EMBO/sd-character-level-ner huggingface dataset:
C Fraction of OCRs having increasing, decreasing, steady and fluctuating accessibility pattern during T-cell development (x-axis). Average number of normalized reads in OCRs (y-axis).
H. Prazosin inhibits AKT phosphorylation in GICs. LY294002 (LY, 30 µM), an inhibitor of the PI3K/AKT pathway, was used as positive control. Terazosin, which does not affect GIC viability, does not alter AKT phosphorylation (see the corresponding cell viability counting in Fig EV4C). Phosphorylated AKT (P-AKT) and total AKT (AKT) expression analysis in GICs by immunoblotting. V: vehicle. kDa: kilodaltons.
B Sequence alignment of the finger loop and N-terminal part of β-strand VI in bovine β-arrestins, color coded as in A. Divergent positions are shown in bold.
Linear correlation plot of the percentage of IFN- γ + NK cells, the mean of fluorescence intensity (MFI) of pSTAT3 for MDSCs, and the expression of TRF2 (r2= 0,5177; Pearson's correlation, p = 0.003) (F) Data information: data are presented by Box plots Min to Max showing all points with median. *p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001; Mann-Whitney test. Pearson correlation and p values are indicated.
C. Bar plot showing the distribution of the number of transitions. Inset: Data from experiments carried out under nutrient-poor conditions in which growth rates are reduced.
Cideb depletion decreases the recruitment of SCAP to the COPII machinery in cells. Primary hepatocytes transfected with control or Cideb siRNA were infected with adenovirus expressing Sar1A-Flag-BirA*. Cells were cultured in sterol-depleted medium and treated with 80 µM biotin, prior to pulldown with streptavidin-conjugated beads. The biotinylated proteins were detected by IB with the indicated antibodies following SDS-PAGE.
(H) Analysis of the conversion of LC3B-I to LC3B-II and the levels of p62. WT and NDST3 KO RPE1 cells were cultured with or without 100 nM rapamycin (Rapa) for 6 h and LC3B-I, LC3B-II, p62, and GAPDH levels in the total cell lysates were determined by SDS-PAGE and immunoblotting. The ratios of LC3B-II to LC3B-I (n = 3 independent experiments, *P = 0.0451, **P = 0.0042) and the p62 levels normalized to β-tubulin (n = 3 independent experiments, *P(Rapa-) = 0.0158, *P(Rapa+) = 0.0183) are quantified. Data information: Error bars represent ± standard deviation. Scale bar, 10 μm.
(E,F) HepG2 were treated with 30 µM 3,4-DC for 8 h in the presence or absence of chloroquine (CQ, 50µM) for 4 h. SDS-PAGE and immunoblot were performed Band intensities of LC3-II and GAPDH were assessed and their ratio (LC3-II/GAPDH) was calculated in (F). Data are means ± SEM of at least three independent experiments (* = p < 0.05, ** = p < 0.01 versus untreated control; ##=p<0.01 versus CQ).
Flow cytometry of intracellular Foxp3 staining in CD4+ naïve T cells transduced with control retrovirus (MigRI) or retrovirus encoding C/EBPβ and cultured for 2 d under conditions as indicated. Representative experiments are shown in the left panel and pooled data fro 5 (D are shown on the right. Dot plots are gated for CD4+GFP+ and numbers indicate percent Foxp3+ cells in the gate
D) NGS plot showing H3K9/14ac comparative patterns in the experimental conditions described in B).
F Quantified MAO basal activity (Y-axis) in MEFs as depicted in the inset (grown in DMEM 10% FBS; replicates/condition n = 3 and independent experiments n = 2). The inhibitors (depicted on the X-axis) were added to cell extracts prior to activity assay to assess the contribution of each MAO isoform. The significant difference corresponds to two-way ANOVA test (P = 8x10-4). CPM: counts per minute. The bars indicate mean ± standard error of the mean (SEM).
