EMBO/sd-character-level-ner huggingface dataset:
BioLayer Interferometry experiments were performed to evaluate the binding capacity of APOE3 on soluble TREM2-Fc pre incubated with Aβ42 oligomers. Schematic representations of two sequential associations of ligand on soluble TREM2-Fc are represented by I and II. Soluble TREM2-Fc loaded on a sensor is first incubated with APOE3 (association I) followed by a second incubation with Aβ42 oligomers (association II) or the reverse- incubated with Aβ42 oligomers (association I) followed by a second incubation with APOE3 (association II). All experiments were replicated 3 independent times
F Measurement of DNMT3A-ADD thermodynamic stability upon ATO or AsPh3 treatment. Purified DNMT3A-ADD was mixed with ATO or AsPh3 at the indicated ratios in 20 mM HEPES (pH 7.5) for 1 hour. Chemical structure of AsPh3 is shown. Melting curves were measured by DSF (upper panel), and the calculated ΔTm values are shown (lower panel).
Cell viability was measured using CellTiter-Glo in NSCLC cell lines treated with volasertib, tepotinib, or a 1:2 ratio of both for 72 hours. Left: representative cell viability graph of cells treated with the indicated drugs. Right: heatmap depicting the calculated combination indices at Fa = 0.25 and Fa = 0.5.
Location of known Parkin ubiquitination sites within the Miro structure. MiroS is shown in a 180°‐rotated top view (relative to Fig 2). Four residues in dMiro correspond in sequence and structure to residues that are ubiquitinated by Parkin in human Miro1 (hMiro1) or human Miro2 (hMiro2) [22]. These are shown as spheres (K220, R240, K439 and K600; labelled with dMiro numbering). An alignment of a 10‐aa segment of dMiro, hMiro1 and hMiro2 flanking each ubiquitinated site is shown. The large blue 'K' within each sequence alignment indicates the ubiquitinated residue in hMiro1 or hMiro2, and a smaller, underlined K indicates that dMiro has a lysine in the same position. Residue S324, a candidate Pink1 phosphorylation site, is also shown in spheres. Note the side‐chain of K439 is disordered in the MiroS structure, so it is modelled as an alanine. dMiro/Dm Miro, Drosophila Miro; Hs Miro 1/2, human Miro1/2.
(A) Immunoblotting of NSC34 cells expressing AR-24Q or AR-97Q (NSC24Q and NSC97Q, respectively) that were treated with DHT for Dnmt1, Dnmt3a and Dnmt3b, and quantification of the signal intensities of bands corresponding to Dnmt1, Dnmt3a and Dnmt3b (n = 3).
(a) Starvation-induced long-lived protein degradation is impaired after NRBF2 knockdown. Compared with control (con) siRNA, NRBF2 siRNA decreases long-lived protein degradation (proteolysis rate) in NIH3T3 cells under starvation (ST) condition. This difference is diminished when the starved cells are co-treated with wortmannin (WM, 100 nM). (Data are shown as mean±s.e.m., P values are indicated on the figures, one-way analysis of variance with Dunnett as post hoc test, n=4; NS, not significant).
Mice were superinfected as described in Fig 3A (A) Frequency of IFNγ+ and IL-17+ immune cells following intracellular cytokine staining and flow cytometry Data information: Data are representative of at least two independent experiments with 3 mice per group. All data are represented as mean ± S.E.M. Statistical significanc was determined by unpaired student's T-test
(b) Survival determined by clonogenicity during chronological ageing of wild-type yeast (BY4741) cells with (o) and without (▪) addition of spermidine (4 mM) at day 1. Data represent means ± s.e.m. (n = 5).
Apg16p interacts preferentially with Apg12p‐modified Apg5p. Total lysates were prepared from apg16Δ cells (YNM124) or apg12Δapg16Δ cells (YNM115) harboring both pHA‐APG5 and pMyc‐APG16 CEN plasmids. The lysates were immunoprecipitated and analyzed as described in Figure 3.
Representative epi-immunofluorescent images of L6 cells transfected with myc-RHEB Q46L and immuno-stained against LC3B and myc after iron treatment (FeSO4, 250 µM) for 24 h.
