EMBO/sd-character-level-ner huggingface dataset:
Oscillatory p53 leads to greater diversity of both mRNA and protein dynamic profiles compared to rising p53 (a-d) Induced p53 targets were subjected to fuzzy c-means clustering according to their mRNA expression profiles under (a) oscillatory or (c) rising p53 expression.Figure 2Induced p53 targets were subjected to fuzzy c-means clustering according to their mRNA expression profiles under (a) oscillatory Differential mRNA expression was defined as fold change > 1.5 and FDR < 0.2 (t test, Benjamini-Hochberg corrected) based on two independent experiments. mRNAs were clustered based on their normalized time traces (z score) into 5 expression clusters under oscillatory or rising p53. Only upregulated clusters are shown. The proteins corresponding to the mRNAs in each group were clustered according to their expression profiles. Differential protein expression was defined as fold change > 1.15 and FDR < 0.2 (t test, Benjamini-Hochberg corrected) based on two independent experiments. The mean expression of each cluster is shown in black. Induced p53 targets were subjected to fuzzy c-means clustering according to their mRNA expression profiles under rising p53 expression. Differential mRNA expression was defined as fold change > 1.5 and FDR < 0.2 (t test, Benjamini-Hochberg corrected) based on two independent experiments. mRNAs were clustered based on their normalized time traces (z score) into 5 expression clusters under oscillatory or rising p53. Only upregulated clusters are shown. The proteins corresponding to the mRNAs in each group were clustered according to their expression profiles. Differential protein expression was defined as fold change > 1.15 and FDR < 0.2 (t test, Benjamini-Hochberg corrected) based on two independent experiments. The mean expression of each cluster is shown in black. (e) Schematic showing all observed mRNA and protein dynamics under each p53 dynamical condition.
YAP signalling drives ectopic progenitor cell expansion in EML1-deficient organoids.Figure 4(A) Quantification of mitotic cells located at the basal side of VZ areas in control and EML1-deficient conditions (3 batches, 3 organoids analyzed per batch). Data information: Data in graphs are represented as means ± SD. Significance based on Kruskal-Wallis-test Dunn's post hoc test for multiple comparisons were performed to define statistical differences between genotypes. P-values: ***<0.001, **<0.01, *<0.05. (B) Quantification of BrdU+ KI67+ cells located at the basal side of VZ areas in control and EML1-deficient conditions (3 batches, 3 organoids analyzed per batch). Data information: Data in graphs are represented as means ± SD. Significance based on Kruskal-Wallis-test P-values: ***<0.001, **<0.01, *<0.05. (C) Normalized expression of YAP1, TEAD2, CCND3, and CYR61 in RG to bRG cells. Separate violins show expression in control and EML1-heKO. Asterisks indicate Bonferroni corrected p-values. Percentages indicate amount of cells in each group expressing the respective gene (3 pooled batches, 3 organoids each). Data information: Data in graphs are represented as means ± SD. Significance based on Wilcoxon Rank Sum test P-values: ***<0.001, **<0.01, *<0.05. (D, E) Immunofluorescence staining for (D) p-VIM and YAP1 or (E) Pax6 and YAP1 in control and EML1-heKO-derived organoids (day 20±2, squares indicate areas enlarged in adjacent part of the panel respectively). (F) Quantification of basally located pVim+ cells with nuclear YAP1 signal per VZ area in control and EML1-heKO-derived organoids (3 batches, 3 organoids analyzed per batch). (G) Quantification of PAX6+ cells with nuclear YAP1 signal per VZ area in control and EML1-heKO-derived organoids (3 batches, 3 organoids analyzed per batch). Data information: Data in graphs are represented as means ± SD. Significance based on Kruskal-Wallis-test Dunn's post hoc test for multiple comparisons were performed to define statistical differences between genotypes. P-values: ***<0.001, **<0.01, *<0.05. Scale bars: (D and E) 50µm, enlarged 10µm (H) Immunofluorescence staining for p-Vim in EML1-heKO and control derived organoids in DMSO, Verteporfin or Fluvastatin treated conditions, counterstained with DAPI. Dotted line indicates VZ areas. (I) Quantification of p-VIM+ cells located at the basal side of VZ areas in control or EML1-heKO-derived organoids under DMSO, Verteporfin or Fluvastatin treated condition (3 batches, 3 organoids analyzed per batch). Data information: Data in graphs are represented as means ± SD. Significance based on two-way ANOVA (on log10 normalized data) Tukey post hoc test for multiple comparisons were performed to define statistical differences between genotypes. P-values: ***<0.001, **<0.01, *<0.05. Scale bars: (H) 50µm (J) Quantification of ectopic neural rosettes per VZ area in EML1-heKO-derived organoids in DMSO, Verteporfin or Fluvastatin treated conditions (3 batches, 3 organoids analyzed per batch). Data information: Data in graphs are represented as means ± SD. Significance based on Kruskal-Wallis-test P-values: ***<0.001, **<0.01, *<0.05.
