EMBO/sd-character-level-ner huggingface dataset:
D. DMSO or Z-VAD-FMK treated HEK293/MycBioID-FAM111A cell lines were induced with doxycycline for 24 h. Total protein from an equal number of cells was incubated with magnetic streptavidin beads, and pull downs were analyzed by western blotting using GANP and Myc antibodies.
C. Representative images for co-localization between p-RPA32S33 (green) and telomeres (red FISH probe) in U2OS-hTR cells treated with control siLuc or siNHP2, 72 h post-transfection. White arrows indicate co-localization events. Quantification of images is shown on the right. Each dot indicates the number of co-localization events per nucleus (n=2). Statistical analyses were performed with Student's two-tailed unpaired t-tests with Welch's correction for normality of distribution. Data information: All scale bars: 5 μm. Data are shown as mean ± SD, with the red line marking the mean value.
B. Representative time-lapse spinning disk confocal images of n = 3 of live COS7 cells stably expressing mCherry-KDEL (green), transiently expressing EB3-GFP (yellow) and SiR-lysosome-stained LE/Lys (magenta). Zoom insets show regions of interest containing a mobile LE/Lys as denoted by the white arrow. First row shows ER (mCherry-KDEL) (green), the second row shows SiR-lysosome-stained LE/Lys (magenta), the third row shows EB3-GFP (yellow) and the fourth row shows the merge of the three preceding channels. Scale bar: 30µm and zoom insets 3µm. All images acquired for 90 frames at 1Hz. Images related to movie EV6.
G Immunofluorescence of TBC1D5 (shRNA#1) cells transiently transfected with shRNA‐resistant myc‐TBC1D5, treated with KU0063794 (6 h). 20 h post‐transfection cells were treated, subsequently fixed, and immunostained with anti‐myc and anti‐LAMP1 antibodies.
A Analysis of reactive oxygen species (ROS) accumulation in frozen liver sections from 120-day-old (P120) and 240-day-old (P240) PR-SET7ΔHepA mice and control littermates (WT).
DC3.2-Rab39a cells were induced to express Rab39a overnight. Cells on coverslips were washed, fixed and stained according to the listed protocol and antibodies. Green = Rab39a (HA tag), Magenta = Lamp2. Arrows highlight punctate structures with colocalization. Scale bar = 10 µm. Representative image of two independent experiments.
Binding of Ronin to the promoters of genes encoding cytoplasmic ribosomal proteins.
F. Quantification of 13C3-Lactate using LC-MS/MS in media of siCON or siSlc16a1 treated BMECs after 13C6-glucose treatment (12hr). (n=3 from independent experiments. Error bars indicate the standard error of the mean (SEM) from unpaired Student's t test.)
Average S-score Pearson's correlation between the same genes (orthologs, solid line) and random gene pairs (shaded distribution) across all the 38 conditions and four strains. Genes are stratified by the number of conditions in which they show a significant phenotype across the four strains.
RNAscope ISH showed an apparent expression of TMEM16A mRNA co-localized with ChAT and Tac1 signals in the mHb compared to the ones in lHb. Enlarged images showed that signals of TMEM16A and Tac1 in the dorsal mHb or ChAT in the ventral mHb reside together at the single cell level.
A. Western blot analysis of Tace using lysates from WT and TaceFl/Fl;P0-Cre mousesciatic nerves at P10 (65ug of total protein lysate loaded). The arrowhead indicates the pro-protein and the arrow shows the cleaved active form of Tace. Representative of two independent experiments.
Western blot analysis of the indicated FAO enzymes in the EV secreted by primary adipocytes from lean (ND) and obese (HFD) mice (top panel) and from human individuals with varying BMI (normal weight, NW; overweight, OW and obese, OB) (bottom panel). For each blot, extracts from 3 independent batches of murine samples or 3 independent individuals for human samples (1-3) are shown. Flotillin 1 (FLOT1) is used as a loading control.
A-B. 3D7 (A) or parasites overexpressing EK-GFP (B) were used to prepare protein lysates followed by immunoprecipitation using anti-PfCDPK7 or anti-GFP antibody, respectively. Subsequently, protein lysate or indicated IP were subjected to Western blotting using anti-PMT (A), anti-GFP (B). PMT was found to co-IP with PfCDPK7 (A, lane 2) and EK-GFP with PfCDPK7 (B, lane 3).
