EMBO/sd-character-level-ner huggingface dataset:
(D, E) The indicated MEFs were treated with etoposide (10 µM) for 6 hr and then immunostained with an anti-LC3 antibody (green). Representative images are shown in (D). LC3 puncta are observed in etoposide-treated PPM1D+/+MEFs. (E) The proportion of cells with LC3 puncta was calculated (n > 100 cells in each experiment). Data are shown as the mean  ±  SD (n = 3 experiments). *p < 0.05, **p < 0.01.
(C, D) Impairment of Sp1-mediated transcription by Htt-Q97 is modified by HOIP. (C) Shown is the relative luciferase activity in HEK293T cells expressing Htt-Q97-HA, a Sp1 luciferase reporter construct and HOIP cDNA. Data are displayed as mean ± SD and were analyzed by Kruskal-Wallis test followed by Dunn's Multiple Comparison Test, n = 9-11. (D) Shown is the relative luciferase activity in HEK293T cells expressing Htt-Q97-HA, a Sp1 luciferase reporter construct and HOIP siRNA. Data are displayed as mean ± SD and were analyzed by one-way ANOVA followed by Tukey's Multiple Comparison Test, n = 9. *p ≤ 0.05, ***p ≤ 0.001.
(E). Serum E level in AKG treated male mice 3 hrs after i.p. injection (n = 8 per group).
B. Analysis of collagen degradation on cross-bow-shaped micropatterns coated with the DQ collagen substrate. Mean intensity projection images of the DQ collagen fluorescence (top panel; cell area is outlined by dashed line; original integer range 0 − 255) and single confocal images of the fibrinogen-Alexa647-marked micropatterns (bottom panel) are shown (n ≥ 53 from four independent experiments). Scale bars represent 10 μm.
Two different AT1aR mutants were generated by site-directed mutagenesis to mimic the spatial distribution of Ser/Thr as in V2R with respect to double-Thr cluster. The red dotted box highlights the resulting sequence upon either the delition of Leu337 or Leu337Thr mutation.
(C) Quantification of cyclin B specific substrates by phospho-proteomics. Phosphorylation sites deemed significantly changing are shown in red
A graphic interpretation of the positioning of PEs at the bottom of LTR-valleys in panel (A).
(B and C) Inguinal lymph node-injected BALB/c mice were sacrificed at the indicated week after initial injection. Total cells isolated from inguinal lymph node tissues were transwell co-cultured with 4T1 cells. Inguinal lymph node tissues came from un-injection BALB/c mice as control. After 5-day co-culture, 4T1 cells at lower well were examined in the RT-qPCR (B) analyses of Il-17rb expression. Gapdh was used as an internal control or as a loading control.
(A) FXR1 expression in PBMC of individuals carrying the rs2601 GG (low risk; N=16) versus AA genotype (high risk; N=27).
Carbonate-washed total membrane fractions from wild type and SPPL2c-/- testis (n=5) were subjected to a label-free quantitative proteome analysis. For each quantified protein, a log2 intensity ratio between SPPL2c-deficient and wild type samples was calculated. Negative values indicate a depletion and positive values an enrichment in SPPL2c-/- samples. In the depicted volcano plot the negative log10 of the p value (y axis) is plotted versus the log2 intensity ratio of SPPL2c-/- versus wild type samples (x axis). All proteins above the significance level of p<0.01 (unpaired Student's t test) are coloured in red. The hyperbolic curve indicates the threshold for a permutation-based FDR correction for multiple hypotheses with p = 0.05 and s0 = 0.1.
Heatmaps showing changes in gene expression levels upon senescence (log2FC) of genes encoding selected chromatin-associated factors. For each gene shown, statistically significant expression changes were recorded in at least one cell type.
C. Leukocidin octamer highlighting stem domain motifs deleted from LukE and LukD (LukEmut1, LukEmut2, and LukDmut), as shown from the side view (left), and top view (right) of the pore. The monomeric structures of LukE (3ROH) and LukD (4Q7G) were obtained from the Protein Data Bank, and superimposed and colored on the structure of the HlgA-HlgB octameric pore (3BO7) using PyMOL. The location of where the deleted residues would be in the pore structure is shown from a side view on the left, and the location within an oligomeric pre-pore structure are shown from a top view on the right.
