EMBO/sd-character-level-ner huggingface dataset:
H Percentages of viability of Tsc2-dificient and wild-type MEFs exposed to DMEM 10% or 0.5% FBS with or without L-histidine for 72 hours. The asterisks indicate significant differences with one-sided t-test (10% FBS, P = 0.017; 0.5% FBS, P = 5x10-4; replicates/condition n = 4-6, independent experiments n = 2). The bars indicate mean ± SD.
Left panel, analysis of clonogenic growth of HCT116 cells assessed 15 days after transfection with non-targeting (siCTRL#1), VPS4A- or/and VPS4B-targeting siRNAs used in various duplex combinations as indicated. Right panel, images of HCT116 clones taken at the day of clonogenic growth assessment. Data are means of 3 independent experiments ±SEM. NT - non-transfected. All values were normalized, clonogenic growth of siCTRL#1-transfected cells was set as 1. One-sample t-test; ns - non-significant (p≥0.05), ****p<0.0001.
D Relative activity of BRE luciferase assay. 293T cells were depleted with serum overnight followed by a treatment with BMP2 for 6 hours. Results are shown as mean ± SEM; n=3. ***: p<0.001 by t test
(D) Wild-type, ULK1/2 dKO and IKKα KO MEFs were infected with Salmonella for 1 hour. Xenophagy rates were examined through Colony Forming Unit (CFU) assays. Quantification of infection rates by immunofluorescence is demonstrated in the right panel. Data information: Unless otherwise indicated experiments were performed three times. Data are represented as mean ± standard deviation and p values were determined by Student's T-Test.
(I) Multifunctionality analysis (IFN, TNF, IL-10) of cytokine+ T cells responding to MutuDC loaded with PbA pRBC.
F. YAP Ser 128 phosphorylation is required for disruption of YAP-14-3-3 interaction by osmotic stress. Flag-YAP WT or Flag-YAP S128A constructs were co-transfected with Myc-14-3-3 into HEK293A cells. Cells were serum starved for 16 hrs, and treated with 0.2 M sorbitol for the indicated time points or refreshed with serum containing medium for 1 hr. Refreshing medium served as a control to disrupt YAP and 14-3-3 binding.14-3-3 was immunoprecipitated with Myc antibody, and the associated YAP was detected with Flag antibody. Data are presented as mean ± SEM. *p < 0.05 (two tailed student's t-test, n = 3).
A) U2OS cells were grown on glass coverslips and synchronized in G2 after thymidine release. Cells were fixed at the indicated times after adriamycin-induced DNA damage and stained for DAPI and γH2AX. Scale bar indicates 10 μm. B) Quantification of cells in 3A. Each dot represents the total signal of γH2AX in the nucleus of a single cell. Bars indicate the mean and error bars indicate the SD of the data points in each condition. Whiskers represent 5-95% of data points. Statistical significance was tested using an unpaired two-tailed t-test (NS for P>0.05, **** for P≤0.0001).
(C) Top: PC3 cells treated with 100 μM NaHS for 30 min at 37°C. Cells were lyzed and treated with or without DTT for 10min at 4°C before subjecting to the modified biotin switch assay with the antibody against p65 to detect S-sulfhydration. Down: quantitative analysis of SSH-p65 protein level, and normalized with total p65 level Data information Data shown represent the means ± SD (n=3 biological replicates) ANOVA followed by Tukey's post-hoc tes was used for the statistical analysis (*P<0.05; **P<0.01; ***P<0.001)
(B) qRT-PCR analysis validates the altered Hippo-related genes in control and cKO mice. n=4.
(A) Acyl-RAC assay of HEK293T cells transfected with the indicated plasmids for 24 h. Data information: EtOH: ethanol; HAM: hydroxylamine; 2-BP: 2-bromopalmitate
D. Western blot of total protein extracts from control S2 cells and cells treated with dsRNAs against GFP or CG2968 (cV), probed for NDUFS3 (cI), ATP synthase subunit α (cV) and GAPDH (loading control).
B The Venn diagram depicts the number of common genes dysregulated by both shDDX3 and RK-33 treatments.
