EMBO/sd-character-level-ner huggingface dataset:
Figure 2OptoCAR‑T cells were loaded with a ratiometric Ca2+indicator (Indo-1) before conjugation with ligand-presenting cells. Conjugated cells were gated on by flow cytometry and the Indo-1 ratio, as a readout of intracellular [Ca2+] was measured over time. Vehicle addition after 60 seconds caused no detectable change in this ratio. Bounded line shows mean ± SEM (n=5). An equivalent experiment was setup as in (A) but now 2 μM dimerizer was added after 60 seconds. Conjugated cells were maintained in the dark to maintain OptoCAR signaling. Bounded line shows mean ± SEM (n=3). An equivalent experiment was setup as in (B) but the conjugated cells were continuously illuminated throughout dimerizer addition to disrupt OptoCAR signaling. Bounded line shows mean ± SEM (n=3). An equivalent experiment was setup as in (B) but now conjugated cells were illuminated 3.5 minutes after dimerizer addition. Bounded line shows mean ± SEM (n=4). An equivalent experiment was setup as in (B) but now cells were illuminated 165 seconds after dimerizer addition, for 30 seconds. Bounded line shows mean ± SEM (n=3). Representative density plots of Ca2+ flux from conjugated OptoCAR‑T cells over time at 37ºC. Conjugated cells were stimulated with dimerizer addition for 0 (F), 5 (G), 10 (H), 15 (I) or 30 (J) minutes prior to disrupting signaling by illuminating cells. The decrease in intracellular [Ca2+] on illuminating conjugated cells after different intervals during activation was plotted as the mean Indo-1 ratio (n=3). The relative [Ca2+] was calculated by removing background Indo‑1 ratio before scaling to maximal output. Inset legend delineates the datasets. A single plot combining all datasets from (K) was fit by the survival function of a gamma distribution, with indicated half-life calculated from the time taken for the relative [Ca2+] to decrease to 50% of maximum.
Figure 3(A, B) Micrographs depicting the cellular composition in the SEZ 7 days after injection of aCSF (control, A) or AntimiR204 (B). Boxed areas correspond to higher magnifications in adjacent panels (A', B'). Dot plots depicting the abundance of LRCs (BrdU+, Ki67-, DCX-) in the SEZ 7 days after aCSF or AntimiR204 injection. Dot plots depicting the abundance of aNSCs and TAPs (BrdU-, Ki67+, DCX-) in the SEZ 7 days after aCSF or AntimiR204 injection. Dot plots depicting the abundance of neuroblasts (DCX+ Ki67+ or DCX+ Ki67-) in the SEZ 7 days after aCSF or AntimiR204 injection. (F, G) Micrographs depicting the elbow - the most ventral region of the RMS (F) and OB neuronal layers (G) upon injection of artificial CSF (control) or AntimiR204. (H) Dot plot showing the density of BrdU+ cells (progeny of NSCs) in the RMS and in the OB 7 days after artificial CSF or AntimiR204 injection.
Figure 2A . The chemical structure of KY-02061. B . A competition curve for the Dvl-CXXC5 interaction by KY-02061. C-E. NMR titration analyses for Dvl PZD domain with KY-02061. 1H-15N-HSQC analyses were performed to analyze the interaction of 15N-labelled Dvl-PDZ domain with KY-02061. The 1H-15N-HSQC spectrum of different molar ratios (Dvl PDZ domain:KY-02061) are displayed as red (1:0), orange (1:10), purple (1:20), cyan (1:40), green (1:60) and blue (1:80) (C, residues with meaningful chemical shift change are indicated by arrows). Plot of chemical shift changes (Δδ) as a function of residue number in molecular ratio 1:80 (D, a red-colored line indicates the line for Δδ=0.05). The residues with Δδ greater than 0.05 are visualized as a stick model on the ribbon representation of the Dvl PDZ domain structure (E). F. Molecular docking of Dvl binding motif (DBM) or KY-02061 to Dvl PDZ domain were analyzed by in silico experiments. The superimposed structure of DBM (green) and KY-02061 (yellow) on the surface of Dvl PDZ domain (gray) was visualized.
