EMBO/sd-character-level-ner huggingface dataset:
D, Immunoblot analysis of protein lysates from ARPE-19 cells overexpressing TFEB-FLAG-WT or TFEB-FLAG-C212A treated with 100 μM NaAsO2 for the indicated times and run under non-reducing conditions. E, Quantification of immunoblot data from TFEB-WT shown in (D). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, (**)p< 0.01, and (***)p<0.001 from three independent experiments. Some error bars are the same size or smaller than the data point symbols.
A-D. Correlative light and electron microscopy (CLEM) on A. untransfected (WT) HeLa cells harbouring endogenous CD63 labeled with GFP as compared to those ectopically expressing B. HA-RILP, C. HA-SKIP or D. both HA-RILP and HA-SKIP. Left panels: wide field fluorescence images of fixed cells showing endogenous GFP-CD63 (green) and SiR-lysosome-stained endosomes and/or lysosomes (red). Cell and nuclear boundaries are demarcated with solid and dashed lines respectively, zoom insets highlight regions selected for EM imaging. Middle panels: overview electron micrographs of perinuclear (PN) and peripheral (PP) cell regions selected for further analysis. Various endolysosomal subtypes are designated by arrowheads: MVBs (white), lysosomes (black), endolysosomes (yellow) and abnormal/enlarged endolysosomes (red). Right panels: graphical representations of endosomal distribution under the indicated conditions based on ultrastructural characterization.
(D-E) Developmental rates of (D) control (control-MO injected), Tet1-, Tet2- and Tet3-single KD and Tet-TKD embryos and (E) control (control-MO + Tet2 mRNA co-injected), Tet2-KD2 (Tet2 MO2 injected), Tet2-KD2 + GFP mRNA and Tet2-KD2 + Tet2 mRNA embryos until blastocyst stage. Indicated p-values were calculated using log rank (Mantel-Cox) test against (D) control embryos or (E) Tet2-KD2 embryos or as indicated (ns = non-significant, * = p < 0.05, **** = p < 0.0001; numbers of analyzed embryos for each condition are indicated in parenthesis).
C-F. NaCl dose-response analyses of Pbs2 phosphorylation. (C-D) KT005 (ste11Δ pbs2Δ) and (E-F) TM280 (ssk2/22Δ pbs2Δ) were transformed with YCplac22I'-Pbs2-HA, and were treated with the indicated concentrations of NaCl for 5 min. (C and E) S514 phosphorylation was analyzed using the Phos-tag band-shift assay. PC; Positive Control. (D and F) T518 phosphorylation was analyzed using anti-phospho-T518 immunoblotting. G-H. Average values of three independent experiments from (C-D) and (E-F), respectively, were plotted. AU, arbitrary unit. Data information: (C-F) Representative results from three independent experiments. (G-H) Error bars are SEM (n=3).
(F) Cross section from cartwheel-containing region of N. gruberi, (G) Zoomed side view of cartwheel from N. gruberi displaying the central hub (CH) and several cartwheel inner densities (CIDs), white arrow.
B. Scheme of the experimental approach to quantitatively capture gene expression changes during mESC differentiation towards the three germ layers at high temporal resolution.
(E, F) Time-lapse observation of the structure of NuMA and microtubules upon centrosome removal. Centrinone-treated HCT116 TetOsTIR1 NuMA-mAID-mClover-FLAG cells were observed with a 60× objective. Red and green represent NuMA and SiR-tubulin, respectively. Z-projections of 17 sections, every 1 μm. Scale bar, 5 μm. Time zero corresponds to NEBD. (F) An example of NuMA formed several asters (arrowheads) at the time of NEBD.
D. Recruitment assay using GFP-LukD (156.3 nM) and a dose response of unlabeled dominant-negative (DN) toxin subunits evaluated by flow cytometry on the surface of primary human PMNs. Emut1 = LukEmut1, Amut1 = HlgAmut1. Data is presented as total fluorescence emitted from GFP-positive PMNs. A representative of three independent experiments each done with hPMNs isolated from different donors is shown.
(B) Full-length EphA2 and serial truncations of EphA2 with deletion of various domains.
