the cell boundaries and oscillation angles observed in the first and last frames (not to scale). C. Quantitative data obtained from the cell shown in A and B. The maximum and minimum Feret diameters (green and blue), and the measured MinD oscillations (red) were expressed in terms of length (top) and angle (bottom) and plotted against time. The number of cells that fit this category was 41/97. Arrows indicate the time when cell width reached the chamber width of 5 μm.
A Time lapse images of E. coli fabA(Ts) cells expressing FOF1 a-mNG. For corresponding controls, see Appendix Fig S10. Cells were grown at 30°C and shifted from 30°C to non-permissive 37°C. See Movie EV2 for full time lapse.
C S. cerevisiae wt cells or hsp42Δ cells expressing mCherry‐VHL (red) and GFP‐Nup49 (green) were treated as in (A). Three‐dimensional reconstructions of respective cells are given.
(J) The percentage of Ki67 positive cells in each HG; n = 3 mice and 10 HGs were counted in each mouse.
Model for the role of Gadd45a in regulation of DNA damage response initiation. In G3Terc-/- mice, Gadd45a promotes the accessibility of telomeric heterochromatin and enhances the DNA damage response initiation (e.g. pATM activation) through a BER-dependent pathway. The initiated DNA damage response further causes upregulation of p53 and p21, which leads to impairment of intestine stem cell maintenance and function. In G3-dKO mice, the telomeric heterochromatin is condensed and damage response initiation is reduced upon Gadd45a deletion, which further leads to decreased expression of p53 and p21. Therefore, the maintenance and function of intestine stem cell is improved.
C. Immunoblot analysis of U2OS WT, U2OS/SCAI KO and U2OS/SCAI KO cells stably reconstituted with Strep-HA-SCAI (U2OS/SCAI KO/Strep-HA-SCAI).
A Relative mRNA expression levels of H2A.B in L1236, L428, HDLM2 HL cell lines and Germinal centre B lymphocytes (GC) compared to the Ramos B lymphocyte cell line. The results are presented as the mean ± SD of three biological replicates.
C4-2 cells were treated as depicted in Figure 2. ChIP was performed using the indicated antisera, and the subsequent DNA was isolated and used in qPCR reaction using primers designed to amplify the indicated genomic loci: BRCA2 enhancer, RAD51 promoter, or TOP3A promoter
A. Immunostaining of non-infected cells (day 13 in differentiation medium) and cells infected with a mutant form of TFEB (TFEBmt) with LAMP1 (red) and Flag (green). Ad-TFEBmt was added to the myotubes for 72 h on day 10 in differentiation medium. Ad-TFEBmt-infected cells show massive accumulation of TFEB in the nuclei and significant reduction in lysosomal size, similar to that seen with TFEB.
(b) Serum and amino-acid starvation of HeLa cells for 5 h reduces LC3-II levels (above), but increases autolysosome numbers detected using tfLC3 (below). With tfLC3, GFP- and RFP-positive puncta represent autophagosomes before lysosomal fusion, whereas RFP-positive/GFP-negative puncta represent autolysosomes-GFP is more rapidly quenched by low lysosomal pH (see Methods). Increased autolysosomes indicate enhanced starvation-induced flux of LC3 to lysosomes.
C. Quantification of mTREM2 expression in whole-cell lysates of WT and GRN-/- hiMGL (data shown in B). mTREM2 levels were normalized to WT (n = 3, biological replicates).
A Schematic representation of the UPF3A and UPF3B protein domains. Below are the respective mutated rescue constructs.
Cell viability of HCT116KO following different treatments as indicated. siCtrl: control siRNA; siPLXNB2-1 and siPLXNB2-2: PLXNB2 siRNAs
A Simplified phylogenetic tree showing mainly the Dikarya subkingdom and including the external branches Homo sapiens (Hs) and Arabidopsis thaliana (At). The Id. column indicates amino acid sequences percentage identities, ranging from 100% (green) to 84% (magenta) identity for eukaryotic actins to S. cerevisiae's actin, and 60-62% (black) for archaeal actins. Squares outlines are solid or dotted for sequences deriving from existing species or ancestral reconstruction, respectively. The "coded by" column indicates which coding sequences were originally used to code genes of interest. Nucleotide sequence identities are ranging from 100% (blue) to 76% (orange) compared to S. cerevisiae's actin coding sequence.
(A) Representative results of Western blot analysis of RAD21, E-cadherin, and VIM are shown for luminal and basal-like breast cancer cell lines. The images were analyzed using ImageJ (National Institutes of Health, USA). Protein levels from the Western blots were quantified and normalized relative to α-tubulin. Bars represent the mean ± S.D for four independent experiments.
