EMBO/sd-character-level-ner huggingface dataset:
(B) Immunoblotting of CD4 T cell lysates treated as in (A) for 3 to 4 days. * indicates an unspecific band. (C) Quantification of immunoblots shown in (B) and additional donors (n= 9 for all but GSDMD, GSDMD n=8, for one donor quantification of the GSDMD band was considered unreliable due to uneven substrate distribution). Individual data points ± SEM are shown. Statistics indicate significance by one-way ANOVA of log2-transformed fold change values of band intensities (activated/non-activated T cells) normalized to β-Actin: ***p ≤ 0.001; *p ≤ 0.05; ns, not significant. P-values were corrected for multiple comparisons (Sidak). All band intensity ratios were tested against β-Actin. CARD8 full length was additionally tested against CARD8 cleaved.
(f) Expression analysis of TFEB target genes in liver of mice with indicated genotypes. Bar graphs show mean ± s.d. for n = 3 mice. *P≤0.05;**P≤0.01; ***P≤0.001.
B HTS. HEK293T cells were co-transfected with the constructs in (A) and, negative (DMSO, orange dots) and positive (CHX 5 µM, purple dots) controls added 3 h after reverse co-transfection with compound-libraries (5 µM). Images and data acquisition collected after about 30 h of treatment. Scatter plot shows distribution of compounds. On the Y-axis reported the Z-score values of cells expressing AUG-RFP and on the X-axis the Z-score values of cells expressing polyGP-GFP. Grid lines represent the thresholds arbitrarily set up around DMSO distribution (polyGP-GFP ±1.5 on the X-axis and ±1.5 for AUG-RFP on the Y-axis) to select compounds for the confirmatory screening and eliminate the ones without effect (orange square).
C) Cumulative frequency curve [%] of the observed coefficient of variation (CV) of proteins that were identified and quantified with a minimum of three valid values within each day. Here, the ten raw files of each day are evaluated individually. The resulting median and average CV for each day are shown.
A. Domain organization of IFT81, IFT74 and IFT22. Numbers refer to the T. brucei protein sequence and indicate different constructs used in this study. The part of the IFT81/74 sequence shown in shaded colors is not part of the construct used for structure determination. Coiled-coil boundaries are depicted based on the structure (ccI-ccVI) or prediction from the PCOILS webserver (cc). (CH = calponin homology, cc = coiled-coil).
(G) Predictions from the 0.782 cABN. Light green regions indicate where some, but not all, concrete networks allow stable conversion to the naïve state. Sall4 is indicated in green, as this was imposed as a constraint and therefore is not a model prediction
Analysis of TFEB binding to the VEGFR2 promoter in human ECs. ChIP was performed using digested chromatin from human control ECs and TFEBS142A ECs incubated with IgG (indicated in the bar graph as "+IgG") or with Ab anti-TFEB (indicated in the bar graph as "+Ab anti-TFEB"), followed by qPCR for VEGFR2.
(C and D) H&amp;amp;E staining of the hippocampus shows reduced pyramidal cell numbers in Epg5−/− mice. Means ± SEM of three mice are shown in B and D.
F Immunoblot of MAPK pathway-related proteins ERK1/2 and phosphorylated ERK1/2 in A549, H1299, and H3255 (RK-33 resistant) cells 24 h after treatment with RK-33 (7.5 μM or 10 μM). ERK2 and especially ERK1 become dephosphorylated after treatment with RK-33 in A549 and H1299 cells but not in H3255 cells. Outlined boxes indicate spliced lanes.
B FAK-expressing SCC cells were grown on glass coverslips, fixed and stained with anti-pSrc Y416 , anti-Paxillin and DAPI Representative immunofluorescence images are shown. Scale bars, 20 μm. The graph (B) shows the quantification of internalized active Src from three independent experiments. Student's t-test was carried out to calculate the statistical significance. Error bars, s.d. * = p < 0.01.
D Oncomine analysis showing median rank for genes in the PROSTATIC NEOPLASMS network across 16 analyses of overexpression in prostate cancer (PCa) versus normal. The right column represents fold change in gene expression detected by RNA‐seq in the LNCaP cell line after DHT treatment (Log2 FC DHT/Vehicle, P 0.01).