B Phos‐tag gel following complete conversion of Ub to phosphoUb, using GST‐PhPINK1. A Ser65 mutant Ub is not modified.
(a) HEK293 cells were transfected with TFEB and AR25Q or AR125Q, as indicated. Immunoprecipitation (IP) was performed with anti-TFEB antibody, followed by immunoblotting (IB) for TFEB and AR. Untransfected HEK293 cells served as a negative control, and no AR was observed with IgG antibody only (not shown).
(E) Tumor spheres formed by 22Rv1-P cells stably expressing ESM1-NLS with either β-catenin or NFκB shRNAs. Scale bar: 100 μm. Bars are the mean ± SD of three independent experiments. ## P< 0.01 when compared to vector-shScramble cells, ** P< 0.01 when compared to ESM1-NLS-shScramble cells by two-tailed Student's t test.
(E) Western blot analysis of FLIP, Caspase 8, and FADD recruitment to the TRAIL-R2 DISC in PC3 cells stably expressing an empty vector (EV) or flag tagged FLIP L (FL) treated with 2.5µM MS-275 for 48 hours, followed by a 90 minute DISC IP. * modified form of FLIP, potentially mono-ubiquitinated p43-FLIP(L).
B PBMC of a healthy donor were incubated with the indicated concentrations of bsAbs, and after three days cytokine release (B) were measured using a 3H-thymidine uptake- and a Legendplex assay, respectively. Mean values and standard deviations from triplicate samples.
K) Mitochondria mass from purified CD25+/CD54+ and CD25-/CD54- NK cells measured by flow cytometry (n=5, biological replicates).
View of nucleolus‐enriched Alu elements exemplified at two genomic loci. Mapping of normalized reads from total (red), nucleoplasmic (orange) and nucleolar (blue) RNA to intron #5 of DCP1A and intron #22 of UBE4B.
(C) Intestine tissues from 8-week-old Rest+/+ and Rest-/- mice (n=6 per group) were collected for immunohistochemistry staining. Typical images were shown in left panel and statistical ISC numbers from 120 crypts (20 crypts per mouse) were shown in right panel as means± S.D. ** P < 0.01 by two-tailed Student's t-test. Scale bar, 50 μm.
H. Immunofluorescence of H3P and cTnI and quantification of the heart tissue sections from control or doxycycline injected mice.
B, After 24 h LDL depletion in 5 % LPDS, cells were loaded with 50 μg/ml LDL for the indicated times. Plasma membrane cholesterol was labeled with D4H and imaged with widefield epifluorescence microscopy. C, Analysis of B. Mean ± SD, n = 15 fields of cells as in B pooled from 2 independent experiments.
H. Immunoblotting for phosphorylated ERK1/2 (P-ERK1/2) and total ERK1/2 following prazosin treatment for 30 min. Prazosin induces ERK1/2 phosphorylation in GICs.
(H) Model of the mechanism of action of Gas6/AXL in cancer cells (left panel) and modulation by OPCML and PTPRG (right panel)
Figure 7. IL-17RB expression is correlated with the elevated prevalence of regulatory T cells in the tumor-draining lymph nodes of breast cancer patients. (A and C) Representative IHC images of IL-17RB (A) and Foxp3 (C) were taken in breast specimens from primary tumors and their paired LN metastasis specimens. Scale bar: 10 μm.
(D). Immunoblot analysis of mitochondrialDRP1 oligomers in control and subject fibroblasts, overexpressing an EGFP-tagged version of wild-type SLC25A46. Crude mitochondria were treated or not with the BMH cross-linker. SDHA was used as a loading control. Monomers (asterisk), dimers (arrowhead) and oligomers of DRP1 are indicated.
F. Percent of left lung lobe parenchyma infiltrated by white blood cells was quantitated by Image J analysis.
C Venn diagram showing the number of H3K122succ peaks detected overlapping with H3K4me3 peaks and H3K9ac peaks.
Effects of CLIP170 on fusion of viral particles by endocytosis. CHME3 cells either treated with control or CLIP170 siRNAs (A) were infected with Mock or HIV-1-VSV-BlaM-Vpr. Representative FACS analysis of cells (n = 3) showing ∼ 9% (Control siRNA) and 3% (CLIP170 siRNA) shift (A) from green (uncleaved CCF2) to blue (cleaved CCF2). % infected cells is indicated at the top left corner of each plot and quantification from independent repeats is presented as bar graphs at the right side of each FACS plot.