S9.6 signal in the nucleoplasm of U2OS cells. An average from 3 independent experiments performed in triplicate. In total, ~90 images for each condition (10 images/slide and 9 slides for each condition) were taken and two cells per image were quantified. Statistical significance was assessed using one-way ANOVA t-test. *: p < 0.05; ****: p < 0.0001.
E A375 parental or DCAF12 KO cells were treated Cell lysates were collected and immunoblotted for the indicated antibodies to confirm MAGE-A3/6 depletion and monitor the autophagy adaptor/substrate p62/SQSTM1.
Gene ontology (GO) analysis of biological processes associated with common (D) up-regulated genes in Zfp207-depleted ESCs (sh1 and sh2); NHEJ: Non-homologous end joining.
(D, E) Rat liver lysosomes incubated or not with 1 mM CaCl2 (+Ca2+) were subjected to Triton X‐114 extraction and sucrose density gradients centrifugation. Levels of cathepsin A (D) and carboxypeptidase A activity (E) were measured in each fraction by immunoblot or using a standardized colorimetric assay, respectively. Values in (E) are expressed as times the activity in untreated membranes and are mean+s.e. of two experiments.
D. Schematic illustration for the lactate uptake assay after chamber removal. At 24hr after chamber removal, the fluorescence of mTFP and Venus were observed in the middle of the wells (yellow area)
C. Cdc5 interacts with Ime2. Cdc5-ha3, Ime2, and Spo13 were detected by immunoblotting in whole-cell extracts and α-Ha immunoprecipitates from the indicated strains. Data information: data are representative of two independent experiments.
(C) Confocal microscopy images of nuclei assembled with N-terminal Nup50 truncations and an minimal N-terminal fragment. Samples were analyzed as in (A). (D) Average percentage of mAB414 positive nuclei for 100 randomly chosen chromatin substrates in each of at least three independent experiments shown in (C). Data points from the individual experiments are indicated.
A. Exemplary normalized current responses a -120mV pulse showing fast Ca2+ dependent inactivation (FCDI).
Whole-mount AP staining of the blister roof epidermis from Col7a1-/- mice (right) and their littermate controls (left) at P1. Scale bar: 500 μm.
C. Cells were treated with doxycycline along with DMSO or Z-VAD-FMK for 24 h. They were fixed with formaldehyde, and stained with NPC antibody Mab414 and DAPI. The images were captured by confocal microscopy. Scale bar: 10 μm.
I-J. Immunofluorescence images of control (FRT40A, I) and Las RNAi (J) MARCM clones (green, outlined by white dotted lines) 10 days after clone induction (ACI). Pdm1 staining (red) was used to visualize ECs. K. Quantification of the ratio of Pdm1+ cells per clone of MARCM clones with indicated genotypes. n is indicated. Each dot corresponds to one clone. L. Quantification of MARCM clone size of experiments in (I-J). n is indicated. Each dot corresponds to one clone. Data information: DAPI stained nuclei are shown in blue. Scale bars represent 10 μm (D Error bars represent SDs. Student's t-tests, ****p < 0.0001, and non-significant (NS) represents p > 0.05.
D p53+/+ and p53 -/- HCT116 cells were transfected with control or SQLE siRNA for 48 hours as indicated. Relative cholesterol concentrations were examined as described in methods. Protein expression were determined by western blot.
(B) The Slx5-SIMs and Slx5-Md are required for Euc1 ubiquitylation. NiNTA-PDs for His-ubiquitin as in (A). All strains expressed wild-type 3FLAGEuc1 from plasmids under the ADH promoter and His-ubiquitin. Slx5 constructs were expressed from plasmids under control of the endogenous promoter. WBs were probed as described in (A), Slx5-levels were probed using an HA antibody. Asterisk denotes a non-specific band. Slx5-SIM*: SIMs 1-4 were mutated as described (Xie et al, 2010), for SIM5, aa 477-479 (IIV) were mutated to alanines. To avoid possible mislocalization by deletion of the Slx5-Md, which overlaps with a putative NLS (Westerbeck et al, 2014), an N-terminal NLS was fused to all constructs. See Appendix Fig. S7C for HU complementation.