(F) FACS-derived expression of indicated surface and intracellular markers in miR-181a versus empty transduced (GFP+) and untransduced (GFP-) γδ T thymocytes, cultured with IL-7 plus IL-2 for 11 days (n=7-11 independent biological samples).
(G) Gene expression analysis in liver tissues from NAFL and NASH patients using GEO datasets (GSE48452 and GSE61260) from NCBI database. In the dataset GSE48452, n=17 for NAFL patients and n=15 for NASH patients. In the dataset GSE61260, n=21 for NAFL patients and n=24 for NASH patients. Data information: * P <0.05, ** P <0.01-. Results are shown as mean ± SD. Student's t test G).
(d) Cell lysates from stable PINK1-Flag-expressing SH-SY5Y cells were treated with 6-OHDA (120 μM), rotenone (1 μM) or FCCP (2 μM), and analysed by Flag immunoblot for PINK1-Flag levels.
B. Size Exclusion Chromatography (SEC) and SEC-MALS (SEC combined with Multi Angle Light scattering) analyses of the recombinant wild-type (wt) and dimer disrupting mutant (I31A) of spMis181-120. Measured molecular weights from SEC-MALS confirm that spMis181-120 wt is a dimer in solution and spMis181-120 I31A is a monomer.
D) Representative WB showing the effect of TRAIL stimulation on endogenous BID cleavage in HCT DKO cells. n=3 independent experiments.
Analysis of apoptotic cells at the indicated time points. SK-Mel-147 cells were treated with vehicle (V) or 0.5 µg/ml BO-110 (BO) for the indicated time points. Cells were collected and apoptosis was analyzed by flow cytometry as indicated in methods. Data correspond to the mean ± SD of 3 experiments. Statistical significance was determined by t-test.
(d) U2OS cells expressing GFP-FOXO3A were incubated with the indicated compounds for 2 h, fixed, stained with Hoechst 33342 and imaged by automated epifluorescence microscopy. The ratio of nuclear to cellular GFP fluorescence intensity was calculated and used to determine IC50 values for each compound. Data points represent values from three wells of one experiment.
F C-circle assays shows increased C-circle formation in ATRX-deleted U87-T cells at day 9 post control or Ad-Cre infection.
E Plasma membrane stained with an αSAG1 antibody is separated from the IMC stained with anti-Ty after aerolysin treatment, indicating that PKAr-3Ty localises at the IMC.
B Levels of β-catenin in response to HTL treatment in NE4C cells were detected. Band intensities were normalized to that of untreated cells. Data are presented as the mean ± SEM and were compared using an unpaired Student's t test. nsnot significant, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001. One-way ANOVA with Dunnett's correction was used for multiple comparisons.
Groups of BALB/c female mice (n = 5) were inoculated twice (four weeks apart) i.m. with 10 µg/dose of the proteins indicated at the top of each panel, folded in their respective postfusion conformations. One week after the last dose, mice were sacrificed and their blood collected. B) Mouse sera were also tested in microneutralization assays with hRSV and hMPV, as indicated at the bottom. The results represented in the y axis correspond to the inverse dilution that inhibited 50% of the viral infectivity (IC50 (log10)). P values: *P = 0.01001 and **P = 0.00271. Long horizontal lines in A and B indicate detection limits.
B-D, Averaged poststimulus time histograms ±SEM from OtofI515T/I515T mice (n=25-32) and Otof+/+ littermate SGNs (n=13-27) to stimulation with 50ms tone bursts at the characteristic frequency (CF) of each fiber, 30dB above threshold at indicated stimulus rates.
The percentage of cells where EYFP-Parkin translocates in response to A&O (both 10 µM, B is decreased in Miro1-EF-expressing PC6 cells. **P < 0.01, ****P < 0.0001 compared with the indicated group (n = 3-6 dishes, 20 fields per dish, one way ANOVA for
(a) Scheme illustrating de novo synthesis of polyamines from ornithine. * indicates the rate-limiting enzyme for de novo synthesis of polyamines. PMO, polyamine oxidase; SMO, spermine oxidase; SSAT, spermidine/spermine N(1) acetyltransferase.
