EMBO/sd-character-level-ner huggingface dataset:
Figure 7Crystal structure of the free D4-5 subunit. Note the different relative orientations of domains D4 and D5 with respect to the structure of full-length CNFY (top, thin grey lines). The domain D4 forms a large interface area (1100 Å2) with the catalytic domain D5 involving several polar interactions (8 hydrogen bonds and 8 salt bridges), whereby the active crevice is extended and fully solvent-exposed as can be seen in the right surface plot at the bottom of the panel. The hypothetical NAD+ binding site of the ART-like D4 domain is located on the opposite face (left surface plot). Note that the deamidase active site of domain D5, unlike in the full-length structure (Fig 1), is fully accessible and that its extended shape is also determined by domain D4. Comparative analysis of RhoA activation in HEp-2 cell lysate by CNFY and the recombinant D4-5 protein. Purified CNFY or the D4-5 fragment (1 µM) was added to extracts of HEp-2 cells and incubated for 10, 20, 30 or 60 min. De­amidation of RhoA was analyzed by the shift of the modified Rho GTPase band in SDS PAGE gels after detection with anti-RhoA antibodies. Comparative analysis of recombinant RhoA deamidation by CNFY and the D4-5 fragment. Recombinant RhoA was incubated with purified CNFY or the D4-5 fragment and samples were separated by SDS-PAGE after the indicated times before subjecting to trypsin digestion and quantification of deamidation of Q63 by mass spectrometry. Error bars represent standard deviations of triplicate measurements.
FFAT-containing proteins recruit the ER-resident MOSPD2 protein to inter-organelle contact sites A: GFP-MOSPD2 (green) expressing cells were stained with an anti-Lamp1 antibody (red) to label late endosomes/lysosomes.Figure 6 A: GFP-MOSPD2 (green) expressing cells were stained with an anti-Lamp1 antibody (red) to label late endosomes/lysosomes Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm  B-F: GFP-MOSPD2 (green) expressing cells were transfected with Flag-STARD3NL (B and labelled using anti-Flag (B-E; magenta) and anti-Lamp1 (B-D; red as markers of late endosomes/lysosome Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm  B-F: GFP-MOSPD2 (green) expressing cells were transfected wit Flag-STARD3 (C and labelled using anti-Flag (B-E; magenta) and anti-Lamp1 (B-D; red as markers of late endosomes/lysosome Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm  B-F: GFP-MOSPD2 (green) expressing cells were transfected wit Flag-ORP1L (D and labelled using anti-Flag (B-E; magenta and anti-Lamp1 (B-D; red as markers of late endosomes/lysosome Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm  B-F: GFP-MOSPD2 (green) expressing cells were transfected wit ), Flag-STARD11 (E and labelled using anti-Flag (B-E; magenta an GM130 (E; red as markers o Golg Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm  B-F: GFP-MOSPD2 (green) expressing cells were transfected wit HA-PTPIP51 (F), and labelled usin anti-HA (F: magenta), an OPA-1 (F; red) as markers o mitochondri Data information: In (A-F), the subpanels on the right are higher magnification (3.5x) images of the area outlined in white. The Overlay panel shows merged green and magenta images. Scale bars: 10 µm
Figure 6A Hexbin plots comparing the regulatory effects (number of regulated genes) between GATA-1, Alas2-enhancer, and 5-ALA. All upregulated genes (FDR < 0.05 and log2 fold change > 0) and downregulated genes (FDR < 0.05 and log2 fold change < 0) were tested. Coefficient of determination (r2) was shown in each plot.B Pie charts display the percentage of activated and repressed genes (FDR < 0.05 and expressed differentially >2-fold) in GATA-1-, Alas2-enhancer, or 5-ALA-regulated genes determined by RNA-seq analysis.C Venn diagram depicting relationships between GATA-1-, Alas2-enhancer, and 5-ALA-regulated genes.D Pie chart depicting the gene numbers in 4 categories for genes regulated by GATA-1, Alas2-enhancer, and 5-ALA.