(a-e). ATG5 is dispensable for PINK1-mediated mitochondrial fission. MEF cells derived from ATG5-null (ATG5KO) and their wildtype control mice (ATG5WT) were co-transfected Δ110-PINK1-GFP-FKBP /FRB-MTS with either PINK1WT or a kinase-dead mutant PINK1D384N. Cells were induced with 250 nM rapalog (Rapalog) for 2 hours to activate PINK1 kinase, followed by immunodetecting mitochondria (TOM20, red) and Δ110-PINK1-GFP-FKBP /FRB-MTS (PINK1-FKBP, green). Cells treated with solvent (DMSO) were used as a treatment control. Representative images are shown, scale bar=25 μm (a). Higher magnification images are also included (panels beneath the large cell images, scale bar=10 μm). Mitochondrial morphology in different transfections was quantified (b-e). Student's test. *p<0.05, ***p<0.001, ns: no significance. Data was presented as mean ± SEM of three independent experiments. For each condition, >100 cells were analyzed.
B Protein lysates of WT and USP19 KO A549 cells infected with SeV at indicated time points were immunoblotted with indicated antibodies.
D Expression of TAM family members and their knockdown by siRNA in cultured astrocytes. Tyro3, Axl and Metk in cultured astrocytes or hippocampal tissue were examined by western blotting. Arrows indicate the corresponding protein band(s). The secondary antibody against goat IgG generates a ~115 kDa non-specific band. The ~115 kDa band in the Axl image is the Axl-specific band overlapping the non-specific band. The ~115 kDa band in the Mertk image is the non-specific band. Axl and Mertk expression was knocked down by two siRNAs. GAPDH expression is shown as an internal control. E Quantification of siRNA knockdown in cultured astrocytes (n = 3 independent experiments, one-way ANOVA with repeated measures). Knockdown of Tyro3 mRNA was examined by qPCR because of the very low expression level even in control astrocytes. Knockdown efficiency of Axl and Mertk was quantified from western blots. Results are normalized to Gapdh or GAPDH, and are shown as ratios to the value of non-transfected astrocytes.
(B) LDH and IL-1B release from siRNA-treated primary human Monocyte-Derived Macrophages (hMDMs), primed with IFNγ (10UI/mL) and PAM3CSK4 (100ng/mL) and then stimulated with Nigericin (20µM) for 4 hours. When specified, the caspase-4/5 inhibitor Z-LEVD (25µM) or the NLRP3 inhibitor MCC950 (10µM) were added to the experiments.
Ni-NTA His-ubiquitin pulldown in control, FLAG-USP28 or FLAG-USP28 C171A transfected HEK293 cells, followed by immunoblot against exogenous ΔNp63. The input corresponds to 10% of the total protein amount used for the pulldown. Relative ubiquitination of the representative immunoblot was calculated using ACTIN for normalization.
(a) The Bcl-2 binding domain of Beclin1 is sufficient to bind vBcl-2. At 48 h posttransfection with wild-type and mutant Beclin1-V5, together with HA-vBcl-2, WCLs of 293T cells were used for immunoprecipitation with anti-V5 followed by immunoblotting with anti-HA.
(A) Surface calreticulin on MM cells was evaluated by flow cytometry after exposure to 800 nM BoxA or 1 μM MTX for 4 or 6 hours. Statistics: One-way Anova with Dunnett's post-test; n=3. Data information: columns represent the average and bars standard deviation of a representative experiment out of at least 2 performed. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.
The relative expression of each interferon-stimulated gene (ISG) indicated in the figure in sorted 4SP thymocytes isolated from Ifih1-/- and Cd4cre A1fl/fl Ifih1-/- mice. Values represent the relative gene expression normalized to GAPDH mRNA and are displayed as the mean ± SEM (n = 3; Mann-Whitney U-test, *p < 0.05). The mean value for the relative expression of each ISG in Ifih1-/- 4SP thymocytes was set as one.
LRRK2 WT or LRRK2 KO macrophages were pre-treated or not with 1 μM GSK2578215A and treated with 1 mM LLOMe for 30 min. The number of LRRK2 per cell were monitored by immunofluorescence and high-content imaging. Scale bar = 10 μm. Data represent the mean ± SEM of three to four independent biological experiments. One-way ANOVA followed by Dunnett's test against the untreated WT control. ns=non-significant; **p≤0.01.
(c) Apoptotic U937 cells expressing H2B-mCherry phagocytized by J774 macrophages expressing GFP-LC3. Confocal images show LC3 recruitment to an engulfed corpse (arrow; see Supplementary Movie S8). Scale bar, 15 μm. (d) Quantification of LC3 recruitment to apoptotic phagosomes in control and Atg5-shRNA-expressing cells; ***P0.0001, chi-squared test.