PINK1 KO HeLa cells stably expressing mito-Keima were infected with control or shATAD3B. Five days later, cells were treated with DMSO, 200 µM H2O2, or 4 mM 3-NPA for 2 h. Cells were then analyzed and imaged with 458 nm (measuring mitochondria with a neutral pH) and 561 nm (measuring mitochondria with an acidic pH) laser excitation for mito-Keima by confocal microscopy. Scale bar, 10 µm. Quantification of the relative ratio of red to green fluorescence intensity (561nm/458nm) of the cells described in (A). Data are presented as mean ± SD (n = 3 independent experiments, 20 cells per experiment), statistical significance was assessed by two-tailed student's t-test, N.S., not significant, **p < 0.01.
(G) Representative examples of estrus cycles in WT and L-HRD1 KO mice as determined by vaginal cytology.
(C) Upper, the linear map of the pRF bicistronic report plasmid. The SV40 promoter (purple box) was used to drive firefly luciferase (Fluc) and Renilla luciferase (Rluc) gene transcription. Different p53 5'UTR fragments were inserted between the Fluc and Rluc genes. Lower, at 24 h after transfection, cells were lysed and luciferase activities were measured using a dual-luciferase kit and the ratio of Fluc/Rluc was calculated. Data shown as means±SD (n=3). Statistical significance was determined by two-tailed Student's t-test. * P<0.01.
Relative expression of Prolactin-induced protein (PIP) mRNA (D) normalized to GAPDH in PF and PF-PH patients.
Representative confocal images of wt L6 and RHEB-Q64L L6 cells pulsed with Lysotracker Deep Red and immuno-stained against LC3B after iron treatment (250 µM, 24 h).
A Time-lapse microscopic images showing that H1299 shCtr or shM2 cells stably expressing Mito-GFP were connected by TNTs containing mitochondria. Cells were seeded in collagen-coated dishes and cultured for 24 h before time-lapse life cell images were acquired by confocal microscopy (also see Movies EV1 and EV2). Fluorescent images (with bright field) at different time points show the mitochondria moving along the TNT and entering in one of the two connected cells. Scale bar: 20 μm. The moving mitochondria are marked by the white arrows. Boxed areas (shCtr in yellow dotted boxes; shM2 in red dotted boxes) are enlarged and shown in the middle panel. B Confocal microscopic images of control and shM2 H1299 cells following immunofluorescence staining with anti-Tom20 antibody (green) and staining using phalloidin (red). The white arrows marked TNTs, whereas the white triangles marked mitochondria inside TNTs. Scale bar: 25 μm. C Data information: nuclei were stained with Hoechst 33342 (blue).
F, Summary figure integrating transcriptomics (normal, n=8 mice; tumor and residual, n=4 mice each), intracellular (Int.) metabolomics (normal, tumor, and residual, n=8 mice each; WT, n=2 mice), extracellular (Ext.) metabolomics of spent growth medium (normal, n=4 mice; WT, n=2 mice; tumor and residual, n=3 mice each) and DNA methylomics (normal, n=2 mice; tumor and residual, n=3 mice each) from the three populations. Color depth represents the normalized Euclidean distance of the respective omics layer in reference to normal. Distances between the centers of the three populations correspond to the normalized mean Euclidean distances across all represented omics layers.
D-E. (D) Maximal EAE clinical scores (individual plots with mean) of selected mice
E GST‐fused propeptide of Ape1, Atg19 as indicated, and a His‐tagged fragment of Atg11 (amino acids 685-1178) were purified from Escherichia coli, mixed, and propeptide‐bound proteins were isolated and analyzed by Coomassie staining. Quantification of three individual experiments is shown. Error bars represent standard deviation.
A. MCU silencing reduces HIF-1α protein levels. Cells were transfected with siMCU or siControl. HIF-1α protein levels were detected 48 hours later.
Quantitative RT-PCR measurement of Fgf21 and GDF15 expression in GC muscle of 4-month-old control, HSA-Cre untreated and HSA-Cre Bezafibrate-treated mice.
