EMBO/sd-character-level-ner huggingface dataset:
Figure 6(A) Root-mean-squared fluctuation (RMSF) of the HP2 backbone in the open and closed state for WT and L448A EAAT1 (GltPh A360). Values are given as mean ± SD of 10 blocks. Amino acid conservation in the indicated stretch (residue range 444-452 (GltPh 356-368) is shown as sequence logo for all K+-independent (GltX and ASCT1/2, upper row) and all K+-dependent (EAAT1-5, lower row) transporters. (B) HP2 closure is accompanied by deformation of the backbone structure around residue L448 (GltPh A360). (C) Probability density distribution for HP2 opening in WT L448A or L448T EAAT1 (GltPh A360); the shaded area indicates closed states. (D) Probabilities of HP2 closure from the integrated densities (Mean ± SD from 1000 bootstrap samples). Statistical significance was tested by comparing bootstrapped confidence intervals (*, 95% confidence intervals do not overlap). (E) Representative transport current recordings from cells expressing L448A or L448T (GltPh A360) or WT EAAT1 upon voltage jumps to ˗140 mV before and after superfusion with 1 mM l-glutamate. Cells were intracellularly dialyzed with a K+-free solution without permeating anions to isolate K+-independent transport currents. Transient capacitive currents during the first 5 ms after a voltage jump were blanked. (F) Current-voltage relationship for net transport current amplitudes for WT and L448A/T EAAT1 (GltPh A360) for choline-based pipette solutions. Values are given as mean ± SD with indicated numbers of experiments. (G) Time-dependent changes in L448A EAAT1 (GltPh A360) uptake currents during two repetitive glutamate applications and subsequent removals. Current amplitudes were measured at the end of voltage jumps to -140mV and plotted versus the time. Glutamate application is indicated by green bars. Values are given as mean ± SD of the last 100 data points in the current trace. (H) Net transport current amplitudes for WT and L448A/T EAAT1 (GltPh A360) for different pipette solutions, Values are given as mean ± SD with indicated numbers of experiments (***, p<0.001 Student's t-test).
Figure 5In vitro binding analysis of tsGlnCTG to Igf2bp1 in the presence and absence of antisense oligo (ASO) against tsGlnCTG. ASOs effectively disrupt the binding of tsGlnCTG to Igf2bp1. (n = 2), Error bars represent S.E.M. Quantification of the Igf2bp1 bound c-Myc mRNA between LIF- versus RA-treated mESCs (n = 2; see Fig. EV5F for duplicate data). Association of c-Myc transcript in different translating pools in RA treated mESCs as compared to LIF condition. (n = 2), Error bars represent SD, significance was calculated by one-tailed unpaired t-test. Relative enrichment of c-Myc mRNA in translating (80S and Polysome) and non-translating (mRNPs) pools (fractionated from polysome profiling) between ASO-treated and Mock-treated RA-induced differentiating mESCs. (n =2) Error bars represent SD, significance was calculated using one-tailed unpaired t-test. Relative levels of c-Myc mRNA in ASO-treated and Mock-treated RA-induced differentiating mESCs compared to LIF treated mESCs. (n = 3) Error bars represent SD, significance was calculated using one-tailed unpaired t-test. Epistatic analysis of 5'-tsRNAs and IGF2BP1 in regulating Myc transcriptional reporter activity. (n = 3), Error bars represent SD, significance was calculated using unpaired t-test. Schematic representing tsRNA based cMyc transcript regulation.