Bissected E10.5 mouse PSM explants were cultured in the absence (-) or presence (+) of 10 µM of RO-3306 for 4 hours and then analysed by in situ hybridization for mLfng mRNA expression. RO-3306 treated explant is two phases behind in the oscillation cycle of dynamic mLfng mRNA expression indicating there is a delay in the oscillation compared to the "-" explant. n=10. Scale bar is 100 µm.
(C) Representative Western blots, stained with the indicated antibodies of HeLa cells pre-treated o/n with an S-phase inhibitor (PHA 767491 hydrochloride, 10 μM) and lysed at indicated time points after UV (16 J/m2)
B. KV10.1 is also present in microsomal fractions. Cytoplasmic fractions obtained as in A were ultracentrifuged to sediment membranes. In those preparations, immunoprecipitation revealed the presence of full-length KV10.1 in both the centrosomal and the microsomal fractions (obtained by ultracentrifugation).
Distributions of piRNA precursor length in ints-11 RNAi-treated nematodes compared to empty vector-treated nematodes, obtained from deeply sequencing one pair of libraries (inserts sequenced up to 75nt). Distributions of total sequences (E) and total read counts (F) are shown.
(b,c) Mitochondrial (Mito) targeting of LC3B-II and C18-ceramide, generated by induction of wild-type CerS1 (b), and that of the products of the catalytically inactive mutant CerS1 (+ tet) (c) was visualized by colocalization of ceramide (Cer; green), CFP-LC3B (blue) and MTR using anti-ceramide and confocal microscopy. Noninduced cells (− tet) were used as controls. Scale bars, 10 μm.
Quantification of average fluorescence intensity of BRD7 of cells in (E). Error bars indicate SEM; n>100.
Representative confocal images of cell-type-specific promoter::GFP transcriptional fusions in hyl1 vs WT roots.
D Log2 fold changes in viable cell count in a panel of five IMT1-resistant cancer cell lines in presence of IMT1 + CAP (1 μg/ml). Data represent mean values of n=5 (resistant RKO cells) and n=3 (PANC1, Capan2, HCT-29 and Calu6) ±SD independent experiments. Paired t-test of IMT1 vs IMT1+CAP: resistant RKO cells **p=0,0014; PANC1, CAPAN2, HCT-29 and Calu-6: non-significant.
Graphs present pre-mRNA (B, C) assays of Krt86 using skin from mutant mice (Foxn1 null, Aff4 cko) or corresponding wild-type controls at P6 (B, C) All values are derived from 3 independent experiments and presented as mean ± S.D. Pre-mRNA measurements were analyzed statistically using two-way ANOVA with post-hoc Sidak's multiple comparison test at a 95% confidence interval.
(b-e) Glucose tolerance test (GTT) (b,d) and insulin tolerance test (ITT) (c,e) performed for WT, Nlrp3−/− (b,c) and Pycard−/− (d,e) mice on HFD for 12 weeks.
Micro-CT analyses of sham and OVX model of AdipoqCre; ranklfl/fl (Rankl-/-) and ranklfl/fl (Rankl+/+) mice (n=5 independent biological replicates). Scale bar=1 mm. Data were compared using an unpaired t‐test (** indicates P < 0.01), error bars are standard deviations.
Representative images of H&E stainings of tumors formed by SCC13 EV and Act cells (clone 2) at day 35 (asterisk indicates cartilage, arrow indicates site of invasion). An inset panel indicates the area where tumor cells invade into the cartilage. Tumor sections were evaluated for invasion through the basement membrane. N=6 (EV), N=6 Act (3 tumors from clone 1 and 3 tumors from clone 2). Tumors from Act mice were pooled in the graph).
B) Motif distribution in all DHSs ordered by increasing DNase-Seq tag count signal for CD4 TB relative to TN as in Fig 2B.
F-H HSF1WT or HSF1S121A plasmids were transfected into HSF1-deficient HEK293T cells. Transfected cells were heat-shocked at 43°C for 40 min. HSP mRNAs were quantitated by qRT-PCR following overnight recovery. HSF1-DNA binding was measured immediately after heat shock (mean ± SD, n = 3, Student's t-test, **P < 0.01, ***P < 0.001).
