EMBO/sd-character-level-ner huggingface dataset:
a, Cellular viability of super-repressor- or empty-vector-infected control;Bcl2 lymphomas treated with ADR or untreated for 5 days, then exposed for 2 days to the inhibitors 2DG or bafilomycin A1, compared to no inhibitor (ut) in vitro. Results represent means ± s.d. (n = 5 each).
D Chromatins extracted from Control (WT) and GS∆44 myoblasts were immunoprecipitated with anti-MyoD and control IgG antibodies. Regions corresponding to the MyoD binding sites present in Peak A (see Figure EV3D) and in the MAFA and LSTZ2 genes were analysed by qPCR. Fold enrichments of PeakA over MAFA or LSTZ2 immunoprecipitation (IP) are plotted for WT and GS∆44 samples. The mean ± SEM of triplicates from one representative among three experiment is shown. Statistical significance of differences between means was assessed by a two-tailed t-test and P<0.05 was considered significant. *P=0.0339, **P=0.0067.
Immunofluorescence analysis of p62 was done in scrambled siRNA transfected HFFs and TRAF6 knockdown HFFs with and without IFNγ using indicated strains (n=3).
Validation of the effect of miRNA inhibition in MEFs by q-PCR. Error bars indicate s.d. (n=3).
Relative cellular viability following Cas9-induced gene editing. Reported number of colonies was normalized based on viability of HAP1 cells transfected with an Empty Vector (EV) control (dots). Error bars indicate mean of SD from three biological replicates (number of colonies for EV rep1=242/rep2=238/rep3=201; CHMP1B rep1=134/rep2=151/rep3=128; NUP58 rep1=11/rep2=13/rep3=15; NUP85 rep1=0 /rep2=0/rep3=5; NUP153 rep1=61/rep2=55/rep3=60)(Ordinary one-way ANOVA ****P ≤ 0.00001; ns = non-significant).
Heat map of Egg ChIP-seq within a 10kb window centered on Egg peaks. Note that ChIP peaks was called using the ChIPed and Input samples Metaplots showing Egg ChIP-seq signals for genomic regions around Egg peaks. Displayed are fold-changes in Egg occupancy normalized to input.
(A) Schematic representation of the experiments used in this figure. See details in main text and Methods.
The mRNA levels of genes related to energy consumption (PGC1α, UCP1 and Dio2). (n=4 biological replicates, per group)
A GST pulldown binding assay of in vitro transcribed/translated 35S-Myc-CALCOCO1 with recombinant GST-tagged ATG8 family proteins. GST and GST fusion proteins were visualized by coomassie brilliant blue staining (bottom panel), and the co-precipitated Myc-CALCOCO1 was detected by autoradiography (upper panel). The numbers below the AR represent % binding in the shown AR .
A Three pairs of small interfering RNA against TRAF3IP3 were transfected into HEK293T cells respectively and immunoblotting were then employed to examine the knockdown efficiency. A pair of scrambled RNA oligoes was transfected and signified as con.
A. Representative DAPI-stained spreads of pachytene, diakinesis, metaphase-I, dyad and tetrad meiotic stages, for wild type and msh2, in Col×Ler or Col×CLC hybrid backgrounds.
e) Histogram charts showing the distribution of clones at D1-3 following tamoxifen induction according to total number of cells and number of basal cells.
Quantification of leaf glucose (Glc). Data for two independent experiments are shown; means ± SEM; n ≥ 17 per genotype and time point.
(G) Primary root length of the plants tested Data are means ± SE (n = 25, individual plants). Student's t-test (**P < 0.01) was used to analyze statistical significance.
(A) Experimental design for demonstrating metabolite production via macromolecule degradation. Yeast cells were grown on 2% unlabeled glucose, and then switched to U‐13C‐glucose for 70 min, which completely labels free metabolite but only partially labels macromolecules. Thereafter, glucose was removed and the metabolome analyzed by LC‐MS.
B-B2 Transient overexpression of secretagogin in immortalized CRH‐expressing mHypoE‐N44 hypothalamic cells significantly reduced CRHimmunofluorescence microscopy intensity in secretagogin+cell bodies, indirectly supporting enhanced CRH release. All experiments were performed in triplicate. Scale bar: 50 nm.