Figure 1(A and B) THP1-derived macrophages were treated with the TLR ligands Pam3, lipopolysaccharide (LPS), or FSL-1 and analyzed by RNA-seq 8 h later. Circos plots show genome-wide differential expression of protein-coding genes (A) and non-protein-coding genes (B) between untreated and treated macrophages. The outermost to innermost circles show downregulated (blue) or upregulated (red) genes in cells stimulated with FSL-1, LPS, and Pam3, respectively. Chromosomes are indicated by the numbers 1-22 and X and Y. (C) Schematic showing cis-acting lncRNAs and protein-coding genes within 5 kb. (D) Classification of lncRNA- and protein-coding gene pairs modulated by TLR ligands according to their biotypes. Pie charts show the proportion of each pair of genes classified as XH (antisense head-to-head), XT (antisense tail-to-tail), XI (antisense inside), XO (antisense outside), SD (sense downstream), and SU (sense upstream). (E) LncRNA- and protein-coding gene pairs commonly upregulated or downregulated in THP-1-derived macrophages stimulated with FSL-1, LPS, or Pam3 for 8 h. (F) RT-qPCR validation of differentially expressed lncRNAs and mRNAs in THP1-derived macrophages stimulated for 8 h with flagellin (FLA), FSL-1, imiquimod, LPS, Pam3, or poly(I:C). Heat map shows the fold change in expression of 13 mRNAs (left) and lncRNAs (right).
a) Western blot analysis showing reduced levels of LC3-II in KO MEFs.f7a) Western blot analysis showing reduced levels of LC3-II in KO MEFs. (b) Quantification of protein signal intensities from western blots showing LC3-II levels in MEFs extracts after normalization to the control protein GAPDH. In a and b, MEFs were grown in normal medium (Nm), treated for 2 h with 50 nM bafilomycin in normal medium (B), or grown in nutrient-deprived medium in the absence (St) or presence of 50 nM bafilomycin (St+B). Values denote means±s.e.m. (n=4). (c) Immunostaining of endogenous LC3 in Pld1 WT and KO MEFs showing a reduced number and size of the LC3-positive compartment in KO MEFs. The DAPI staining is shown in blue. Cells were deprived of nutrients for 2 h in the absence (St, upper panels) or presence of bafilomycin (St+B, lower panels). Scale bar, 5 μm. (d) Quantification of the average size (top panel) and number of LC3-positive compartments per cell (bottom panel) in Pld1 WT and KO MEFs starved for 2 h. Values denote means±s.e.m. (n=6, number of cells=26-45 for all the conditions). **P0.01; *P0.05.
Identification of C99-Bpa interaction sites in γ-secretase.Figure 2A Schematic representation of the substrate-photocrosslinking strategy. B Identification of C99 interaction sites with γ-secretase. C99-Bpa substrates were irradiated with UV light in the presence of CHAPSO-solubilized γ-secretase. Crosslinked substrates were captured by Ni-NTA affinity pulldown and bound subunits were identified by a ~10 kDa molecular weight increase compared to the input. Results are shown for the major substrate crosslinking residues. Crosslink formation was not observed in the absence of UV irradiation or in the presence of Triton X-100, which dissociates the γ-secretase complex, proving crosslink specificity. Bracket indicates the molecular weight range of putative C99-APH-1 crosslink bands. APH-1aL, long splice variant of APH-1a. C Excess amounts of parental C99 WT substrate compete crosslinking of C99-Bpa substrates to γ-secretase. Single and double asterisks indicate antibody crossreactivities to C99 monomer and dimer, respectively.
