EMBO/sd-character-level-ner huggingface dataset:
(B) Immunoprecipitation/immunoblot of phosphorylated E3 ubiquitin ligases and mitofusin 2 in MEFs in the of tunicamycin (5 μg/ml). LRRK2(G2019S) MEFs were also pretreated with LRRK2-IN-1 (1 μM). MEFs were transfected with each E3 ubiquitin ligase and biquitin and lysates were immunoprecipitated with antibody against the Myc or HA epitope. Precipitated proteins were subjected to SDS/PAGE, and blots were stained with antibody against the Myc epitope, phosphoserine or mitofin2 as indicated to the right of each panel. Mfn2: mitofusin 2, Ub: ubiquitin. Data represent ratios of phosphorylated to total E3 ubiquitin ligase and mitofusin2 to actin. Error bars represent ±SD from four independent experiments. Data information: For graph B, the P values was determined by a Mann-Whitney U test. ns = not significant, *P < 0.05, **P < 0.01
Micropeptide CIP2A-BP levels were determined by western blot with anti-CIP2A-BP antibody in MCF-10A and TNBC cells either mock treated or treated with TGF-β.
B Normalized probability of protein-protein contacts by phosphomimicking aspartates in 12D and serines in Wt resolved by amino acid type from coarse-grained simulations. Error bars smaller than symbols. Inset: Distributions of the number of water molecules within 5 Å of side chains of phosphomimicking aspartates of 12D and corresponding serines in Wt from 15 µs of coarse-grained molecular dynamics simulations.
(C) Anti-Myc IP of Plat-E cells expressing different Myc- and HA-B-Raf proteins. Note that B-RafAVKA is not impaired in its ability to interact with other B-Raf proteins, while homo-dimerization of B-RafEVKD is enhanced.
J Quantified fluorescence of UbG76V-GFP normalized to mRFP in the hypodermis of animals from the indicated time point (in hours) after the L4 stage. Animals were exposed to the indicated knockdown RNAi bacterial strains or a strain that only contained the empty RNAi vector since hatching. *P<0.001 using ANOVA with Dunnett's posthoc comparison to the wild-type control equivalent time point, with N=20 animals per genotype and time point. Error bars indicate SEM.
A) Venn diagram showing the overlap between genes down-regulated or up-regulated in response to the indicated perturbations.
D. Western blot of the expression of Vimentin, N-cadherin, and E-cadherin in MDA-MB-231 cells with the indicated treatments.
H. Levels of Axin-2 and cyclin D1 protein in LN229 cells transfected with FoxM1 siRNA#1 or #2 were determined by Western blotting.
A. ERK7 overexpression leads to phosphorylation of PWP1, as shown by appearance of an extra band on Phos-tag Western blot (marked with red arrow). Insulin-inducible bands (dark blue) remain unresponsive to ERK7.
(G and H) NIH3T3 cells stably expressing mStrawberry (mStr)-Gal3 and GFP-LC3, mStr-Gal3, and GFP-p62, or mStr-Gal3 and GFP-Ub were transfected with Effectene-coated latex beads for 30 min and then washed. Live cells were observed at 1-min intervals by fluorescence microscopy. Bar, 3 µm. The time after galectin3 localization was measured for at least 30 cases for each combination (H). Statistical analysis was performed by Student's unpaired t test. *, P < 0.05; NS, not significant.
FRAP experiments of Tau:PEG, Tau:polyA, and Tau:PEG:polyA condensates at 1h, 4h, and 24h after LLPS induction. Tau:PEG and Tau:PEG:polyA condensates show a decrease in recovery rate and mobile fraction over time. Tau:polyA coacervates maintain fast recovery and a large mobile fraction even after 24h. Time until half maximum recovery: t(y1/2)polyA: 1h≈2s, 4h≈3.5s, 24h≈4.5s; t(y1/2)PEG: 1h≈6s, 4h≈8s, 24h≈11s; t(y1/2)PEG+polyA: 1h≈4.5s, 4h≈9.5s, 24h≈9s). Scale bars=5 μm for Tau:polyA and Tau:PEG, and 10 μm for Tau:PEG:polyA. Data presented as error bars (SEM), N=15 to 20 condensates per condition and time point.