Co-depletion of Wapl and separase has a synergistic effect on HDR. U2OS DR-GFP cells were transfected with the indicated siRNAs, treated as in (A+B) and analysed by GFP-flow cytometry (D Shown in (D) are averages (bars) of 2-3 independent experiments (dots)
D IGF-1 protein levels in serum measured by ELISA (mean ± SEM; 3-8 mice per genotype and age group).
(C)BMDMs were infected with JR32 or the legA9 mutant for 6 h and the level of p62 protein was quantified in cell lysates by ELISA (performed in triplicate and the SD are too small to detect).
Relative expression of Oct4 and Rex1, determined by qRT-PCR. The data represent the means ± SD (n=3, from biological repeats). ***P＜0.001. T-test was used to determine the significance.
B. H149, N152 and D160 in MavQ are critical for its kinase activity. Purified wild-type and mutant MavQ were reacted with PtdIns and ATP. The kinase activity was determined by the ADP-Glo assay.
C The protein pulldown experiments identified 56 proteins putatively associated to the Shigella-septin cage, and then categorised into 6 groups based on analysis from the Gene ontology database. See also Dataset EV1.
A Schematic representation of the group A carbohydrate (GAC) gene cluster. Biogenesis of GAC is obtained with 12 gac operons. Genes from gacA to gacC are responsible for rhamnose backbone biogenesis, and those from gacI to gacL are involved in GlcNAc side chain addition.
Representative images of the distribution of Fib-Tau (red) 8 h and 24 h after injection. Dendrites are immunolabeled using MAP2 antibody (green). 8 h after injection, both diffused and clustered distribution of exogenous Fib-Tau is observed in stratum oriens The number of Fib-Tau clusters per µm2 within stratum oriens decreases between 8 h and 24 h post-injection (G). A similar decrease in the total fluorescence (clusters + diffused) is observed indicating the diffusion and/or clearance of exogenous Fib-Tau (H). Paired-t-test performed between '8/24h-right' and '8/24h-left'; Unpaired t-test performed between '8h-left' and '24h-left' columns (n is number of animals: 6 for 8h and 5 for 24h); ***p<0.001, **p<0.01, *p<0.05, ns= not significant. Panels (E and F) are shown for same exposure and are used for quantification. Data information: Scale bar 10µm
(A) Immunoblot analysis with anti-LC3 antibody shows that DETA NONOate reduced autophagosome synthesis in bafilomycin A1-treated WT Bcl-2 and AAA Bcl-2 MEFs.
(D) Relative crosslinking frequencies among CTCF/RAD21 binding sites on the genes were measured with a 3C assay in the control (blue line) or RAD21-overexpressing (red line) MDA-MB-231 cells on day 10 after lentiviral transduction. BamH1 or XbaI restriction sites on the TGFB1 or ITGA5 gene, respectively, appear as gray shaded bars. Black shading indicates the anchor fragment. Each value was normalized to crosslinking frequency at the ERCC3 gene. The maximum crosslinking frequency was set at 1. (means ± S.E.M, n = 3). *P < 0.01, **P < 0.05. Statistical significance was validated by Student's t-tests.
H Laggard chromosomes were observed in the metaphase I of Tex11-/Y spermatocytes. The arrowhead indicates the laggard chromosome.
NPC2 immunoblots of un-concentrated CM (containing 10% FCS) from fractionated IMC and AT2 cultures. Total protein staining of the membrane by Amido black (bottom) serves as a loading control showing serum albumin at 70 kDa, derived from FCS in the media.
A Immunofluorescence for total MLX protein detection (red signal) performed on C2C12 cells treated with siRNAs against Mlx-γ (si-Mlx-γ), lnc-SMaRT (si-SMaRT), DHX36 (si-DHX36) or with a control siRNA (si-SCR). Cells were fixed two days after the switch to differentiation medium. Representative binary images of single confocal planes for MLX immunofluorescence are shown for each treatment. The inserts on the topside of the images sketch the MLX signal peaks in the nuclei (white stars) to highlight the variation of MLX staining in each condition. Dashed lines indicate the edge of the nucleus.
(J) Acetylation level of endogenous IDH1 K224 when treated with SIRT1 or SIRT2 by Western blot analysis.
A) Control and irradiated murine small intestines analysed with RNAscope in situ hybridisation for Myc mRNA (black arrows point to mRNA granules in the control) and by immunostaining for Sox9 protein reveals an expansion in the expression domain of both genes. Nuclear localisation of beta-catenin is readily detectable after irradiation. n = 3 samples per condition, quantification shown in the insets, error bars = 1 SD. Statistical analysis was performed with an unpaired student's t-test. Similar results were obtained at 3dpi and 5dpi.