a) Treatment with CCCP (10 μM) results in an increase in LC3-II in the mitochondrial but not the cytosolic fraction of cells stably expressing the empty shRNA vector (control), and this is delayed in stable Fbxo7 knockdown (KD) SH-SY5Y cells.figf7(a) Treatment with CCCP (10 μM) results in an increase in LC3-II in the mitochondrial but not the cytosolic fraction of cells stably expressing the empty shRNA vector (control), and this is delayed in stable Fbxo7 knockdown (KD) SH-SY5Y cells. (b) As in a, an accumulation of LC3-II was observed in the mitochondrial fraction of healthy control fibroblasts following 1 and 5 h CCCP treatment, but this was reduced in fibroblasts from a patient carrying the R378G mutation. Western blots were performed a minimum of three times. (c) Mitochondrial accumulation of p62 following CCCP treatment is inhibited by Fbxo7 siRNA. Flag-Parkin overexpressing SH-SY5Y cells were transfected with scrambled (scr) or Fbxo7 siRNA as indicated and treated with either DMSO or CCCP (10 μM) for 6 h. Colocalization of p62 with HtrA2, a mitochondrial marker, was assessed by Pearson's correlation coefficient (Rr) on a cell-by-cell basis. Histogram shows the percentage of cells in which Rr was greater than 0.5. Data are represented as mean ± s.e.m., *P 0.05. Scale bars, 10 μm. (d) Mitophagy was analyzed in untransfected (UT) SH-SY5Y cells or in stable Flag-Parkin overexpressing SH-SY5Y cells transfected with either scrambled (scr) or Fbxo7 siRNA. Histogram indicates the percentage of cells with no remaining mitochondria following 24 h treatment with CCCP (10 μM) for each condition. Complex Vβ subunit (CxVβ) was used as a mitochondrial marker. Data are presented as mean ± s.e.m., **P 0.01. Scale bars, 10 μm. (e) Mitochondrial mass was measured in Flag-Parkin overexpressing SH-SY5Y cells transfected with scrambled (scr) or Fbxo7 siRNA and treated for 24 h with either dimethylsulfoxide (DMSO) vehicle or CCCP (10 μM). For representative images, see Supplementary Figure 8d. Full-length blots are presented in Supplementary Figure 9.
FXR1 protein localizes to the NE and interacts with NUPs.Figure 1A Lysates of HeLa cells stably expressing GFP alone or GFP-FXR1 were subjected to immunoprecipitation using GFP-Trap beads (GFP-IP), analysed by Western blot and quantified (shown a mean value, *P < 0.05, **P < 0.01; N = 3). B Lysates of HeLa cells stably expressing GFP alone or 3xGFP-Nup85 were immunoprecipitated using GFP-Trap beads (GFP-IP), analysed by Western blot and quantified (SE, short exposure, LE, long exposure) (shown a mean value, *P < 0.05; N = 3). C Immunoprecipitation from HEK293T cell lysates using FXR1 antibody or IgG analysed by Western blot. The arrow points to the heavy chain of IgG (IgG HC) (shown a mean value, *P < 0.05; N = 3). D HeLa cells were treated with indicated siRNAs, synchronized by double thymidine block, and released for 12 hours and analysed by immunofluorescence microscopy for the lamin B receptor (LBR) to label the NE, and FXR1. E HeLa cells stably expressing GFP-FXR1 were analysed by immunofluorescence microscopy for GFP and mAb414, which labels FG-Nups. The magnified framed regions are shown in the corresponding numbered panels. The arrowheads indicate NE and cytoplasmic localization of GFP-FXR1. F HeLa cells stably expressing GFP-Nup107 were synchronized by double thymidine block and released for 12 hours, permeabilized with Triton/SDS or digitonin for antibodies to access the nuclear and cytoplasmic or cytoplamsic side of the nucleus, respectively, and analysed by immunofluorescence microscopy.