C Western blot results of PABPN1L in MII oocytes of wild type (WT) and Pabpn1l−/− females. DDB1 is blotted as a loading control.
LDH release assay from primary monocyte-derived macrophages (MDM) left untreated or treated with IFNγ, transfected with siRNA against GBP1 or non-targeting control (CTRL) and infected with indicated strain of Tg for 24 hours. Mean ± SEM of n = 4 donors shown.
H. Full-length FUNDC1-MYC, FUNDC1-MYC (AA 96-155), FL (Δ129-138) or vector control were transfected with Mito-DsRed (red) into FUNDC1 KD MEFs. 12 h post- transfection, cells were exposed to hypoxia for 12 h and then stained with anti-MYC (blue) and anti-DRP1 (green). Bar = 10 µm. Numbers at the right show the average length of mitochondria in Mito-DsRed-positive cells.
MOAP-1 deficiency promotes tumor initiation. Frequency of mice with macroscopic tumor nodules of the 4.5-month-old WT and MOAP-1 KO mice (n=18 and 23 respectively) injected with a single dose of DEN (25 µg/g body weight) at 15-day-old.
Bar graphs showing the genomic distribution of HMGB1 ChIP-seq peaks in HUVEC and IMR90. The number of peaks (N) analyzed per each cell type is indicated.
Excitatory GABAergic response in WT, but not in CAVII KO neurons. Picrotoxin blocked the biphasic response (PiTX 90 μM; n=3/genotype). The mean initial slope of the depolarizing shift was faster in WT than in CAVII KO (13.53±1.27 versus 6.39±0.96 mV/s, P<0.001, Student's t-test). The resting membrane potential of WT and CAVII KO neurons did not differ (−71.0±1.1 versus −72.3±1.4 mV, respectively; n=15 neurons for both genotypes, P=0.47, Student's t-test). *P<0.05, **P<0.01 and ***P<0.001. Error bars denote s.e.m. and n is the number of slices (C) or cells tested (one cell per slice) (A, B and D)
A. Confocal microscope images of ROCK1:ER-expressing cells co-stained for F-actin, MMP10 (left) or MMP13 (right) and DAPI following treatment with vehicle or 1 µM 4HT for 24 h. Multiple z-planes were used to generate x-z and y-z images. Scale bar = 5 µm.
A Gene set enrichment analysis (GSEA) of transcriptome changes in TFEB−injected vs. uninjected wild−type (WT + TFEB vs. WT) mice. GSEA of genes annotated as participating in the lysosomal function are reported. The upper panel shows the enrichment generated by GSEA of ranked gene expression data (left in upper panel and red in middle panel: upregulated; right in upper panel and blue in middle panel: downregulated). Peak value corresponds to the point (along the ranked microarray) where the enrichment score (ES) meets its highest value, while the zero value indicates the boundary (along the ranked microarray) between positive fold change and negative fold change. In the middle panel, vertical blue bars indicate the position of lysosomal genes within the ranked list. Lower panel shows the cumulative distribution of lysosomal genes within the ranked lists. The ranking positions that include 50% of lysosomal genes are indicated. The analysis shows that lysosomal genes have a significant global shift toward upregulated genes in TFEB−injected mice compared with uninjected littermates (ES = 0.56, P = 0.0029). n = 4 mice/group.
I. Interactions between Mfn1/2 and OPA1 occur independent of hFis1. WT 293T cells were treated with control or hFis1 siRNA, followed by in vivo crosslinking with 1% FA. Cell lysates were used for co-IP with Protein G beads bound to mouse normal IgG (negative control) or mouse anti-OPA1 antibody as indicated, followed by immunoblotting with indicated antibodies.
C. Time-averaged mean-square displacement (MSD) versus lag time (tlag) of 10-15 tracks from WT and IFT-kinesin mutant animals. Proteins used for tracking experiments are CHE-3 tagged with 7x GFP, CHE-3 tagged with 7x GFP plus DYF-11 tagged with 3x GFP, CHE-2 tagged with 7x GFP, or DYF-11 tagged with 3x GFP.
Western blots showing ASC-oligomerisation after DSS-mediated crosslinking in supernatants from THP1s treated as in A. Blots show membranes that were probed with an anti-ASC antibody. The input represents non-crosslinked supernatant. Data is representative of at least three independent experiments.