Quantification of THP-1 ∆GBP1+Tet-mCH-GBP1 cells stably expressing YFP-CASP4C258S and infected with STm for 2 h. Graph shows percentage of STm vacuoles targeted by GBP1, CASP4 or both from n = 3 experiments. n.d. not detected.
(d) Self-association pattern of Beclin 1 in vivo. Both GFP- and AsRed-tagged full-length Beclin 1 are co-transfected into HEK 293T cells and their self-association is probed by co-IP using GFP antibody for immunoprecipitation, followed by western blot with anti-AsRed antibody.
(a) Comparison of AMP/ATP ratios between PCF trypanosomes grown in the absence (−Tet) or presence (+Tet) of tetracycline to induce RNAi, and with or without glucose (Gl), expressed as fold increase (means±s.d., n=3; *P0.05; **P0.01, Student's t-test).
2D-tSNE representation of all single cells included in the study (n = 1,247) depicting the separation of microglia isolated from LPS-injected mice (770 cells in red) and steady state (477 cells in blue) in 2 main clusters.
D) with the abundance of the species in the gut microbiome Asterisks indicate statistical significance based on Spearman correlation analysis. p < 0.05; Cor.Coeff: Correlation coefficient; Log2FC: log2(fold change).
A We speculate that under normal conditions, ERF8 represses the other network genes. Upon mild osmotic stress (indicated with a red arrow), some genes of the network can be phosphorylated (PTM), a hypothesis based on literature and the abundance under normal conditions. Subsequently, the expression of the network genes increases during four groups of transcriptional induction. The direct transcriptional regulations of the core network members (ERF6, ERF8, ERF9, ERF59 and ERF98) are depicted with green and red arrows, representing activation and repression respectively. The regulatory connections of the core network members that were identified when evaluating the effect of two TFs together on a shared target gene, are depicted with dashed green and red arrows, representing activation and repression respectively. The latter regulatory interactions occur in the presence of a necessary transcriptional partner. The color of the nodes represents the strength of the induction. FC = Fold change. B Schematic representation of the feed forward loop the network is composed of. Upon input, such as mild osmotic stress, activators of the core network are induced (A) and activates downstream TFs (C). These TFs or another unknown component could induce the expression of the repressors of the core network (B), which in turn leads to the repression of the downstream TFs (C), restoring the original state of the network.
(G) Torin 1 increases binding of TFEB to mTORC1. HEK‐293T cells that express TFEB-3 × FLAG along with HAGST‐Rap2A or HAGST‐RagsDN were treated with vehicle or with Torin1, lysed and subjected to FLAG immunoprecipitation followed by immunoblotting for mTOR and raptor. FLAG-Metap2 served as negative control.
(b) Cytokine array analysis of conditioned medium from U2OS cells subjected to glutamine deprivation in the presence of 1 μM BrefA or 100 nM WYE-125132.
Cells were treated as in F and whole cell protein lysates were collected and analyzed by capillary electrophoresis for the expression of ATF4 and Vinculin (G) and by the standard western blot for the expression of Noxa and Actin.
G Flow cytometry analysis of splenic CD8+ T cells in WT and KO mice with or without anti-PD-1 treatment after tumor challenge.
(d) The 8-hydroxy-2′-deoxyguanosine (8-OHdG), popular marker for oxidative DNA damage, in mitochondria DNA from DOX-treated hearts. Results are shown from 6-8 hearts analysed in duplicate.
E Immunoblotting of ATF4 and SLC1A5 in melanoma cell lines mock- or si-ATF4-treated for 24 hr and then incubated with LDHAi for 48 hr. Data information: Statistical analysis was performed by ordinary one-way Anova for the comparison of more than two groups.
(A) Western Blot detection of ATM pS1981, SMCHD1, TRF2 and hnRNPA1 loading control in wild-type and two SMCHD1 knockout (KO1 and KO2) HeLa cells transfected with shTRF2 plasmid or EV control.