Fig.Figure 4Dendrogram of the Arabidopsis SnRK2 protein kinases belonging to three subgroups. Expression of the SnRK2s in yeast revealed ABF2-mediated transactivation by SnRK2.1, SnRK2.4, SnRK2.5, and SnRK2.10 in addition to OST1. Inset: Immuno-detection of FLAG-tagged SnRK2s expressed in yeast. The data in (B) are normalized to control samples expressing ABF2 but no SnRK2 The active subgroup I members together with SnRK2.2 and SnRK2.3 were analyzed for differential activation of ABFs. The OST1 data shown in Fig. 1A is included for better comparison. The induction level of ABF4 relative to the empty vector control (ev) is indicated. The OST1 domain structure comprises the kinase domain (KD) containing the activation loop and the two carboxyterminal regulatory domains DI and DII/ABA-box. Potential phosphorylation sites within the activation loop are underlined. Analysis of the phosphorylation status in the activation loop of SnRK2s expressed in yeast. Phosphorylation occurred at serine 175 or threonine 176, and at threonine 179. The relative abundance is given as the ratio of signal intensity of phosphorylated versus non-phosphorylated peptide for OST1, SnRK2.3, and SnRK2.4 in the mass spectrometric analysis. The numbering of phosphosites corresponds to OST1. Inhibition of OST1 and SnRK2.4 by clade A PP2Cs. The data in (G) to cells expressing inactive ABI1D177A instead of ABI1.
Lactobacillus acidophilus NCK2187 and its SlpA do not prevent immune infiltration and activation during DSS‐induced colitis in Signr3−/− miceSignr3+/+ (WT) or Signr3−/− (KO) mice were orally gavaged with NCK56, NCK2187, or SlpA on days −1 and −3, and 3% DSS was given in the drinking water.Figure 7Signr3+/+ (WT) or Signr3−/− (KO) mice were orally gavaged with NCK56, NCK2187, or SlpA on days −1 and −3, and 3% DSS was given in the drinking water. Mice were gavaged with bacteria or purified SlpA every other day for an additional three times, and immunity was analyzed by flow cytometry at day 10.A Representative plots indicate the frequency of neutrophils in the colons of untreated or DSS‐treated WT (left) and KO mice (right). Empty bars = WT; lined bars = KO; white bars = untreated; purple bars = DSS; red bars = DSS + NCK56; green bars = DSS + NCK2187; blue bars = DSS + SlpA. Signr3+/+ (WT) or Signr3−/− (KO) mice were orally gavaged with NCK56, NCK2187, or SlpA on days −1 and −3, and 3% DSS was given in the drinking water. Mice were gavaged with bacteria or purified SlpA every other day for an additional three times, and immunity was analyzed by flow cytometry at day 10.B, C Colonic DCs and macrophages (MΦs) were analyzed by flow cytometry for the production of IL‐1β (B), and colonic FoxP3+ Tregs were evaluated for co‐expression of RORγt+ (C). n = 5 mice/group. Gray tinted line = isotype control; black = untreated; purple = DSS; red = DSS + NCK56; green = DSS + NCK2187; blue = DSS + SlpA.
Figure 5Scheme of the encapsulation procedure followed for the PEG/DNA LLPS system and illustration, on the right, of the distribution of species within the encapsulated LLPS system. B,C. Representative confocal images of the microdroplets stabilized by the E. coli lipid mixture containing the biphasic PEG/DNA mixture and the FtsZ·SlmA·SBS complex without and with 2 mM GTP, respectively. Last image in (B) focuses in the lipid interface to show the high density of condensates. Scale bars: 20 μm except in images on the far right (10 μm), which are an independent image at higher magnification (B) or a magnification of the indicated region in the merged image (C).