Hek293T cells were transfected with the indicated constructs. MICU1-GFP immunocomplexes were precipitated with a GFP antibody and analyzed with PAS (phospho-Akt substrate) antibody by western blotting
c. Mitophagy quantification for experiments in 5a. Mitophagy index per cell is measured as a ratio of the number of mitolysosomes per mitochondrial content. Graph generated using the mean of each biological replicate ± SEM (n=7 with at least 70 cells quantified per biological replicate for each condition). Mean area of mitolysosomes per cell and mean number of mitolysosomes per cell for experiments in 5a.
C Venn diagram of DHS sites and ARBEs. Right panel shows DNase‐seq tag distribution around ARBEs within ± 1 kb.
Representative images of B‐cell cultures of the indicated genotypes 48 h after being stimulated in vitro with LPS. Insets provide a magnified view of an individual cell (arrowhead) for better comparison of the observed differences in cell size. Scale bar, 100 μm.
Representative maximum intensity z-stack-projected images of DAPI-stained (blue) AC16 exposed to 18°C and returned to 37°C for the time periods indicated, probed by RNA-FISH for TP53 (C)
(A) Cell extracts were generated after the indicated treatments and then incubated with anti-GFP beads. Factors co-purifying with GFP-SLD5 were monitored by immunoblotting.
mRNA in situ hybridization and immunofluorescence in E9.5 or E10.5 wildtype and Slc7a5-null embryos for key marker genes. (M-N) Wholemount Tubulin-β-III (Tuj1) immunofluorescence performed on (M, m1) wildtype and (N, n1) Slc7a5-null E9.5-E10.5 embryos (n=5 each condition). Dorsal root ganglia (DRG) and Sympathetic Chain Ganglia (SCG) are indicated with open arrowheads together with cranial ganglia IX, X and XI. Images in (m1, n1) show higher magnification of the cranial ganglia V, VII/VIII (open arrowheads). Data information: Scale bars 200 µm
d) Immunofluorescence with an anti-calnexin (ER) antibody. A549 cells transfected with adenovirus-based GFP-Atg14L. After 48 h, the cells were fixed and immunostained with calnexin (ER).Scale bar, 10 μm.
(J) ELISA analysis of IL8 production upon Shigella infection. HeLa scrambled control cells and SPHK1/2 double KD cells were infected with Shigella flexneri M90T for 3 h and 6 h, or stimulated with TNFα for 6 h.
(D) Quantification of Ki-67+ and AURKB+ double positive cells was performed using five confocal images per mice in control and TGFα mice. Images were obtained at 40X magnification. Scale bar: 50µm. ****P < 0.00005, unpaired t-test, (n=4).
D. Potential H+-bond network between protonated E406386 (TM7b) and residues in HP2b in EAAT1CRYST transmitter-bound structure. HP2 is depicted in yellow and the 398NMDG motif in green. Residue numbering corresponds to EAAT1WT, and the interatomic distance (angstrom) is parenthesis.
Thioperamide treatment of tissue culture cells. BHK, CHO and HeLa cells were treated with thioperamide and processed as in Fig 1C. Low magnification views are shown after staining with anti-LBPA antibodies (green) and DAPI (blue). n=3 independent experiments with > 200 images acquired and analysed automatically.
E. EM picture of a negatively stained 2D crystal of Dln1M5 on the lipid monolayer. The inset represents the computed diffraction pattern Fourier transform of the image.F. Projection map with a P4 symmetry.
A-D) Analysis of Tbk1-/- MEFs complemented with the indicated Flag-tagged TBK1ΔC-adaptor fusion proteins. C) TBK1-Nap1-NDP52 complex. N- and C-termini, domains and binding partners are indicated.
Analysis of pHistone H3, HER3, pHER3 and cleaved caspase-3 in tumors of mice injected with BTRH and treated (T) or not (C) with EV20/MMAF (3.3 mg/kg for 24 hours or two weeks). Numbers below the blots show the quantitation of the signal of each band, referred to that of a control untreated tumor.
(C) The ultrastructure of the muscular exopher and its schematic view. Arrows: red - morphologically changed mitochondria inside the exopher, yellow - normal, elongated mitochondria inside the muscle cell. c - cuticle, m - muscle, e - exopher.