(A) Experimental paradigm performed in all the trials: mini-pumps were implanted in the striatum of 7-week-old mice. A battery of behavioral tests was performed before and after mini-pump implantation. In bold, a list of R6/2 phenotypes (above) and read-outs analyzed following cholesterol (chol) infusion (below).
(C) Cross-sectional analysis of the gastrocnemius muscle calculated from 250 fibers per mouse from four mice per cohort (n = 4). Cross-sections were stained by H&E. Scale bars represent 100µm Data information: Error bars represent the SEM. Significance between means was first determined using ANOVA. Significance p-values were calculated using Fisher"s LSD. *, p < 0.05; **, p < 0.01 from saline controls; †, p < 0.05; ††, p < 0.01 from C26 controls
(A) A bacterially purified catalytic domain of DAPK was incubated for 15 min at 30°C with increasing concentrations (5-10 nmol) of a peptide corresponding to the BH3 domain of beclin 1 (aa 108-127) and with the same peptide in which Thr 119 was substituted by alanine. An in vitro kinase assay was performed, and the reactions were applied to Whatman filters. Total levels of TCA‐insoluble counts were measured and plotted against substrate concentration. Data are the mean±s.d. of three experiments.
(C) Western blot of BN-PAGE of quadriceps of 6-month-old mice. We analyzed: CI (NDUFA9, NDUFB8), CIV (COX1), CII (SDHA). (D) Quantification of the BN-PAGE western blots from female mice shown in panel C.
(D) RT-qPCR analysis of relative Fancc mRNA levels from whole brains or testes in mock- and ZIKV Paraiba infected Ifnar-/- mice six days post-infection. Mean ± SEM of n=3 biological replicates, *p < 0.05 and ns by Student's t test.
(A-C) 3D rendering of Z-stack confocal image showing Isolectin B4-stained laser-burn with FITC-dextran-labeled CNV and IBA-1-stained MPs.
(E) WASH attenuates the interaction of Beclin 1 with Vps34. MEFs stably expressing vector or Flag-WASH were starved with EBSS for 1 h, followed by immunoprecipitation with anti‐Beclin 1 antibody. Ratio of Vps34/Beclin 1 was shown in the right panel.
G. m6A enrichment of Stat1 and Irf1 was examined by m6A RIP-qPCR in control, Mettl3, or Mettl14 depleted CT26 tumors as indicated. Ctla4 functioned as a m6A negative control (Wang et al., 2019) Data are mean ± s.d. * * P < 0.01 by Student's t-tests.
C. Hierarchical clustering of mRNA expression during differentiation of mESCs towards endoderm (yellow) or EpiSCs (green).
Relative accumulation of gRNA3, RNA3-derived vsiRNA, and vsiRNAs/gRNA3 ratios calculated from hybridization signal intensities of northern blotting in D, E. Values in Col-0 plants were set as 1. Data points represent means of three biological repeats. Error bars indicate SD. Letters indicate significant differences (ANOVA, P < 0.05).
The 173 genes where the transcriptomic and proteomic expression data (Exp) are consistent (see Appendix Figure S8) are compared with flux states predictions generated by parsimonious flux balance analysis (Flux). True Positives (TP) and True Negatives (TN) correspond to genes where both predictions and observations are consistent, while False Positives (FP) and False Negatives (TN) where they are inconsistent. Genes considered expressed or with an active flux are represented in dark blue, and silent in light blue.
D Western blot showing association of mitochondrial membrane with lysosomal membrane. The presence of membranes of each organelle was assessed using anti-MFN1, VDAC (as mitochondria markers), and LAMP1 (as a lysosomal marker). n = 3.
C. Netrin-3 high expression is a marker of poor prognosis in NB. 280 months overall Kaplan-Meier survival curves in a panel of 181 patients of all NB stages. The cohort was dichotomized base on netrin-3 median expression (in green and red). Statistical treatment of the data: Mantel-Cox; p value is indicated below the graph.
Quantification of LPS insertion sites from (D). Numbers are indicating mean percentages. n = 566 bacteria (3 biological replicates).