(Top) Lack of editing in Filamin A pre-mRNA produces a Filamin A isoform that only encodes a glutamine residue (Q) at position 2431. This leads to mislocalization of p190RhoGAP, misregulation of PLC and ROCK signaling, increased MLC phosphorylation, aortic hypercontraction, thickening of the smooth muscle layer and increased perivascular collagen deposition. Loss of FLNA editing leads to persistently elevated diastolic blood pressure, left ventricular hypertrophy and cardiac remodelling. (Center) Filamin A pre-mRNA editing by ADAR2 triggers a Q to R codon exchange at the end of exon 42. (Bottom) Edited FLNA helps localize p190RhoGAP to the cellular cortex where it can inhibit RhoA and regulate the activity of key smooth muscle contraction regulators like PLC and ROCK machinery. This can maintain normal aortic function and normal diastolic blood pressure to preserve normal heart morphology.
HeLa cells were transfected with plasmids to visualize ER (in green) and mitochondria (in red), and then were treated with ApoE3- and ApoE4-ACM for 24 hours. Shown is a box-and-whisker plot comparing co-localization in ApoE3 ACM (n = 15 images analyzed [gray circles, average of co-localization in ~5 cells in each image field]) compared to that in ApoE4 ACM (n = 18 images).
An equivalent experiment was setup as in (B) but the conjugated cells were continuously illuminated throughout dimerizer addition to disrupt OptoCAR signaling. Bounded line shows mean ± SEM (n=3).
(D) atg1Δatg13Δ cells expressing GFP‐tagged wild‐type Atg13, and either wild‐type Atg1 or the Atg1‐VE mutant (left panel) and wild‐type cells containing Atg13‐GFP and either wild‐type Atg13 or Atg13‐FV were grown to mid log phase followed by starvation for 6 h in SD‐N media. GFP cleavage was analysed by western blotting. Figure source data can be found with the Supplementary data.
Immunisation with VLPs induces germinal centre formation. Germinal centre B cells were identified by flow cytometry as B220+IgD-CD95+GL7+ cells. The gating strategy is shown in (D)
Waveforms of ABRs are compared in WT, untreated Kcnq1−/−, and treated Kcnq1−/− mice, as labeled above data traces. Series of averaged ABR data traces were evoked from tone-burst sounds with intensities ranging from 20 to 90 dB SPL.
(C-F) Tissue levels of soluble and aggregated mutant ataxin-3 were measured in cerebellum (C, E) and frontal lobe (D, F).
A Phb2-deficient mouse podocytes showed higher levels of phosphorylated S6 ribosomal protein than control podocytes, which could be blocked by treatment with rapamycin (n = 5, bars represent mean ± SEM, Student's t-test, **P = 0.0036). pS6RP, phosphorylated S6 ribosomal protein; S6RP, S6 ribosomal protein.
). Immunohistochemistry staining for monocytes/macrophages (F4/80) and neutrophils (Ly6G; arrowheads) in brain tumors derived from WT or Ang-2 overexpressing mice (E), Quantitative analysis of C (F) (WT n=14; Ang-2 DT n=10)
(D) SDS-PAGE of cell lysates derived from 293T cells transfected in (B) followed by immunoblotting with anti-FLAG M2 and anti-actin antibodies.
J. MM029 (left) and MM099 (right) were pre-treated with either siCTL or siGATA6 for 48h and treated with increasing concentrations of THZ1 for 72h. Mean growth is shown relative to vehicle (DMSO)-treated cells. Data information: data are presented as mean values + SD for six replicates (n=6). The p-value (Student's t-test) is indicated, **<0.01, ***<0.005 and ns=non-significant
Quantification of miRNA and pre-miRNA expression levels using the 2-ΔCt method and U6 as normalizer, upon Sema3A (B) stimulation. Data are normalized to PBS control. Each data point represents a single RT-qPCR reaction. n=3 (B) independent experiments. Values are mean ± SEM. Abbreviations: ns, not significant.
Frequency of WT donor (CD45.1), DKO donor (CD45.2) and recipient (CD45.1/2) cells among the BM LSK cells, splenic monocytes and alveolar macrophages of the recipients that were irradiated with lung shielding, treated with one dose of busulfan as described in Materials and Methods, transferred with a 1:1 mixture of lineage-depleted BM cells from WT and DKO mice and analyzed ≥ 8 weeks after transfer. Representative flow cytometry results for alveolar macrophages (left) and quantification (right) is shown.