a) Representative co-immunoprecipitation results showing the effect of thioperamide on the interaction of H3R with CLIC4 during OGD/R.f7(a) Representative co-immunoprecipitation results showing the effect of thioperamide on the interaction of H3R with CLIC4 during OGD/R. Full-size blots are shown in Supplementary Fig. 14. (b) Cell viability was tested by MTT assay after administration of Tat-H3RCT414-436 (10−6 mol l−1 at reperfusion) under OGD/R (data are presented as mean±s.e.m.; n=7 per condition; *P0.05, **P0.01, with analysis of variances (ANOVAs) followed by the Bonferroni/Dunn post hoc test). (c) Representative western blots showing the effect of Tat-H3RCT414-436 on phosphorylation of Akt/GSK-3β/mTOR/P70S6K signalling and LC3 expression in OGD/R. Full-size blots are shown in Supplementary Fig. 14. (d) Representative images of the effect of Tat-H3RCT414-436 on LC3-positive autophagic puncta (red) in 4',6-diamidino-2-phenylindole-stained (blue) neurons after OGD/R (scale bar,10 μm). (e) Western blots showing the effect of siRNA for CLIC4 on phosphorylation of Akt/GSK-3β/mTOR/P70S6K signalling and LC3 expression in OGD/R. Full-size blots are shown in Supplementary Fig. 14. (f) Representative western blots and bar graph showing the expression of LC3-I and LC3-II on steady-state autophagy and autophagic flux under OGD/R when CLIC4 was knocked down (data are presented as mean±s.e.m.; n=6; *P0.05, **P0.01; , with ANOVAs followed by the Bonferroni/Dunn post hoc test). Full-size blots are shown in Supplementary Fig. 14. (g) Cell viability was assessed by MTT assay after siRNA for CLIC4 under OGD/R (data are presented as mean±s.e.m.; n=8 per condition; ***P0.001, with ANOVAs followed by the Bonferroni/Dunn post hoc test).
Figure 7A Bacterial pathogens invade leaf tissues through stomata. B Pathogens proliferate in the extracellular space or the apoplast. C, D Virulent pathogens spread over the infection site (C) and develop disease (D). E, F Avirulent pathogens are restricted to the infection site as a result of the construction of the CASPL- and lignin-deposited structure (E) and are eventually eliminated from the infection site of the leaf (F).
Figure 1A. Endogenous localization of VAPA or VAPB and Syt in hippocampal neurons (DIV16) expressing GFP-Sec61β. Zooms represent (1) an axonal structure with presynaptic boutons (arrowheads), and (2) a dendritic structure. Scale bars: 10 µm (full size) and 5 µm (zoom). B. Schematic illustration of the Syt antibody uptake assay: live neurons were stimulated with bicuculline and incubated with primary Syt antibodies, next neurons were fixed and stained with secondary antibodies. C. Representative image of Syt antibody uptake at axons of hippocampal neurons (DIV18) co-expressing RFP and pSuper empty vector or VAPA/B shRNAs. Yellow and grey arrowheads mark presynaptic boutons with and without internalized Syt, respectively. Zooms represent typical boutons. Scale bars: 5 µm (full size) and 2 µm (zoom). D. Quantifications of fluorescence intensity of internalized endogenous Syt (B, C) at single presynaptic boutons of hippocampal neurons (DIV18) co-expressing RFP and pSuper empty vector or VAPA/B shRNAs. N=2, n=288-541 boutons. Data information: Data represent Mean ± SEM; NS: not significant; *P < 0.05; **P < 0.01; ***P < 0.001, by Mann-Whitney U test. E. Western blot of endogenous SCRN1 expression in indicated adult rat neuronal and non-neuronal tissues. Cereb., cerebellum. Hippoc., hippocampus. Spin., spinal. F. Pull down assay of HEK293T cells co-expressing Myc-VAPA with BioGFP or BioGFP-SCRN1. G. Pull down assay of HEK293T cells co-expressing Myc-VAPB with BioGFP or BioGFP-SCRN1.Data information: Data represent Mean ± SEM; NS: not significant; *P < 0.05; **P < 0.01; ***P < 0.001, by Mann-Whitney U test. H. Scaled representation of SCRN1-associated proteins identified with pull-down assay followed by mass spectrometry analysis of purified BioGFP or BioGFP-SCRN1 from HEK293T cell lysates with adult rat brain extracts. All candidates showed >10 enrichment of PSM compared to control. I. Endogenous localization of SCRN1 and Syt in cortical neurons (DIV18) expressing GFP. Zoom represents an axon structure with presynaptic boutons (arrowheads). Scale bars: 10 µm (full size) and 5 µm (zoom). J. Representative image of Syt antibody uptake at axons of hippocampal neurons (DIV18) co-expressing RFP and pSuper empty vector, SCRN1 shRNA or SCRN1 shRNA with GFP-SCRN1. Yellow and grey arrowheads mark presynaptic boutons with and without internalized Syt, respectively. Zooms represent typical boutons. Scale bars: 5 µm (full size) and 2 µm (zoom). K. Quantifications of fluorescence intensity of internalized endogenous Syt (B, C) at single presynaptic boutons of hippocampal neurons (DIV18) co-expressing RFP and pSuper empty vector, SCRN1 shRNA alone or SCRN1 shRNA with GFP-SCRN1. N=2, n=201-300 boutons. Data information: Data represent Mean ± SEM; NS: not significant; *P < 0.05; **P < 0.01; ***P < 0.001, by Mann-Whitney U test.