A: Immunoblot analysis of wild type (WT), FAM83D-/- knockout (KO) and FAM83DGFP/GFP knockin (KI) U2OS cell lines. Data information: All blots are representative of at least 3 independent experiments.
(g) Quantification of posterior cell division outcome for apical and non-apical divisions. Outcomes are 2 epiblast daughter cells (epi/epi), one epiblast and one mesoderm daughter cells (epi/meso) or 2 mesoderm daughter cells (meso/meso).
(d) TEM of mitochondria in cultured neurospheres from Ctrl and FIP200GFAP cKO mice (n = 2 mice for each, >30 cells for each mouse). Arrows mark mitochondria.
(N) OCR in Sh3gl1-/- or WT B cells after overnight culture in CD40L, supplemented with 5 μM hemin chloride. N = 6 mice. P, two-way ANOVA with matched samples for basal and maximum OCR
(I) IF for βIII-tubulin (Tuj1) of control (Ccny+/-; Ccnyl1-/-; shRNA Co) and mutant (Ccny+/-; Ccnyl1-/-; shRNA Ccny) NPC monolayer cultures grown in differentiation medium for seven days. Insets depict neurons with representative neurite morphology for controls and mutants. Scale bar 20 μm. (J) Quantification of the percentage of Tuj1+ cells from (I). Data are means ± SEM (experiment performed twice in triplicates, representative experiment shown).
Both RQC and NGDRQC+ are triggered by a (CGA-CCG) dicodon containing arrest sequence in vivo. (C) Northern blot for the 5' NGD-IM derived from the (CGA-CCG) reporter in ski2∆ cells. 5'-NGD-IMs were detected with a DIG-labelled GFP probe. SCR1 was used as a load control.
A-B. Gene track of CDK7 occupancy at MITF (a) or SOX10 (b) loci in 501melBIO-FLAG:CDK7 cell line. Gene tracks of H2A.Z, BRG1, MITF, SOX10 and H3K27ac (GSE94488 and GSE61967) at the same loci in parental 501mel are indicated. SE is denoted by a red opened square. H3K27ac deposition is also shown in Hair Follicle Melanocytes (HFM) (GSE94488).
E. The bar diagram quantifies frequencies of SARS-CoV-2-positive cells in different brain organoid sections derived from two donor iPSC lines (IMR90 and Crx-iPS, see methods). Please note that each point represents one organoid section. SARS-CoV-2 shows an enhanced tropism for Day-60 organoids. Note, comparative statistics are shown between different age groups and respective days post infection (dpi) of organoids, and the significance is given as Asterisks in Day-60 groups. There is no significant difference in SARS-CoV-2-positive cells between 2- and 4-dpi within each age groups. At least twelve organoids sections from four (n=4) independent batches, from each donor and day post infections (dpi) were analyzed. One-way ANOVA, followed by Tukey's multiple comparisons test, ***P<0.001. Data presented as mean + SEM.
F. Box plots (the minimum, first quartile, median, third quartile, and maximum are shown, with data >1.5 interquartile ranges denoted with circles) for relative reporter activity (LUC/REN) in plants expressing different reportors and effectors. The LUC/REN in plants expressing ProFT alone is set to 1.0. Immunoblots were done to show GFP-CRY2W374A, Myc-CIB1 and Myc-CO protein levels in the transient Dual-LUC assay. Samples were separated on 10% SDS-PAGE, blotted, probed with the anti-Myc antibody, and with the anti-GFP antibody. Data information: the relative LUC activities normalized to the REN activity are shown (LUC/REN) with their standard deviation (n =6). The letters "a" to "f" indicate statistically significant differences determined by Tukey's HSD test (P≤0.05). Two biological replicates were performed.
NF-κB and JAK/STAT signalling drive innate immune responses. (A-C) Fold induction at 24 hpi of (A) CXCL10, (B) IFIT1 or (C) IL-6 mRNA (qRT-PCR) after Calu-3 infection with SARS-CoV-2 over a range of MOIs (0.004, 0.04, 0.4 TCID50VERO/cell) in the presence of 10 μM TPCA-1 or DMSO as shown.