Urine glucose levels of cInsulin immunized (red, n=3 for d0, d26; n=5 for d14) and CI mice (grey, n=3) were monitored at indicated days post immunization. Upper panel showing visualization of glucose standard (top lane) and representative images of tested animals (middle and bottom lanes), respectively. Dots represent individual mice, mean ± SD. Statistical significance was calculated by using repeated measure ANOVA test, *P < 0.05.
(N) Diagram showing the experimental procedure to obtain media from cerebellar organotypic cultures dissected from the brain of wt and ASMko mice orally treated or not with PLX.
Images and comparisons of absolute and relative lacteal lengths in duodenum (DD), jejunum (JJ) and ileum (IL) of WT and VR3iΔLEC mice. Note that, in addition to the alterations in the lacteal lengths in VR3iΔLEC mice, diameters of the lacteal also shrank by VEGFR3 ablation. Each dot indicates mean value of 5-10 villi in a mouse (n = 6 mice/group). Scale bars, 100 μm.
B Percentage of full-length Cdc24 and cdc24 ph domain mutant (cdc24 ph) associated with liposomes containing 75% PC 20% PS and 5% PI4P.
E. Mitochondrialiron in humancardiac tissue sample from non-failing hearts and from hearts with ischemic cardiomyopathy (ISCM). * P=0.041 with two-tailed unpaired T-test. N=4 independent samples in each group.F. Cytosoliciron in humancardiac tissue sample from non-failing hearts and from hearts with ischemic cardiomyopathy (ISCM). Two-tailed unpaired T-test was performed. N=4 independent samples in each group. All data are expressed as mean ± SEM.
C. Quantification of MT-flux upon shRNA-mediated depletion of KIF4A alone or in combination with conditional expression of the RNAi-resistant mCherry-KIF4A variants. The bars in graph represent mean ± SD. N (number of cells, number of independent experiments): uninduced shKIF4A (48, 4), shKIF4A (60, 4), uninduced KIF4A WT (36, 5), shKIF4A + KIF4A WT (43, 5), uninduced KIF4A K94A (42, 7), shKIF4A + KIF4A K94A (38, 6), uninduced KIF4A ΔZip1 (32, 4), shKIF4A + ΔZip1 (30, 4).
5E) HeLa cells and HeLa cells stably overexpressing LC3 were transfected with scrambled siRNA or PINK1 siRNA. After 48 h cells were starved in HBSS for additional 24 h and afterwards DEVD cleavage as parameter for caspase-3 activity was analyzed and normalized to 1,000,000 cells. PINK1 knockdown increased caspase-3 activity significantly, which was prevented by LC3 overexpression; n = 3; p<0.05.
E-F: Endogenous MOSPD2 (green) staining in HeLa cells expressing Flag-STARD3NL (E (anti-Flag; magenta) Data information: (A, D-F): The subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. The Coloc panel displays a colocalization mask on which pixels where the green and the magenta channels co-localize are shown in white. Right: Linescan analyses with fluorescence intensities of the green and magenta channels along the white arrow shown on the subpanel Overlay. Black rectangles indicate the positions of late endosomes (E). Scale bars: 10 µm
(B) Bar chart visualising the top kinases recurrently associated (absolute effect-size>0) with drug resistance (top seven bars) and sensitivity (bottom seven bars) in the GDSC and CCLE drug sensitivity datasets.
CDK5 mediated cilia length decrease can be completely blocked by expression of the phospho-mimetic DBN1S142D in hCDC14APD cells. Three independent experiments; 150 cells per experiment and condition. Mean ± SD; ns = not significant; **** P≤ 0.0001.
The ERMIT assay relies on the signaling properties of IRE1, one of the three ER stress sensors and reports for a dimerization event occurring in the lumen of the ER. The assay can be applied to heterodimerization or homodimerization events. Western blot showing the expression of AGR2 dimers (D) and monomers (M) in HEK293T subjected to DSP-mediated cross-linking and that were previously transfected with either a control siRNA (siCTL) or a siRNA targeting AGR2 (siAGR2) for 24 h (C) Reduced or non-reduced samples were resolved by SDS-PAGE and immunoblotted using anti-AGR2, anti-ERK1 antibodies (for loading control).