(A, B) The ap-Gal4 driver induces dS6K in combination with either (A) the DHR3-EP transgene or (B) the EP with the EMS DHR3 mutation
(D) Neuronal cell death was evaluated in the cerebellum (Calbindin immunostaining) and in different layers of cortex (NeuN immunostaining of frontal cortex) of 10 months-old mice belonging to each experimental group of mice. Number of cells was quantified from 20 different images (taken from 5 mice for each group) and expressed as % of WT. Values are means ± s.e.m. *P<0.05, **P<0.001 Student's t-test. Scale bar: 50 µm.
(D) Quantitation of percentages of autolysosomes (autolysosomes/total autophagic structures) in nondegenerating ovaries from fed w1118 and SesBOrg flies. 10 egg chambers were manually examined per genotype. Error bars represent the means ± SD. Statistical significance was determined using a two-tailed Student's t test (***, P < 0.0001).
F Examples of electrophysiological properties of humanneurons: control individual (top); PKAN patient (bottom). Traces on the left represent trains of action potentials induced by injection of a suprathreshold current step through the patch electrode in current-clamp mode. Middle traces show Na+ and K+ currents (down- and upward-deflecting from baseline, respectively) in response to a 60 mV step from a holding voltage of -70 mV in voltage-clamp mode. Insets on the right display enlarged portions of the traces to magnify fast Na+ currents. G Histogram with percentages of recorded cells showing repetitively firing (z-test).
(B) Western blotting of WT and Drp1-KO MEFs with or without FCCP treatment (30 min) using the indicated antibodies.
Adult Swiss mice received a single i.c.v. injection of vehicle or 10 pmol AβOs, and hypothalamic levels of mRNA for AgRP (C; n = 6 Veh; 5 AβOs), NPY (D; n = 6 Veh; 5 AβOs) and POMC (E; n = 7 animals/group) were analyzed 7 days after injection. In (C), *P = 0.0191; in (D), *P = 0.0115; Student's t-test.
Exosomes were isolated from the media of BMDMs (BMDMs-exo), BMDM-polarized M2 macrophages (M2-exo), BMDM-polarized M2 macrophages transfected with si-CRNDE (M2-si-CRNDE-exo), and BMDM-polarized M2 macrophages transfected with the negative control of si-CRNDE (M2-si-control-exo). Then, MFC cells were treated with indicated exosomes and cisplatin (CDDP; 1μg/ml) for 48 h. (D) Cell apoptosis was measured by flow cytometry. Data information: The results shown are from three biological replicates. Data are expressed as mean ± SD. one-way ANOVA. **P<0.01.
(D) The expressions of PTEN and NEDD4-1 were measured using western blot in SGC7901 cells transfected with pcDNA-CRNDE or Mock. GAPDH was served as an internal control.
(F) Hepatic Igf1, Cyp2d9 and Cyp4a12 mRNA in the WT and L-HRD1 KO mice 5 weeks after AAV-shCrebh injection. (n=5 for each group).
HeLa cells stably transfected with tetracycline (Tet)-inducible Bcl-xL expression vector were incubated in DMEM containing Tet or not for 12 hours, then treated with DMSO or 10 μM MG132 for 8 hours, then washed and incubated in fresh medium for another 3 days, Cells were then subjected to SA-β-gal staining. The induced Bcl-xL was detectable even 3 days after treatment withdrawal, because this protein is stable.
Mice were superinfected as described in Fig 3A (D) Pneumococcal cfu in BAL following αLy6G antibody mediated neutrophil depletion. Control T-bet-/- mice were administered with IgG isotype control antibody Data information: Dat are combined from two independent experiments (n = 8-12 mice per group) All data are represented as mean ± S.E.M. Statistical significance was determined wit Mann-Whitney U test in (D
A Schematic presentation of the approach. Cytosolic translation is inhibited with Harringtonine (Harr) while mitochondrial ribosomes are allowed to incorporate the alkyne-containing methionine homolog (HPG, orange) into newly-synthesized proteins. In fixed cells, HPG moieties are clicked to an azide-conjugated fluorophore (star, magenta), through copper-catalyzed Huisgen cycloaddition (click), and can be visualized microscopically.