Activity dependent synaptic plasticity was investigated in cDKO after injection of AAV-Venus (red) or APPsβ (blue). F, G AAV-mediated overexpression of APPsß (n=23 slices, blue symbols) failed to rescue the LTP defect of cDKO mice (n=24 slices, red symbols). G TBS induced strengthening of fEPSPs resulted in similar potentiation levels for the last 5 minutes of recording (t75-80). The LTP induction rate is shown as percentage % of mean baseline slope. Data points were averaged over 6 time points. H PPF paradigm yielded no significant differences between viral vector injected cDKO mice. Age: 5-6 months. n=number of recorded slices. N=number of animals. Data represent mean ± SEM. Data were analyzed unpaired two-tailed Students t-test.
(B) Similar transfection as in (A) was performed, and transfected cells were processed for qRT-PCR analysis. The mRNA levels of lysosomal and autophagic genes were quantified and normalized relative to GAPDH. The data from three independent experiments are presented as means ± S.E.M., *, p<0.05; **, p<0.01, one-way ANOVA.
D Tail-clip assay reported as bleeding time in seconds. Significant differences between groups were assessed using One-way Anova test p value is = 0.11.Data are presented as mean±SEM. Each dot corresponds to a different animal. Knockout (K.O.) group: n=4; CodopV3: n=4; N6 intein: n=4; CodopN6 intein: n=
C, Flow cytometry analysis of phosphorylated histone H2AX (H2AX) at the indicated time points after IR (5 Gy, fold vs. non-irradiated cells, ctrl). *: one-way ANOVA, P=0.03.
A, B Quantification to the number of mitotic cells of dissected gonads from wild-type animals stained with either anti-phosphohistone H3 (PH3) (A), anti-CDK-1 (CDK-1) (B) transformed with either empty vector (con) or vector-expressing dsRNA against frh-1 (frh-1) or isp-1 (isp-1) and treated with gamma radiation 125 Gy. Quantification is expressed as fold changes of positive stained cells treated over the positive untreated cells Data information: Data are presented as mean ± SD (A, B N=2 (A, B * P < 0.05, ** P < 0.01, *** P < 0.001 and **** P < 0.0001 vs untreated; # P < 0.05, ### P < 0.001, ## P < 0.01, ### P < 0.001 and #### P < 0.0001 vs con. Not significant if nothing is specified. One-way ANOVA Tukey"s multiple comparisons test
H qRT-PCR analysis of total TERRA and polyA+ TERRA in cellular fractions as in (G). Values are expressed after normalization to total cellular extracts. Bars and error bars are averages and SD from 3 independent experiments.
Representative images of Olfm4 staining in small intestine sections from mice treated with or without APE1 inhibitor (CRT0044876, CRT). Arrows indicate Olfm4 positive intestine stem cells (I). Quantification of Olfm4 positive intestine stem cells (n=4 mice per group) (J).
Protein abundance correlation of affinity-purified complexes analyzed by mass spectrometry analysis applied to the proteasome complexes and their interacting proteinsHeat-map representing the correlations (expressed as the R2) between the abundances of 73 known proteasome-interacting proteins (PIPs) and the abundances of 8 reference proteins or protein complexes, PA28γ, β2i (representing the iP20S), PA28αβ, ncP20S (median of α1-α7, β3, β4, β6, and β7 profiles), 19S (median of Rpt1-6, Rpn1-3, 5-14 profiles), PI31, β5 (representing the sP20S), and PA200. For protein complexes, the median PAI of their subunits in each of the 24 AP-MS experiments was used: α1-α7, β3, β4, β6, and β7 subunits for the ncP20S, Rpt1-6, Rpn1-3, 5-14 for the 19S RP, and PA28α and PA28β subunits for the PA28αβ RP. The R2 values were hierarchically clustered. Three distinct clusters of composition detailed hereafter could be obtained. Cluster 1 (from top to bottom): Rpt3, Rpn13, α2, Rpn7, USP14, hHR23B, α1, β6, β3, α4, α7, Rpn6, Rpn3, Rpt4, Rpn10, Rpn5, Rpt5, Rpn1, Rpn11, Rpt1, Rpn9, Rpn2, Rpn8, α3, α6, β4, Rpt6, PDC6, Rpn12, APEH, Ubiquilin-1, α5, β7, CCT7, CCT4, CCT2, CCT3, CCT5, CCT6A, DNAJA1, HSP90AB1, HSP90AA1, PNP, Rpt2. Cluster 2 (from top to bottom): 14-3-3ζ/δ, CAND1, GSR, UBE3C, β2, ATP5A1, ATP5B, β1, PA200, β5, FBXO7, UCHL5, TXNL1, ECM29, PI31. Cluster 3 (from top to bottom): PA28β, PITH1, β2i, PA28α, PA28γ, β5i, β1i.Principal component analysis (PCA) of the abundances of 73 known PIPs. The circles represent the main clusters observed (iP20S, ncP20S/19S, sP20S and the 20S assembly chaperones).Plot of the R2 values between the iP20S or the sP20S and 193 protein correlating (R2 > 0.8) with the iP20S, the sP20S, or the ncP20S.