A. Differential protein expression and phosphorylation was determined in NB1 cells in response to ALKAL2 stimulation. See Table S2 for detailed results.
a, The CD loop sequences of PROPPINs from a wide variety of organisms. Potential amphipathic helices were predicted using the online tool Heliquest and are plotted in red. Amino acids removed in Atg18DLoop (aa 328-384) are printed in bold.
(B) Heatmap comparison of the top 25 enriched Diseases and Biological Functions in AhR KO (KO/WT) and TCDD (TCDD/vehicle in WT mice) treated cells as identified by IPA.
HepG2 cells were treated with bovine serum albumin (Con or (−)), PA (500 μM), PA+verapamil (Ver, 50 μM) or PA+nicardipine (Nic, 100 μM) for 9 h. (a-d) After each treatment, cells were loaded with a calcium indicator X-Rhod-1-AM (a,b) or Fura-2-AM (c,d). Calcium levels were visualized by laser confocal microscopy (a) or by dual fluorescent microscopy (c, 340/380 nm ratio image) and quantified (b,d; n=8 and 30, respectively).
In vitro differentiation of Ronin fl/del, ecto-Ronin (grown in the presence of Dox) and Ronin del/del ESC.
Immunoblot analysis of PMEPA1 levels in 1603MED (G) cells 48 hours after transfection with either siCTRL (blue) or siPMEPA1 (red). Bar graphs on the right represent the quantification of the relative level of PMEPA1 protein level normalized to β-actin. The p-values were determined by unpaired t-test *p < 0.05, **p < 0.01, ***p < 0.001. Bars represent the mean ± SD. Number of replicates is n≥3.
(C) Knockdown of FIS1 does not increase ROS production. ROS production in FIS1 RNAi and control cells as measured by DHE FI under different glucose levels. Cells were loaded with dye and FI was measured before and 10 min after changing of glucose concentration (NS, non‐significant; P=0.5; *P0.001).
(B) TDP-43 immunostaining of quadriceps skeletal muscle from 3-mo-old mice treated with saline or 50 mg/kg/d intraperitoneal chloroquine (Chlq) for 4 wk. (A and B) Insets show one myonuclei from each field. Bars, 30 µm.
Percentage (%) of chymotrypsin-like (CT-L), trypsin-like (T-L) and peptidylglutamyl peptide hydrolysing or caspase-like (PGPH) proteasome activities in brain extracts of killifish of different ages. n >=6 per sample group; error bars indicate standard error of the mean. ** p < 0.005, *** p=0.0001, **** p < 0.001, one-way ANOVA, Holm-Sidak's multiple comparison test. For each sample group, the mean value of each activity in young samples (5 wph) was set to 100%.
(D) Chart comparing the expression of eight homeodomain TFs in the four anterior MNs (MN2c, MN2a, MN4, MN3) as identified by scRNA-Seq (Seq) and immunohistochemistry (IF/HCR), with absence of expression color-coded in magenta and presence of expression in green. The genes were selected for validation, as they allowed an unambiguous distinction of the four anterior MNs according to the scRNA-Seq data.
(c) MEFs stably expressing EGFP-Parkin were treated with CCCP (30 μM) for the indicated times, fixed, permeabilized and subjected to immunofluorescence analysis with antibodies to FKBP38 and to cytochrome c and with a confocal microscope. The fluorescence of EGFP-Parkin was monitored directly. (d) Higher-magnification views of the boxed areas in c. Scale bar (c,d), 10 μm.
(B) Comparison of arch domains conformations from Mtr4 (green) and FRH (large cell in cyan, small cell in marine) after superposition of the most N-terminal helical region. The isolated KOW modules of FRH, Mtr4 (with FRH superimposed), and Ski2 have considerably different loop configurations.
AT-3-OVA mammary tumour cells (1x106) were injected into the fourth inguinal mammary fat pads of female Ly5.1+ mice. Seven days after tumour injection FACS-purified 2x106 naïve CD8+CD44loCD62Lhi lymph node T cells from Ly5.2+;OT-1;Ptpn2fl/fl versus Ly5.2+;OT-1;Lck-Cre;Ptpn2fl/fl mice were adoptively transferred into tumour-bearing Ly5.1+ mice. d) for survival over 86 days.