Higher magnitude of endogenous SIINFEKL-specific CD8+ T cell responses induced by a sporozoite antigen.Figure 4(A) Schematic overview of experimental design. (B- C57BL/6 mice received 10,000 γ-radiation attenuated WT, CSPSIINFEKL or UIS4SIINFEKL sporozoites, either intravenously (n=6-8 mice per group) or (E- intradermally (n=4 mice per group). Additional control mice did not receive sporozoites (n=6-8 intravenously, n=4 intradermally). Spleens and livers were harvested either at day 14 (n=5 livers and n=6 spleens for intravenous immunisation) or day 42 (n=8 for intravenous immunisation), and IFN-γ-secreting lymphocytes following restimulation ex vivo with SIINFEKL peptide were quantified. Flow cytometry plots show representative percentages of CD8+ T cells co-stained with IFN-γ and CD11a (B, E). C57BL/6 mice received 10,000 γ-radiation attenuated WT, CSPSIINFEKL or UIS4SIINFEKL sporozoites, either intravenously (n=6-8 mice per group) Additional control mice did not receive sporozoites (n=6-8 intravenously, n=4 intradermally). Spleens and livers were harvested either at day 14 (n=5 livers and n=6 spleens for intravenous immunisation) or day 42 (n=8 for intravenous immunisation), and IFN-γ-secreting lymphocytes following restimulation ex vivo with SIINFEKL peptide were quantified. The upper panel of bar charts (C, show the percentage of CD11ahi IFN-γ+ CD8+ T cells and the lower panel (D the absolute cell counts. Bar charts show mean values (±SEM) from representative experiments (*, p<0.05; **, p<0.01; ***, p<0.001; one-way ANOVA with Tukey's multiple comparison test). See Appendix Table S4 for exact p-values. C57BL/6 mice received 10,000 γ-radiation attenuated WT, CSPSIINFEKL or UIS4SIINFEKL sporozoites intradermally (n=4 mice per group). Additional control mice did not receive sporozoites (n=6-8 intravenously, n=4 intradermally). Spleens and livers were harvested either at day 14 (n=5 livers and n=6 spleens for intravenous immunisation) or day 42 (n=8 for intravenous immunisation), and IFN-γ-secreting lymphocytes following restimulation ex vivo with SIINFEKL peptide were quantified. The upper panel of bar charts show the percentage of CD11ahi IFN-γ+ CD8+ T cells and the lower panel the absolute cell counts. Bar charts show mean values (±SEM) from representative experiments (*, p<0.05; **, p<0.01; ***, p<0.001; one-way ANOVA with Tukey's multiple comparison test). See Appendix Table S4 for exact p-values.
Figure 6A. The interaction partners of YTHDF1 were identified by pull-down assays and mass spectrometry. The Flag-YTHDF1 pull-down assay was performed in CNE2EBV cells transfected with a plasmid encoding Flag-tagged YTHDF1. The proteins in the differentially abundant bands were identified by mass spectrometry. The interactions between YTHDF1 and ZAP (B) were measured by co-IP in 293T cells. The results were detected using specific antibodies. C. The interactions between YTHDF1 DDX17 (C) were measured by co-IP in 293T cells. The results were detected using specific antibodies. D. The interaction between YTHDF1 and DCP2 was detected by co-IP in 293T cells. E. The interactions between endogenous YTHDF1 and ZAP, DDX17 or DCP2 were detected by endogenous IP using a YTHDF1-specific antibody in CNE2EBV cells. F. The interactions between endogenous YTHDF1 and ZAP, DDX17 or DCP2 were detected by endogenous IP assays in CNE2EBV cells transfected with YTHDF2-, YTHDF3-specific siRNAs or the siNC control. YTHDF1, YTHDF2, YTHDF3, ZAP, DDX17, and DCP2 were detected in the immunoprecipitated complexes using specific antibodies, respectively.