(E) MCF10A cells grown at high density on PDMS membranes and were stretched (or not) at 17% elongation for 2 hours and lysed while under tension. Anti-TRIP6 antibodies were used to isolate immune complexes. Immune complexes and lysates were probed by western blotting for LATS1 and TRIP6. Quantification is shown. (Mean ± SD; n=3; *P≤0.05, T-test).
(C) Genome edited XPO1 mutant HAP1 cells were transiently transfected with siRNA targeting endogenous STK38 (siSTK38#206 or with non-targeting siRNA (siNT)) at low or high confluence. 72 hours later, cells were fixed and stained for endogenous YAP1. Quantitative representation of YAP1 nuclear fluorescence intensity (n = 150 cells from 2 independent experiments). Here, phosphomimetic XPO1 (S1055D) potentiated YAP1 nuclear exit at both low and high confluences even in the absence of STK38 as compared to cells carrying wt XPO1
F. Frequency (in percentage) of MB-labeled pre-miR-181a-1 co-trafficked with CD63-eGFP-positive vesicles. Each data point corresponds to one axon. Total number of puncta and axons analyzed: 253 puncta, 22 axons (MB+), 306 puncta, 22 axons (CD63+). n=5 independent experiments. Values are mean ± SEM. Abbreviations: CD63, CD63-eGFP; MB, molecular beacon.
(J) Staining for DAPI in P3 WT and hFOXM1-cKI mice. Asterisks indicate gyrus and arrows indicate sulcus structures. Scale bars, 500μm .
E. HeLa cells were transfected with miR-586 mimics or inhibitors as indicated. Forty-eight hours after transfection, total RNA was extracted and subjected to real-time RT-PCR analysis. Data shown are mean ± SD (n = 3; **P < 0.01, two-tailed t-test).
(E and F) RAW264.7 cells transfected with miR-155 mimic (E) or inhibitor (F) was incubated with DMSO or bafilomycin A1 (BafA.) at a concentration of 100 nM for 2 h, and then LC3 levels were detected by Western-blot. The ratios of LC3-II/β-actin were calculated as shown below the representative blot. ***, p<0.001.
B Sensitivity analysis for integrated pS6 in mCFU-E, 32D-EpoR and BaF3-EpoR cells. Measured, cell-type-specific protein abundance and estimated global kinetic rates were taken into account. Proteins were grouped according to the network modules. Integrated pS6 exhibited high sensitivity towards RSK in 32D-EpoR and BaF3-EpoR cells but not in mCFU-E cells.
(B) Fluorescence time‐lapse images of F-actin bundling in an in vitro bundling assay. A mixture of unlabeled and Rhodamine labelled non-muscle actin was polymerized in the absence (PBS control, upper panel) or presence of mCA7 (lower panel). Numbers in images indicate the time after the onset of the experiment (0, 5 and 23 min). Intensity based Fire-coloring (Fiji) was used to visualize intensity changes. Scale bar 10 µm. (C) Quantification of the mean increase in filament length (n = 10 filaments at each time point) and the mean relative fluorescence intensity values of cross-sections for individual filaments /bundles (n = 30 - 31) in the absence and presence of mCA7 (1.12 µM). The data were analyzed using a general mixed model with time as a within-unit factor and the presence of CA7 as a between-unit factor. n = 3 independent repetitions, experiment repeats were included as a covariate and were non-significant.
E. Long-term potentiation in the mossy fiber pathway demonstrates no differences in long-term plasticity between control (black) and hTauAT expressing mice (red).
(A) Tau directly acetylates β-cat in test-tube. Both Tau40 and β-cat were purified from E. coli using Ni-NTA resin (n = 3 from three independent experiments).
Representative data of western blot for nuclear HDAC1 levels and semi-quantification of the expression levels. N = 6 mice per group. ** p = 0.0022 (Tg)/ 0.0012 (Tg+5104434) by multiple comparison. Data information: All data are presented as mean ± SEM (%)
EAE was induced by immunization of naive B6 mice with MOG (35-55), and the mice were randomly divided into five groups. BMDCs transduced with either lentiviral vector encoding GFP and NLRC3 (LV-NLRC3) or only GFP (LV-Ctrl) were administered i.v. 4 times, once every 4 days, starting at day 10 after EAE induction. Mean clinical scores of EAE (n= 5 mice per group ). Arrows indicate DC vaccine administration.