BMDMs from wild type, Traf3ip3-/- and Mavs-/- mice were treated with or without VSV, SeV, poly (I:C) or HSV-1 respectively as indicated for six hours. Il6 (C), Isg54 (D), and Cxcl10 (E) induction was measured respectively by qPCR. All data are presented as the mean values based on three independent experiments, and error bars indicate s.d. P values were determined by unpaired two-tailed Student's t-test. *P<0.05 and **P<0.01. NS indicates no statistically significant difference.
(F-J) The crucial role of Ulk1 dephosphorylation at Ser637 in etoposide-induced autophagy. (F-H) Ulk1/Ulk2 DKO MEFs that were stably transfected with HA-Ulk1 or its mutants, S637D and S637A, were treated with etoposide (10 µM). Then, the cell lysates were collected time-dependently, and the expression of each protein was examined by western blotting. Asterisks in the Ulk1 blots are nonspecific bands. Semiquantitative analyses are shown in Fig. EV2E.
(A) Illustration of the linearized expression cassette containing the CAG promoter, 3xFLAG-tagged DIA1(R1204X), and SV40 poly(A) sequence used for injection into fertilized eggs.
A Gene set enrichment analysis of the IL-6/JAK/STAT signaling pathway in G-MDSCs from tumor-bearing Tet2-/- (KO) versus WT mice (n=2). p-value was calculated by permutation.
D Electrostatic surface potential of the alkaline groove in partial Ca2+-bound structure. The gradient shown is from -3 (acidic, red) to +3 (basic, blue) kT/e.
A The complex between uPA and PAI-1 is an agonist of the uPAR-mediated cell adhesion to VN. RTCA analysis of 293/uPAR cells seeded on VN and treated with 10 nM sc-uPA, PAI-1 and uPA•PAI-1 or a combination of 10 nM tc-uPA and 10 nM PAI-1. The time of the addition is indicated by a vertical dashed line.
Representative human brain metastases showing different percentages of nuclear HSP90. Scale bars: (J) 50 µm Black arrows point to cancer cells positive for HSP90 in the nucleus.
Quantification of Vegfr3Luc emission in the Lymph nodes, spleen of mice in experiment (I), respectively. N= 4 mice per condition. Statistical significance was determined by Two-Way-ANOVA.
Co-IP from siRNA-transfected HUVECs. The graph shows relative levels of PlexinD1-VEGFR2 association, in which the values at 0 min are set to 1. n = 3 per group.
(C) An assembly of IgG (first row), DVL3 (second row) ChIP-Seq in MDA-MB-468 cells, IgG (third row), and, DVL3 (fourth row) ChIP-Seq data in MCF7 for the KMT2D gene, visualized by IGV. Each column is 20000 bp wide. The fifth row show the location and orientation of the KMT2D gene.
B: PIKfyve is selectively downregulated in mice infected with ME7 prions. Right: Quantification. Each dot represents one individual (n=4/group). ***: p<0.001; unpaired t-test. Error bars represent s.e.m.
F Comparison of tumor weight of each group at end point. Data were shown as Mean ± SEM (n = 6). Statistical comparasions were performed using a one-way ANOVA with Dunnett test. *P = 0.0177, **P = 0.002.
(a) Representative images of rat liver (FAO) cells immunostained with TSC2, TSC1 or Rheb (green) and PMP70 (peroxisome marker) or LAMP1 (lysosome marker) (red) antibodies. Scale bars, 10 μm. (b) Pearson's correlation coefficient for TSC1 or TSC2 co-localization with PMP70 or LAMP1 calculated using Imaris software. Quantification was performed on 8-12 cells from each of the 4 independent experiments, giving rise to a total of 40 cells. All error bars represent s.e.m., ***P0.001.
A, B RNF123 inhibits the activation of the NF-κB (B) promoters induced by the overexpression of RIG-I, RIG-I CARD, MDA5 CARD, and VISA, but has no effect on the TAB2- and IKKβ-induced activation of the NF-κB promoter (B). HEK293T cells (2×105) were transfected with the indicated plasmids (100 ng), NF-κB promoter luciferase plasmid (100 ng), and pRL-TK Renilla luciferase plasmid (100 ng), together with an empty vector and RNF123 construct (100 ng). Luciferase assays were performed 24 h after transfection.
(A) In vitro ubiquitination assay. Purified His-RNF114 was subjected to ubiquitination assays with the indicated components and the samples were analyzed by immunoblotting with anti-FLAG (Ub) antibody. Lanes 1-4 indicate negative controls. The results show that recombinant RNF114 efficiently catalyzed the formation of polyubiquitin chains.
B) HEK293A GFP-LC3B (2GL9) cells were transfected with mCherry or mCherry-TBR, treated with EBSS for two hours, fixed and analysed by confocal microscopy.
