EMBO/sd-character-level-ner huggingface dataset:
Fat body ERK7 inhibits cell autonomous and systemic growthA.Figure 4A. Weight of control and ERK71 mutant larvae at 72h after egg deposition (AED) (N=4 replicates of 12 larvae/replicate for each genotype). B. Pupariation kinetics of control and ERK71 mutants (N=4 replicates of 30 larvae/replicate for each genotype). p<0.0001, analyzed by Log-rank test. C. ERK71 mutants fail to suppress growth upon 24 h of starvation (N≥4 replicates of 15 larvae/replicate for each genotype). D. ERK71 mutant larvae (N=4 replicates of 24 larvae/replicate for each genotype, p<0.01 analyzed by Log-rank test) are sensitive to nutrient deprivation. Fat body specific knockdown of ERK7 by RNAi (BDSC 56939) leads to a moderately accelerated pupariation rate (E, p<0.001 analyzed by Log-rank test, N=8 replicates of 30 larvae/replicate for each genotype) F. Fat body specific knockdown of ERK7 by RNAi (BDSC 56939) leads to an increase in larval weight at 72h AED (F, N=5 replicates of 15 larvae/replicate for each genotype). G. A representative image showing that fat body-specific overexpression of ERK7 by the CG-GAL4 driver (CG>ERK7) impairs larval growth. Scale bar: 0.5 mm. H. Ectopic expression of ERK7 leads to delayed pupariation (p<0.0001, analyzed by Log-rank test), while kinase-dead (K54R) and activation loop phosphorylation-deficient (T190A/Y192F) mutants of ERK7 do not influence pupariation rate (N=4 replicates of 30 larvae/replicate for each genotype). I. In contrast to ERK7 wt, kinase-dead (K54R) and activation loop phosphorylation-deficient (T190A/Y192F) mutants of ERK7 do not influence the pupal volume (N=4 replicates of 10 pupae/replicate for each genotype).
Figure 2Immunoblots showing CIB1 (A) protein levels in WT and co-1 backgrounds (A) over the course of 20 hours under LDs. Samples were separated on 10% SDS-PAGE, blotted, probed with the anti-Myc antibody, stripped and re-probed with the anti-actin antibody (ACT). B. Immunoblots showing CO (B) protein levels in WT and cibh backgrounds (B) over the course of 20 hours under LDs. Samples were separated on 10% SDS-PAGE, blotted, probed with the anti-Myc antibody, stripped and re-probed with the anti-actin antibody (ACT). The relative levels of CIB1 in WT and co-1 backgrounds over the course of 20 hours under LDs were calculated Data information: Error bars represent the standard deviation of three biological replicates of the Western blots. D. The relative levels of CO proteins in WT and cibh backgrounds over the course of 20 hours under LDs were calculated Data information: Error bars represent the standard deviation of three biological replicates of the Western blots.
Model for the role of Sinhcaf in germ plasm aggregation and subsequent PGC specification is mediated by Kif26ab.Figure 8(A) Sinhcaf is the Sin3-HDAC complex associated factor. In WT eggs, Sinhcaf promotes deacetylation of histone 3, which is essential for activating transcription of kif26ab. In turn, Kif26ab promotes recruitment of germ plasm to furrows and subsequent PGC specification. (B) In sinhcaf -/- eggs, transcription of kif26ab is blocked, resulting in failed germ plasm recruitment to specify PGCs.
Figure 2A: Wild type and mps1 mutant cells were synchronised in G1 with α-factor at 25°C and then released at 34°C in the presence of nocodazole (t=0). Cells were collected at the indicated time points for FACS analysis of DNA contents. B: Cells were treated as in (A) and after 2 hours from the release α-factor was re-added to prevent cells from undergoing a second cell cycle. Cell samples were collected at the indicated time points for western blot analysis of the indicated proteins. Pgk1 was used as loading control. Cyc: cycling cells. C: Top: in vitro kinase assays with recombinant Mps1 and Mps1-3 purified from E. coli and incubated at 34°C for the indicated times in the presence of radioactive ATP and a recombinant Spc105 N-terminal fragment (aa 1-320) as substrate. Bottom left: kinase activity was quantified on the autoradiographs by Image J and normalized to the levels of the full length protein on the Coomassie Blue-stained gel (right). Bottom right: 1μg of the recombinant Mps1 kinases used for kinase assays was loaded on SDS page and stained with Coomassie Blue for normalisation of kinase assays. D: Wild type and mps1-3 cells were synchronised in G1 with α-factor at 25°C and then released at 34°C in the presence of nocodazole. Cells were collected after 90 minutes and fixed with formaldehyde for ChIP-seq analysis. ChIP sequence reads were normalised against sequence reads from corresponding input samples, and relative enrichment is plotted for chromosome III around the centromere (see the centromeric regions of all 16 yeast chromosomes in Fig. S2). Y‐axis shows enrichment values (linear scale, range is 0-10). Values below 1.5 are shown in grey, and values above 1.5 (i.e. sequences enriched in ChIP samples) are red coloured. E: Cells with the indicated genotypes were synchronised in G1 with α-factor at 25°C and then released in fresh medium at 34°C (t=0). Cells were collected at the indicated time points for FACS analysis of DNA contents.
