EMBO/sd-character-level-ner huggingface dataset:
(E and F) Freeze-substitution fixation image of Cvt bodies in the vacuole (marked with arrows). A Cvt vesicle is marked with an arrowhead.
F. The heatmap of T cell activation biomarkers for RNA-seq analysis. The unit of heatmap scale is expression value after centering and scaling by genes.
Quantitative phosphoproteomic analysis defining the role of Dna2 (WY-AA mutation impairs Mec1-activating function of Dna2) in the set of Mec1-dependent phosphorylation in rad9∆ cells. Dna2 dependency (y axis) is plotted against Mec1 dependency (x axis). These phosphoproteomic analyses were conducted in the presence of 0.02% MMS for 2 hours.
The following experiments were performed in glial shNf-1/shTp53 glial cells serum starved 4 hr before assaying. Cells were expressing either Cas9 alone (control) or Cas9 and sgRNA targeting Atg7 (sgAtg7 #1 or #2). G: Pearson's colocalisation coefficient between mCherry-Rab5 and EGFR Data information: Statistical analyses were performed on at least 3 independent experiments, where error bars represent SEM and p values represent a two-tailed Student's t test: NS p>0.05, * p<0.05, ** p<0.01.
B. ShRNA#1 and 2 downregulate Nrg1 type III expression also in myelin-forming Schwann cell/DRG neuron co-cultures analyzed after 7 days of ascorbic acid treatment, as shown by western blot analysis, representative of two independent experiments. Clx is calnexin.
F MAGE-A3/6 protein instability upon nutrient deprivation of HeLa cells is not altered by EGF 0.2 ng/mL, testosterone, or bovine pituitary extract 25 µg/mL.
Mitochondrial membranes from fibroblasts of control (WT) and patient fibroblasts (i, immortalised) were solubilised with digitonin and separated on native gradient gels. A BN-PAGE gel stained with Coomassie showing molecular weight marker (BHM) and samples from control and patient (primary fibroblasts, (I)=immortalised fibroblasts) B BN-PAGE gel stained with NADH:NTB reductase activity stain showing molecular weight marker (BHM) and samples from control and patient (primary fibroblasts, (I)=immortalised fibroblasts) C Further complexome analysis of the BN-PAGE gels shown in A and B. Each native lane was cut into even fractions from high to low molecular mass. Proteins in each slice were digested with trypsin and analysed by quantitative mass spectrometry. Shown here are average IBAQ values of all identified subunits from complexes III, I and IV
The duration of EB3-positive comets presence in the bottom plane was measured in DMSO and CK666 treated cells (left). Error bars correspond to standard deviations. The number of EB3-positive comets exiting the a 2-µm wide centrosomal area was also compared between the two conditions (right). In both cases P values were calculated with Mann-Whitney test.
A, HCT116 cells were treated with 100 nM Thapsigargin for the indicated times in the presence or absence of shRNA targeting USP22. Cells were harvested and whole-cell lysate was subjected to immunoblotting with the indicated antibodies.
BLI sensorgrams of intermediate VHHs A SARS-CoV-2 RBD-loaded sensors were incubated with nanobodies at the indicated concentrations for 300 s, followed by dipping in assay buffer for 600 s. The response curves (black) were fitted using either a 1:1 (for VHH-72) or 2:1 (for Re6B06 and Re9F06) binding models to obtain KDs. Fitted curves are shown in red. For Re6B06 and Re9F06, some secondary binding with weaker affinity at high VHH concentrations was observed (denoted as KD2).
(a) The amount of glycogen in the whole larvae measured by biochemical assay (n=4), normalized to RasV12; ctrl at day 6, and per number of larvae.
Experimental setup to determine lung colonization of in vivo vehicle-treated and JNK-inhibited MDA231-LM2 cancer cells after intravenous injection. In vivo treatment with JNKi commenced at the time of implantation and was repeated every 3 days thereafter until analysis at day 17.
Evoked and spontaneous release in autaptic munc18-1 null hippocampal neurons expressing M18Y473D, M18Y473D/P335A or wild-type Munc18-1 as control.(B) Typical traces of spontaneous release.