(A and B) WB analysis (A) and protein quantification (B) in WT (lane 1; blue), pex3 atg1 (lane 2; red), and pex3 atg1-PAOXPEX19 (lane 3; green) cells grown for 6 h on MM-M/G. The protein levels in WT were set to 100%.
(d) Subcellular fractionation and immunoblotting. Jurkat T cells were transduced to express ATG5-ΔNES, ATG5, or GFP. Although increased levels of ATG5 were observed in both cytoplasm and nucleus, ATG5-ΔNES accumulated following lentiviral gene transfer in the cytoplasm only. Full-length immunoblots are provided in Supplementary Fig. S11.
E) Cells generated in (D) were transfected with siCtrl or siADAR1. The type I IFN response and ADAR1 knockdown efficiency was monitored 78h post-transfection as in (B). Data are means ± s.d. from a representative of four biological replicate experiments.
(B) Gene Ontology (GO) analysis of the differentially expressed genes identified in the previous panel. Shown is a list of process networks with statistical significance of enrichment in Black HKCs, total or with < 20% genomic admixture (upper and lower panels, respectively). The complete list is provided in Dataset EV1.Fisher's Exact p-values are reported.
Automated microinjection of mouse neurons. Automated microinjection was performed on organotypic slices of mouse E16.5 12 wpc dorsal telencephalon using Dx-A555 (not shown) and LY (green, Slices were fixed after microinjection and stained for Reelin (white, C) Left: overview image of a mouse microinjected area (arrow indicates a pyramidal neuron, arrowhead indicates a Cajal-Retzius neuron; image is MIP of 20 focal planes). Microinjected neurons show different morphology and positivity for the Cajal-Retzius marker Reelin (white). Right, top row: the pyramidal neuron indicated with an arrow in the overview image is negative for Reelin. Right, bottom row: the Cajal-Retzius neuron indicated with an arrowhead in the overview image is positive for Reelin. Images on the top and bottom row left (LY MIP) are a MIP of 2 and 3 focal planes, respectively. The three images on the top and bottom right (LY, Reelin and DAPI) are single optical sections. Scale bar is 20 µm for MIP and 10 µm for single focal planes. VZ, ventricular zone; SVZ, subventricular zone; CP, cortical plate.
Western blot against CB1 and GAPDH (used as loading control) and graph showing mean ± SEM CB1 protein levels in cerebellar extracts of WT and ASM-KO mice (n = 3 mice per group).
GC-MS-based quantification of intracellular asparagine (Asn) and alanine (Ala) levels in MeWo and 1205Lu cells treated with LDHAi for 72 hr. Data information: Statistical analysis was performed by ordinary one-way Anova for the comparison of more than two groups. Data are shown as the mean ± SD, n = 3. ns: not significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001
C Quantification of collagen fibers using SHG microscopy in tumor sections from (A). Error bars represent mean ± s.d. of 4 independent fields per section, n=2 tumors per condition. ***P=0.0002, Mann-Whitney test.
C Percentage of indicated animals with different levels of SQST‐1::GFP aggregates. S: strong. M: medium. N: none. >30 animals were examined for each genotype.