Figure 1Scheme of the targeting vector for intragenic deletion of the mouse Mex3a gene. The insertion of the Velocigene cassette ZEN-Ub1 created a deletion of 1125bp in exon 2 of the Mex3a locus. Representative images of the size of Mex3a mutant mice and control littermates at postnatal day (P) 15. Scale bar, 1 cm. Genotypes were confirmed by Mex3a mRNA ISH in intestinal tissue (right panels). Scale bars, 50 μm. The offspring number (n) and observed genotype frequencies (%) resulting from heterozygous crosses are indicated below. Absolute weight of Mex3a KO mice and control littermates at different ages. Data is represented in a box and whiskers plot as mean (middle line) with the minimum and maximum distribution values. Each point depicts one animal (WT: P1, n = 15; P8, n = 36; P15, n = 43; P22, n = 14; Adult, n = 14; Mex3a+/-: P1, n = 24; P8, n = 90; P15, n = 90; P22, n = 37; Adult, n = 21; Mex3a-/-: P1, n = 9; P8, n = 24; P15, n = 26; P22, n = 14; Adult, n = 8) ***P = 0.0003, ****P < 0.0001, two-way ANOVA test. Kaplan-Meier survival curves for the different Mex3a genotypes (n = 80 for each genotype) ****P < 0.0001, Log-rank (Mantel-Cox) test.
Figure 5A Topology and cytosolic C-terminus (210-284 residues) of DHHC2. Lower panel indicates the conserved cysteines (residues 255, 258, 260, and 262) of DHHC2 from six Plasmodium species. Western blot of episomally expressed DHHC2::6HA or variants containing truncated segment (C1:210-232, C2:233-262, C3:263-284 residues) in WT zygotes. C IFA of episomally expressed DHHC2::6HA and three truncation mutants in zygotes and retorts. Scale bar = 5 μm. D Acyl-RAC method detecting palmitoylation of endogenous DHHC2 in the dhhc2::6HA zygotes. E Palmitoylation of DHHC2::6HA and C4A mutant episomally expressed in zygotes. C4A: substitutions of C-terminal four cysteine to alanine. Right panel indicates the quantification of protein band intensity from two replicates, values are means ± SD. F Cytoplasmic distribution of C4A mutant during zygote to ookinete differentiation. Scale bar = 5 μm. G IFA of DHHC2 at the dhhc2::6HA zygotes treated with 100 μΜ 2-BP. Scale bar = 5 μm. H In vitro ookinete differentiation of the dhhc2kd parasites complemented with either WT DHHC2 or C4A mutant. Values are means ± SD from three independent tests. I Palmitoylation of the episomally expressed DHHC2::6HA with mutation in respective individual cysteine (C255A, C258A, C260A, and C262A). Representative of three independent repeats. J Palmitoylation of HA-tagged and human codon optimized DHHC2 ectopically expressed in human HEK293T cells. Both C4A and C128A mutations ablate DHHC2 palmitoylation. K DHHC2_C128A-HA mutant protein could be palmitoylated by the co-transfected DHHC2WT-Myc, but not DHHC2_C128A-Myc in HEK293T cells. Representative of two independent repeats. L Co-immunoprecipitation of the DHHC2_WT-HA and DHHC2_WT-Myc co-expressed in HEK293T cells. M C-terminus 75 residues peptide (Cter75) of DHHC2 could be palmitoylated as an independent substrate by co-transfected DHHC2WT-HA in HEK293T cells. Cter75 is fused with a BFP::Myc for easy detection via western blot. Cter75* peptide contains C4A mutation. N Palmitoylation of tagged DHHC2 ectopically expressed in bacteria E.coli. O Palmitoylation of DHHC2 ectopically expressed in bacteria treated with 2-BP. P Model showing self-palmitoylation of DHHC2 is catalyzed by its own PAT activity, which directs proteins for IMC targeting at zygotes.