A. Representative Per2::Luciferase traces before (black) and after vehicle (grey) or Prok2RA (pink) topical application of SCN slices.
MCIDAS (A) regulatory elements (see materials and methods) cloned upstream of the luciferase gene were co-transfected into HEK293T cells with vectors expressing GemC1, McIdas and Geminin, as indicated or an empty vector (-) as a control. All luciferase experiments (A, B, D) were normalized for transfection efficiency with an expression vector for Renilla luciferase. Fold induction is the ratio between the normalized luciferase activity induced by the expression constructs and that induced by the empty expression vector. Data are the mean values of at least three independent experiments and error bars indicate ± SEM. **p<0.01, ***p<0.001. P-values were calculated by the non-parametrical two-tailed Mann-Whitney test. Abbreviations: RLF: Relative Luciferase Fold induction.
Under ambient temperatures, CBF1 transcript and protein levels are induced by phyA and phyB in the light. CBF1 promotes PIF4 and PIF5 protein accumulation in the light by directly binding to PIF4 and PIF5 promoters to induce their expression, and by inhibiting phyB interaction with PIF4 and PIF5. Thus, CBF1 promotes hypocotyl elongation under ambient temperatures by positively regulating PIF4 and PIF5 protein accumulation in the light through transcriptional and post-translational mechanisms. Lower levels of PIF4 and PIF5 proteins accumulate in the cbf1 mutants in the light, therefore cbf1 mutants develop shorter hypocotyls under light conditions.
C In vitro HMTase assay using disulfide-linked ubiquitylated histone H3. H3K14CUb and control histone H3 (H3C110A) were used as substrates in the HMTase assay, and analyzed
D, E. mRNA stability of Stat1 and Irf1 were measured by qRT-PCR in tumor cells treated with IFN-γ and actinomycin D. Mean ± SD of n=3. * * P < 0.01; * * * P < 0.001 by Student's t-tests.
D. Volcano plot showing fold change (log2 values) in transcript levels of PC3 cells upon BAZ2A knockdown. Gene expression values of two replicates were averaged and selected for 1.5 fold changes and P <0.05.
D Inhibition of interactions formed with DDX39B in the presence of MTX using co-IP assay. DDX39B is fused to the C terminus of a V5 tag, and the interaction partner is fused with a Strep-HA tag. The diagram (upper) shows the interaction pairs formed with DDX39B used for co-IP assay. Dot-blot results (lower) show the interaction pairs (arrangement as upper panel) in presence of different concentration of MTX as indicated on the top of the image. The image displayed is representative of 3 replicates (n = 3).
(F) Effect of the Bad knock‐out on autophagy. Wild Type (WT) or Bad−/− MEF were transfected with LC3‐GFP and then subjected to nutrient depletion (NF) and/or treatment with ABT737 (18h; means±s.d., n=3). The asterisks denotes a significant (P0.05) effect of Bad deficiency.
(S-V). Serum levels of E (S), NE (T), T3 (U), and T4 (V) in male mice supplemented with AKG for 11 weeks (n = 8-9 per group).
T-DNA insertion site in the PLDα6 gene and the complementation construct introduced into the T-DNA insertion mutant. Boxes denote exons and lines introns.
P-R Functional analysis demonstrated the effective ability of the PF and Mabs-PlGF combination to generate a complete and functional artificial TA replacing the ablated TA injury. (P) The fore-hind limb grip test diagram reveals functional grab force recovery for mice treated with PF-embedded Mabs-PlGF (TA ABL-PF+MABS) after only 4 weeks post-TA injury, being comparable with not-ablated wild-type (WT) mice and significantly higher than control ablated mice (TA ABL-CONT) or mice implanted with void PF (TA ABL-PF CONT) (n = 10 mice per group). (Q, R) The treadmill analysis of the treated mice relative to wild-type not-ablated (wt) shows both exhaustion time and distance indicative of a full recovery of motor functionality already at 3 weeks after TA ablation with an increase in time and distance run for mice treated with PF-embedded Mabs-PlGF, while control mice (TA ABL-CONT or TA ABL-PF CONT) showed a much lower resistance (n = 10 mice per group). The values shown in the diagrams are expressed as means ± standard error and statistical significance was tested using Student's t-test (P < 0.05 was considered significant).
K Quantification of GFP fluorescence in the QC of pSCR::GFP and pSCR::GFP-SCR transgenic seedlings, as shown GFP signal intensity of each genotype was normalized relative to that of the WT. Data information (K), n denotes the total number of scored samples. Individual values (black dots) are shown. Data represent mean ± SD of three independent replicates. (K), **P < 0.01 (Student's t-test). Scale bars: 20 µm.