Protein abundance correlation of affinity-purified complexes analyzed by mass spectrometry strategy applied to proteasome complexes.Figure 3A Proteasome complexes were immunopurified from nine formaldehyde-crosslinked human cell lines and analyzed by nano-LC-MS/MS. Protein abundance indexes (PAIs) were used to represent the abundance of proteins in purified proteasome samples. The correlation between two different proteins was quantified using coefficients of determination (R2). B-E Correlations of abundances of α7 and α6 (B), Rpn3 and Rpn1 (C), PA28β and PA28α (D), and Rpn3 and PA28β (E).
Figure 3A, B BIC1 and BZR1 synergistically promote hypocotyl elongation. The Col-0, 35S:BZR1-MYC, 35S:BIC1-YFP and 35S:BZR1-MYC/35S:BIC1-YFP plants were grown for 5 days under long-day conditions. Images of the representative seedlings are shown in (A), and the hypocotyl lengths of the indicated genotypes were measured and are shown in (B). Data are means ± SD (n>20). **p < 0.01, as determined by Student's t-test. Scale bar, 2 mm. C-E BR signaling is enhanced in the 35S:BZR1-MYC/35S:BIC1-YFP plants. Seedlings were grown for 6 days on medium supplemented with 1 μM brassinazole (BRZ) plus a gradient of concentrations of epibrassinolide (eBL) under long-day conditions. Images of the representative seedlings when grown with 500 nM eBL (+eBL) or not (-eBL) are shown in (C), and the hypocotyl lengths of the indicated genotypes were measured and are shown in (D) and (E). Data are means ± SD (n>20). **p < 0.01, as determined by Student's t-test. Scale bar, 2 mm. F, G BZR1-mediated hypocotyl elongation phenotype is dependent on the function of BIC1 and BIC2. Seedlings were grown for 5 days under long-day conditions. Images of the representative seedlings are shown in (F), and the hypocotyl lengths of the indicated genotypes were measured and are shown in (G). Data are means ± SD (n>20). **p < 0.01, n.s. indicates no significant difference, (Student's t-test). Scale bar, 2 mm. H BZR1-mediated hypocotyl elongation phenotype is dependent on the function of BIC1 and BIC2. The Col-0, 35S:BZR1-MYC, bic1bic2, 35S:BZR1-MYC/bic1bic2 seedlings were grown on medium supplemented with 1 μM BRZ plus different concentrations of eBL for 6 days and then the hypocotyl lengths were measured. Data are means ± SD (n>20).
Figure 2A Sec63-mNeon images of cells of the indicated genotypes harboring the inducible system. Cells were treated with 800 nM estradiol for 6 h. Two examples of each phenotypic class are shown. B Violin plots of Z scores from the three metrics for ER size determined for each of the 4800 mutant strains. The untreated WT is shown for reference. C Sec63-mNeon and Rtn1-mCherry images of ∆lnp1 and ∆sey1 cells harboring the inducible system and treated with 800 nM estradiol for 6 h. D Classification of hits. Numbers in brackets indicate the number of mutants in each class. Striped areas indicate mutants belonging to two classes.