(a) Manually segmented rosettes on 3D reconstructions from Z-stacks of 25 μm of an E5.75 embryo imaged using lightsheet microscopy. AVE cells are in green, and all cell membranes are labelled in grey. Left: anterior view; Middle: lateral view (anterior to the left); Right: posterior view. Scale bars: 20 μm. (a') Mouse line strategy used to perform live imaging. (a") Left: Number of rosettes normalized by the total number of cells in the epiblast region in focus expressed in percentage. Right: Frequency of rosettes, normalized by the area in mm2 of the epiblast region in focus and the time of observation in minutes. Data information: Mean ± SEM, n=41 frames from 3 embryos. (b, c) Rosettes outlines on optical slices from E5.75 (b) and E6.25 (c) embryos imaged by confocal microscopy. AVE cells are in green, and all cell membranes are labelled in grey. Left: anterior view; Middle: lateral view (anterior to the left); Right: posterior view. Scale bars: 20 μm. (b') Mouse lines strategy used to perform live imaging. (b'', c') Left: Number of rosettes normalized by the total number of cells expressed in percentage. Right: Density of rosettes, normalized by the area in mm2 of the epiblast region in focus. The posterior region includes the PS region, as it could not be precisely discriminated. Data information: Mean ± SEM. E5.75: n anterior=19 frames from 3 embryos, n lateral=23 frames from 5 embryos, n posterior=20 frames from 3 embryos. E6.25: n anterior=16 frames from 2 embryos, n lateral=12 frames from 1 embryo and n posterior=16 frames from 2 embryos.
(G-I). Motor function tests. Rotarod (G; wt, n= 7; LowOXPHOS, n= 6), Open Field (H; wt, n= 4; LowOXPHOS, n= 4) and gripforce (after fatigue, I; wt, n= 7; LowOXPHOS, n= 6) tests. In H, lines represent the movement of the mice in the cage. The histogram represents the relative time that wt and and LowOXPHOS mice expended in the corners. Data information: Bars are the mean ± s.e.m. of the indicated (n) mice/genotype *p <0.05 when compared to wt by ANOVA or Student's t-test.
f, Xbp1 mRNA splicing (Xbp1u, unspliced; Xbp1s, spliced) of crypts; densitometry in g (n = 7; unpaired Student's t-test; mean ± s.e.m.).
E. Parental U2OS and derivative lines stably expressing GFP-RADX alleles or all three RPA isoforms at near-endogenous levels (U2OS/Super-RPA) (Fig EV4A) were transfected with control (−) or RADX siRNA. Replication fork speeds were determined as in (A) (bars, mean; n=400 fibers, pooled from two independent experiments, per condition).
P Gln levels in the medium after treatment of MeWo cells with mock- or si-GCN2 for 24 hr and subsequent incubation with LDHA-i for 48 hr. Data information: Statistical analysis was performed by ordinary one-way Anova for the comparison of more than two groups. Data are shown as the mean ± SD, n = 3. ns: not significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001
UMAPs of single cell transcriptome data. (B) UMAPs of epithelial, immune, and stromal cell transcriptomes, color-coded by tissue origin as assessed by marker genes. For marker genes, see Table EV3.
Staining, and quantification of p62 in ECs from healthy subject and patients either treated with vehicle (DMSO) or BAY. Scale bars, 50 µm; (n=3 patients per group; n=80 cells; one-way ANOVA; **p<0.0001 vs healthy DMSO; ##p=0.003 vs patients DMSO).
DAF-FM staining, a vital dye for nitric oxide (NO), showed that 57.56% of kugeln were positive for nitric oxide reactivity (2.5µM incubation from 96-102hpf; n=22 4dpf embryos; 118 of 205 kugeln filled; 3 experimental repeats). Images show representative "filled" (DAF-FM positive) and "unfilled" (DAF-FM negative) kugeln.
(C) Serum ROS of the mice from (A) were determined. Data information: Data from 3 experiments more 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.
I) Representative immunoblot and quantitative analysis (N=5 per condition; P<0.0001) of AMPK demonstrating loss of function following transfection with an shAMPK construct in C2C12 myotubes.