(E) Smyhc1 immunohistochemistry stain of 24 hpf (hour post fertilization) zebrafish larvae. Filamentous actin is stained with phalloidin (red). Nuclei are stained with DAPI (blue). Smyhc1 is stained with the F59 antibody (green) (Elworthy et al, 2008). smyhc1+/+, smyhc1R673H/+, and smyhc1R673H/R673H larvae display Smyhc1 in the muscle fibers at 24hpf. smyhc1-/- larvae at 24 do not stain positively for Smyhc1. Scale bar represents 50 µm.
(C) Dissected brains of w1118, alix1, γ−Tub23CPI and γ−Tub23CPI;alix1 larvae were subjected to MT regrowth assay and stained with anti-α-tubulin (red), anti-Cnn (white) and Hoechst (blue). Representative images of NBs after 0 and 60 seconds regrowth are presented. Scale bars, 5 μm
F Representative image of proliferating astrocytes (SOX2+/MCM2+; cyan and magenta, respectively; arrowheads) of all DG layers. G Quantification of the percentages of proliferating astrocytes in distinct DG layers over all proliferating DG astrocytes. Data information: All data are represented as mean ± SEM; number of experimental animals (indicated by red dots): (G) n = 4, dotted lines border GZ; scale bars = 50 µm F,
A BAK-6H on liposomes was activated or not with cBID for 60 mins prior to deuteration for 10s, 60s, 600s, or 3600s. Incorporation of deuterium was assessed by mass spectrometry. Change in deuteration at each time point is expressed relative to BAK on liposomes without cBID.
H-I. Rag1-/- mice which received 1 x 105 IL-17A+cells of indicated genotypes and (I) total cell numbers (mean +SEM) of infiltrates isolated from CNS and used for further analysis.Data (H-L) consist of Rag1-/-mice which received WT (n=6), IL-1R1ΔT (n=3) and IL-1R1-/- (n=4) cells polarized in the presence of IL-23 prior to transfer. All experiments were performed at least twice with similar results.
(C) EC were seeded on gelatin-coated Transwell inserts. RFP-labeled LLC cells were allowed to transmigrate for 8 h through the endothelial layer. Thereafter, transmigrated tumor cells were counted. The dot plot presents the normalized data (mean ± SD, n = 9 inserts from three independent experiments). ***, P<0.001 (two-tailed Mann-Whitney U test).
A. A diagram showing two groups of ER-resident proteins: those that become macro-ER-phagy cargos (in green), and those that do not (in blue or grey). Like overexpressed Snc1-PEM or Snq2, the ER-resident membrane proteins Sec61 (translocon subunit) and Hmg1 (sterol biogenesis) are transported to the vacuole via macro-ER-phagy. In contrast, the ER-to-Golgi exit regulators Sec12 and Sec13 and the ER lumen chaperone Kar2 are not co-transported to the lysosome through macro-ER-phagy. Four or six different strains were used for this analysis: WT (YPT1), pep4∆, and pep4 prb1∆, ypt1-1, ypt1-1pep4∆, and ypt1-1pep4∆ prb1∆. In each strain, one ER-resident protein was tagged at its C-terminus (except for Kar2). (The pep4∆ strains were not used in all experiments because they do not show the full defect). The level of ER-resident proteins was determined by immunoblot analysis in cells that either do not (B-D) or do overexpress GFP-Snc1-PEM (F-H). B-C. Sec61-3xHA (B) and Sec13-3x HA (C) expressing cells (2 independent un-transformed colonies) were tested by immunoblot analysis (using anti-HA antibodies). Shown from top to bottom: strain genotype, HA-tagged protein, G6PDH (loading control), quantification of HA-tagged protein expressed as average fold of WT (p-value = 0.025 for Sec61).
(d) U937 cells were transfected with either control siRNA or siRNA specific to MFN1/MFN2, endogenous LC3 puncta per cell were quantified.