nChIP-seq reveals chromosome-wide absence of H3K27me3 and H2AK119ub enrichment, but residual enrichment at initially active genes in the Xist ΔB+C Plots showing H3K27me3 and H2AK119ub accumulation over the X chromosome in Xist FL and Xist ΔB+C with upon DOX induction at day 2 (D2) of differentiation; Each dot represents a single 10Kb window and its enrichment relative to noDOX condition; Black line is a loess regression on all windows; Xist locus is represented by a blue long line, active genes by green lines.Figure 3Plots showing H3K27me3 and H2AK119ub accumulation over the X chromosome in Xist FL and Xist ΔB+C with upon DOX induction at day 2 (D2) of differentiation; Each dot represents a single 10Kb window and its enrichment relative to noDOX condition; Black line is a loess regression on all windows; Xist locus is represented by a blue long line, active genes by green lines. Violin plots quantifying H3K27me3 and H2AK119ub enrichment over intergenic regions, active promoters and active gene bodies in the X chromosome in Xist FL and Xist ΔB+C cell lines at D2 upon DOX induction; Shown is the log2 fold change of DOX vs noDOX conditions; violin plots represent the distribution of the values, the horizontal band is the median, the lower and upper hinges correspond to the 25th and 75th percentiles; n = indicates the number of regions/genes analyzed; p-values were calculated using paired Wilcoxon test, comparing Xist FL and Xist ΔB+C cell lines. Average plots showing the mean enrichment of H3K27me3 (top) and H2AK119ub (bottom) over all X-linked initially active transcriptional start sites (TSS); Shown is the mean of normalized log2 enrichment of DOX vs noDOX in both Xist FL and Xist ΔB+C cell lines. Genome browser plots showing H3K27me3 (top) and H2AK119ub (bottom) enrichments in a region encompassing the inactive Atp1b4 and the initially active Lamp2 genes within the XqA3.3 region and the initially active Rlim/Rnf12 gene at the XqD region; Region around the promoter of Rlim/Rnf12 is highlighted in yellow.
Binding of USP14 C114A to ubiquitin chains in cells.Figure 5Polyubiquitin chains were co-immunoprecipitated with FLAG-PSMD4, an ubiquitin receptor for the 26S proteasome, from cells expressing either USP14 wild-type, C114A, or C114R. One representative experiment of two is shown.
Figure 5A Human phospho-RTK array displaying the activation of phosphokinase-related proteins in CDK12-knockdown BT474 cells. The bar graph shows the fold change of spot intensity. Cell line information: a/a, CDK12-amplified/HER2-amplified cells S, trastuzumab-sensitive. B Immunoblots showing the levels of the indicated proteins in different types of HER2+ breast cancer stably expressing CDK12 or with CDK12 knocked down. Cell line information: a/a, CDK12-amplified/HER2-amplified cells; n/a, CDK12 non-amplified/HER2-amplified cells; R, trastuzumab-resistant; PR, trastuzumab-partial responsive; S, trastuzumab-sensitive. Co-IP analysis showing the amounts of bindings between ErbB receptors. Cell lysates from CDK12-overexpressing ZR-75-30 cells were immunoprecipitated with anti-HER2 antibody and immunoblotted using the indicated antibodies. IgG, negative control. Cell line information: ; n/a, CDK12 non-amplified/HER2-amplified cells S, trastuzumab-sensitive. D Western blot and qRT-PCR analysis of the phosphorylation levels and expression of IRS1, respectively, in the indicated cell lines. Mean ± s.d. (n = 3). P-values were calculated using a two-tailed Student's t-test. Cell line information: a/a, CDK12-amplified/HER2-amplified cells; n/a, CDK12 non-amplified/HER2-amplified cells; R, trastuzumab-resistant S, trastuzumab-sensitive. E Co-IP analysis indicating the amounts of IRS1 protein interaction with ErbB receptors and p85 in CDK12-overexpressing ZR-75-30 cells. Cell line information n/a, CDK12 non-amplified/HER2-amplified cells S, trastuzumab-sensitive. F IRS1 effect on CDK12-induced phosphorylation of ErbB receptors. Lysates from cells treated with either control siRNA or IRS1 siRNA (siIRS1) for 24 h were subjected to immunoblotting with the indicated antibodies. Cell line information: n/a, CDK12 non-amplified/HER2-amplified cells S, trastuzumab-sensitive. G Proposed model for the mechanism of action of CDK12 regulation of cancer stemness, cell growth, and trastuzumab response in HER2+ breast cancer.