(C) PBneutrophil production and nuclear segmentation were assessed in control and Am80-GCSF mice surviving on day 13 (section i).
(I) Intracellular ROS levels in colon tissues from wild-type (WT) or Otud1─/─ mice treated with AOM/DSS (n = 4 biological replicates, mean ± s.e.m., **P = 0.0072, two-tailed unpaired Student's t-test) were detected by DCFDA staining. MFI, mean fluorescence intensity.
(A) ASPC-1 cell without treatment (800 fold). (B) ASPC-1 cell undergoing apoptosis upon 160 µg/ml OMP after 24 hours (800 fold). Vacuolisation of the cytoplasma and condensation of the nucleus are visible. (C) Phagophores and autophagosomes in a segment of an ASPC-1 cell treated with omeprazole 80 µg/ml for 24 hours (2800fold enlargement). The phagophores are characterised by a cup-like shape (white arrows). Autophagosomes are closed particles, the number of which is increased in treated cells (black arrows). (D) Early phagophores and autophagosomes are also found in MiaPaCa-2 cells treated with OMP 80 µg/ml after 24 hours in a perinuclear region containing lysosomes and the Golgi complex. In contrast to ASPC-1 cells, early signs of apoptosis such as vacuolization, are also present.
(C). Flow cytometric analysis (FACS) of surface ICAM-1 (α human mAb, see methods) expression in control non-treated PLB985, ICAM-1kd and EPCRkd PLB985 cells.
(A) Schematic of pulses of p53 expression in response to DNA double strand breaks. p53 dynamics are differentially decoded to induce target genes with diverse downstream functions, including mediating cell cycle arrest and p53 regulation.
H. Percentages of apoptotic cells (AnnexinV+/7-AAD-) in donor-derived LSK cells in control and cKO chimera mouse BM at 24 weeks after transplant and 16 weeks after pIpC injection. Data are compiled from two independent experiments (total of n=7-8 mice per genotype). Data information: Data presented as mean ± SEM. *p<0.05, **p<0.005 and ***p<0.001 by two-tailed Student's unpaired t-test analysis.
(G-I) Confocal images of Cnga3-/- treated (G), wildtype (H) and Cnga3-/- untreated (I) retinal sections stained for Cnga3. Note some non-specific signal detected in INL and adjacent synaptic layer. Scale bar: 50 μm. OS, outer segment layer; ONL, outer nuclear layer; INL, inner nuclear layer.
f alpha-KG does not further extend the lifespan of ogdh-1 RNAi worms, mveh = 21.2 (n = 98), mα-KG = 21.1 (n = 100), P = 0.65 (log-rank test).
For each time point of the IFN-γ treatment, proteasome complexes were purified and analyzed by LC-MS/MS. Proteasome complexes dynamics was measured by label-free quantitative proteomics. The normalized abundance index of each protein or protein complex obtained at each time point was compared to the one obtained at the 0 h time point to obtain a regulator relative normalized PAI (n = 3).
Body weight change in the mice in (O,P) over the 24 h after the first treatment (Q) or after the 3 week course of treatment (R); n=9.
B Ub chain interaction of the LUBEL-NZF and UBA2 domains. Immobilized GST-NZF or GST-UBA2 was incubated with Lys (K) 48-, K 63-linked, or linear di-Ub (Ub2) chains and bound Ub chains were subjected to immunoblotting using anti-Ub antibody or a linear linkage-specific Ub antibody (anti-linear Ub antibody). Loading of GST-fusion proteins was visualized by Ponceau S staining.
E. HeLa cells were co-transfected with either Flag-c-Myc or control vector plus the indicated reporter constructs and Renilla luciferase plasmid. Twenty-four hours after transfection, reporter activity was measured and plotted after normalizing with respect to Renilla luciferase activity. Data shown are mean ± SD (n = 3; **P < 0.01, two-tailed t-test).
qRT-PCR analyses of normalized miR-9 expression, expressed as fold over the untreated condition, in control (shCTR) and antimiR-9 FaDu cells treated with Gefitinib (GEFI) for up to four hours. Data represent the mean (±SD) of three independent experiments performed in duplicate and two-way ANOVA test was used to verify the statistical significance.