Growth assay of wildtype and Rsp5F618A strains containing GID2 or GID5 deletions on SD-Lys (-) and SD-Lys containing 0.75 μg/mL thialysine (+). Spots represent 1:2.6 serial dilutions.
The presence of GFP-PSF-1 on mitotic chromatin (indicated by white arrows) was monitored by spinning disk confocal microscopy (see Materials and Methods), in embryos derived from worms that were fed on bacteria containing a single plasmid expressing the indicated RNAi ('Control' = empty vector). The scalebars correspond to 5 µm.
B. Western blot analysis of puromycin (Puro)-labeled fibroblasts (CAF, normal human mammary fibroblasts, WI-38, and NIH3T3) after 2-h starvation and EAA add-back in the presence or absence of cycloheximide (CHX). C. Western blot analysis of Puro-labeled fibroblasts cultured in growth medium.
(B) Quantifications of the corrected total cell fluorescence (CCF) for genotypes in (A). CCF = Integrated Density - (area of selected cell × mean fluorescence of background readings). At 0h ALH, CCFs were 735.7±220.7 (n=168) for control, 302.1±150.0 (n=160) for 716/Df. At 24h ALH, CCFs were 2238.3±628.4 (n=158) for control, 310.4±207.1 (n=113) for 716/Df. At 48h ALH, CCFs were 3834.0±924.2 (n=124) for control, 465.9±295.0 (n=126) for 716/Df. Data are presented as mean ± SD. Statistical analyses were done comparing between the CCFs at later time points with the CCFs for control at 0h ALH using Mann-Whitney test . ***P < 0.001. Data information: Quantification data are presented in as mean ± SD (B Statistical analyses were done comparing two different genotypes using Mann-Whitney test (B, ns: non-significant.
(M) Enzyme-linked immunosorbent assay (ELISA) for interleukin (IL)-6 and IL-1β from colon tissue. Data are mean ± SEM from seven biological replicates, **P<0.01 using one-way ANOVA.
D Kinase activity of PIPKIγi5 is required for LAPTM4B association. HEK293 cells expressing LAPTM4B and wild‐type (WT) or kinase‐dead (KD) PIPKIγi5 were starved overnight, stimulated or not with 100 ng/ml EGF for 15 min, and harvested for immunoprecipitation with anti‐myc. The co‐immunoprecipitatedLAPTM4B was detected by immunoblotting.
E Learning in the hippocampus-dependent IntelliCage serial reversal task (ANOVA: day F2,56=26.71 p<0.0001). Scores provided no evidence for a mutation effect on learning rate or performance.
Figure 1. Comparison of genome structures recovered from five synchronized stages over the cell cycle. (C, D) Comparison of contact maps. The 16 yeast chromosomes are displayed atop the maps. Black arrowheads: inter-telomere contacts. Yellow arrowheads: inter-centromeric contacts. Left panels: Hi-C maps obtained from two G1 cell populations synchronized independently (C) . Right panels: log-ratio between each pair of maps. Insets display magnifications of chr4. Blue to red color scales reflect the enrichment in contacts in one population with respect to the other (log2).
(c) RagAT21N and RagCQ120L suppress insulin-induced stimulation of S6K phosphorylation. pcDNA3 (200 ng, lanes 1 and 2) or each indicated Rag construct was co-transfected with HA-S6K (20 ng) or GST-Akt (100 ng) into HeLa cells. Cells were serum-starved overnight and stimulated with insulin (400 nM) for 30 min. Phosphorylation and protein levels were determined by immunoblotting with the appropriate antibodies, as indicated. Full scans of blots are provided in Supplementary Information, Fig. S6.
E Drug-response curves showing increased resistance of shTINCR-WM902B cells after treatment with BRAF (Vemurafenib; from 10 to 10,000 nM) and MEK (Trametinib; from 4 to 2,000 nM) inhibitors. The shSCR- (control) and shTINCR-WM902B cells were treated with increasing concentrations of drugs and their viability assayed after 72 hours using CellTiter-Glo luminescent kit (Promega). Error bars represent s.e.m. of three biological replicates. The dose response curve was fit with nonlinear regression (GraphPad Prism).