Figure 6A Schematic illustration of treatment scheme. B, Hemizygous K18-hACE2 mice were treated intranasally with 20 µg of NM1251 (n = 8), NM1267 (n = 9) or NM1268 (n = 9) seven hours prior to infection with 3*103 PFU SARS-CoV-2 B.1.617.2 (Delta). Weight loss were monitored for 14 days. Dashed line indicates humane endpoint and symbols represent mean ± SEM ****P < 0.0001 in orange between NM1251 and NM1267 and in purple between NM1251 and NM1268, *P < 0.1 in black between NM1267 and NM1268, by two-way ANOVA with Sidak's multiple comparison test C Hemizygous K18-hACE2 mice were treated intranasally with 20 µg of NM1251 (n = 8), NM1267 (n = 9) or NM1268 (n = 9) seven hours prior to infection with 3*103 PFU SARS-CoV-2 B.1.617.2 (Delta). survival were monitored for 14 days. ***P < 0.001 in orange between NM1251 and NM1267 and ****P < 0.0001 in purple between NM1251 and NM1268, by log-rank test
expression in human subjects with hepatic steatosis and NASH Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and BMI (A), plasma alanine aminotransferase (ALAT) (B) and liver fat content (C).Figure 1Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and BMI Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). Scatter plot and correlation coefficient (r) of ASK1 mRNA expression , plasma alanine aminotransferase (ALAT) (B) Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and liver fat content (C). Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). ASK1 mRNA expression in patients with different NASH scores Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). *** Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and ATG5 (E) mRNA expression. Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). *** Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and ATG7 Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). Scatter plot and correlation coefficient (r) of ASK1 mRNA expression and ATG12 mRNA expression. Data were collected in lean subjects (n=14), obese subjects (n=23) and obese subjects with type 2 diabetes (n=19). Values are expressed as mean ± SEM (D). ***
Figure 1A Schematic of ApoER2 expression and signaling via ligand binding (eg. Reelin, ApoE). ApoER2 alternative splicing of exon 19 produces two ApoER2 protein isoforms with distinct functions. While both isoforms of ApoER2 (+ex19 and Δex19) can bind ligand, a protein domain of ApoER2 encoded by exon 19 interacts with PSD95 to mediate association of ApoER2 with the NMDAR complex following reelin binding. This signaling leads to calcium influx through NMDARs and the induction of long-term potentiation (LTP). B RT-PCR analysis of RNA from mid-temporal autopsy brain sections obtained from NCI, MCI or AD subjects. Representative gel of results is shown.C Quantitation of RT-PCR products of all samples analyzed from post-mortem brain tissues (mean s.e.m.). An omnibus significant effect was obtained using the one-way analysis of variance (one-way ANOVA) with Tukey post-test to determine the p-value. Error bars are s.e.m. D RT-PCR analysis of RNA from the hippocampus of 1-2 and 3-7 month old non-transgenic (WT) or TgCRND8 (AD) mice.E Quantification of the relative ratio of exon 19 inclusion for TgCRND8 (AD) and WT mice (mean s.e.m.; one-way ANOVA with Tukey post-test). The number of samples analyzed (n) is indicated.