(g-j) Representative cut views from three-dimensional reconstitution images from wild-type (g,h) and Atg1 knockdown (i,j) cells at early third instar larval stage (g,i) and at puparium formation (h,j). (k) Quantification of cell volume (μm3) from g-j, n = 5 animal intestines per genotype with 1-5 cells measured per intestine per stage. Black columns: early third instar larval stage. White columns: puparium formation stage. Quantification is shown as mean ± s.d. NS, not significant. Scale bars, 20 μm.
(B), same as (A), in HepG2 cells stably expressing pCYP1A1 luciferase plasmid (AhR reporter). Chemical structures of FKK5, FKK6 and FKK999 compounds are overlaid. Indole is colored blue. The bar graph(s) depicts one representative experiment of a series of experiments (n > 3) performed in four consecutive passages of cells. *p < 0.05, one-way ANOVA with Dunnett's post hoc test. *significant over vehicle (DMSO) control.
B Experimental workflow for the establishment of a SARS-CoV-2-host interactome to identify potential drug targets and treatment. C Virus-host protein interaction networks were constructed by AP-MS (top) and BioID-MS (bottom). Only protein-protein interactions (PPIs) with a Bayesian false discovery rate (BFDR) ≤0.01 (assessed by SAINTexpress) are shown.
D. HuH7 cells were transfected with increasing amounts (0.03, 0.06, 0.125, 0.25, or 0.5 μg per M6 plate) of a plasmid expressing PKR (pPKR) and collected 48h post-transfection.
(C) Western blot analyses of the NKX2.2 expression levels in the WT and KO cortices at E13.5.
ChIP analysis of enrichment of ac-H3 (F) on promoters of Acta1 and Nppa in ventricular samples. n=3:3
(A) A predicted N-terminal extension (green) of the ROD β-propeller (red), slightly rotated from the view in B. The propeller proper begins with strand β7d and ends with strand β7c, which leads into the helical domain. The N-terminal extension augments the sixth and seventh blades with external β-strands (β6e and β7e). The position of T13 and S15 on the extension is shown.
Clonogenic assay on FaDu cells silenced for Sp1 (shSP1#3 or #5) or scramble sequence (shCTR). Left graph reports the number of colonies. On the right, WB analyses of the indicated protein expression in FaDu cells. Tubulin was used as loading control. Bottom panel, representative images of clones are shown. In the graph data represent the mean (±SD) of three independent experiments performed in duplicate and one-way ANOVA was used to verify the statistical significance. Sphere forming assay of the cells described in E. Left panel reports the mean number of spheres formed in first- and second-generations. Right graph reports the area of second-generation spheres. Data represent the mean (±SD) of three independent experiments performed in duplicate and one-way ANOVA was used to verify the statistical significance. Each dot represents one analyzed sphere. Bottom panels show representative images of the spheres.
D Immunoblotting of resected tumors (4 from each group) for the indicated proteins.
(D) Western blot of P. falciparum and P. knowlesi parasites showing that host RBC β-spectrin truncation during egress (red arrowhead) is inhibited by all three P. falciparum SERA6-maturation inhibitors (concentrations as in panel B), matching the egress inhibition in panel E (reproducible in 2 independent experiments).
E, F Immunoblotting using anti-P-Akt, anti-P-S6, anti-4EB-P1 or anti-GAPDH (loading control) antibody of protein extracts from CAF siCTR- or siSHP2- (E) or siPTPε- (F) transfected CAF treated or not with SOM230 for 30 min (representative of n = 3).
C Mice (n = 5 or 6 per group) were infected with viruses (1000 pfu) carrying PR8 PB1-F2 with the I68T, L69P, or I68T+L69P mutation. Body weight changes and survival rate were monitored daily. Detailed analyses on the virulence necessitated the monitoring of body weight below the standard level (80%) of mortality.
E Immunohistochemistry for SOX9 (brown) in control (Sox9fl/fl;RosaCre-) or Sox9-null (Sox9fl/fl;RosaCre+) mice following fibrosis. Bile ducts (bd) and hepatocytes (arrowheads) indicated. SOX9 is absent, even in bile ducts in Sox9-null liver. Size bar, 25 μm (A, B and E), 10 μm (C).
B. Western blot showing PRC1 protein from unlabeled cells, tubulin-GFP HeLa cell line and PRC1-GFP HeLa cell line. Detailed measurements are in Expanded View 4.