Chromatin immunoprecipitation (ChIP) analysis of AhR and FoxM1 promoter interaction using WT colonic organoids treated with DMSO or TCDD, n=3 or 4 per group. A DNA fragment amplified in a gene desert on mouse chromosome 6 was used as a negative control.
Representative images of ear, tail and paw of c-Jun/JunB Δ/Δ K5-Cre-ERT2 at the start (Day14) and end (Day 28) of treatment with JNK Inhibitor or vehicle.
G and H IHC staining of submucosal glands (SMG) (G) or bronchi (H) with an anti-H1N1 polyclonal antibody (bar: 100 µm; red: influenza antigen (neufuchsin); blue: nuclei (haematoxylin)).
e, Line scan analysis of fluorescence intensities in b. Red: FM4-64. Green: Atg18-GFP. Scale bar: 1 µm.
Mitotic numbers during different time points of regeneration in starved conditions for xbp1/atf6 RNAi and controls. n ≥ 7 planarians per time point.
Schematic representation of the AQUA-MS/MS-based proteomics approach for quantification of cellular Ub linkage composition.
D. Interaction of ERα and 14‑3‑3β in a GST pull-down assay. GST‑tagged 14‑3‑3β was expressed in E. coli and immobilized on beads. PA‑tagged ERα was co-expressed with either wildtype PAK5 or NEK6, the kinase‑dead (KD) version or without a kinase in HEK293 cells. Immobilized 14‑3‑3β was used to pull PA‑tagged ERα from the HEK lysates. Co-precipitation of ERα was successful in the presence of wild kinases, but not in their absence or with kinase‑dead mutant versions.
(B) Identification of highly variable genes using the method by (Brennecke et al, 2013). Scatter plot depicts for each gene the mean expression and squared coefficient of variation across cells using the Poxm+ cluster as input. The solid line indicates the fit, dashed lines the 95% confidence interval. Genes with a significantly elevated variance are shown as triangles, other genes as circles. Different gene classes are color coded.
E. Cellular ROS was measured by flow cytometry and shown in both curve (left) and bar (right) charts (n = 3, biological replicates, mean±SEM, Dunnett-t test) Data information: * for P < 0.05; ** for P < 0.01; NS for no significance. Control referred to the corresponding plasmid vector and si-NC was the abbreviation of commercial normal control of siRNA.
F-I. Effect of ZD7288 (ZD, 10 μM, n = 10 from N=3), DDOA (15 μM, n=9 from N=4) or ODQ (10 μM, n=12 from N=4) on intrinsic properties of PFC pyramidal neurons. THC and vehicle conditions are similar to those on panel A-D. (F) resting potential membrane: THC + ZD: -74.14 ± 1.70 mV, p < 0.001 vs. THC condition, THC + DDOA: -70.47 ± 1.19 mV, p < 0.001 vs. THC condition, THC + ODQ: -63.10 ± 0.81 mV, p < 0.001 vs. THC condition,
Oncosphere formation in JNKi-treated cancer cells from breast cancer patients (pleural effusions or ascites). (I) Quantification of oncospheres per well in 4 different patients. Values are mean from 10 wells ± SD. (J) Representative oncospheres from patient samples. Scale bar, 50 μm.
Genetically induced titration of PdhR in a pdhR mutant reintroduced lag commensurate with expression level for a given TI feedrate (f = 0.38 mmol/g/h). Induction level corresponds to the amount of doxycycline (max - 50 ng/uL, half - 10 ng/uL, and zero 0 ng/uL) added at the onset of starvation. The TI feedrate is abbreviated as f (units: mmol glucose/g dry cell weight/hour). Wild-type lag (from Figure 2A empirical fit) is indicated by the dotted grey line. Lag time reduced with synthesis levels of titrated FtsZ in the wild-type strain at f = 0.31-0.32 mmol/g/h. Lag time increased with titrated synthesis of ClpX in wild-type cells at f = 0.43-0.45 mmol/g/h
Genome organization of SARS-CoV and schematic illustration of Nsp3 domains. Ubl1: ubiquitin-like domain 1; Mac: macrodomain; DPUP: Domain Preceding Ubl2 and PLpro; PLpro: papain-like protease; NAB: nucleic-acid binding domain.