G) Influence of Palbociclib on MCF-7 after 80 hours. Data was chosen from one representative time point during the exponential growth phase. Statistical significance was analyzed with a one-way ANOVA with Dunnett's multiple comparisons test and compared to the vehicle substance dimethyl sulfoxide (DMSO)-. The mean values from three technical triplicates (one experiment) with standard deviation are shown.
F. Co-immunoprecipitation between FLAG-γ-COP and TRAPPII in control (siFFL) and Rab18 (si-Rab18) siRNA depleted cells. TRAPPII was precipitated with Myc-TRAPPC10 and the presence of FLAG-γ-COP was determined by immunoblotting. The molecular weight of endogenous Rab18 overlaps with IgG light chain, and therefore, the amount of endogenous Rab18 precipitated with TRAPPII cannot be determined in this experiment.
Membrane overlay assay of Taxol-stabilized MTs using the apical junctional complex (AJC)-enriched fraction. The AJC-enriched fraction was further fractionated using ultra-centrifugation and ion-exchange chromatography with an SP-Sepharose and a Q-Sepharose column (see also Fig EV1C). The bands (marked by an arrowhead and red dots) correspond to LUZP1. 1: Ho, homogenate; 2: Sup, supernatant; 3: Pel, pellet; 4: SP-Pass, passing material through an SP-Sepharose column; 5: SP-E3(2), 2nd elution fraction through an SP-Sepharose column; 6: Q-Pass, passing material through a Q-Sepharose column; 7 to 9: Q-E3(1) to Q-E3(3), 1st to 3rd elution fractions through a Q-Sepharose column.
(J) Immunofluorescence staining against Iba-1 (green) and CathB (red) in lobes of the Cb sensitive or resistant to Purkinje cell death. Asterisks indicate Purkinje cells in the resistant lobes. Scale bar, 50 μm.
(B) Oxygen consumption rates (OCR) quantification show a decreased OCR in old whole heads compared to young flies. Data were normalized to 1-week-old flies (young flies). and N=14 young versus N=12 old. (C) At midlife, flies show a reduced physical activity compared to young flies. (D) Oxygen consumption rates (OCR) quantification show an increased OCR in midlife whole heads compared to young flies. Data were normalized to 1-week-old flies (young flies). N=21 young versus N=24 midlife.
(A) Schematic describing KP and KPΔNF1 cell transplants into immunocompromised mice.
Heatmap showing unsupervised hierarchical clustering of human differentially expressed genes between human scr-shRNA and sh-TFEB ECs. Red: up-regulated genes; green: down-regulated genes.
A. Extracts of RPE-1 cells treated for 72 hours with either non-targeting or Fbxw5-directed siRNAs (mix of siFw5-1&3) either grown asynchronously or arrested using 2 mM HU (S-phase), 75 ng/ml nocodazole (M), 10 µM RO-3306 (G2) for 18 hours or via serum-starvation for 24 hours (G0). Indicated antibodies were used for detection. Asterisk indicates an unspecific band detected by the Fbxw5 antibody. Uba2 served as a loading control (same for all following blots).
Schematics (F) of the experiment to test the function of Ifnar1 in bone marrow (BM) derived cells. WT C57BL/6 mice were transplanted with either Ifnar1+/+ or Ifnar1-/- BM cells. Recipient mice were allowed to recover followed by the injection of WT MC38 cells, before treatment with 5-FU or PBS.
(a,b) BJAB and Jurkat cells stably expressing mCherry-GFP-LC3 were flow sorted for high and low autophagic flux, treated with Fas ligand (1.5 ng ml−1) for 24 h and viability was assessed by MTS (a; percentage of no ligand control, mean ± s.e.m., n = 3 wells, *P = 0.0046). (b) Immunoblots following sorting of the samples in a.
(B) Co-stimulation of monocytes by GM3 species (16:0, 24:0, 24:1) plus synthetic TLR4 ligands LA506 (15 ng/mL), LA505 (150 ng/mL), or LA504 (150 ng/mL).
Representative PET image of the upper body of a human subject who received the PDE10A radioligand [18F]-MNI-1869. Supraclavicular brown adipose tissue (BAT) is highlighted by the crosshairs. The mean standardized uptake value (SUV) of [18F]-MNI-1869 in supraclavicular BAT and skeletal muscle was calculated in human subjects (n = 5). ** P = 0.00210 (5 mins), **** P < 0.0001 (15 mins), **** P < 0.0001 (30 mins), **** P < 0.0001 (50 mins) and ** P = 0.00462 (75 mins) using unpaired 2-tailed Student's t tests.