G. Schematic representation illustrating that in contrast to ARMC8α, incorporation of ARMC8β prevents hGID activity towards GID4 substrates.
(C) HIV-1-infected (strain NLAD8) CD4 T cells were cultured with the indicated antibody and NHS. After 24h, the surface levels of C3 and the frequency of infected cells (Gag+) were determined by flow cytometry. One representative experiment is shown.
Quantitation of the BODIPY signal shown as normalised LD size (normalised to DMSO) for cells corresponding to each treatment and donor mentioned =15 cells for each treatment. The box end points represent the upper (75%) and lower (25%) quartiles, the whiskers define the maximum 95th percentile and minimum 5th percentile values respectively, the central band is the median, the square is the mean, and the diamond an outlier. Data were analysed using the Kruskal Wallis test followed by Dunn's test for post hoc analysis for significance due to unequal sample sizes. ***p<0.001.
effect of in vivo TGFBRi and MMPi treatment of mice for two weeks on colon tumor formation. Apcfl/fl-Cdx2CreERT2 mice were treated with Tamoxifen and, one day post treatment, injected with either MMPi (n=10) or the TGFBRi SB431542 (n=10) at 40mg/kg for two weeks with five injections per week. Vehicle treated mice: n=11. Average tumor size per mouse was assessed during colon resection one week after the end of treatments (E).
c. Representative dot plots of the frequency of IFN+Foxp3+ Tregs in the co-cultures in a at a 1:2 Treg:Tresp ratio after adding GolgiStop for the last 6 hours.
Intracellular ROS levels detected in H1299 cells transfected with the indicated shRNA in normal medium or after 12hr glucose starvation (Glu-)
G. Determination of viral progeny. Supernatants of SARS-CoV-2 exposed Vero cells, and brain organoids were analyzed for viral RNA assessed by qRT-PCR. While an increase in viral RNA was detected in the supernatants of Vero cells, no apparent increase was identified in brain organoid supernatants. Data are obtained from five technical replicates from four (n=4) independent batches of organoids. Data presented as mean + SEM.
(B) A summary visualization of the enrichment of the top 10% MTA-predicted targets from each dataset for the antiviral hits (i.e. those whose KO inhibits SARS-CoV-2 infection) identified in the two published CRISPR-Cas9 screens (Wei et al. 2021 and Daniloski et al. 2021; Methods). Y-axis represents the odds ratio of enrichment, the dot color corresponds to the negative log10 adjusted one-sided P value from Fisher's exact tests, and dot size corresponds to the number of enriched target reactions. The datasets are ordered by P values, the first 8 datasets have FDR<0.1.
The heatmap representation of gene expression pattern in miRNA mimic-transfected Dgcr8-/- ESCs.
(D) Similar BRET experiment showing the Rab7-pool of PI4P both in control HEK293-AT1 cells and in two clones of cells with PI4K2A deletion (#20 and #26). Means ± S.E.M. are shown from three experiments performed in triplicates.
(A) Cells transduced with the indicated shRNAs were fixed and localization of α-catenin and β-catenin was determined by imaging with a confocal microscope after staining with antibodies against α-catenin and β-catenin. Representative images from three independent experiments are shown. Scale bars, 10 μm. (B) Quantification of plasma membrane (PM) to cytoplasmic ratio of α-catenin intensity per each cell from figure 4A was plotted (n=40 cells). Error bars indicate SD. ***p < 0.001, (one-way ANOVA, post hoc test: Bonferroni's multiple comparison test).