HER-2 CAR-T cells generated from C57BL/6 and Lck-Cre;Ptpn2fl/fl splenocytes were transfected with GFP versus Ptpn2 siSTABLE™ FITC-conjugated siRNAs and intracellular PTPN2 levels were determined by flow cytometry.
D: Classification of primary response genes. For each gene, response time r is calculated as the sum of deduced transcriptional delay ∆t and mRNA half-life t1/2 and used for ranking. In general, immediate-early genes (IEGs) have short response times, whereas both immediate late genes (ILGs) and delayed early genes (DEGs) have longer response times. For ILGs, long response times are due to long mRNA half-lives. For DEGs, long response times are mainly due to long transcriptional delays (cf. boxplots).
A: Presence of putative AREs (AUUUA) in transcripts of WT oocyte. Transcripts containing no ARE or transcripts containing 1 or more AREs were subdivided into frequency groups according to the log2 fold change (MII/GV) in WT oocytes
B) For each population, equatorial Z stacks of two representative nuclei are shown in the left panel (chromatin is shown in blue (DAPI staining) and nuclear envelop in white (Sun2 staining). Scale bar: 5 µm. The corresponding 3D reconstructions are presented in the right panel. Convex and non-convex surfaces are shown in blue and yellow, respectively. C) Nucleus-invagination ratio is defined as the ratio of the non-convex area over the total nucleus surface area. Nuclei are larger and deformed in CD33+ (magenta, n=106) than CD38- and CD19+ cells (blue [n=120] and green [n=115], respectively; 3 independent experiments; Bars correspond to mean ± SD; ****: p<0.0001, Mann-Whitney test; n.s: non significant).
B BreastMark microarray platform; Kaplan-Meier curves of overall survival (OS) of ER- breast cancer. A log rank test was used to calculate P=0.4481 (n= 322, number of events= 130).
Quantification of fibroblast proliferation (left; n= 4 E16.5, P10; n= 3 E10.5, E18.5, P2, P21, P44, P50; n= 2 E17.5, P0 biological replicates) and collagen density (right; n= 4 E16.5, P2, P6, P10, P14, P16, P21, P50; n= 3 E10.5, E18.5, P2, P21, P44, P50 biological replicates) with age.
a-e, Typical autophagic structures in the tissues of an Atg5-/- embryo (E14.5): a, midbrain; b-d, liver; c, d, developing autophagosomes; e, heart. Arrows indicate the autophagic vacuoles.
(d) Peak to peak distance in auto-correlation plots of de-trended NGN2::mScarlet-I spatial profile in motorneuron (MN) and interneuron (IN) domains as a measure of spatial period in NGN2 expression along dorsal-ventral axis of spinal cord; Grey data points represent mean peak to peak distance in a single slice, n=33, left and right ventricle analysed separately in 4 experiments; black line shows overall mean, error bars show SD; 2-tailed Mann-Whitney test with exact p-value *** p=0.0003.
B. Western blot analysis of TFEB, and phospo-TFEB (Serine 142) in RCS chondrocytes stably expressing human TFEB-3XFlag protein. Cells were treated with vehicle (5% ABS) or FGF18 (50 ng/mL) for 16h. β-actin was used as loading control. Representative of three independent experiments.
(b) Confirmation of the expression of GFP-tagged human SAMHD1 (WT, and DN, DA, TA, and RE mutants). Histone H3 expression is shown as a loading control.
(E) Gene ontology analysis of common genes undergoing AS in sh1 and sh2 mouse ESCs according to the type of AS event.
(C) Ribosome-protected fragment (RPF) counts in each sample plotted along ife-1 and pat-10 CDS. Vertical grey lines indicate UUG codons. Data information: In , n = 3 biological replicates.
(F-H) Examples showing old (green) versus new (red) H3 distribution patterns and percentage of signal strength at a single genomic locus labeled with fluorescent probes (gray) for dad gene (F), bgcn gene (G), and ss gene (H) in bam mutant GSC-like cells at prometaphase marked by anti-H3S10p (blue). Scale bars: 1μm. Zoomed-in images of each probe are displayed to the right of each figure panel. Scale bars: 0.1μm.
Immunoblot after a full length JARID2 isoform-1 was expressed from an exogenous vector in HaCaT cells showed an increase (2-3 fold) in ~80kDa level.