(e) Steady-state levels of p62, LC3 and GAPDH visualized by western blotting in PSN-1 cells treated for 48 h with the indicated doses of PIK-III. (f) Steady-state levels of p62, LC3 and GAPDH visualized by western blotting in Panc10.05 cells treated for 48 h with the indicated doses of PIK-III.
C TCGA FPKM values of interferon-related genes in tumours grouped in TERThigh (n = 500) and TERTlow (n = 500) groups. Heat map showing mean FPKM values of each gene.
B OCR rates of cardiac mitochondria after administration of pyruvate, malate and ADP, n= 7 per genotype.
(G) EpiSC resetting in 2i+LIF measured by Oct4-GFP+ colony formation after Stat3 siRNA in wild type EpiSCs (left), or Klf4 KO EpiSCs transfected with Tfcp2l1 and Gbx2 siRNAs. n=3 independent experiments in wild type cells (box plots indicate min, median, max), n=2 for Klf4-/- cells (box plots indicate min, mean, max)
E. Proliferation of melanoma cells over indicated times after treatment with siASNS, siATF4, or both.
E Comparison of oxygen consumption rates (pmol O2/s, normalized for cell numbers by Hoechst staining) of basal, proton leak, maximum and non-mitochondrial respiration in WT and MIC10 KO cells obtained from 3 independent experiments. The data is normalized to basal respiration from HAP1 WT. Bar and error bar represent mean ± SEM from 3 independent experiments. *P = 0.034 for basal (using one sample t-test) and *P = 0.018 for maximum respiration (using unpaired Student's t-test).
(A) Western blot from whole cell lysate of munc18-1 null mutant neurons rescued with M18WT and incubated with WIN55,212-2 (5 M) for 30 s. Cells were harvested after the indicated time. CB1R activation leads to a rapid and transient increase in pERK activity.
C, D TEM of dorso-lateral tracts of spinal cords of 6-7 week old WT (n = 4, 1209 mitochondria) and NYKO (n = 4, 1792 mitochondria) mice, 17-18 week old WT (n = 4, 442 mitochondria) and NYKO (n = 4, 724 mitochondria) mice and 31-32 week old WT (n = 3, 764 mitochondria) and NYKO (n = 3, 817 mitochondria) mice. Scale bars, 200 nm. Unpaired t-test, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ns=not significant. Data are means ± SEM.
H. qPCR quantification of selected genes reflecting ER-stress or protein methylation-related processes in CamkIIδc TG mice (n=4) and control mice (n=5). *p<0.05, ns = non-significant, Unpaired t-test; two-tailed. Data is normalized to Hprt1 expression.
(D, E and F) Polyphenols repress T3SS induction in P. syringae in MM. Transcripts of the indicated genes in the presence of 100-μM TA (D), 100-µM PGG (E) or 2-mM EGCG (F) were determined by RT-qPCR 6 hours after culturing in MM. Data information: , * indicate significant differences between mock group and strains under the different test conditions. (Student's t test. n.s., not significant, *, P ≤ 0.05, **, P ≤ 0.01, and ***, P ≤ 0.001). Error bars represent standard deviations (n=3 biological replicates).
(C) WT and PAQR3-deficient MEFs were treated with 80 mM chloroquine (CQ) for 2 h to induce LC3-II accumulation. Whole-cell lysates were harvested for IB analysis of LC3-II.
C: Expression of typical liver stage protein markers: Circumsporozoite protein (CSP) top left; Exported Protein 2 (EXP2) top right; Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) bottom left; and Merozoite surface protein 1 (MSP1) bottom right. Scale bar is 25 µm.
B Fraction of GFP-Pho8 structures classified as stacks, cytosolic whorls, uptake intermediates or vacuolar whorls. The number of GFP-Pho8 structures in WT, ∆vps23, ∆snf7 and ∆vps4 cells was 562, 345, 367 and 449, respectively. The table shows the results of Chi-Square tests for independence for all pairs of strains. ***, highly significant (P < 0.00001); ns, not significant (P > 0.05).