A, Transcriptional regulation of WOX5 (red) and PLT (green) expression by negative feedback regulation in the Arabidopsis RAM. WOX5 is expressed in the QC and indirectly promotes PLT expression (dashed arrow), whereas PLT expression is directly regulating WOX5 expression confining it to the QC position (solid barred lines).
Anti-GFP immunoblot analysis of streptavidin pulldown of the indicated biotin-conjugated H3 peptides following incubation with 293T-GFP-UBR7 cell lysate after mutation of residues highlighted in (C).
A) Representative western blot of autophagy markers at steady state in skeletal muscle. GAPDH was used as a loading control. B) Densitometric analysis of immunoblots similar to that shown in A (n=4/group). Black: untreated WT; grey: rapamycin-treated WT; red: untreated Cox15sm/sm; blue: rapamycin-treated Cox15sm/sm. Error bars represent SEM. The asterisks represent the significance levels calculated by one-way ANOVA with Tukey post-hoc multiple comparison test **p=0.0068 (LC3-II WT vs. Cox15sm/sm), **p=0.0063 (P62 WT vs. Cox15sm/sm) *p=0.025 (P62 Cox15sm/sm vs. Cox15sm/sm+rapamycin), **p=0.0098 (S6 WT vs. Cox15sm/sm), *p=0.034 (P-S6 WT vs WT+rapamycin), ***p=0.0002 (Cox15sm/sm vs. Cox15sm/sm+rapamycin), *p=0.014 (P-S6 WT vs. WT+rapamycin) ***p=0.0007 (Cox15sm/sm vs. Cox15sm/sm+rapamycin)
G SDS-PAGE analysis of purified WT WNV or WNV-poly(A) particles. Bands corresponding to E and Capsid proteins are denoted.
qRT-PCR analyses of Shp2mRNA levels in tumor tissues from MLN4924;Shp2fl/fl (n = 8), MLN4924;MMTV‐Cre;Shp2+/fl (n = 6), and MLN4924;MMTV‐Cre;Shp2fl/fl (n = 6) mice. Error bars represent upper and lower quartiles.
A-B. Involvement of filamentous cytoskeletal structures in BiFC foci polarization. Time-lapse micrographs of agarose-embedded cells expressing the Dsl1pVN•ε-COPVC BiFC combination at RT with fresh PM Glc+ura medium supply. (A) Upon disruption of actin filaments with 1,25 μM latrunculin A, COPI•Dsl BiFC signals dispersed. Arrowheads: polarized signal, asterisks: unpolarized localization. Fluorescent images are displayed with inverted brightness values. Scale bar 10 μm.
B, B1 Large axon terminals in the median eminence were immunopositive for secretagogin with silver‐intensified immunogold particles (open arrowheads) associated with axonal membrane and dense‐core vesicles (see Fig D for quantitative data). Solid arrowheads denote silver deposit particles proximal to the plasmalemma.
(c) RPE cells stably expressing GFP-myosin VI were processed for immunofluorescence microscopy to evaluate GFP-myosin VI and endogenous Tom1/Tom1L2 or Rab5 co-localization. The insets show higher magnifications of the areas outlined in the main panels. The arrows indicate areas of co-localization.
LPS-primed Abro1+/+ and Abro1−/− BMDMs were left untreated or treated with ATP. Immunoblot analysis of NLRP3 and ASC protein in cell lysates immunoprecipitated with anti-ASC antibody.
E. HDAC6+/+, HDAC6-/- MEFs, and HDAC6-/- MEFs reconstituted with HDAC6 wild-type were subjected to immunoprecipitation using an anti-RIG-I antibody and immunoblotted with anti-acetyl-lysine and anti-RIG-I antibodies. Acetyl-tubulin was measured in total lysates to determine HDAC6 deacetylase activity.
(F) c10orf76 binding induces differences in HDX throughout multiple domains of PI4KB. Regions of >7% difference in deuterium exchange in the presence of c10orf76 are colored onto the structure of PI4KB according to the legend (PDB: 4D0L). The N-lobe linker of the kinase domain is disordered in the structure and is represented by a dotted line.
Period analyses of WT and CKO PER2::LUC cells in the presence or absence of GSK3 inhibitor CHIR99021 (5 µM; CHIR).