H19 promotes Igf1r expression in endometrial stromal cells by inhibiting let-7.Figure 2A-D Endometrial stromal cells from patients #166 and #80 were each transfected with a mixture of siCon (control siRNA) and microRNA inhibitor control (iCon), siH19 and iCon, or siH19 and iLet7 (let-7-specific inhibitor). RNA and proteins were analyzed at 48 h post-transfection. Combined results from the two patient cells are presented. A-D Endometrial stromal cells from patients #166 and #80 were each transfected with a mixture of siCon (control siRNA) and microRNA inhibitor control (iCon), siH19 and iCon, or siH19 and iLet7 (let-7-specific inhibitor). Western blot gels from #166 cells are shown in (C). Quantitation of Western blots combining both patient cells are shown in (D). E-H Endometrial stromal cells from patients #98 and #212S were each transfected with empty vector or pH19. RNA and proteins were analyzed 48 h post-transfection. Combined results from the two patient cells are presented. E-H Endometrial stromal cells from patients #98 and #212S were each transfected with empty vector or pH19. RNA and proteins were analyzed 48 h post-transfection. Combined results from the two patient cells are presented. Western blot gels from #98 cells are shown in (G). Quantitation of Western blots combining both patient cells are shown in (H).
Figure 3(A) Visualization of the overlap of the top 10% MTA-predicted target reactions between each pair of datasets analyzed using Fisher's exact tests (Methods). The dot size corresponds to the effect size of the overlap as measured by odds ratio, and the color corresponds to the negative log10 adjusted one-sided P value (grey means below 0.05). (B) A summary visualization of the enrichment of the top 10% MTA-predicted targets from each dataset for the antiviral hits (i.e. those whose KO inhibits SARS-CoV-2 infection) identified in the two published CRISPR-Cas9 screens (Wei et al. 2021 and Daniloski et al. 2021; Methods). Y-axis represents the odds ratio of enrichment, the dot color corresponds to the negative log10 adjusted one-sided P value from Fisher's exact tests, and dot size corresponds to the number of enriched target reactions. The datasets are ordered by P values, the first 8 datasets have FDR<0.1. (C) cases of significant enrichment (FDR<0.1) of top 10% MTA-predicted targets from each dataset for the experimentally validated anti-SARS-CoV-2 drug sets from previous studies (compiled by Kuleshov et al. 2020). "Union" means the union of all drug sets. Axes and the meanings of dot color and size are similar to (B) but the axes are flipped and the adjusted P values are not log-transformed. (D) significant cases of significant enrichment (FDR<0.1) of top 10% MTA-predicted targets from each dataset for the host proteins involved in host-SARS-CoV-2 protein-protein interactions (combined from Stukalov et al. 2020 and Gordon et al. 2020). Axes and the meanings of dot color and size are similar to (C). (E) A summary of the area under ROC curve (AUROC) value of the MTA prediction using each dataset, based on positive and negative sets (i.e. genes whose KO inhibits or promotes the viral infection, respectively) from the two published CRISPR-Cas9 screen data (Wei et al. 2021 and Daniloski et al. 2021; Methods). The error bars (vertical lines through the dots) represent 95% confidence intervals obtained with bootstrapping. The horizontal dashed line corresponds to AUROC of 0.5 (i.e. performance of a random predictor). (F) Example ROC curves from two of the best-performing datasets, one in vitro (Vero) and one in vivo (SC.Liao). The curves are colored by color gradients corresponding to the threshold of top MTA predictions. (G) Summary of the metabolic pathways significantly enriched (FDR<0.1) by the final list of consensus candidate targets identified across datasets (Methods). The axes and the meanings of dot color and size are similar to (B) except for that the axes are flipped.