Figure 7(A) Dipeptides accumulation under stress conditions depends on the autophagy and protein recycling Thirumalaikumar et al., 2020(). Interestingly, GAPC1 was reported to inhibit autophagy by an interaction with the ATG3 protein Han et al, 2015Henry et al., 2015(; ). ROS mediated disruption of the GAPC1-ATG3 complex promotes autophagy. (B) Inhibition of the GAPC activity redirects the glycolytic pathway towards PPP, resulting in the production of NADPHRalser et al., 2007(). While oxidative inactivation of GAPC is a fast event Ralser et al., 2009Zaffagnini et al., 2019(; ), we speculate that Tyr-Asp inhibition becomes critical under prolonged stress to maintain high levels of NADPH production. In contrast to GAPC, GAPN is unaffected by Tyr-Asp and less sensitive to oxidation. (C) NADPH is used to replenish the pool of reduced glutathione (GSH), needed to mitigate oxidative stress damage. Tyr-Asp binding to chaperones suggests its role in protein folding, possibly further contributing to the Tyr-Asp related stress protection. Figure was prepared using Biorender.com
Figure 4A B (top). Sequences interacting with ChiX (A) or ompA (B) were analyzed for conserved motifs using MEME. Motifs that were enriched in the target RNAs and are shown above the heatmap with proportion of target RNAs carrying the motif (N) and the expected value for the motif (e-val). The complementary sequence motifs within ChiX or ompA are shown below the logo. (Middle) Heatmaps showing the position of predicted basepairing for each interaction within ChiX or ompA. Grey bars boxed below indicate the position of basepairing with experimentally verified mRNAs (known interactions). (Bottom) Hfq (blue) and RNase E (grey) binding sites within ChiX and ompA. Hfq and RNase E-bound sequence, and non-genomically encoded oligo(A) tails within ChiX and ompA are shown as line plots where the x-axis position correlates with heatmaps above. C, D E. Cumulative plots of oligoadenylation (C), Hfq (D) and RNase E (E) binding at predicted seed motifs.
Divalent cation bindingA.Figure 5A. Superimposition of Rb+/Ba2+ and Ba2+ bound EAAT1CRYST structures with tranD depicted in pink and green, respectively, and TM4 removed for clarity. Black mesh depicts anomalous-difference map in the absence of Rb+ contoured at 4σ around the tranD, and inset shows details of Ba2+ coordination at Na3. Residue numbering corresponds to EAAT1WT, and the coordination distance (angstrom) is parenthesis. B. Ca2+ binding isotherms of EAAT1CRYST at pH=7.4 in the presence (pink) and absence (blue) of UPCH101. Mutation D400380N at Na3 strongly impairs Ca2+ binding (green). Symbols represent average and s.e.m. values of at least three independent experiments. Solid lines represent fits of Hill equation and dashed line is the fit in Fig. 1C (blue line) corresponding to Na+ binding. C. Log-log plot of L-asp KD as a function of Ca2+ (n=13).