(A) Representative confocal images and Quantification of STX within the lysosome in U2-OS and HeLa cells after acute silencing of CLN3 (siCLN3) in DMSO or treated 48h with Tamoxifen. (*** vs siSCR DMSO, °°° vs siCLN3 DMSO). Data are presented as mean ± SD, °°°/***: P ≤ 0.0001, as determined by ANOVA (n=3 biological replicas in duplicate). Scale bars: 20 µm.
F Western blot analysis of liver lysates (100 µg) from either CodopN6 intein-treated mice (n=6) or untreated haemophilic mice (n=2). A lysate from HEK293 cells transfected with the N6 full-length plasmid was used as positive control (50 µg). Arrows point at N6 full-length protein (N6), the 5' half of N6 (5'N6) and 3' half of N6 (3'N6).
A. Apc1-loop500 is the hub for both Plx1 and PP2A-B56 phosphatase. CDK sites (T or S) and B56-binding site (green) on Apc1-loop500 (yellow bar) are shown. Bindings of Plx1 and B56 to Apc1-loop500 depend on phosphorylation of CDK sites. The PP2A-B56 associated on Apc1-loop500 may dephosphorylate CDK sites.
(e) Immunofluorescence of HeLa cells expressing a Flag-tagged truncated form of ATG16L1 (Flag-ΔN85-ATG16L1; green). Blue, DAPI.
Fluorescence polarization assays performed with 50 nM 6-FAM-ds20, a 6-FAM-labeled 20 nucleotide double-stranded DNA, and serially diluted Siz2∆CT or Siz2∆CT mutant.
D. Mouse genotyping by multiplex PCR. Top: A 221 bp PCR product is present in wild-type mice (+/+); the Rnaseh2bA174T allele (also) give a 460 bp product. Bottom: Position of forward (x) and reverse primers (y, z).
Middle panel: ribbons representation of the sE trimer (PDB 1URZ) colored according to domains with high HDX regions a and c boxed and highlighted in light grey. The central box marks the domain I-II hinge. Residues 401 and 403, which occupy the
(A) UBQLN4 interacts with non-glycosylated (and thus defective) IL-2Ra WT in SRP54-knockdown cells. After treatment of HeLa cells with SRP54 siRNA (10 nM) for 48 h, Flag-tagged IL-2RaWT and T7-tagged UBQLN4 proteins were expressed, and the cells were treated with (+) or without (-) 10 µM MG-132 at 4 h before harvesting. UBQLN4 was affinity-purified from cell extracts and probed with an anti-Flag antibody. Note that the levels of non-glycosylated IL-2RaWT (indicated as Defective) increased, while the glycosylated form of this protein (indicated as Assembled) decreased in SRP54 knockdown cells.
(C) Three fractions representing the 150S and 75S peaks were pooled and run on SDS-PAGE, and the 35S-Methionine labeled bands were visualized. The bands are labeled according to expected relative migration pattern, and VP2 is identified by its absence in the 75S peak. The ratio of VP0 to VP3 bands was analyzed from four independent experiments and plotted in arbitrary units. * p<0.05, ** p<0.01, *** p<0.0001.
(b) FoxO1(WT) or Flag-FoxO1(ΔDB) was transfected into H1299 cells; 48 h later the autophagic vesicles were observed under an electron microscope. Enlarged images (right) show clearer autophagic vesicles indicated by arrows. Scale bars, 2 μm.
A Robust immunofluorescence detection of ubiquitination on LAMP1-positive compartments with pan-ubiquitin FK2 or K48 chain-specific antibodies. HeLa cells were treated with 250 µM LLOMe or EtOH (untreated) for 3 h, fixed and stained with indicated antibodies. Confocal microscopy was followed by automated image analysis. Graphs represent data from three independent experiments with ≥ 50 cells per condition (mean ± SD). Scale bar: 20 µm.
c, Scanning electron microscopy (SEM) of primary cilia. Arrows, cilia-associated vesicles; arrowheads, ciliary pocket.
Log-rank (Mantel-Cox) survival analysis based on high and low p-T172-BCL9 expression scored by immunohistochemical staining in two tissue arrays with (G) 75 lung cancer samples or (H) 145 breast cancer samples.