(C) Co-immunoprecipitation of exogenous expressed Hsc70 with Htt-552 from PC12 cells. Upper panel: PC12 cell were transfected with PCDNA4-Hsc70 for 72 h. Cell lysates were subjected to immunoprecipitation with anti-Hsc70 antibody or no primary antibody and Western blot analysis was performed with anti-Htt antibody. Lower panels:PC12 cell were transfected with Htt-552 for 48 h. Cell lysates were subjected to immunoprecipitation with anti-Htt antibody or no primary antibody and Western blot analysis was performed with anti-Hsc70 antibody. Ab: Antibody.
(B) Box-and-Whisker plot displaying the sample specific expression of three FBW7 isoforms; FBW7α (NM_033632), FBW7β (NM_018315) and FBW7γ(NM_001013415'). Each box represents the expression distribution over three replicates.
The Bd1971 cNMP dimer (light/dark blue) situates its output domain (EAL) on the opposite side to that of CRP (DNA-binding subdomain, dimer coloured pink/red). Both Bd1971 and CRP dimerize via a central C-helix:C-helix interface. Close-up of hinge helix to C-helix crossover (circled). Tilt of the hinge helix relative to the C-helix allows Bd1971 to bridge from the cNMP domain body into the C-helix of the opposing monomer; the arrangement in CRP differs by pairing the C-helix with the cNMP body of the same monomer. Despite this difference, the respective C-helix dimers remain in the same orientation, as observed in panel (A). Overlay of the cAMP-binding pocket of Bd1971 and CRP (aligned on cAMP ligand, Bd1971 residue numbering given first), ligand-contacting sidechains shown in stick form, residues from the opposing monomer labeled by a '.
A. Amino acids of exon A. Positive charged amino acids are marked blue, negative charged ones are shown in red. Mutated regions are in bold and different mutant combinations investigated in Fig. EV5 are depicted below.
A Lentiviral knock‐down of THEMIS expression in primary humanCD4+T cells. Following transduction, cells were expanded on CD3/CD28 beads in the presence of exogenous IL‐2 and analysed for THEMIS KD by immunoblotting on day 4 post‐transduction. Data shown are representative of three independent experiments.
At day 26 (d26) the numbers of activated tumour infiltrating lymphocytes (TILs) were determined.
WT and Mfn2 KO cells were transfected with plasmids encoding mt-RFP and ChiMERA or control plasmid, and after 48 h they were fixed. N) Representative images. Scale bar= 5 μm. (P) aspect ratio and (Q) circularity were calculated (n= 300 mitochondria analyzed in 3 independent experiments). Data are presented as mean ± SEM.
B ACE2 competition experiments with selected VHHs from A. 50 nM RBD was mixed with indicated nanobodies (at 500 nM). Binding to ACE2 (immobilized on the sensors) was monitored by BLI.
(H) Measurement of Ki67+ cells (*p<0.05, ***p<0.001 compared to Apoe-/-; unpaired two-tailed Student's t test).
(E) Graphic depicting the participation of NAT in triggering mitohormesis in animal cells.
(F) Image of a ThP1 cell infected by HIV-1 with EU labeled RNA in absence (left) or presence (right) of nevirapine (NVP, 10 µM). Blue: DAPI staining. Green: vRNA (RNA-FISH). Grey: EU-labeled vRNA. See Appendix Fig. S19 for a larger image region. (G) Intensity profiles of EU and RNA-FISH along the dotted lines in panel F (top).
E. Representative TEM images of RCS chondrocytes treated with 5% ABS (vehicle) and FGF18 (50 ng/mL) for 16 hours. BafA1 (100 nM; 4 hours) was used to inhibit lysosome activity. Arrows indicate lysosomes. Higher magnification insets showed presence of ER membranes decorated with ribosomes (arrowheads). Scale bar 500nm. Quantification shows average number of ER membranes and mitochondria number/ lysosome vesicle (Lys). Mean +/- standard error of the mean (sem) of N=3 biological replicates/treatment. ER fragment/vesicle n=60 (vehicle) and n=72 (FGF18) cells were analyzed; Mitochondria number/vesicle: n=40 (vehicle) and n=72 (FGF18) cells were analyzed. Student paired T-Test ***p<0.0005; NS not significant.