The mRNA profiling on shRNA-meditated Prom1 knockdown (3 different shRNAs were used, marked as NO.1, 2 and 3) in cultured CLESCs. qRT- PCR results are in arbitrary values after normalization for GapDH.
(a,b) Midguts dissected from either early third instar larvae (a) or at puparium formation (b) that express GFP-labelled mitochondria in enterocytes (larger nuclei) and contain Uba1H33 loss-of-function mutant cell clones (lacking RFP). Wild-type control cells possess stronger RFP and heterozygous cells have weaker RFP. Representative images are shown.
F. HumanHEK293T cells were transfected with plasmids expressing the GFP-TatA (A8L)-Flag and recoded S. pomberbd2. C-terminal cleavage products were purified using anti-Flag antibodies and purified protein was analyzed by mass spectrometry.
(D) The indicated plasmids of HA-tagged full-length NLRP3 and NLRP3 truncation mutants were respectively transfected into HEK293T cells along with Myc-tagged EphA2, and then cell lysates were immunoprecipitated with anti-Myc antibody, followed by western blot analysis with anti-HA antibody and anti-Myc antibody. Data information: The position of protein markers (shown in kDa) is indicated on the right-hand side. Data are representative of three independent biological replicates.
qRT-PCR for INHBA relative to RPL27 using RNA from SCC13 cells transduced with a lentiviral vector allowing expression of INHBA in a doxycycline (DOX)-inducible manner (SCC13 Act clone 1 and 2) or empty vector (EV) (N=3). SCC13 Act and EV cell lines were generated from lentivirally transduced cells upon single cell clonal expansion.
(b) Weight in adult male MSUS offspring and the offspring of males injected with MSUS serum compared to respective control groups. MSUS offspring n = 22, Control offspring n = 24, one-tailed Student's t-test (data reproduced) P = 0.045, t = 1.734, df = 44. MSUS serum-injected offspring n = 30, Control serum-injected offspring n = 31, two-tailed Mann-Whitney U = 334.5, P = 0.059.
A, B Ex vivo cell cultures from Ptch1+/−mice MBs were treated with GlaB (5 μM), GANT61 (10 μM) or DMSO only. After the indicated times, a trypan blue count was performed (A) to determine the growth rate of viable cells.
L. Skm iTRAQ ratio of FADH2-binding proteins (wt, n= 12; LowOXPHOS, n= 12). A higher intensity of red color represents a higher LowOXPHOS/wt expression ratio.
(H) Ratio of nuclear to somal AIF signal was measured as mean fluorescence intensity in the center of nuclei divided by the mean fluorescence intensity in the cell body, outlined by the NeuN staining (excluding the nucleus). The bar diagram summarizes the mean±s.e.m. of 282 and 262 neurons measured in n=19 and 20 image stacks in three independent experiments for aged SNOC and SNOC, respectively (P∼0.99 not significant by Student's t‐test).
Quantitative analysis of absolute splenic B‐cell numbers from Tam‐treated mb1‐CreERT2 (filled circles) or mb1‐CreERT2;Sykfl/fl (open circles). The animals were treated either with the isotype control, non‐blocking anti‐BAFF‐R antibody clone 5A12 (open squares) or the anti‐BAFF‐R‐blocking antibody clone 9B9 (open diamonds). Mice were analyzed 2 weeks after antibody injection. Each circle represents an individual mouse.
(A) TNF-induced mRNA expression in indicated L929 cell lines measured via qRT-PCR (mean of three independent experiments ± standard deviation, two-tailed Student's t-test *P<0.05, **P<0.01).
F. Quantification of area under curve for insulin tolerance test in E. Data represent mean ± SEM, Welch's t-test.
(B) TBC1D15−/− cells prepared as in (A) were subjected to immunoelectron microscopy with anti-GFP antibody. White arrowheads indicate YFP-LC3-labeled tubules. High magnification image is shown in the lower panel. Scale bars, 500 nm.
(B) HeLa cells were transiently transfected with GFP-ATG16L1 for 20 hr. Cells in basal conditions were fixed and labeled with anti-RAB11 antibody.