C and D. Immunoprecipitations of YAP protein and western blot analysis for p53 binding are performed from lysates of cancer cell lines expressing different mutant p53 proteins (described in Materials and Methods) (C) and from HCT116 cells expressing wild type p53 protein (D). YAP protein was immunoprecipitated with a rabbit polyclonal antibody and the same amount of rabbit IgG was used as negative control of IP.
Scheme of API transport pathways to the vacuole. As described in the text, proAPI forms a complex in the cytosol. This Cvt complex is then targeted to the vacuole by two pathways depending upon the nutrient conditions. In growing conditions, a double membrane vesicle of ∼140 to 160 nm forms around the Cvt complex and excludes cytosol. In contrast, when cells are starved for nitrogen, the Cvt complex is taken up into larger autophagosomes (300-900 nm) together with cytosolic components. Vacuolar delivery is achieved by fusion of the outer membrane of the Cvt vesicle or autophagosome with the vacuolar membrane. Finally, the resulting Cvt bodies or autophagic bodies are broken down by vacuolar hydrolases to release proAPI into the vacuole lumen. The precursor form of the protein is then processed into the mature hydrolase.
Unilateral AAV-mediated human P301L tau expression in the EC and DG of age-matched WT mice (n=3). Representative images of brain sections show donor neurons in the injected EC and DG, and a few tau recipient neurons (white arrows) adjacent to the AAV injection site. Scale bar 50 µm.In the contralateral hemisphere of the same brain section, some tau recipient neurons (white arrows) were also present in the (non-injected) axonal projection areas in the contralateral EC (GFP-filled terminal ends). Scale bar 100 µm.
E. Growth curve for POT1 WT and POT1 knockout cells. Population doublings (PDL) are represented as mean ± SD (three biological replicates).
(F) IFNγ-primed WT, Irgm2-/-, Casp1-/-Casp11-/- and Irgm2-/-Casp1-/-Casp11-/- BMMs were treated with LPS (1 µg/ml) for 24 h and cell lysates and supernatants collected. Protein levels in cell lysates (caspase-1, and actin) and supernatants (caspase-1 p20) were visualized via immunoblotting.
C. Rosa26_Alkal2;Th-MYCN animals treated with lorlatinib did not display any significant loss of body weight compared with vehicle controls. Data shown represents mean ± SD.
(E) Heatmap depicting mRNA levels of SMAD3-signature in BRAF(V600E) non-treated melanoma patients (dataset from TCGA; SKCM, BRAF(V600E) mutated: n=118). Three pigmentation genes (MITF, MLANA and TYR) have been added to highlight the differentiation states of tumors. ~20% of tumors are considered as dedifferentiated tumors (red box) with a high SMAD3-signature.
(g) Quantification of larval motility measure by crawling distance and crawling pattern in RasV12; ctrl (n=14), RasV12; scrib-/- day 6 (n=15), day 8 (n=16) and day 10 (n=14), each colored line represents a single larva.
(c) Twenty-four human sera were then two-fold diluted and incubated with the ISA D614 and mCherry D614 strains and nAb titers were recorded at 5 days post-infection. nAb titers were defined as the highest dilution that inhibited the production of distinct cpe with the ISA D614 SARS-CoV-2 or fluorescence with the fluorescent mCherry D614 SARS-CoV-2. Each black dot represents results from a given number of sera. Statistical analyses were performed using univariate linear regression. The error band (in grey) represents the 95% confidence interval of the regression line. The Pearson correlation coefficient (R2) and P value analyses are shown.
(B) Immunoblot analysis of the immunoprecipitated HA-SOX9 or FLAG- FBW7α and their pull-down products. The corresponding Western blotting from the whole cell extract (WCE) of the transfected HEK293 showed the levels of exogenous HA-SOX9 or FLAG-FBW7α proteins in the total input. Cells were treated with 10 µM MG132 for 4 hours prior to harvesting for the immunoprecipitation. Blots are representative from 3 independent experiments.