Figure 4A IBD GWAS results (Liu et al, 2015) at a multi-disease-associated locus on chromosome 6q23. B Fine-mapping results (Huang et al, 2017) (posterior probabilities) for candidate SNPs at this locus. C A single variant (rs6927172) has the largest and most significant expression-modulating activity in resting (left panel) and stimulated CD4 T cells (right panel) with the risk allele reducing transcription. Plots represent median and IQR (box) and min-to-max (whiskers). FDR-corrected meta-analysis P value shown. D Sequence logo for an experimentally-validated NF-κB binding motif (Wong et al, 2011). The genomic sequence around rs6927172 is aligned below. E Allele-specific NF-κB binding in CD4 T cells from rs6927172 heterozygotes, demonstrating reduced NF-κB binding to the risk allele following stimulation (n=8; one-sample t-test, two-tailed). F Allele-specific expression of enhancer RNA in heterozygous CD4 T cells. DNA used for technical control (n=6; paired t-test; two-tailed). G Genome-wide H3K27ac ChIP-seq in stimulated CD4 T cells from major- and minor-allele homozygotes at rs6927172 (n=6). Upper panels show input-normalised H3K27ac signals plotted against enhancer rank. Super-enhancers are defined above the inflection point of the curve. Lower panels show H3K27ac reads from a major (left) and a minor (risk) allele homozygote (right) in a 9kb window around rs6927172. H Promoter-capture Hi-C plot depicting interactions of the 6q23 super-enhancer in stimulated CD4 T cells. I Expression of genes on 6q23 in CD4 T cells from 131 patients with active IBD, stratified by rs6927172 genotype (qPCR; one-way ANOVA). Error bars represent SD. Expression of IL20RA and IL22RAR2 not detected.
InlK interacts with the Major Vault Protein.Figure 3A. Bacterial pull-down of purified GST-MVP with the L. monocytogenes strains WT+pPRT-empty, ΔinlK+pPRT-empty, ΔinlK+pPRT-inlK and WT+pPRT-inlJ. GST-MVP bound to InlK over-expressing bacteria (ΔinlK+pPRT-inlK) but not to other bacteria. GST-ScarA was used as control of the specificity of the MVP precipitation by InlK over-expressing bacteria. B. Bacterial pull-down of MVP-GFP from transfected HeLa cell lysates with the L. monocytogenes strains WT+pPRT-empty, ΔinlK+pPRT-empty, ΔinlK+pPRT-inlK and WT+pPRT-inlJ. MVP-GFP bound to InlK over-expressing bacteria (ΔinlK+pPRT-inlK) but not to other bacteria. C. Co-immunoprecipitation (Co-IP) of purified InlK (20 µg) with endogenous MVP of HT29 cells. The right panel shows anti-MVP co-IP performed on HT29 cell lysates. The left panel shows the control anti-MVP co-IP performed on lysis buffer. D. Detection of MVP recruitment at the surface of InlK over-expressing bacteria. HeLa cells were transfected with MVP-GFP (green), infected with L. monocytogenes EGD-e wild-type (WT), ΔinlK or ΔinlK+pPRT-inlK for 4 h, fixed for fluorescence light microscopy, and stained with anti-Listeria antibodies (red). Inset regions are magnified. The scale bar represents 1 µm.
Figure 6A PCR amplification efficiency at the telomere in the mock- and FPG-treated DC and age-matched healthy control fibroblasts supplemented with vehicle or 3 mM NR. All values are presented as mean ± SD of three replicates. The relative telomere PCR amplification in each sample was normalized to the 36B4 reference gene. B Telomere restriction fragment analysis in DC and control fibroblasts treated with vehicle or 3 mM NR. Genomic DNA was isolated from the indicated cell lines at 0 d or 14 d, with or without 3 mM NR treatment. At left, DNA molecular mass markers, in kilobase pairs (Kb). C, D Representative images of γH2AX (green), telomere immuno-FISH (red) in DC cells treated with vehicle or 3 mM NR. White arrows: co-localization of γH2AX foci and telomeric DNA (or TIF). Enlarged views of co-localizing foci are shown at the right panel and in Figure EV1. Percentage of DC and control cells with indicated TIFs per nuclei. ~ 100 cells per condition were scored.
Fig.Figure 1(A) Schematic diagram of study organization. Metabolome profiles were collected from yeast that were treated with 1280 drugs as well as yeast where 6 genes were independently expressed. (B) The Spearman correlation coefficients for the comparison of z-scored drug and overexpression metabolome profiles are shown in a heat map format with a call out of the indicated cluster. In that cluster a pattern of decreasing correlation was observed as is indicated by the trend in average -1 to 1 scaled correlations shown in the line plot. (C) The relationship between the Spearman correlation of the metabolome profiles of yeast treated with drugs and each GPR1 overexpression condition is shown for two example drugs across the different concentrations of inducer that were used. (D) The fit of a logistic curve to the normalized correlation data (scaled between 0 and 1 for minimum and maximum values) from C is shown. (E) A summary of the analysis of the two example drugs.