Fig.Figure 6(A) CASP1-GFP and lignin deposition in WT, sgn3, rbohD, rbohF and rbohDF. CASP1-GFP and lignin (fuchsin) are presented in green and magenta respectively. Pictures were obtained from more than 10 roots from each background with similar results. Scale bar = 10 µm. (B) Time lapse imaging of single- or cotreatment of 10 nM CIF2 with 25 µM cycloheximide (CHX) on CASP1-GFP in cif1 cif2. Seedlings were pretreated with or without CHX for 30 min and transferred onto each medium. Scale bar = 10 µm (See also movie EV1). (C) Quantification of (B). Relative numbers of holes in CASP1-GFP domain after single- or co-treatment with CIF2 or CHX from the pictures in (B). Bars are S.D. * indicates statistical significance from all other conditions (p<0.01) after one-way ANOVA and Tukey test. 6 roots in total for each condition were observed during two independent tests. (D) Fold-change of 930 genes (p<0.05 and log2(Fold change) ≥1 or ≤ -1 at least one time point in one genotype) after CIF2 treatment at indicated time points in WT, cif1,2 and sgn3. Degree of the fold changes are shown in color code as indicated. (E) Relative expression levels of PER15 to CLATHRIN control in each genotype with or without 2-hour CIF2 treatment. Bars are S.D. (n=3). Different characters indicate statistically significant differences (p < 0.01, ANOVA and Tukey test). (F) Fold changes of PER15 in each genotype with or without 2-hour CIF2 treatment. Bars are S.D. (n=3). Different characters indicate statistical significance differences (p < 0.01, ANOVA and Tukey test). (G) Relative expression levels of PER49 to CLATHRIN control in each genotype with or without 2-hour CIF2 treatment. Bars are S.D. (n=3). Different characters indicate statistically significant differences (p < 0.01, ANOVA and Tukey test). (H) Fold changes of PER49 in each genotype with or without 2-hour CIF2 treatment. Bars are S.D. (n=3). Different characters indicate statistically significant differences (p < 0.01, ANOVA and Tukey test). (I) Relative fold changes of PER15 in pCASP1::SGN1-Citrine and pCASP1::myrpalm-SGN1-Citrine compared to the expression level in WT. Bars are S.D. (n=3). Different characters indicate statistically significant differences (p < 0.01, ANOVA and Tukey test). (J) Relative fold changes of PER49 in pCASP1::SGN1-Citrine and pCASP1::myrpalm-SGN1-Citrine compared to the expression level in WT. Bars are S.D. (n=3). Different characters indicate statistically significant differences (p < 0.01, ANOVA and Tukey test).
Figure 7A IFNAR1+/+ and IFNAR1-/- MEF cells were stimulated with IFN-β or infected with SeV, and the expression of IFNAR1 and RBM47 was determined using western blot. The gray values of the blots were analyzed using ImageJ software and normalized to GAPDH. The relative density of the control group was set to "1." B IFNAR1+/+ and IFNAR1-/- MEFs were infected with shmRBM47 or control lentivirus, and the knockdown efficiency of RBM47 was determined using western blot. The relative band densities were analyzed using ImageJ software and normalized to GAPDH. C RBM47-deficient or control IFNAR1+/+ and IFNAR1-/- MEFs were infected with VSV or HSV-1. The cells were harvested at 12 hpi. (VSV) or 24 hpi. (HSV-1), and viral RNA levels were analyzed using qRT-PCR.The PCR results are represented as the means ± SD of n = 3 biological replicates. Data information: The data shown are representative of n = 3 independent experiments. NS, non-significant; **P ≤ 0.01, and ***P ≤ 0.001 (Student's t-test). D The mRNA levels of VSV and RBM47 in blood samples of IFNAR1 deficient (IFNAR1-/-) mice (n = 6 per group) infected with VSV. Mice were infected with shmRBM47 or control lentivirus for 7 days and then challenged with VSV for another 24 h. Each dot represents one mouse. The PCR results are represented as the means ± SD of n = 6 biological replicates. Data information: The data shown are representative of n = 3 independent experiments. NS, non-significant; **P ≤ 0.01, and ***P ≤ 0.001 (Student's t-test).
Figure 4(A) Responding cells were characterized based on the indicated metrics of target promoter activation (timing, magnitude, and rate). (B-G) Effects of p53 duration modulation on target promoter activation in terms of the mean timing (B, E), magnitude (C, F), and rate (D, G) for the MDM2 (B-D) and CDKN1A (E-G) promoters in response to short (light green) and long (dark green) p53 durations. N = at least 45 cells per condition, error bars = SEM. (H) Mean nuclear MDM2-YFP levels in single cells at times corresponding to the first pulse (5.5 h) or second pulse (11 h) of p53 expression in response to natural and long duration Nutlin-3 dosing regimens. N = at least 40 cells per condition, error bars = SEM.