C) Dietary supplementation of 3-methyl-L-his for 4 days significantly increased adult nerfin-1 clonal growth, and rescued nerfin-1 growth inhibition due to his dietary withdrawal (n =18,14, 7,9).
(B) The integrity of the isolated lysosomes. The release of β-hexosaminidase from lysosomes was indicated with incubation of lysosomes with para-nitrophenylphosphate (PNPP). The reaction was terminated with 0.25 mol/L NaOH. Release of all lysosomal β-hexosaminidase was induced by incubation with 0.1% Triton X-100. Release of all lysosomal β-hexosaminidase was induced by incubation with 0.1% Triton X-100. Values of β-hexosaminidase activity are expressed as percentage of total activity in the lysosomal fraction at time 0, 10, 20 and 30 min and are means±SE of three independent experiments.
(E Detection of steady-state conversions of endogenous MgtE by YqgP variants bearing single-point mutations in putative Mn-binding region (BS184; 187; 196-203, Table EV2), cultivated in modified M9 minimal medium supplemented by low (1 μM) or high (100 μM) MnCl2. Black arrow marks full-length MgtE, and red arrow marks its N-terminal cleavage product by YqgP.
Histopathological examinations (F) in lungs from all the inoculated macaques at day 7 post infection. Lung tissue was collected and stained with hematoxylin and eosin.
C. Close-up views of the DCX binding site focusing on loop 3 density, which differentiates DC domains. Refined atomic models in predominantly backbone representation are fitted to their respective density maps (wireframe). Selected residues are labelled with single letter codes and selected side chains are shown as sticks. Nucleotide in tubulin subunit designated as β1 is shown as sticks in the top left corner of each panel. Colours as in (B), or oxygen atom, red; nitrogen atom, navy blue.
At steady state, nuage and piP-body accommodate piRNA factors both stably localized at the condensate or transiently docked to it. Siwi, while does not accumulate in piP-body, transiently enters piP-body and interacts with BmSpnE and BmQin. An active slicer is required for Siwi to segregate from piP-body and go back to nuage, where BmVasa uses its ATPase to remodel RNP interaction and modulates the ping-pong cycle. Some Siwi-interacting factors like BmMael and BmArmi can enter multiple compartments where Siwi is present. When Siwi-D670A is expressed, the slicer mutant is expelled from nuage by RNP remodeling activity of BmVasa and is trapped in piP-body while co-aggregating with piP-body factors, as well as those freely shuttling between compartments. A hypothetic fidelity determining factor could be similarly trapped in piP-body by the mutant, resulting in the up-regulation of non-TE piRNAs. This production could be done in nuage, where native piRNA biosynthesis machineries may still function. Of note, the length distribution and the presence of 2′-O-methyl modification in non-TE piRNAs suggest that these piRNAs are likely to be processed on the outer mitochondrial membrane and in the cytoplasm. It is also possible that non-TE piRNA is directly generated from piP-body, where a battery of piRNA biosynthesis factors abnormally accumulated.
(A) Glu529 (purple) forms a hydrogen bond with the C2-amino group of the primer terminal templating 8-oxo-dG (orange) in the anti-conformation. (B) This interaction is abolished when 8-oxo-dG (orange) adopts the syn-conformation. An overlay with the 8-oxo-dG(anti)-containing structure reveals that Glu529 (black) would likely clash (red-dotted line) with the C8-carbonyl of 8-oxo-dG(syn). Repositioning of Glu529 (purple) places the side chain 4.5 Å away from the C8-carbonyl. (C) Comparison of catalytic efficiencies for extension past an 8-oxo-dG(anti):dC, 8-oxo-dG(syn):dA and dG:dC base pairs at the primer terminus using WT and E529A Pol λ.
p53-/- C57BL/6N male mice (n=7) were injected with a single dose of diethylnitrosamine (DEN) (25mg/kg) at two weeks and then fed with HFD diet at 4 weeks. Oral administration of terbinafine were given at 13 weeks old as indicated. Body weight (J) of mice are shown.