(B) Binding kinetics of the GFP-tagged Chs5 and ChAPs at the TGN is different. Data processing was carried out as in (A).
(H-I) EPG-7::GFP is expressed in pharyngeal muscles (H, arrow) and cells in the tail region (I) at larval stages.
A X-ray cocrystal structure of XMU-MP-5 with wild-type ALK (PDB ID: 7BTT). Specific interactions are depicted as dash lines, hydrogen bonds are shown in yellow and salt bridge is colored red.
WT and Bax-/-Bak-/- Hoxb8-FL cells were treated for up to 8h with 500 nM CHIR and p21 mRNA levels were compared to Hprt in response to CHK1-inhibition by real-time RT-qPCR. Bars represent means ± S.D. of independent experiments (n=4-6), performed in duplicates.
E-G. Resistance to 2DG induced cell death in Rab25 cells is mediated through the PI3K/AKT pathway. Inhibition of the PI3K/AKT pathway was achieved by addition of 10 µM of PI103 or MK2206.G. Or down-regulation of Rab25 by shRNA specific to Rab25 enhances 2DG induced cell death. *p < 0.001 versus RNAi control cells.
qRT-PCR detection of IFNB1 and IL6 mRNA in THP-1 cells (H, I) and mBMDMs (J, K) stimulated with ecGAMP (5 μg/ml) and icGAMP (0.1 μg/ml), respectively, for 4 h in presence of DMSO or bafilomycin A1 (BafA) (1 μM).
G Proposed model of polarized co-localization and expression compensation between ISP1 and ISP3.
I) Quantification of March5 at disease end stage. Results are expressed as mean ± SEM and percent of Non Tg; n= 8 (4 males and 4 females) mice per group. No statistically significant differences were found between G93A and PKO/G93A by paired Wilcoxon's test (p= 0.742). Protein levels are normalized by Complex V. March5 levels are unaffected by Parkin knockout.
F. STRING-based and Cytoscape-visualized network of phosphorylated proteins belonging to the Recycling Adaptors cluster (light green in E) and found in TPA1. Kinases are highlighted in burgundy. The border is color-coded based on the substrate motifs.
B Immunoblot analysis of the serine/threonine phosphorylation (pS/T) status of HIF2α in GSCs after inhibition of ERK1/2 activity. GSCs were cultured under 20% or 5% oxygen for 24 hours, then treated with the MEK inhibitor U0126 (10µM) for 30 minutes to inhibit ERK1/2 activity, and harvested for immunoprecipitation with anti-HIF2α antibodies. Immunoprecipitated endogenous HIF2α were immunoblotted with anti-pan-pS/T antibodies. In comparison with 20% oxygen treatment, the treatment with 5% oxygen elevated pS/T of HIF2α in GSCs, but the increased pS/T of HIF2α was abolished by U0126 treatment.
GSEA plot showing normalized enrichment scores (NES) for cholesterol homeostasis, positive regulation of cholesterol efflux, SREBP targets and stem cell proliferation signatures using RNA-seq data from 24OHC and DMSO treated cells.
D) ykt6ts pho8∆60 cells containing an empty plasmid, 3HA-Ykt6, or ADH1 promoter driven 3HA-Ykt6 wild type or 3HA-tagged Ykt6 mutant variants as indicated were grown and Pho8∆60 activity was monitored as described in A).
When cancer cells enter mitosis, BCL9 will bind to the β-catenin/Axin1 complex via its N-terminal, which includes HD2. As CDK14 and CDK1 kinases are activated by Cyclin Y/Y-L and Cyclin B1, LRP6 is phosphorylated by CDK14, and BCL9 is phosphorylated by CDK1 on its N-terminal. These changes will stabilise BCL9 and LRP6 signalosome components and prevent clathrin from binding and subsequent clathrin-mediated-endocytosis with mechanisms not fully explored, which further maintains mitotic Wnt/STOP signalling, modulates efficient cell division and promotes cancer cell growth.
(F) Schematic representation of wild-type and deletion mutants of SLFN11 used in this study.