Relative luciferase activities of the indicated promoter with/without E-box mutations in differentiating C2C12 cells by TGF-β treatment. n = 3, mean ± SD. *p < 0.05. **p < 0.01. n.s., not significant (unpaired two-tailed Student's t-test).
C) Overview of differentially expressed MXEs for the Embryonic Development, ENCODE, and Human Protein Atlas datasets.
(a) Principal Component Analysis (PCA) of microbiome samples from soil (S) and the endosphere of wheat roots without (R-mock) and with SA187 treatment (R-187). The first component (PC1) explains 67% of the total variance, while PC2 represents 10%.
Immunoblot analysis of EA.hy926 cells transiently transfected with control siRNA or KRIT1 siRNA. Left, evaluation of siRNA efficiency with antibody directed against KRIT1. Right, immunoblot for p62 and LC3 I/II. Actin was used as a loading marker. Quantification of total LC3 on actin is reported (*P = 0.02527). The results are representative of three independent experiments.
MIPs taken 20min apart showing examples of different kugel behaviour including shape changes (asterisk), expansion (circle), retraction (triangle), or little change (pentagon).
F. ELISA of serum IFN-β (left) and IL-6 (right) after poly(I:C) injection. HDAC6+/+ and HDAC6-/- mice were intravenously injected with poly(I:C). Serum was collected at the indicated time points and analyzed (200 μg/mouse; n = 3 per group; Student's t-test).
(E) HCT116 cells were treated with Embelin as indicated. LC3‐II accumulation was determined by western blot analysis with anti‐LC3 antibody. The data are representative of three biological replicates. The ratio of LCII/LC3I to actin is presented in Supplementary Figure S1B.
(E) Cumulative histograms based on fractional anisotropy (FA) values. (F) show the CC volume and the number of tensors relative to FA values. As FA values increased, the structural (CC volume and the number of tensors) differences increased between the control and Plcg1f/f;Nes-Cre mice. Depending on the number of tensors, the length in the CC appeared to be shortened (E, F). (n = 4; Shapiro-Wilk and t-test; *p < 0.05). Data are presented as the mean ± s.e.m.
B. HeLa cells stably expressing CHMP4B-eGFP were treated as in (A) and stained for p62. CHMP4B is recruited before p62 on the damaged lysosomes. The number of LAMP1-positive foci appears stable independently of LLOMe treatment Data information: Quantification graphs are normalized to the cell number where error bars represent 95% confidence intervals. Data were obtained from >150 cells per condition from three independent experiments
A. Expression pattern of Nmnat2 common and SATS isoforms in mESCs and somatic cells detected by qRT-PCR. Grey triangles represent independent biological replicates; red dots and blue intervals represent the mean values and (mean + SEM), respectively.
F. Time course of ear swelling elicited by topical application of 3% iodine in 0.15% Oxa-challenged mice, following intraperitoneal injection of RP67580 or vehicle, and bar graph highlighting the response at 24 h.
(B) From top to bottom: experimental design of 4D induction, fluorescence pictures of the SVZ of a 4D+ mouse and quantification of the proportion of RFP- (black) and RFP+ (red) progenitors among B1, C and A cells identified with markers as indicated.
A, B The period of DD locomotor rhythm of Nipped-A RNAi flies over-expressing tim (A) or carrying heterozygous tim01 mutation (B). Ptim is a tim cDNA construct driven by tim promoter. Data information: Error bars represent SEM. Digits on the bar are the number of flies tested. Percentage of rhythmicity is indicated above the bars. Statistical difference is measured using one-way ANOVA, P<0.001, Tukey's multiple comparison test, *P<0.05, **P<0.01, ***/###/+++/$$$P<0.001, * compared with the G4 control, # compared with the UAS control, + compared with the Nipped-A RNAi flies, $ compared with the over-expression or mutant flies. White bar indicates UAS or GAL4 controls. Dashed bar indicates flies with genetically manipulated core clock gene. Gray bar indicates flies with Nipped-A knocked down. Checked bar indicates flies with both genetically manipulated core clock gene and Nipped-A knocked down. G4, GAL4; U, UAS.