M238P cells were transduced with a miR-143/-145 construct and selected for stable expression of the cluster or transduced with a control vector, followed by RNA-seq analysis (dataset 2, n = 2). (D) Heatmap highlighting the common predicted upstream regulators altered in cells stably expressing the miR-143/-145 cluster and M238R cells compared to control M238P cells. A subset of common regulators (out of the top 50 scores) corresponding to transcription factors (TFs), cytokines and growth factors (GFs), transmembrane receptors, kinases and phosphatases is shown. Red arrows indicate annotations related to the TGF-β pathway.
(A)-The total activity of flies recorded on the second day of experiment (mean +/- SD). Silencing the Tsc1 gene in per-positive cells increased the total activity of the flies [U Mann-Whitney Test, * for p≤0,05].
(E and F) Immunostaining assay of SOX1 (red) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of dorsal (E) and ventral (F) organoids differentiation. Scale bar, 100 μm. (G) Immunostaining assay of MAP2 (green) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of dorsal organoids differentiation. Scale bar, 100 μm. (H) Immunostaining assay of GABA (green) after SOX1-OT V1 overexpression in V1-PAKI cells on day 40 of ventral organoids differentiation. Scale bar, 100 μm.
Flow cytometry analysis of CD69 expression in P/I-stimulated CD3+ T cells treated with or without FK506 or CX5461.
Solubilized mitochondria from MITRAC15FLAG- or C12ORF62FLAG- expressing cells were subjected to anti-FLAG immunoisolation and analyzed by SDS-PAGE and immunoblotting. Total 0.4 %, Eluate 100 % (MITRAC15FLAG isolation), 70 % (C12ORF62FLAG isolation).
(A-F) Coronal brain sections obtained from mice at 4-6 (A-C) or 20-23 (D-F) months of age were immunostained for NeuN. The number of NeuN-positive cells with neuronal morphology was determined in dentate gyrus (A, D), CA3 (B, E), and CA1 (C, F). n = 11 (A-C) or 18 (D-F) NTG; 11 (A-C) or 5 (D-F) CamKII-tTA; 7 (A-C) or 9 (D-F) hTau-WT(L32); and 14 (A-C) or 10 (D-F) hTau-A152T(L1) mice. *P < 0.05, **P < 0.01, ***P < 0.001 by one-way ANOVA and Tukey test. Values are means ± SEM.
K. Schematic diagrams showing the glucose or lactate derived carbon flow in pericytes (HBVPs) when [13C6]-glucose or [13C3]-lactate was used as labeling substrate(s). (Unlabeled carbon : black circle, labeled carbon : red circle)
Scatterplot of proteins identified by mass spectrometry analysis identifies NEDD8 and components of the NEDD8 conjugation pathway as being the most abundant proteins in HEK 293 NEDP1 KO lysates following NEDP1-CA pulldown. iBAQ analysis of proteins identified in A) are plotted as the log2 value of the enrichment ratio (mass spectrometry intensity of the HALO-NEDP1 CA pulldown over HALO-NEDP1 DAGC pulldown) versus the log10 value of the iBAQ intensity from the HALO-NEDP1 CA pulldown. Blue markers indicate known components of the NEDD8 pathway and red markers indicate proteins that have been identified as substrates of PARP-1 in the database of ADP-ribosylated proteins, ADPriboDB (Vivelo et al., 2017).
WT and Mfn2 KO cells were transfected with plasmids encoding: GFP and ChiMERA or control plasmid. After 24 h, the transfected cells were sorted and plated for another 24 h, when (K) oxygen consumption was measured (n= 4 independent experiments). Data are presented as mean ± SEM. Data information: *p<0.05, one-way ANOVA followed by Tukey's post hoc test. Note that experiments were performed at the same time, so the values of WT and Mfn2 KO transfected with control plasmid are the same but the figures have been split in two for the sake of linearity.