E. Representative docked vesicles (arrows) in a DRG neuron. Quantitative data are on the right. Scale bar, 200 nm.
Effects of VPS18 depletion on the recruitment of TBC1D15 to the SKIP complex. D. Representative confocal images of fixed HeLa cells harbouring endogenous GFP-TBC1D15 (G-e1D15, green), transfected with either control siRNA (siC) or oligo pool targeting VPS18 (siVPS18) and ectopically expressing HA-SKIP (red), immunolabeled against endogenous VPS18 (eVPS18, blue) and HA, scale bar: 10µm. Cell and nuclear boundaries are demarcated with solid and dashed lines respectively, zoom insets (3.5x) highlight select peripheral (PP) and perinuclear (PN) cell regions. E. Colocalization (Mander's overlap) between SKIP and endogenous TBC1D15 as a function of VPS18 depletion, n siC=9, n siVPS18=11 images (2≥ cells per image) analysed from 2 independent experiments.
c Morris watermaze. In the standard Morris watermaze task (left graph) the path length to reach the hidden platform decreased across training blocks for Grin2a+/+ and Grin2a+/S mice but not for Grin2aS/S mice. Two probe trials were conducted after 12 and 24 training trials (P1 and P2, respectively) during which the platform was removed from the pool. At both P1 and P2 (right bar graphs) the Grin2aS/S mice failed to search for the platform in the target quadrant. AdjL adjacent left, Target fixed location of the hidden escape platform during acquisition, AdjR adjacent right, Opp opposite of target quadrant. Dashed lines indicate chance levels.
E. Ribbon diagram of the superposition of DBD C-term with the highly similar DNA-binding domains of telomeric protein TRF1 (PDB ID 1W0T) (Court et al., 2005) and the bacterial cell-cycle regulator GcrA (PDB ID 5Z7I) in complex with their DNA substrates. A similar DNA-binding mode by DBD would cause a steric overlap of helix H4 with the phosphate backbone of dsDNA. DBD C-term is in light blue, TRF1 and GcrA proteins in brown and their DNA substrates in khaki.
D. UMAP plots with color-coded intestinal stem cells identified by transcriptional signatures of intestinal stem cells or by expression of Lgr5 or Gkn3 genes.
Schematic of STIM1 domain architecture is shown, with the AMPK consensus motifs for S257 and S521
RT-qPCRs to detect polyA+ histone mRNA levels (H) in Flag-Cntl (eIF4A3) or siRNA-resistant Flag-ALYREF stable expression cells treated with Cntl or ALYREF siRNA. The bars show the relative abundance of histone mRNAs to the GAPDH mRNA. Data information: Error bars represent standard deviations from biological repeats (n=3). Statistical analysis was performed using Student's t-test. *P < 0.05, **P < 0.01, ***P< 0.001.
Representative IAHCs of IgG and αDll4 treated embryos. Kit is shown in white in the image, in black in the label. Multistack reconstruction of confocal images. Note the increased IAHC size in the right panel. Scale bars: 50μm (overview), and 10μm (magnification).
E) The number of cells that have invaded from the top to the bottom chamber of the Boyden chamber was quantified by automated segmentation in non-treated (NT), in ROCK inhibited conditions (Y27632, H1152) or in cells transfected with siRNA against ROCK1 and ROCK2 (siROCK1+2). The number of cells that have invaded was normalized to control conditions and reported as the average invasion rate from at least three independent experiments (Means ± SEM). P values were calculated using unpaired t-test (****p<0.0001, **p<0.01).
Pull-down of His-Rab22AWT using HeL lysate the bead bound His-Rab22A/His-KIF13A domains were shown as coomassie stained gel separatel
C) Effect of AST and LUF7346 in the presence of AST on APD90, APD70, and APD50 in WT- (grey), LQT1R594Q- (red) and JLNSR594Q-CMs (black) paced at 1 Hz. * = p<0.05 vs respective baseline; ° = p<0.05 vs AST. N: 11-16.
(B) Mean fluorescence intensity quantification of whole cell phalloidin signal in MDA-MB-231 cells overexpressing PTPRN2, PLCβ1, or control vector. N = 40 cells/group.
Cryo-EM density is shown at a low threshold to highlight the almost continuous density connecting the Rab1/Ypt1 nucleotide-binding domain (NBD) to its HVD (hypervariable domain). Inset is a closeup of the density corresponding to the Rab1/Ypt1 HVD region. Density is colored by subunit as in Figure 2, Rab1/Ypt1 is yellow. Density is depicted as it is likely oriented at the surface of organelle membranes.
3 mo old C57BL/6J mice after EdU application (experimental setting see in E) for 4 weeks in drinking water