(C) Overexpression of an p97/VCP ATP hydrolysis-defective mutant, E305Q/E578Q (p97QQ), blocked MITOL redistribution from mitochondria to peroxisomes, while overexpression of wild-type p97/VCP had no effect on MITOL redistribution. HeLa cells stably expressing GFP-Parkin were transfected with MITOL-HA and Flag-p97/VCP wild-type or p97QQ, treated with 15 µM CCCP for 3 hours, and then subjected to immunocytochemistry with anti-HA and anti-catalase antibodies. Higher magnification images of the boxed regions are shown in the small panel. Scale bars, 10 µm. Arrowheads indicate representative examples of MITOL-peroxisome co-localization that was only observed in the presence of a functional VCP.
(F to H) Comparison of standard differential expression and CNV approach for the three datasets shown in Figure 1-B:D. Volcano plots based on fold changes and adjusted p-values (limma) obtained for (F) lung vs. liver cells (Geiger et al, 2013)
(A-C) Immunostaining of TUNEL assay (red), active Caspase-3 (magenta, A'-C') and Repo (green, A"-C"). (A) GMR-wIR; repots>lacZ. (B) GMR-wIR; repots>Shits1. (C) GMR-wIR; repots>DERDN. GMR-wIR was used to reduce the autofluorescence of the eye pigments. Scale bar: 20 μm. (D-F) The in vivo fluorescent sensor of caspase activity (Apoliner) indicated there was no active caspase activity in the glia. (D) repots>Apoliner+lacZ. (E) repots>Apoliner+Shits1. (F) repots>Apoliner+DERDN. All adults were shifted to 28°C for 5 days. nls-GFP (green, D-F); merge of GFP (green) and mRFP (red) in (D'-F'); merge of nls-GFP and DAPI (magenta) in (D"-F"). Scale bar: 10 μm. (G) Percentage of vacuole area in the lamina in repots>Shits1flies that coexpressed the anti-apoptotic factors P35 and Diap1. The adults were shifted to 28°C for 21 days. The P-values were calculated using one-way ANOVA with Tukey's post-test. (H) The cell number of the epithelialglia was not reduced in the vacuolated lamina. The numbers of epithelial glia in the lamina of repo>H2B-RFP and repo>H2B-RFP+Shits1 female adults incubated at 28°C for 14 days were examined by counting the nuclear RFP at the epithelial layer from the entire Z-stacks of confocal images. P-values were calculated via Mann-Whitney tests.
C) Left. Representative image of coronal striatal slices of fresh frozen brains of wild-type and Tor1a+/- mice probed with the DRD2 radioligand 3H-spiperone. Scale bar 1.5 mm. (Right) Graph showing DRD2 binding density in Tor1a+/- striatal sections, obtained from the densitometric quantification analysis, expressed as percentage variation compared to control animals
(e) LC3 interacts with ERK in vivo. Immunoblots showing co-immunoprecipitation of LC3 with ERK, MEK and bRAF in homogenate (Hom) (e, top), and of LC3 with P- and total ERK in nuclear fractions from mouse livers (e, bottom).
E, Changes in ER calcium levels measured with an ER-targeted cameleon probe in serum-starved WT and Nox4KO MEFs after the addition of histamine (100 μmol/L, E) n=3/group (with >30 cells per individual experiment).
Luciferase-based NF-κB gene reporter assays using Myc -HOIP wildtype (WT), a catalytic inactive mutant C885A, ubiquitination site mutants of K454R, K458R, K735R, K784R co-transfected with HOIL-1L-HA and Flag-SHARPIN. Luciferase signal was normalized to an internal Renilla control signal.
B. ABRs from Wrbfl/fl:CreA mice showed a progressive threshold increase, Wrb+/+:CreA mice and Wrbfl/fl mice lacking Cre recombinase had normal thresholds. Statistical comparison of the threshold at 12 kHz was done by Kruskal-Wallis test, p values are from post-hoc Dunn's multiple comparison test. Measurements in which no ABR was observed at the maximal available tone burst level (90 dB) or click level (120 dB) scored as 100 dB threshold. Data are represented as mean ± SEM.