Depletion of RBM14 induces ectopic formation of centriolar protein complexes.Figure 1Depletion of RBM14 induces ectopic formation of centriolar protein complexesA-E Mitotic control U2OS cells or U2OS cells treated with RBM14 siRNA were stained with antibodies against centrin-2 (green) and C-Nap1 (magenta) (A), Centrobin (green) and C-Nap1 (magenta) (B), centrin-2 (green) and CPAP (magenta) (C), acetylated-tubulin (green) and CP110 (magenta) (D), and centrin-2 (green) and γ-tubulin (magenta) (E). DNA is shown in blue. Insets show approximately twofold magnified views of fluorescent foci around the centrosome. Scale bar, 5 μm. Histograms on the right represent frequency of mitotic cells with excess foci of the indicated proteins at spindle poles in each condition. Values are mean percentages ± standard error of mean (SEM) from three independent experiments (n = 30 for each condition). *P < 0.05, **P < 0.01, n.s., not significant (one-tailed t-test). Note that we counted the number of C-Nap1 foci after anaphase in mitosis when C-Nap1 signals recover from the reduction in prometaphase.
Figure 6A, ift74 CRISPR mutants display defective cilia formation. Embryos at one cell stage were injected and fixed at stage 27 with two independent sgRNA targeting ift74. Representative confocal images of the Xenopus epidermis, and plots display defective ciliation. ;60 MCCs from 6 embryos Cell membranes (mGFP), cilia (Ac-tub, green) were labeled with indicated antibodies. Data information: Quantitative data from three independent experiments were scored. Unpaired two-tailed t-test was performed (*** p<0.001; ** p<0.01). Mean ± s.d. values are also presented. B. ift74 CRISPR mutants display defective cilia formation. Embryos at one cell stage were injected and fixed at stage 27 with two independent sgRNA targeting ift74. Representative 3D-SIM images of the Xenopus epidermis, and plots display defective ciliation. ;70 MCCs from 6 embryos for each condition. Cell membranes (mGFP), cilia (Ac-tub, green) and basal bodies (Cep164) were labeled with indicated antibodies. Data information: Quantitative data from three independent experiments were scored. Unpaired two-tailed t-test was performed (*** p<0.001; ** p<0.01). Mean ± s.d. values are also presented. C. Ift74 depletion affects centriole apical migration in MCCs. Representative 3D-SIM images (x-y) and 3D-reconstructions (x-z) of MCCs show failure of apical trafficking of basal bodies. Basal bodies (Cetn1, green) and distal appendages (Cep164, purple) were labeled with indicated antibodies. Quantitative plot shows a slight significant reduction in basal bodies by the depletion of Ift74. >70 MCCs from 6 embryos for each condition. D. Representative 3D-SIM images and plots reveal that apical expansion of MCCs is slightly affected by Ift74 depletion. >70 MCCs from 6 embryos for each condition. Embryos were fixed and stained with phalloidin (purple). Cilia (Ac-tub, gray) were also presented. Data information: Quantitative data from three independent experiments were scored. Unpaired two-tailed t-test was performed (*** p<0.001; ** p<0.01). Mean ± s.d. values are also presented. E. Representative confocal images and plot show a significant reduction in apical F-actin enrichment upon Ift74 depletion. >70 MCCs from 6 embryos for each condition. Cell boundaries and apical actin network were marked with mGFP (green) and phalloidin (purple). Data information: Quantitative data from three independent experiments were scored. Unpaired two-tailed t-test was performed (*** p<0.001; ** p<0.01). Mean ± s.d. values are also presented.