a,b) Primary neurons transfected with TDP43(WT)-EGFP (a) or TDP43A315T-EGFP (b) were treated with autophagy inducers and survival determined by AFM.figf6(a,b) Primary neurons transfected with TDP43(WT)-EGFP (a) or TDP43A315T-EGFP (b) were treated with autophagy inducers and survival determined by AFM. Data were obtained from eight wells per condition, with experiments performed in triplicate. Supplementary Table 2 lists the number of neurons per condition, hazard ratios, 95% CI and P values, as determined by Cox hazards analysis. (c) Autophagic induction improved survival in TDP43M337V HB9-positive human iPSC-derived MNs. (d) FPZ and MTM also reduced the risk of death in MAP2-positive human iPSC-derived MNs Values in c and d were pooled from 18 wells per condition, with experiments performed in triplicate. (e) Astrocytes differentiated from human iPSCs exhibit mutant TDP43-related toxicity that is mitigated by FPZ and MTM. Values were pooled from 18 wells per condition, performed in triplicate. Supplementary Table 3 lists the number of human iPSC-derived neurons and astrocytes per condition, hazard ratios, 95% CI and P values, as determined by Cox hazards analysis.
Fig.Figure 4(A) Wt neurons were kept in culture until DIV5, then the cells were treated with acitretin (4 µM) or vehicle control for 48h, as described earlier (Tippmann et al, 2009). (B) Densitometric quantifications of the Western Blots are shown. Given are mean +/- the standard error of the mean (*** p < 0.001; **** p < 0. 0001, two-sided students t-test n = 9-20). The mean levels of solvent treated cells were set to 1. (C) Detection of sNrCAM and human serum albumin (hSA) in AD patients CSF that had been treated with acitretin (30 mg/d) or placebo for 30 days (n = 9) (Endres et al, 2014). (D) Densitometric quantification of the Western Blots in (C). sNrCAM/hSA ratios were calculated, and then the treatment to baseline ratios (t/b) for every patient were calculated. Two-sided student's t-test (n = 9). Given are mean +/- the standard error of them mean. Representative Western Blots are shown.
x localizes to the nuclear regionA, Representative immunofluorescence images showing the subcellular distribution of AGO1x (red) relative to nuclear and cytoplasmic markers.Figure 2A, Representative immunofluorescence images showing the subcellular distribution of AGO1x (red) relative to nuclear and cytoplasmic markers. DAPI was used to mark the nucleus (blue). The co-stained subcellular marker is indicated in each panel in green. SC 35, Lsm4, α-Tubulin, ERP72, p54 (NRB), and Nucleolin serve as markers for nuclear speckles, cytosol and nucleus, cytosol, endoplasmic reticulum, paraspeckle and nucleolus, respectively. B, Mean (+/- s.d.) pixel intensities of AGO1x staining in nucleolus and nucleoplasm, computed from z-stack images of MDA-MB-231 cells (n=20). The P-value was determined using a paired two-tailed t-test. C, Representative AGO1x and AGO1 blot from MDA-MB-231 cell fractions. GAPDH and hnRNP C1/C2 served as markers of purity of the individual fractions.
Figure 5A-L First column shows the triple immunolabelling of CETN3 (green), PCNT (red) and SYCP3 (grey) on squashed WT mouse spermatocytes. Second column shows the correspondent labellings for Plk1(∆/∆). Third column shows signals for BI2536-treated spermatocytes. Images are shown for (A, E, I) zygotene, (B, F, J) pachytene, (C, G, K) diplotene and (D, H, L) diakinesis. Amplifications at 300% magnification for all labellings are displayed for each image. M-O Quantification of the percentage of cells without PCNT at centrosomes (PCNT-) in (M) zygotene, (N) pachytene and (O) diplotene in control, Plk1(∆/∆) and BI2536-treated spermatocytes. Data for BI2536 represent results for 100µM at 8h treatment on organotypic cultures of seminiferous tubules. Data are presented as mean ± SD for three biological replicates for each condition.