Under the same growth conditions (permitted water, free food, and a 12 h/12 h dark/light cycle at 20±2°C), eight-week-old male C57BL/6 mice were continuously fed with methionine- and choline-supplemented (MCS) or methionine- and choline-deficient (MCD) diets for 6 weeks, and at the same time, gavaged with LicoB, MCC950, or a combination of LicoB and MCC950 (n=8 mice per group (B, C (A) Representative liver section images stained with H&E, Sirius Red, and Masson's trichrome stains. Scale bar: 0.5 cm (top row) and 200 μm (3 bottom ows). (B, C) The serum levels of ALT and AST (n=8 mice per group) were measured using ELISA.
(A) Humanretinal explant, prepared from the far periphery, infected with an AAV8 Y447 733F-CAG :ReaChR-GFP, placed on a multi-electrode array chip (left); GFP epifluorescence of the same explant following 12 days of incubation is shown on the right. Scale bars: 100 µm. The location of two cells that responded to repeated 2 s full field stimulations of orange light (600 nm) are highlighted.
Volcano plot of the RIP-seq data. Each point represents a gene. Red points correspond to the 146 genes with log2(IP/Input)≥0.5 and adjusted P<0.1. P-values computed using Wald test as implemented in DESeq2.
Jurkat T cells were treated with either DMSO (vehicle) or with 100μM of either Dieldrin or PMA for the indicated times. Abundance of mRNA for the indicated genes by RT-qPCR.
(g) Immunofluorescence for NeuN in the olfactory bulb (OB) and DG of Ctrl and FIP200GFAP cKO mice at P31. Lines indicate the layers of the OB and boundaries of the DG. Boxed areas in top panels are shown in more detail for staining of NeuN in insets. CC, corpus callosum; E, ependymal cells; LV, lateral ventricle; ML, molecular layer; ST, striatum; IGL, internal granular layer; MCL, mitral cell layer; EPL, external plexiform layer; GL, glomerular layer.
J. Quantitative analysis of the time constants for TNF-SBP-mCherry to reach the plasma membrane (Tauarrive PM) and the maximal change in fluorescence intensity relative to the initial fluorescence value (Intensity Change TIRF F/F0). Error bars represent SEM. Control: n=6-9; U18: n=8-9; U18+PF: n=5-9.
Western blot analysis of indicated proteins in both KPC (C) and Pan02 (D) cells following radiation and chemotherapy ± AG490, a JAK/Stat kinase inhibitor. Actin represents loading control.
M. Luciferase reporters driven by WT or mutant JBS4 were transfected into HEK293T cells with a JunD expression plasmid or with or without menin expressing plasmid. Luciferase assays were performed 36h after transfection. Three independent experimets (n=3).
(E) Representative immunoblots for 24-h endogenous LC3 lipidation. The asterisk stands for a nonspecific band. GAPDH, glyceraldehyde 3-phosphate dehydrogenase.
(B) Representative intravital image of calvarial bone marrow of Tcirg1EGFP/+ male mice treated with BTPDM1 showing osteoclasts (left, EGFP fluorescence) and 2PLIM image for pO2 changes (right, phosphorescence lifetime of BTPDM1). Scale bar, 20 μm.
(G) Quantification of Western blot bands for COL17A1 (full length) normalized to β actin.
(A) In wild-type cells, the disaccharide pentapeptide subunit linked to the undecaprenyl lipid transporter (Lipid II) is translocated to the outer leaflet of the cytoplasmic membrane by MurJ and polymerized by the glycosyltransferase and D,D-transpeptidase activities of the PBP2-RodA complex. MepM is essential and sufficient for insertion of new material in the PG net, although this endopeptidase can be replaced by overproduction of MepH, MepS, or PBP7. (B) Inhibition of PBP2 by β-lactams leads to the accumulation of uncross-linked glycan chains that are cleaved by the Slt70 lytic transglycosylase. This limits the supply of glycan chains for cross-linking by the YcbB L,D-transpeptidase. Under this condition, MepM or overproduction of MepS is required for insertion of new glycan strands. Production of YcbB leads to a requirement for an additional endopeptidase, MepK. This function of MepK can be bypassed by overproduction of MepS. (C) Deletion of the sltY gene encoding lytic transglycosylase Slt70 prevents digestion of uncross-linked glycan chains leading to a full supply of neo-synthesized glycan chains to YcbB and improved expression of β-lactam resistance. IM, inner membrane; OM, outer membrane; CRO, ceftriaxone.