C. Western blot analysis of mTORC1 activity in MYH10 KO MEF cells with stably re-expressed exogenous mCherry-MYH10 in response to starvation and AA stimulation. MYH10 KO MEF cells were transfected with mCherry-MYH10 and selected with G418 for 2 weeks.
(A) Western blots of cell lysates derived from mock (CTRL), single UBE4B or LSD1, or double UBE4B and LSD1 knockdowns. Supernatant (S) and pellet (P) fractions were probed with indicated antibodies. While the LSD1 or UBE4B single-knockdown reduces the SOD1G85R aggregates in both supernatant and pellet fractions, the combined knockdown produces the strongest reduction in the aggregates. (B) Quantification of SOD1G85R protein levels by western blotting (A). n = 3 (supernatant); n = 8 (pellet).
B BMMΦ were infected with increasing doses of M.bovis BCG for 6 days and MGC formation was determined by hemacolor staining.
D Thermal stability changes in protein complex members. Scores for thermal stability shown for proteins from 26S proteasome (separately for 19S regulatory and 20S core particles) and CCT chaperonin complex. Boxplots indicate median, first and third quartiles with whiskers extended to 1.5 times the interquartile range out from each quartile. Data used in stability score calculation is from two (mock shock) and three (heat shock) biological replicates.
(J) Western blot analysis of endogenous Atg8a from third instar muscle lysate. Dmef2-Gal4 driven knockdown of AGBE but not CG9485 inhibits Atg8a lipidation, indicating reduced autophagy levels.
(A) Increased protein levels of proteasome subunits upon the UBE4B and LSD1 double knockdown in HEK293T cells
E. Bioluminescent imaging of tumor growth in mice bearing xenografts derived from the luciferase-labeled T3359 GSCs treated with C646 or the vehicle control at indicated days after GSC transplantation (n=5 mice per group). F. Quantification of tumor growth from (E) (n=5 mice per group).
(E) CD86 and CD40 expression was quantified on BMDCs plated at high density (2x105 cells/cm2) and stimulated with poly(I:C) in the presence (red dots) or absence (blue dots) of anti‑IFNAR1 blocking antibody. Data are pooled from 2 independent experiments.
Western blots against CYP46A1 and actin-β (ACTB) in total extracts (T) and biotin-streptavidin immunoprecipitates (S) from synaptosomes from wt and NPC1nmf164 mice in which cLTP was induced or not. Graphs show mean ± SEM of the levels of biotinylated-surface CYP46A1 as a percentage of the total amount of each protein (n=3 mice, 3 month-old, 2-way ANOVA , *pCYP46A1 = 0.0371).
Voltage command (top), corresponding whole-cell Ca2+-influx (middle) and the functional fluorescence responses (bottom) (bandstop filtered at 33.3 Hz) from Rhod-FF and iGluSnFR, respectively. A modified Boltzmann function (see Materials and Methods, R2 = 0.81) was fitted to a Rhod-FF fluorescence trace in response to a voltage ramp (C-C'). iGluSnFR-AUC was calculated per depolarization voltage and used for a Boltzmann fit (D', R2 = 0.92). The voltage of half-maximal activation (V1/2) and the dynamic range (10-90%) of synaptic Ca2+-influx and glutamate release were calculated from the fits and depicted as circle and bar in C' and D', respectively. The obtained fits from a Ca2+ "hotspot" (C') and from glutamate release (D') were plotted against each other in a voltage range from -57 mV to -17 mV in 1 mV increments. A power function was fitted to the 25% of the maximum iGluSnFR-AUC (R2 = 0.99) to obtain the m-estimate. (ruptured patch-clamp, 10 mM intracellular EGTA, 5 mM [Ca2+]e) (See also Fig EV4)
(A) Experimental schemes LacI-CENP-B or LacI control (baits) were expressed into CENP-AEA/- lacO-tetO U-2OS cells. Immunostaining against CENP-C or CENP-A (preys) was performed following LacI-CENP-B transfection in untreated, IAA treated and IAA wash-out (WO) conditions.