Figure 1(A-D) Serum NfL concentrations of preataxic (green) and ataxic (red) SCA3 subjects and controls (blue) were measured in two independent cohorts, each with a different Simoa approach: cohort #1, recruited by the ESMI consortium (A, C), and cohort #2, recruited by the EuroSCA/RiSCA consortium (B, D). Boxes show the ranges between lower and upper quartiles, the central bands show the medians, and the whiskers show data within 1.5∙IQR of the median, with dots representing outliers. Groups were compared with Mann Whitney U tests (*** p<.001, ** p<.01, ns p≥.05, two-tailed, Bonferroni-corrected; In the scatter plots, the individual NfL values were plotted as a function of subjects' age. The dashed grey lines visualise the optimal cut-offs for differentiating ataxic SCA3 subjects from controls in each cohort (cohort #1: 20.0 pg/ml, 98.7% sensitivity, 92.2% specificity; cohort #2: 50.9 pg/ml, 92.6% sensitivity, 100% specificity; cut-offs were derived by maximising Youden's index irrespective of age). Note the logarithmic scale of the y-axes. (E, F) Serum pNfH levels of preataxic and ataxic SCA3 subjects and controls were also measured in both cohorts, each with a different Simoa approach (two-tailed Mann Whitney U tests, Bonferroni-corrected;
Figure 4A. Reduced weight (in grams) of adult Rb∆K7 (n=11) vs control littermate (n=17) males. Results are presented as mean ±SD; P values calculated by two-tailed unpaired student's t-test. (B) number of litters from breeding of homozygous Rb∆K7 (n=4) or control wildtype (n=5) males with control wild-type females. Bars represent mean ±SD; P values calculated by two-tailed unpaired student's t-test. (C) Representative images of testes from two Rb∆K7 mice compared with a control littermate. (D) Sections through seminiferous tubules. Arrows point to mature spermatids in control testes that are lacking in Rb∆K7 adult testes. Scale bar, 20 μμ. E. Kyphosis in Rb∆K7 mice. Top, images of eight-month old Rb∆K7 mouse with kyphosis (right) and control wildtype littermate (left). Bottom, radiographs of these mice under anesthesia showing abnormal cervical curvature (arrows) in the Rb∆K7 knock-in mouse. F-I. Delayed hair re-growth in eight-month old Rb∆K7 mice two weeks post hair plucking (F). (G) Abnormal skin histology in Rb∆K7 mice relative to control. FCL, subcutaneous fat cell layer, T, telogen; A, anagen. (H-I) Representative immunostaining for keratin 14 and PCNA showing reduced expression in Rb∆K7 hair follicles vs controls. Scale bar, 50 μm. J. Hyperglycemia in Rb∆K7 but not Rb∆K4 mice. Blood glucose in 10-12-month old fasting Rb∆K7 (left, n=11) or 18-month old fasting Rb∆K4 (right, n=15) mice versus control littermates (n=6 and n=8, respectively). Mean ±SD; P values by two-tailed unpaired student's t-test.
Figure 7HeLa cells were co-transfected with wildtype or K95A GFP-TDP NLS, HA-ubiquitin and iRFP or GA175-iRFP. 24h after transfection, cells were treated with rolipram (30 µM), MG132 (10 µM) or DMSO (vehicle) for 16h. Lysates were immunoprecipitated with Protein G beads coupled with anti-GFP antibody.
Abundance of subunits of protein complexes and kinases in the tetrasomic cell line.Figure 3(A) Density plots of subunits of protein complexes (as defined in the CORUM database) encoded on all disomic chromosomes compared with non‐CORUM proteins (left panel); the same for proteins encoded on the tetrasomic chromosome 5 (right panel). The differences between CORUM and non‐CORUM populations of chromosome 5 are statistically significant (Wilcoxon rank sum test). Dashed lines indicate medians of the populations. (B) Examples of protein complexes with at least one subunit coded on the tetrasomic chromosome 5. Each dot represents abundance changes of one gene (black), its corresponding mRNA (blue) and protein (red). Subunits coded on chromosome 5 are indicated with an arrow and number. (C) The abundance of DNA, mRNA and proteins of kinases coded on chromosome 5. See also Supplementary Figure S3. Source data is available for this figure in the Supplementary Information.