B The RLuc-Mlx-5' construct was co-transfected in N2a cells together with plasmids expressing lnc-SMaRT (+) or with a control vector (-) and treated with control siRNA (+) or siRNA against DHX36 (-). Luciferase activity data, presented as the mean ± s.e.m. of three biological replicates (dots), are shown with respect to RLuc-Mlx-5' vector set to a value of 1. Statistical analysis was performed with ordinary analysis of variance (ANOVA) followed by Dunnett's multiple comparison test. **P < 0.01, ***P < 0.001.
(B) Slc6a20a-ASO decreases the levels of Slc6a20a mRNAs in the brain of WT and PtenΔC/ΔC mice (P20-32), as shown by qRT-PCR analysis performed 1-2 weeks after injection. (n = 5 mice for WT-Sal/saline, 4 for WT-ASO, 3 for ΔC-Sal, and 3 for ΔC-ASO, *P < 0.05, **P < 0.01, ***P < 0.001, Student's t-test for each comparison). The error bars represent SEM.
(A) The degradation of 3HA-Pgc1 was analyzed in cells with the indicated genotype as in Fig. 1A. The graph shows the average of three independent experiments; error bars represent the standard deviation.
(F) Maximal NMDAR mediated currents measured in the dLGN of P28~P34 mice, p = 0.03, N = 14 (Ctrl), 23 (CKO) cells from 5, 7 mice, respectively. Mean ± SEM.
(H) Western blotting for pMst1/2, Mst1, pMob1 and Mob1, pYap1 and Yap1 24 hours after the treatment. β-actin was used as a loading control.
Cumulative distributions of TE change for non-m6A and m6A transcripts as in (C). Kolmogorov-Smirnov test.
Expression of TARDBP (left), total SORT1 (center) and exon 17b SORT1 (right) mRNAs quantified by Q-PCR in human neurons derived from the indicated stem cells carrying either wild type Met337 (grey bars) or disease-associated Val337 (black bars) TARDBP alleles. Results are presented as average ± SEM (N=3 independent experiments; n=3 wells per condition in each experiment). *, p<0.05; **, p<0.01. Data information: Results are presented as average ± standard error of the mean (SEM). All statistical tests were performed using 2-way ANOVA with Sidak post hoc analysis.
Representative WBs of RPE-1 wild-type (WT) cells and the two selected KO clones for AKAP450 (akap KO), PCNT (pcnt KO) and CDK5Rap2 (c5rap2 KO) probed with the indicated antibodies. GM130 was used as a loading control
Population average of ERK activation dynamics cell‐averaged EKAR2G emission ratios (ERs) from n= at least 111 cells. StDev are shown.
(A) Overexpression of Wip1 down-regulates Smad4 phosphorylation at Thr277. MOCK- or Flag-Wip1-transfected HEK293T cells were treated with 1 h pulse of FGF2 (10 ng/ml), followed by addition of U0126 (40 μM) and harvesting for western blotting at the indicated times. - FGF2, no treatment with FGF2.
(C) LacZ staining (blue) to detect gene expression of the knocked-in LacZ cassette from the Abca12 locus in islets (islet outlined, scale bar = 25μm).
A Framework for investigation of effects of translation inhibition on UPF1LL activity.
Purvalanol B treatment reduced the NICD-FBXW7 interaction. 200 µg of HEK293 cell lysates treated with MLN4924 or MLN4924 in combination with 0.1 µM of Purvalanol B were subjected to immunoprecipitation using FBXW7 antibody, or IgG antibody as negative control, and precipitated material was analysed by western blot using NICD antibody. Western blot with FBXW7 antibody served as loading control for immunoprecipitation efficiency. 10% of cell lysate before immunoprecipitation was used as input control and β-Actin has been used as loading control.   Quantification of the density of western blot bands in (A) performed by ImageJ software. Data are expressed as percentage changes compared to MLN4924 treated samples. All data represent the mean ± SEM from three independent experiments. Student's t-test analysis was performed, with ***p≤0.001.