A. Histograms show the luc activity obtained from zebrafish embryos microinjected simultaneously with puORFchop-luc, phRG-TK, and each indicated plasmid followed by analysis of luc activity at 96 hpf. Embryos microinjected with pCS2 vector during normal condition (non-stress; blank column) served as a control group, while the microinjected embryos treated with 40℃ at 72 hpf for 1 hr were the heat-shocked group (solid column). The relative luc activity of microinjected embryos shown on the right was normalized by the amount of RNA in the injected embryos shown on the left. The relative luc activity was represented by the fold increase of Fluc/Rluc ratio over that obtained from the control group (this was normalized to 1). The luc activity mediated by the huORFchop transcript was measured by dual-luciferase assay, and its corresponding huORFchop-luc transcript was measured by RT-qPCR. Values representing the non-stress and stress condition were calculated from three independent experiments and presented as mean ± S.D. (n=3). Student's -t test was used to determine significant differences between each group (**: P<0.005, ***: P <0.001).
(B) Examples of ChIP-seq genome-browser views showing co-localization of uSTAT5 binding with ERG.
Targeted transduction of HSJD-GBM-001 and G26 cells, over a time course, by RGD4C/AAVP-Luc. Non-targeted/AAVP-Luc was used as negative control. Results represent RLU/1μg protein of triplicate wells. Data shown are representative of two experiments, n=3. Scale bar, 100 μm.
D Detail of a porcine ZP filament model, generated by superposition of homology models of adjacent ZP3 and ZP4 subunits (green and purple, respectively) onto the structure of UMODfl (semi-transparent grey), and projection of the residues of the interdomain linker of another ZP3 subunit onto the UMOD ZP-N/ZP-C linker (red). A cluster of experimentally verified glycosylation sites is shown, with side chain atoms depicted as spheres and boxes highlighting the proximity of ZP3 N271 and ZP4 N220, whose N-glycans mediate sperm binding in pig (Yonezawa, 2014). Note how these residues are also close to the conserved N- (dashed underline) and O- (site 1, single underline; site 2, double underline) glycosylation sites in the ZP3 interdomain linker. The invariant ZP3 Trp that follows site 1 is shown in stick representation at the interface with ZP4.
F Zoomed views focusing on interactions of the eIF3j C-terminus with the 40S. The loop following the third helix of eIF3j protomer 2 in a pocket formed by the 40S h33, h34 and eS10. Lys230 of eIF3j C-terminus (protomer 2) and Phe59 of eS10 are sandwiching the flipped-out G1435 base of h34 (upper left); Lys231 and Lys234 interact with h33 (U1266 and G1267) and h34 (G1438) (upper right); salt bridges between Arg220 and Glu70-Glu71 of eS10 further stabilize the loop (lower left). Following the loop, the eIF3j C-terminus bridges the 40S body and head in the latch and contacts are formed with h18 (via Lys236) and via hydrophobic interactions with uS3 (lower right). See Appendix Text 1 for more molecular details.
(A-C) SEC elution profiles and corresponding SDS PAGE gels of IFT172ΔC, IFT57/38, IFT80, IFT54/20 as well as mixtures of IFT172ΔC/57/38, IFT80/57/38 and IFT172ΔC/80/57/38 demonstrating that IFT57/38 associates with both IFT80 and IFT172ΔC. Only the central peak fraction from each SEC experiment is shown on the SDS PAGE gel. Note that the presence of both IFT172ΔC and IFT80 in a single peak in (c) (peak 3) is due to the fact that these two proteins elute in the same position, and not because they physically interact (see also Appendix Fig. S2B).
(G) Quantification of cells with GFP-LC3 puncta after RNAi treatment. Cells with more than three GFP-LC3 punctate dots were considered to be GFP-LC3-positive cells. Cells treated with the RNAi indicated here all showed a significant difference (P < 0.05) in the percentage of GFP-LC3-positive cells compared with the human (Hs) RNAi control. (Pten, P = 0.006; Tor, P = 0.034; Buffy, P = 0.005; debcl, P = 0.003; Bruce, P = 0.003; Dcp-1, P = 0.007; hid, P = 0.002; Ras, P = 0.006; and phl, P = 0.050). Results represent the mean value ± SD from three independent experiments.