Automated microinjection was performed on organotypic slices of mouse E14.5 dorsal telencephalon using Dextran-A555 with slice culture for 24 or 48 h After fixation slices were stained for Tbr2 and TuJI. Fluorescence images of tissue after microinjection with dextran-A555 (magenta). The tissue was stained for Tbr2 (green) and TuJI (white). Microinjected cell at 0 h; injected cells show the typical bipolar morphology of an AP and are mainly negative for the BP marker Tbr2. Two-daughter cell progeny 24 h after microinjection; the top cell shows the bipolar morphology characteristic of an AP and is negative for Tbr2; the bottom cell is a newborn BP positive for Tbr2. Both cells are negative for the neuronal marker TuJI (white). Four-daughter cell progeny 48 h after microinjection; note that daughter 1 and 2 reside in the VZ, while daughter 3 and 4 are positive for Tbr2 and TuJI and reside in the SVZ. Distribution of microinjected cells and their progeny in the VZ, SVZ and CP (cortical plate). n = 147 cells total for 0 h, 151 cells total for 24 h, and 26 cells total for 48 h. Distribution of microinjected cells into 9 bins based on the distance from the ventricular surface (0 µm) and expressed as % of total. n = 147 cells total for 0 h, 151 cells total for 24 h, and 26 cells total for 48 h. VZ, ventricular zone; SVZ, subventricular zone; CP, cortical plate.
G. Eigen value showing the expression of the 3-microRNA signature in the hippocampus of 4 and 8 months old APPPS1-21 mice. The center line indicates the median, while the upper and lower lines represent the 75th and 25th percentiles, respectively. The whiskers represent the smallest and largest values in the 1.5× interquartile range. Number of mice: 5-6/group.
(F) hSgo1 RNAi and rescue with the indicated hSgo1 RNAi-resistant constructs was performed. Representative still images captured during the live cell imaging showing DIC and YFP-hSgo1 WT, 3A, and 4A localization during mitosis. Time (min) from nuclear envelop breakdown (NEBD) is indicated. Scale bar, 15μm.
cTAZ repressed the antiviral activity of ectopic STAT1/2. HEK293A were transfected with indicated plasmids, infected with gVSV (MOI = 0.01) for 12 hours, fixed, and subjected to microscopy (Left). Viral infection and replication were marked by green fluorescence. Scale bar, 100 μm. (Right) Quantification of GFP positive cells. About 300 cells (from 3 different fields) were analyzed. Error bars indicate SD. **P<0.01; ***P<0.001; Student's t-test.
F Density profiles in TDP-43 LCD condensates (peak at center) coexisting with dilute solutions for Wt, 12D, 5pS, 12pS, and 12A from coarse-grained simulations with the implicit solvent coarse-grained HPS model.
B-F: Immunoprecipitation (GFP-Trap) experiments between GFP-MOSPD2 (WT and RD/LD mutant) and Flag-tagge STARD11 (E Approximatively 5 µg of total protein extracts were analyzed by Western blot using anti-Flag (B, C-E anti-GFP (B-F), and anti-actin (B-F) antibodies Immunoprecipitated material was analyzed using anti-Flag (B, D-F and anti-GFP (B-F) antibodies
(E) NMDA treatment (25 min, 100 µM followed by 14h chase) caused 83.6±3.5% cell death in control neurons compared to 60.3±4.6% cell death in neurons with PRR7 expression knocked down. A c-Jun inhibitory peptide reduced NMDA-mediated cell death to 66.2±6.0% in control neurons and to 55.6±5.3% in neurons knocked down for PRR7, but this difference was not statistically significant (n=5 independent experiments). Two-way ANOVA was used in conjugation with Tukey post-hoc test. (Left) Representative images showing transduced neurons (green- GFP) and PI staining (red).
(C, D) Standard replication assays on undamaged (UD), thymine glycol (Tg), and cyclobutane pyrimidine dimer (CPD) templates in the presence (C) and absence (D) of Pol η, Pol ζ, Rev1, Rad6-Rad18, Uba1, and ubiquitin (TLS machinery). Reactions contained 2.5 nM Pol δ.
(B) Quantification of symmetric vs. asymmetric AP division in E13.5 control and DKO neocortices, expressed as percentage of total AP divisions. Four independent embryos per genotype from three litters were analysed. A total of 110 and 99 cells were counted for controls and DKOs, respectively. Data are means ± SEM.