Identification of new LMF1-interacting partners.Figure 1 A). Experimental schematic of DSP crosslinking experiment  B). Schematic of pBPA crosslinkin  C). Western blot of LMF1-partners after DSP crosslinking and affinity tag purification using LMF1's C-terminal His tag. Tetracycline induces LMF1 expression. Loading dye with 50mM DTT was used to break the disulfide bonds between LMF1 and its interacting partners. LMF1 complexes are labeled with asterisks and an arrow points to LMF1  D). Western blot of LMF1-partners after pBPA photo-crosslinking and affinity tag purification. The DMSO panel does not include pBPA and serves as a negative control. Fractions were eluted with increasing concentrations of imidazole. An arrow points to LMF1  E). SYPRO orange protein stain of DSP crosslinked samples. Samples without tetracycline but with DSP and DTT (negative control), and with tetracycline, DSP and DTT were sent for LC MS/MS. LMF1 complexes are labeled with asterisks and an arrow points to LMF1  F). SYPRO orange stained gel of pBPA samples. An arrow points to LMF1
c-Maf knock-down in ILC2s dampens papain-induced lung inflammation.Figure 4(A) Treatment schedule for acute or chronic papain­-induced lung inflammation. (B, C) Number of total ILC2s and percentage of c-Maf+ (T test, ***p<0.001) (B); activation status of ILC2s (2way ANOVA, interaction: F(3,24)=27.44, p<0.0001) (C) from lungs after acute or chronic papain-induced lung inflammation (*p<0.05, ***p<0.001, ****p<0.0001). Bars represent mean ± SEM. Data information: are representative of 2 independent experiments. (D) IL-13 and IL-5 secretion of ILC2s sorted from lungs after acute or chronic papain-induced lung inflammation, stimulated for 48h with IL-33 (Multiple T tests, IL-13: **p=0.0019; IL-5, **p=0.0047). Bars represent mean ± SEM. Data information: are representative of 2 independent experiments. (E) Percentage of eosinophils (Siglec-F+CD11b+ cells) among CD45+ cells was measured after acute or chronic papain-induced lung inflammation (T test, **p=0.0038). Bars represent mean ± SEM. Data information: are representative of 2 independent experiments. (F) Id2-CreERT2- Maffl/fl and Id2-CreERT2+ Maffl/fl mice were treated with tamoxifen and papain following the chronic schedule. (G) Representative histogram and summary of the percentage of c-Maf+ ILC2s from lungs of treated mice (T test, *** p=0.0001). Bars represent mean ± SEM. (H) Activation status of lung ILC2s from 5 miceper condition treated as in (F). Bars represent mean ± SEM. (I) IL-13 and IL-5 secretion of sorted lung ILC2s after chronic papain-induced lung inflammation and 48h stimulation with IL-33 (Multiple T tests, IL-13: **p=0.0026; IL-5: ***p=0.00027). Bars represent mean ± SEM. (J) Representative dot plot and percentage of eosinophils (Siglec-F+CD11b+ cells) among CD45+ cells was determined after chronic papain-induced inflammation in the indicated mouse strain (T test, **p=0.0088). Bars represent mean ± SEM. (K) Lungs from these mice were fixed and used for hematoxylin-eosin staining. Representative pictures from 5 mice for each genotype are shown.
Figure 8(A) Lesion sizes (left) were determined 5 days after C. higginsianum (Ch) inoculation. Invasion rates (right) of Ch on Arabidopsis leaves were determined 3 days after spore inoculation. An average of 3 replicates is shown (±SD) (left) and (±SE) (right). (■, p < 0.05 in two-tailed tests between the indicated samples. *, p < 0.05 in two-tailed tests compared to the corresponding WT values. Two independent experiments were combined for statistical analysis.) B) Immunoblot (left) and qRT-PCR (right) analyses following Ch challenge for the indicated times (dpi). The numbers below the immunoblots represent relative intensities of the indicated bands normalized to the backgrounds, with the values at 0 dpi set as 1.0. Experiments were performed three times with the same conclusions.