Image of homozygous mutant nrpb2-2 fully complemented by NRPB2WT-FLAG (top, NRPB2WT +/+ nrpb2-2 -/-) and partially complemented by NRPB2Y732F-FLAG (bottom, NRPB2WT +/+ nrpb2-2 -/-). Plants were grown for 4 weeks in soil. Scale bars represent 1 cm.
(A) Representative immunoblots of the indicated proteins from whole-cell lysates of MCF7 cells subjected to 100 nM rapamycin or amino acid starvation for 0-24 h.
H, I Meiotic metaphase I spread preparations of Miwi heterozygous (H) and knockout (I) spermatocytes. Scale bars, 5 μm.
(a) Cytosolic calcium levels were evaluated in iPSC-derived neurons at basal conditions by measuring the 340/380 nm ratio of Fura-2 acetoxymethyl ester. Data are represented as mean+s.e.m.; experiments were independently repeated three times in triplicate. *P0.01; Student's t-test. (Number of cells analysed: CTRL, n=58; RecNcil/wt, n=36; RecNcil/wt GC, n=107; L444P/wt, n=82; L444P/wt GC, n=33; L444P/L444P, n=62). (b) Caffeine (10 mM) was applied to iPSC-derived neurons, and the peak cytosolic calcium concentration was measured as a change in Fura-2 fluorescence. Data are represented as mean+s.e.m.; experiments were independently repeated three times in triplicate. *P0.01; **P0.001, Student's t-test. (Number of cells analysed: CTRL, n=58; RecNcil/wt, n=36; RecNcil/wt GC, n=107; L444P/wt, n=82; L444P/wt GC, n=33; L444P/L444P, n=62).
(E) Lgr5GFP-CreERT2;Rosa26lsl-lacZ (LRlacZ) were crossed with Rest-/- mice, followed by TAM administration for lineage tracing for intestinal whole-mount staining for β-gal (upper panel) . Mice were sacrificed after one week and typical sections were shown (middle panel). Scale bar, 200 μm. Numbers of traced crypt-villus units were calculated as means± S.D. ** P < 0.01 by two-tailed Student's t-test in right panel. 50 fields (10 fields per mouse) were calculated for each group (n=5) (lower panel).
G. Pan-neurexin deletion does not decrease the readily-releasable pool of vesicles, but lowers the initial release probability during a high-frequency stimulus train. Summary graphs show the cumulative EPSC amplitudes extrapolated to time zero as an estimate of the readily-releasable pool size (left), and the ratio of the first EPSC amplitude divided by the cumulative EPSC amplitudes extrapolated to time zero as an estimate of the initial release probability (right).
To characterize subcellular distribution of target transcripts, real-time PCR analysis was done on the fractionated MCF7 cells with the indicated experimental treatments.
(D) Tukey box plot of ZEB1, ZEB2, TWIST1, and MITF mRNA expression according to the IC50 of the drug (µM) administered (BRAFi/MEKi), in melanoma cell lines from the CCLE (n=28) (Student's t-Test). High ZEB1, low ZEB2, and low MITF expression levels were correlated with BRAFi (PLX4720) and MEKi (AZD6244) resistance. PLX4720 is an analog of PLX4032.
F SlHAK20 complements the K+ uptake-defective yeast mutant assayed using a dose-response method. The K+ uptake-defective yeast mutant R5421 transformed with SlHAK20Hap1, SlHAK20Hap2, and vector (p416-GPD) were cultured in AP medium containing indicated concentrations of K+ for 24 h. The data points represent three replicates.
B) IFNγ-stimulated MEFs were infected with Pru, PruΔgra15 and RH for 3 h and subsequently stained for TRAF6. 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. Experiments were performed 3 times.
(a) Quantification of numbers of hyperplastic lesions/per lung (mean values±s.e.m.) and (b) representative histological lung sections (haematoxylin and eosin (HE) staining) of KRas;Atg5fl/+ and KRas;Atg5fl/fl mice2 weeks after AdCre inhalation treated in vivo either with either vehicle control or POM1 (intraperitoneally; i.p. 10 mg kg−1 daily) to inhibit CD39 activity. n=4 per group. ***P0.001 (χ2-test assessing the genotype effect in a generalized linear model with log link). Scale bars, 2 mm.