Quantitative data showing the ratio of SA-β-gal positive hOSEs at their 4th passage with or without ectopic expression of LATS2.
(b) Absolute loadings for the genes with the largest absolute weights in the mRNA data. Plus or minus symbols on the right indicate the sign of the loading. Genes highlighted in orange were previously described as prognostic markers in CLL and associated with IGHV status (Maloum et al, 2009; Morabito et al, 2015; Plesingerova et al, 2017; Trojani et al, 2012; Vasconcelos et al, 2005)
RNase treatment of cells before crosslinking results in reduction of long-range interactions specifically at B-type compartments, and in the preservation TAD boundaries. On the other hand, transcriptional inhibition leads to significant weakening of TAD boundaries without drastic alterations in genomic compartmentalization.
A IF of E10.5 embryos showing OCT4+ (red) PGCs with cPARP (green). Arrows indicate apoptotic PGCs. Scale bar, 20 μm.B Quantification of apoptotic OCT4+PGCs in control and PCKO embryos at E10.5. Data are shown as mean ± SEM. Standard error is across visual fields containing 10 PGCs. In total, about 4-5 fields were used for the quantification for each genotype.
Quantification of the level of actin around the centrosome from (C), which shows the reduction in actin accumulation around centrosomes following CK666 pre-treatment. DMSO-STLC= 1 ± 0.02768, n=99, CK666 STLC= 0.776 ± 0.02186, n=87, DMSO-RO-3306= 1.339 ± 0.03048, n=127, CK666-RO-3306= 0.7699 ± 0.02246, n=70, one-way ANOVA, p<0.0001.
293T-FcR-Kb cells expressing Rab5ACA were incubated with opsonized OVA coated latex beads in the presence of varying doses of Epoxomicin. After 6hrs the cells were fixed, incubated with B3Z cells and IL-2 production was measured.
(a-c) HeLa cells were either left untreated, amino-acid (aa)/FBS starved for 5 h, or starved and then recovered in amino-acid/FBS-containing medium, then immunostained, or immunoblotted using antibodies as shown. Co-localization panels show an overlap between mTOR and LAMP1 signals. Note that changes in the positioning of lysosomal mTOR (quantified as the percentage of cells with predominantly peripheral localization of LAMP1-positive vesicles; a,c) correlate with mTORC1 activity (levels of phosphorylated S6K relative to the total S6K; b).
(B) Volcano plot representing the interactors that are enriched for the P182L mutant versus GFP as a negative control, obtained with affinity-enrichment mass spectrometry. Proteins that co-immunoprecipitated significantly more (p<0.05) with the P182L variant are shown in the upper right quadrant. Interactors with one or more [I/V]-x-[I/V] motifs are displayed in red while other significantly enriched interactors are displayed in black.
Clonogenic assay for ESCs in response to etoposide (A) or camptothecin (B). Non-treated cells were set to 1.
Thymocytes isolated from WT and Cd4cre A1fl/fl mice were stained with anti-CD4 and anti-CD8 antibodies and analyzed by flow cytometry. Representative images are shown (B).
C) HIF1α and BNIP3 protein expression were measured in control and Mfn2KD C2C12 myotubes untreated or treated with the HIF1α specific inhibitor NSC-134754 (n=3 independent experiments performed in triplicate). Data were normalized by tubulin as a loading control and expressed as a fold change compared to control C2C12 myotubes.
(a) Eyes from Pald1+/LacZ mice collected at P17 from untreated animals or animals with oxygen-induced retinopathy (OIR). Pald1-promoter driven LacZ expression and X-gal staining generated signals in capillaries, veins and arteries in the OIR retina at P17 compared to the normoxia control with predominantly arterial LacZ expression. Scale bar: 1 mm.
Differential expression of stemness- and differentiation-related genes of triple KD organoids grown in OCM vs. FTM. Fold-changes were derived from microarray data and indicate and an increase in stemness (CD133, SOX2, MYCN) and a drop in differentiation (OVGP1, PGR, FOXJ1) under OCM conditions. Differential expression determined by dual-color microarray for 2 biological replicates was significant for all genes with p < 0.05.