Figure 5A Immunoblot analysis of USP33 and HIF2α protein levels in T387 GSCs expressing USP33-targeting shRNAs (shUSP33) or non-targeting shRNA (shNT). Targeting USP33 by two distinct shRNA clones through lentiviral infection reduced USP33 and HIF2α expression in GSCs. B Representative images of tumorsphere formation of T387 GSCs expressing USP33-targeting shRNA (shUSP33) or non-targeting shRNA (shNT). Twenty-four hours after lentiviral infection, T387 GSCs expressing shUSP33 or shNT were planted in 96 well plates at the density of 2,000 cells per well and cultured for 48 hours under normoxia (20% O2) or hypoxia (5% O2). Disruption of USP33 inhibited GSC tumorsphere formation in normoxia and dramatically suppressed GSC tumorsphere formation in hypoxia. Scale bar, 200 μm. C, D Statistical quantifications of tumorsphere numbers (C) and sizes (D) showing the inhibition of tumorsphere formation after disruption of USP33 in T387 GSCs. Disruption of USP33 inhibited tumorsphere formation of GSCs in normoxia, whereas hypoxia further augmented the inhibitory effects of USP33 disruption on GSC tumorsphere formation. Data represent three independent experiments. The presented bars for tumorsphere numbers and sizes were based on 5 and 10 technical replicates, respectively. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (mean ± s.e.m.; two tailed unpaired t-test). E Cell titer assays showing cell growth of T387 GSCs expressing shUSP33 or shNT in normoxia or hypoxia. GSCs were transduced with shNT or shUSP33 through lentiviral infection for 24 hours, and then split into 96 well plates at the concentration of 2,000 cells per well. Cell titer was determined by the Glo luminescent cell viability assay kit (Promega) at the indicated time points. Disruption of USP33 inhibited GSC growth in normoxia and hypoxia. The experiment has been repeated three times. The presented curves were based on three technical replicates. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (mean ± s.e.m.; two tailed unpaired t-test). F Representative immunofluorescent staining of the apoptotic marker cleaved caspase 3 in T387 GSCs expressing shUSP33 or shNT in normoxia or hypoxia. GSCs were attached to hESC-Qualified Matrix (Corning) in 6-well plates at the concentration of 50,000 cells per well. Forty-eight hours post infection with shUSP33 or shNT lentiviruses, cells were stained for cleaved caspase 3 (green) and nuclei (blue). Scale bar, 80 μm. G Statistical quantification of (F) showing relative apoptosis of T387 GSCs expressing shUSP33 or shNT in normoxia or hypoxia. Disruption of USP33 increased the apoptosis of GSCs in normoxia, which was further elevated in hypoxia. Data represent three independent experiments. *, p < 0.05 (mean ± s.e.m.; two tailed unpaired t-test).
Figure 2Morphology of four-week-old soil-grown plants of Col-0, chs1-2 (A), chs2-1 (B) and two independent transgenic lines of SNIPER1 OE into chs1-2 (A) and chs2-1 (B) backgrounds. Plants were grown at 16 °C under long day conditions. Scale bar = 1 cm. Morphology of four-week-old soil-grown plants of Col-0, chs3-2D (C), snc2-1D eds5 npr1 (D) and two independent transgenic lines of SNIPER1 OE into chs3-2D (C) and snc2-1D eds5 npr1 (D) backgrounds. Scale bar = 1 cm.