(F-H) EM images of neurons. The red dashed line marks the outline of neurons. The red arrows mark LDs. Scale bar: 0.5 μm. The scale bar in enlarged images is 100 nm. (I) Serial sections of atgl-1(xd314) show two LDs (indicated by blue and red arrows) in a neuron cell body. Scale bar: 0.5 μm.
(G) FACS analysis of cell cycle using DAPI. Numbers indicate the percentage of cells in G1, S and G2/M respectively.
A HeLa cells expressing LAMP1-RFP were treated with vehicle alone (control) or LLOMe for 1 h to induce lysosome damage. Cells were fixed 2 h after washout, stained with p62 and galectin-3 (Gal3) antibodies, and analyzed by confocal microscopy.
B The system does not interfere with phage adsorption to bacterial cells. Strains either containing (red) or lacking (black) the BREX system were infected with phage Φ3T and then chloroform‐treated 15 min following infection. The culture was plated on Φ3T‐sensitive B. subtilis cells and plaques, representing extracellular, unadsorbed phages, were counted.
B Boxplot showing gene expression levels among different groups in the E13.5 forebrain. A total of 22075 genes with transcripts per million (TPM) greater than zero were grouped into quartiles according to expression levels (Quartile 1-4, each quartile containing 25% of genes). AP: active promoter, co-marked by H3K4me3 and H3K27ac. There were 11451, 9689, and 9204 genes under the regulation of H3K9ac, H3K9cr, and the active promoter, respectively.
(B) Immunoprecipitation assay showed that Src kinase interacted with PLCγ1. Rac1 is a positive interacting control for PLCγ1 (N = 3).
(c) Beclin-1-shRNA reconstituted lines (WT or mutant) and controls (scramble shRNA or Beclin-1 shRNA) were grown with or without amino acids and assessed for autophagy (left panel) and Beclin-1 levels (right panel).
(F) Western blots for GFP-Buc expression in 4-cell stage WT and MZsinhcaf -/- zebrafish embryos injected with GFP-Buc and RFP mRNA
(A-C) Atg5 KO MEFs were either untreated (NT) or starved (ST) with EBSS (Earle's Balanced Salt Solution) for 30 min. Total membranes (mem) from lysed cells were collected and incubated in a lipidation reaction with cytosols prepared from starved HEK293T cells. Reactions contained the indicated concentrations of PI3K inhibitor (PI3KI) 3-methyladenine (3-MA) (B) or FYVE protein (C). A diagram of the experimental scheme is shown in (A). RPN1, Ribophorin 1
A. HEK 293T cells were transfected with pCS2 vector (black), pCS2-Endouc (red), and pCS2-EndoucK242A (green) for 24 hr and subjected to either DMSO (control) or thapsigargin (TH; stress) treatment for 1 hr. The lysates of transfected cells were applied to polysome profile analysis (PPA). The sucrose gradient from top to bottom was shown from left to right, respectively. The sedimentation of 40S, 60S, 80S, and polysomes was indicated. Lysate from control, Endouc-expressing, and EndoucK242A-expressing cells were applied to PPA. The resulting fractions were TCA-precipitated, analysed by SDS-PAGE, and subjected to Western blot using antibodies against Flag and S6.
Yield reduction assay was performed to determine the inhibitory effects of ACE2-Fc on protein expression. The culture medium and cell extracts were harvested 24 hours post-infection for western blot (C). The NP/PCNA represents the relative NP expression as compared to that of PCNA, which served as a loading control. The NP/PCNA numbers below the panel are the ratios of NP/PCNA normalized to that of the human IgG control group. PCNA: Proliferating cell nuclear antigen.
(B) Characterization of Mrp17 translocation signal using an in vitro import assay: shown are autoradiographs of full-length Mrp17 or its truncations fused to DHFRmut, translated in vitro with radiolabeled amino-acids, incubated with isolated yeast mitochondria for 2, 5, or 10 min, treated with proteinase K (PK) to remove nonimported proteins and visualized by 16% SDS-PAGE/autoradiography. As a negative control, mitochondria were treated with valinomycin, antimycin and oligomycin (VAO) that eliminate membrane potential. For comparison, 20% of the protein used per import reaction was loaded on the first lane.