Immunoprecipitation (IP) with an anti-TRAPPC2 Ab or IgG from cells with or without SA treatment, as indicated, followed by SDS-PAGE and Western blot with anti-RACK1 (G) Abs. Input: cell lysate (40 µg for TRAPPC2, 10 µg for RACK1
On day 15 during reprogramming, the expression of E-cadherin (E-CAD) and N-cadherin (N-CAD), was determined with immunofluorescence together with the morphology in Oct4GFP+ colonies (A, Scale bar, 25 μM).
B Phenotype of wild type, MZgrk2, and grk2-GFP mRNA injected MZgrk2 embryos at 24hpf (n=20 for each sample). The white lines indicate the shape of the somites (middle panels) and the separation of the eyes (right hand panels). Scale bar, 200μm
(A) Experimental set up for the MRI (non-invasive magnetic resonance imaging) experiments.
The baseline IL-6 level was 197.39 pg/mL in a 69-year-old female patient who showed high fever and dyspnea. IL-6 decreased to 9.47 pg/mL after treatment (day 8), while the symptoms were not relieved. The C-reactive protein (CRP) rebounded and procalcitonin (PCT) increased together with disease exacerbation. Follow-up chest computed tomography (CT) assessment was not performed due to poor general condition, whereas chest X-ray showed aggravated pneumonia. Follow-up sputum culture confirmed the exacerbation was caused by bacterial infection.
C. Correlation of the mean LFQ intensities from (A) and YFP+ cells from BiFC assay from (B). Dotted lines show differences in interaction of the POIs with STIM1 in comparison with STIM1A.
(f,g) Representative electron microscopy images (f) and quantification (g) of autophagic vacuoles in control and miR-212/132-overexpressing TG H9c2 cells under normal and serum/glucose-deprivation conditions (n=20).
B Increasing protein age results in increasing tandem reporter ratio. The fluorophore states of the tFT reporter are shown in the inset: (1) immature; (2) only sfGFP matured; (3) only intermediate mCherry matured; (4) sfGFP and intermediate mCherry matured; (5) only mCherry matured; and (6) both mCherry and sfGFP matured. kg, kr1, kr2 denote the maturation rates of sfGFP, intermediate mCherry and fully-matured mCherry respectively. The different curves correspond to sets of values for kg, kr1, kr2 selected from Gaussian distributions with half-time means 27, 40 and 9 minutes and standard deviations 2.7, 4 and 0.9 minutes respectively.
The following experiments were performed in glial shNf-1/shTp53 glial cells serum starved 4 hr before assaying. Cells were expressing either Cas9 alone (control) or Cas9 and sgRNA targeting Atg7 (sgAtg7 #1 or #2). E: Cells transiently expressing YFP-Gal8 were treated 100 μM monensin for 1 hr and stimulated with 2 ng/mL EGF for 15 min before fixation and immunofluorescence staining against EEA1. Scale bar: 10 μm. F: Quantification of percentage of total EEA1 vesicles that colocalise with YFP-Gal8 upon monensin treatment (in E). Data information: White arrows indicate colocalisation. Statistical analyses were performed on at least 3 independent experiments, where error bars represent SEM and p values represent a two-tailed Student's t test: NS p>0.05, ** p<0.01, *** p<0.001.
A Binary food choice assays were conducted after GRNs were ablated by expressing the cell death gene (hid), under Gr28b.c-GAL4 (n = 4).
(I) Early EMT and facilitated OGS form a positive feedback loop to induce pluripotency.
A. Optical coherence tomography (OCT) horizontal B-scan and thickness map of neovascular age related macular degeneration (NV AMD) patient with active lesions (1), cystoid macular edema (2), subretinal fibrin deposits (3) and serous retinal detachment (4). B. Optical coherence tomography (OCT) horizontal B-scan and thickness map of control patient with a medium sized, stage 3, full thickness macular hole (5).
(A, B, M238P cells were transfected with miR-143-3p, miR-145-5p or a control mimic (miR-neg) (72 h, 30 nM). Effect of miR-143-3p or miR-145-5p overexpression on YAP (A) and MRTFA (B) nuclear translocation by immunofluorescence. Data are represented as scatter plots with mean ± SD (n≥30 cells per condition). Each point represents the nuclear/cytoplasm ratio. Mann-Whitney U test was used for statistical analysis. ****P≤0.0001. Scale bar 40 μm.