Maternal ZNRF3 is essential for the dorsal ventral axis formation in Xenopus.Figure 3A qRT-PCR analysis of the efficacies of znrf3 AS normalized to odc. Data are shown as mean ± SEM. n = 3 biological replicates; ****p < 0.0001 by unpaired t test. B Whole mount in situ hybridization of Spemann organizer markers chrd, gsc, and xnr3 in gastrula embryos (stage 10.5). Oocytes were injected with AS or mRNAs as indicated, Dosage: znrf3 AS (3 ng AS4 + 3 ng AS9), znrf3 mRNA 100 pg, znrf3ΔRING mRNA 50 pg. embryos were obtained through Host transfer technology. Scale bars: 1 mm. C Representative phenotypes of tailbud stage Xenopus leavis embryos injected with ctrl or znrf3ΔRING mRNAs as indicated. mRNAs were injected in pre-mature oocytes and embryos were obtained through Host transfer technology. Scale bars: 1 mm. D Western blot analysis of Axin and β-catenin in blastula stage (st7-8) embryos injected with 500 pg znrf3ΔRING or 500 pg hwa at 1-cell stage. Lysates were immunoblotted with Axin, β-catenin, HA, and GAPDH. Blots are representative of 3 independent experiments. E Relative levels of β-catenin (left) and Axin (right) in (D) normalized to GAPDH. n = 3 independent experiments, mean ± SEM, *p < 0.05, ***p < 0.005, ****p < 0.0001 by unpaired t test. F Representative phenotypes of Xenopus embryos at tailbud stage. Embryos were injected with ctrl (20 pg gfp mRNA), 20 pg znrf3ΔRing mRNA at one ventral blastomere of 4-cell stage embryos, or 20 ng β-catenin morpholino (β-cat MO) at 1-cell stage as indicated. Duplicated axes (red arrowhead). Scale bars: 1 mm. G Representative phenotypes of Xenopus laevis embryos at tailbud stage. 20 ng hwa morpholino (hwa MO) was injected in oocytes at stage VI, embryos were obtained through Host transfer technology. znrf3ΔRING mRNA (20 pg) was injected at one ventral blastomere of 4-cell stage embryos. β-gal mRNA 200 pg was co-injected with znrf3ΔRING mRNA as lineage tracer. Duplicated axes (red arrowhead). Scale bars: 1 mm. H Whole mount in situ hybridization of Spemann organizer markers chrd and gsc in gastrula embryos (stage 10.5). Embryos were injected and obtained as descripted in (G). Scale bars: 1 mm. I Representative phenotypes of tailbud stage Xenopus leavis embryos injected with 500 pg mRNAs of ctrl (gfp mRNA), znrf3FL, and znrf3 truncations as indicated at 4-cell stage. Duplicated axes (red arrowhead). znrf3FL, full-length znrf3; znrf3ΔRING, znrf3 lacking the RING domain; znrf31-320, amino acids 1-320 of znrf3; znrf3320-606, amino acids 320-606 of znrf3; znrf3606-854, amino acids 606-854 of znrf3. Scale bars: 1 mm.
Figure 6Comparison of the Hallmark gene expression signature "TNFA_signaling via NFKB" in near-diploid (low aneuploidy) and highly-aneuploid (high aneuploidy) human cancer cell lines from the CCLE. *, p= 5e-08, empirical Bayes-moderated t-statistics. The y-axis represents the ssGSEA expression score. The grey line marks an ssGSEA score of 0, indicating the genes in the hallmark "TNFA_signaling via NFKB" geneset is not differentially regulated. Correlation plot between ssGSEA expression score of "TNFA_signaling via NFKB" and the aneuploid score for human cancer cell lines from the CCLE. The trend line for Spearman's correlation plot is indicated in red. Spearman's ρ= 0.181, p-value = 2.2e-08. HCT116 (left) or DLD1 cells (right) were treated with either DMSO or the Mps1 inhibitor reversine (500 nM) for 48 hours before assessing NK cell-mediated cytotoxicity. The drug was washed out during the NK cell co-culture assay. n=3 biological replicates; mean±SEM. HCT116 vs. HCT116 Mps1 inhibitor, p= 0.44, n.s.; DLD1 vs. DLD1 Mps1 inhibitor, p= 0.44, n.s.; KS test.