is required for cold-induced mitophagy in mouse BAT (A,B) BATs isolated from Fundc1fl/fl/Ucp1cre- (Cre-) and Fundc1fl/fl/Ucp1cre+ (Cre+) mice at 30℃ or following 72 h at 4℃ were analyzed by transmission electron microscopy.Figure 5(A,B) BATs isolated from Fundc1fl/fl/Ucp1cre- (Cre-) and Fundc1fl/fl/Ucp1cre+ (Cre+) mice at 30℃ or following 72 h at 4℃ were analyzed by transmission electron microscopy. Representative images are shown, scale bar, 1μm (A). The number of mitophagosomes per 100 mitochondria were counted and normalized (B). (C,D).Representative fluorescence images of BAT sections from Fundc1fl/fl/Adiponectincre- (Cre-) and Fundc1fl/fl/Adiponectincre+ (Cre+) mito-Keima mice at normal conditions (30℃) or 4℃ for 72 h. Scale bar, 50 μm (C). Quantification of the ratio between red and green signals (D). (E) Fundcfl/fl/Ucp1cre- (Cre-) and Fundcfl/fl/Ucpcre+ (Cre+) mice were maintained at 30℃ (Normal) or exposed to 4℃ for 72 h (Cold). Mitochondrial DNA copy numbers were analyzed by null and normalized to nuclear DNA standards. (F) The relative total mito-keima signal (green signal ) in Fundc1 deficiency and control mito-keima mice under normal condition or cold exposure were quantified and plotted. (G) BATs isolated from Fundc1fl/fl/Ucp1cre- (Cre-) and Fundc1fl/fl/Ucp1cre+ (Cre+) at 30℃ or following 72 h at 4℃ were analyzed by transmission electron microscopy. The percent of mitochondria without cristae is shown in the histogram. Oxygen consumption rate (OCR) (H) were measured in BAT from Fundc1fl/fl/Ucp1cre- (Cre-) and Fundc1fl/fl/Ucpcre+ (Cre+) mice kept at 30℃ or at 4℃ for 72 h. mitochondrial superoxide (I) were measured in BAT from Fundc1fl/fl/Ucp1cre- (Cre-) and Fundc1fl/fl/Ucpcre+ (Cre+) mice kept at 30℃ or at 4℃ for 72 h.
Phenotypes of the double crosses between core network members.Figure 8The gain-of-function lines of all members of the core network were crossed either with each other, referred to as double cross, or with the control line (GFP-GR), referred to as single cross. The projected rosette area of the F1 double and single cross and the GFP-GR, germinated and grown on DEX, was measured at 22 DAS. A representative picture of the single and double crosses is depicted. The values represent the relative increase or decrease in rosette size of the double crosses compared to their parental single crosses. The outcome of every double cross is classified into three groups: additive (green connections), negative (blue connection) and synergistic (orange connections) phenotype. The precise measurements can be found in Fig EV2.
Figure 2Binding of Mz and 47F Fabs to the immobilized proteins indicated at left. Sensorgrams of eight different concentrations are shown in each panel. The calculated binding parameters are shown in the Table. RUs, response units.
Figure 2A. Scheme outlining the mechanism of Ig diversification in DT40 cells, indicating the action of uracil glycosylase inhibitor (UGI). UNG = Uracil DNA glycosylase. B. Diversification in wild type and h3.3 cells expressing similar levels of hAIDup and UGI. Red lollipops = transitions at C/T; Black lollipops = other point mutations; blue bars = gene conversions. C. Mutation rate as a function of hAIDup expression in UGI-expressing wild type and h3.3 cells. Each clone was expanded for 36 cell divisions and hAIDup expression determined by qPCR during the last week of culture.