A. Left, control or SA-treated (500 µM, 120 min) human fibroblasts (HFs) were incubated at 40°C for 180 min and stained for eIF3, PCI, and Sec24C. Right, the cells were imaged 10 min after shifting the temperature from 40°C to 32°C and stained for eIF3, PCI, and Giantin (to label the Golgi). Data information Scale bars, 10 µm.
C. In the public database lncCAR, high expression level of lnc-AP was correlated with long overall survival in CRC patients.
qRT-PCR detection of Il23p19 mRNA expression levels in BMDCs of indicated genotype stimulated with IMQ for 4h and/or pre-treated with JNKi (SP600125, 25µM, 1h). (n = 4-14; 2-5 independent experiments). Data information: Data are shown as mean ± SEM. P‐values were calculated by Tukey multiple comparison test Statistical significance: ns > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001. See Appendix Table S3 for exact P‐values.
A. Representative images of time-lapse spinning disk confocal images of n = 3 of COS7 cells stably expressing GFP-TMD (green) and SiR-lysosome-stained LE/Lys (orange) treated with either siControl or co-treated with siVAPA, siVAPB and siMOSPD2. Zoom insets show regions of interest containing mobile LE/Lys as denoted by the white arrow. First row represents ER (TMD-GFP) (green), the second row shows MDA generated ER movement (magenta), third row shows a merge of the two preceding channels and the fourth row shows a merge of ER (green) and SiR-lysosome stained LE/Lys (orange). Scale bar: 5µm, zoom insets 3µm. Images related to movie EV40 and 47. Images were acquired for 90 frames at 1Hz.
(C) Changes in body weight relative to starting weight during 6 weeks (n = 6). Data information: Data were representative and were the mean ± SD from three independent experiments.
A Model fitting to the experimental tFT-Bcd ratio in early n.c. 14 as function of AP position for the different models Black line represents mean measured ratio of tFT-Bcd reporter with mCherry-sfGFP-Bcd in y/w flies. The shadowed area represents ±1s.d. Inset is same as main panel, except for the tFT-sfGFP-fmCherry-Bcd reporter.
D, E MAGE-A3/6 protein stability is controlled by a <3 kDa, heat stable, lipophilic molecule present in FBS. HeLa cells were treated with EBSS alone, or EBSS containing 10% complete FBS, 10% charcoal-dextran stripped FBS, 10% boiled FBS, 10% dialyzed >3 kDa FBS, or 10% dialyzed <3 kDa FBS for the indicated times. Quantitation represents average of n=3 with standard deviation shown. Data information: Asterisks indicate ** p<0.01; *** p<0.001. ns indicates p>0.05 using 2-way ANOVA.
G Total liver cholesterol concentrations of mice were examined. Protein levels are shown (bottom).
Total cell number of skin pDCs defined as in (B) is shown. (n= 5-6: 2 independent experiments) Data information: Data are shown as mean ± SEM. P‐values were calculated by multiple t-test with the Holm-Šídák method (C) multiple comparison test. Statistical significance: ns > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001. See Appendix Table S3 for exact P‐values.
(B) SMAD3 expression increases with melanoma cell dedifferentiation. PCA analysis of SMAD3 expression in melanoma cell lines in function of their dedifferentiation states. Scale: red color corresponds to a high SMAD3 expression level (unit of the scale : Log2 FPKM).
D CD69 up‐regulation on re‐stimulated THEMIS KD CD4+T cells. CD3/CD28 beads were removed on day 1 post‐transduction, and cells were cultured in IL‐2 and IL‐7. After removal of cell debris at day 4, cells were restimulated with CD3/CD28 beads at the indicated bead to cell ratio. CD69 surface expression was assessed by flow cytometry 24 h later. Data shown are representative of two independent experiments. n = 3, means ± SD are shown; two‐tailed unpaired Student's t‐test, *P 0.05, ***P 0.001.