(F) Schematic of simplified σV circuit with only σV and RsiV, where σV (green) activates its own expression and that of its anti-sigma RsiV (orange, R).
hnRNPK IP enrichment ratios for genes with the indicated number of exonic hnRNPK eCLIP clusters in ENCODE data from HepG2 cells (average of the two ENCODE replicates). Enrichment ratios are log2-transformed fold changes computed by DESeq2 based on four replicates. The box plots show the interquartile range (IQR) and the line indicates the median value. Whisker ends extend to 1.5 times the length of the IQR (unless the data range is smaller, in which case the whisker ends extend to the minimum or maximum value), points indicate outliers. Spearman's correlation R and P-value are indicated.
MDTF cells were then challenged with MMLV-YFP vectors (MOI 1) for 48h. Flow cytometry dot-plots from a representative experiment are shown.
A. Table summarizing affinities (Kd, dissociation constant) of VP-containing peptides versus the COP1 WD40 domain (WT) and COP1Lys422Ala as determined by ITC. All values are μM (± standard deviation).
A Luciferase activity in 293T cells transfected with an ISRE (left) or IFN-β (right) promoter-driven luciferase reporter and together with plasmid encoding RIG-I (N) or cGAS and STING.
D, 293T cells transfected with a p53-specific siRNA, a USP7-specific siRNA, or a control siRNA (siCont.) were treated with 20 μM erastin or untreated for 24 h as indicated. Western blot analysis was then performed.
(B) Schematic representation of two different T6SS apparatus encoding gene clusters (named as T6SS-1 and T6SS-2) of NGJ1. The locus id (as per Burkholderia genome database) of upstream and downstream genes of the T6SS locus is provided. The gene disrupted in the respective T6SS mutants is marked by triangle (△).
(A) Experimental plan. Mice were subcutaneously injected with LLC cells. When the tumor reached 80-100mm3 size, mice were treated by gavage with glufosinate (10, 20 and 40 mg/kg) for 14 days.
J Plasma phospholipid in (H). n=7 for all groups. ***P<0.001 PLTP and PLTPM159E relative to GFP; *P<0.05 PLTP relative to PLTPM159E by ANOVA one-way followed by Fisher's LSD test.
O. Immunostaining of TH and human α-synuclein in SN. Scale bars equal 20 microns
, A549 cells were infected with HIV-luc/2019-nCoV pseudoviruses and then incubated with teicoplanin at various concentrations. The intracellular luciferase activity was measured at 48 h post-infection. The IC50 was calculated using GraphPad Prism software.
Heterofibrils containing mCHERRY-M451-90 and YPet‑RIPK1497‑583 C Heterofibrils containing mCHERRY-M451-90 and YPet-RIPK3387‑518 D Heterofibrils containing mCHERRY-M451-90 and YPet‑ZBP1170‑355 E Heterofibrils containing mCHERRY-RIPK1497‑583 and YPet-RIPK3387‑51
(G) The bars represent the mean running time and distance (± SEM) for 8-week-old male mice on a motorized treadmill. WT, n = 10; mdx, n = 13; mdx/499Tg, n = 8. *p = 0.01406 (Running time, vs. WT), ‡p = 0.005756 (Running time, vs. mdx). *p = 0.004558 (Running distance, vs. WT), ‡p = 0.001668 (Running distance, vs. mdx).
A. Protein phase separation can lead to the formation of protein assemblies with varying dynamics. Soluble proteins may phase separate into liquid-like droplets or insoluble deposits that according to their viscoelastic properties and their ability to exchange components with the cytoplasm vary from being dynamic to static.
(C) qRT-PCR analyses of the indicated genes were performed on RNA extracted from P1's primary or SV40T-transformed fibroblasts. cDNA expression levels were normalized to primary fibroblasts.
C HeLa cells were infected with S. flexneri M90T for 4 h 40 min, labelled with MitoTracker Red CMXRos, and fixed for quantitative confocal microscopy. Boxplots show the length of mitochondrial (μm; whiskers from min to max) surrounding Shigella outside (-) or inside (+) SEPT7 cages from 3 independent experiments (analysis of at least 130 measurements per biological replicate). Student's t-test; *** = p<0.001.
E, Representative confocal images of triple labeling for endogenous ORP2, NPC1-mCherry and integrin β1 after 2 h of LDL loading, and saponin permeabilization prior to fixation. Dashed line indicates cell edge.