Figure 2qRT-PCR detection of IFNB1 mRNA abundance in THP-1 cells treated with ecGAMP and icGAMP at indicated concentrations for 4 h. qRT-PCR detection of IFNB1 mRNA abundance in THP-1 cells treated with ec-di-AMP and ic-di-AMP (B) at indicated concentrations for 4 h. ELISA detection of IFNβ release from THP-1 cells stimulated with ecGAMP and icGAMP (C) at indicated concentrations for 4 h. ELISA detection of IFNβ release from THP-1 cells stimulated with ec-di-AMP and ic-di-AMP (D) at indicated concentrations for 4 h. qRT-PCR detection of Ifnb1 mRNA abundance in mBMDMs treated with ecGAMP and icGAMP (E) at indicated concentrations for 4 h. qRT-PCR detection of Ifnb1 mRNA abundance in mBMDMs treated with ec-di-AMP and ic-di-AMP (F) at indicated concentrations for 4 h. ELISA detection of IFNβ release from mBMDMs stimulated with ecGAMP and icGAMP at indicated concentrations for 4 h. ELISA detection of IFNβ release from mBMDMs stimulated with ec-di-AMP and ic-di-AMP (H) at indicated concentrations for 4 h.
Figure 6(a) Native gel analysis of pre-splicing complexes. Pre-splicing complex assembly was carried out in the test tube with Prp5-reconstituted cell extracts and labeled pre-mRNA. Lane 1, Prp5-depleted wild-type cell extract. Lane 2, Prp5-depleted wild-type cell extract, plus wild-type Prp5. Lane 3, Prp5-depleted snR81Δ pus1Δ cell extract, plus wild-type Prp5. Lane 4, Prp5-depleted wild-type cell extract, plus mutant Prp5 (GAR). Lane 5, Prp5-depleted snR81Δ pus1Δ cell extract, plus mutant Prp5 (GAR). The pre-splicing complexes A and B are indicated. (b) DMS in vivo probing. After being exposed to DMS, yeast cells were lysed, total RNA collected, and primer-extension analysis carried out (left panel). Lane 1, cells that were not exposed to DMS. Lane 2, wild-type cells exposed to DMS. Lane 3, snR81Δ pus1Δ cells exposed to DMS. Lane 4, snR81Δ pus1Δ cells, transformed with a plasmid carrying an artificial guide RNA gene targeting positions 42 and 44, and exposed to DMS. Lane 5, snR81Δ pus1Δ cells, transformed with a plasmid carrying a control guide RNA gene with random guide sequences, and exposed to DMS. The nucleotides in parentheses are DMS-modified C and A residues that are one nucleotide after the actual primer-extension stops (indicated by the arrows). A partial U2 sequence (along with the BSL structure) is shown (right panel). DMS-modified adenosine and cytosine in the BSL are also indicated. (c) Residue accessibility quantification. Relative accessibility (relative band intensity) was quantified based on three independent experiments. The intensity of every band in each lane was normalized against the C46 band (the least changed band; compare lanes 2-5 in (b)) in its respective lane [(intensity of a band) / (intensity of C46 in the same lane)]. Each C46-normailzed band in lanes 3-5 (refer to (b)) was then further normalized against its respective counterpart in lane 2 [(the value of a C46-normalized band in lanes 3-5) / (the value of its counterpart in lane 2)]. The error bars represent the standard deviations of the measurements.
Atg8 and Other ATG Genes Are Necessary during UPR Induction.Figure 8Atg8 and Other ATG Genes Are Necessary during UPR InductionSerial dilutions for wild-type, hac1Δ, atg1Δ, atg8Δ, atg9Δ, atg16Δ, and atg20Δ deletion cells and vps4Δ pep4Δ double deletion cells were grown on rich-media plates with no drug (YPD) or with different concentrations of tunicamycin (TM; 0.2 or 1.0 μg/ml). atg19Δ gave an identical result to the other autophagy genes shown here (unpublished data).