Figure 1A) Box blot of vitrealANG-1 levels from newly diagnosed patients diagnosed with wAMD, DR, proliferative DR, and RVO compared to controls (macular whole).B) Box blot of vitrealANG-2 levels from patients as in A). The interquartile range of the data is indicated by the box. A non-parametric Kruskal-Wallis analysis (**, P = 0.029) followed by Dunn's method for multiple comparisons was used to show significant differences of the groups to control which are indicated by an asterisks. ANG-1 levels did not differ significantly, but ANG-2 levels were significantly different:control vs AMD (*, P = 0.0451), vs DR (*, P < 0.0001), pDR (*,P < 0.0001) and RVO (P < 0.0001). C) Schematic of cellular experiment to measure transendothelial resistance in human endothelial cells using transwell filters and CellZscope technology. D) 24-h concentration of ANG-2 in supernatants of the basal site of the culture stimulated with 5 ng/ml of VEGF-A, 200 ng/ml of ANG-1 or the combination. Error bars show SEM with one sided ANOVA (****, P < 0.0001) and Tukey's multiple T-test for five independent experiments indicating significance for control vs VEGF-A (**, P = 0.0028); VEGF-A vs ANG-1 (****, P <0.0001); ANG-1 vs VEGF-A ANG1 (**, P = 0.0061); VEGF-A vs VEGF-A ANG-1 (ns, P = 0.0625). E) Humanendothelial cells plated on filters were assessed for endothelial barrier function over time after addition of 5 ng/ml VEGF-A alone or in combination with 200 ng/ml ANG-1 or 10 μg/ml anti-ANG-2 or all three test items. The final time point was used for statistical analysis. Error bars show SEM with one sided ANOVA (****, P < 0.0001) and Tukey's multiple T-test for three independent experiments indicating significance of VEGF-A vs untreated (****, P < 0.0001); vs VEGF-A ANG-1 anti-ANG-2 (***, P = 0.0004); vs VEGF-A anti-ANG-2 (P = 0.038); vs VEGF-A ANG-1, (P = 0.040). Furthermore, VEGF-A anti-ANG-2 ANG-1 is significantly different vs VEGF-A ANG-1 (*, P = 0.0451) and VEGF-A anti-ANG-2 (*, P = 0.0474) and untreated is significantly different from vs VEGF-A ANG-1 (*, P = 0.0009) and VEGF-A anti-ANG-2 (*, P = 0.0008).
Figure 1HUVECs transfected with control or CEP41 siRNAs were scratched (0 h) to induce wounding and then incubated for 12 h to allow wound closure. The wound margins were observed every 4 h in the CEP41#1 and #2 siRNA-transfected cells and compared to those of control cells. Representative images of cells subjected to the wound closure assay in (A). Scale bars, 600 μm. Quantification of the extent of wound closure in (B) presented graphically by measuring the distance between the dotted lines at each time point. The siRNA-transfected HUVECs were plated inside a transwell chamber and incubated with serum for 18 h. The cells that invaded were observed after staining with crystal violet (CV) solution. Scale bars, 600 μm. The numbers of cells that invaded in each field of view was counted with the ImageJ software in (D). Tubulogenesis of control and CEP41-knockdown cells for 18 h was compared via an in vitro angiogenesis assay. Scale bars, 600 μm. Quantification of tube node numbers in (F) and tube length in (G) from each field of view using the ImageJ angiogenesis analyzer at the indicated time points. The graph compares the relative length of control cells and CEP41-knockdown cells. Data are shown as mean ± SD of five independent experiments with ≥ 5 tubulogenesis regions per condition.
Figure 2(A) Schematic representation of the arrangement and number of PPxPxY motifs present in the indicated host proteins (left panel). Multiple sequences aligned using ClustalW, of peptides containing PPxPxY motif derived from cellular proteins and from the EBOV NP protein (right panel). (B) Co-immunoprecipitation between HA-VP30 and either GFP or GFP-fused to peptides derived from the host proteins was performed. Western blots of the co-immunoprecipitation and whole-cell lysate (WCL) are shown. (C) Equilibrium dissociation curves of FITC-RBBP6 from eVP30130-272 in the presence of increasing concentrations (0.13-500 μM) of the PPxPxY containing peptides. Fluorescence polarization was determined with constant concentrations of FITC-RBBP6 and eVP30130-272, at 0.50 μM and 3.8 μM respectively. Experiments were performed in two independent replicates. Error bars represent standard deviation.
Figure 6(1) Under predivision conditions, SlmA·SBS may partially recruit FtsZ forming FtsZ·SlmA·SBS condensates, which may accumulate in transient microenvironments. Condensation into liquid droplets may aid in FtsZ·SlmA·SBS localization nearby the membrane. (2) Under division conditions, the amount of SlmA at midcell decreases and other elements would compete for the interaction with FtsZ, that would leave the condensates and, (3) in the presence of GTP, form a membrane anchored FtsZ ring. In non-central regions, FtsZ would still be under the control of SlmA, which protects the chromosomes from scission by aberrant division ring formation. Scheme of the self-association of FtsZ. In its GDP form, FtsZ is found as an ensemble of species of small size. GTP binding induces its assembly into fibers that disassemble upon depletion of the nucleotide by FtsZ GTPase activity.