Telomere dysfunction increases with age in mouse cardiomyocytes.Figure 1Representative images of γH2AX immuno-FISH in troponin-C/α-actinin (top left and bottom left panels, respectively) positive mouse CMs (white - troponin-C/α-actinin; red - telo-FISH; green - γH2AX). Images are Z-projections of 0.1μm stacks taken with a 100x oil objective. Scale bar: 20μm. Right panels represent a single Z-plane where co-localisation between a γH2AX foci and telomere was observed, Scale bars 1μm. (Below) 3D reconstruction of Immuno-FISH using STED microscopy for γH2A.X and telomeres in a CM from 30 month old mice. Scale bars as indicated and scale the same for both individual TAF examples using STED. Mean number of TAF α-actinin positive CMs from C57BL/6 mice. Data are mean ± SEM of n=4-7 mice per age group. α-actinin positive CMs from C57BL/6 mice. Data are mean ± SEM of n=4-7 mice per age group. 100 α-actinin positive CMs were quantified per mouse. Mean number of total γH2A.X foci in α-actinin positive CMs from C57BL/6 mice. Data are mean ± SEM of n=4-7 mice per age group. 100 α-actinin positive CMs were quantified per mouse. % of γH2A.X foci -positive nuclei (E) in α-actinin positive CMs from C57BL/6 mice. Data are mean ± SEM of n=4-7 mice per age group. 100 α-actinin positive CMs were quantified per mouse. Fold enrichment of γH2AX at telomere repeats by real-time PCR. Graph represents fold enrichment of γH2AX at telomeric repeats between IgG control, 3 and 30 month old whole mouse hearts. Data are mean ± SEM of n=3 mice per age group. Quantitative PCR-ELISA TRAP assay comparing telomerase activity of 3 and 30 month old C57BL/6 mice whole heart lysates. Data are mean ± SEM of n=4 mice per age group. Histograms displaying distributions of telomere intensity in CMs analysed by 3D Q-FISH in young (4 months) and old (30 months) wild-type mice. Data are from n=3 mice. > 100 CMs were analysed per mouse. % of telomere FISH signal loss in 30 months old wild-type mice and late generation Terc-/- mice (6 months old) in comparison to 4 months old wild-type mice. Data are from n=3 mice. >100 CMs were analysed per mouse.
AML-EV impair ribosome biogenesis in hematopoietic cells and suppress protein synthesis in LT-HSC.Figure 3A Venn diagram showing the most highly up- and downregulated proteins between Molm-14-EV and HL-60 EV -treated c-kit+ HSPC determined by isobaric TMT labeling and edge-R analyses to calculate the false discovery rate False (FDR) analysis to obtain ~10% of the differentially regulated proteins. We obtained 325 and 221 upregulated proteins in the HL-60 dataset and the Molm-14 dataset, respectively. We also obtained 166 and 173 downregulated protein in the HL-60 dataset and the Molm-14 dataset, respectively. Among them, 54 proteins were commonly downregulated. A bar graph showing the functional annotation enrichment analysis by DAVID biostatistical plate form of the 54 commonly downregulated protein identified the indicated pathways, among them the ribosomal biogenesis pathway showed the highest enrichment. P-values are indicated in white. A heatmap showing the highly-deregulated ribosomal proteins in AML-EV exposed cells. Flow cytometric analysis showing the histograms and MFI of O-propagyl-puromysin (OPP) incorporation in LT-HSC in: Molm-14-xenografts, red n=6 vs. non-engrafted controls, black n=6, Flow cytometric analysis showing the histograms and MFI of O-propagyl-puromysin (OPP) incorporation in LT-HSC in: IF injection of EV from Molm-14, U-937, HL-60 (red, n=5,43), AML patients plasma (Orange, n=6), or human CD34+ cells (blue, n=3) normalized to vehicle-injected contralateral femurs after subtracting the background fluorescence.
Figure 6Transcriptional profiles reflect the dynamic immune response in COVID-19. GSEA (FDR < 0.25; 1,000 permutations) was used to identify positive (red), negative (blue) or no (white) enrichment of BTMs (gene sets). The graph shows the normalized enrichment score (NES) of each selected BTM in the different time points (T1, T2 and T3) for patients with mild or severe COVID-19 illness, in comparison to healthy controls. The timepoints 1, 2, 4 or 1, 3, 5 were used as T1, T2 and T3 for patients with four or five sampling timepoints, respectively. Expression levels of NETs' markers for individual transcripts in severe (n=37) versus mild (n=179) comparisons. Data were represented as means ± SEM and differences between groups were estimated using a Student's t test. *p < 0.05; ** p < 0.01; ***p < 0.001. Heatmap of IFNG, GZMB and PRF1 gene expression in COVID-19 patients. Average expression values were centered and scaled. Red indicates a higher expression and blue indicates a lower expression. Heatmap of the expression of genes enriched in IL-17 signaling pathway between healthy control and COVID-19 patients. Average expression values were centered and scaled. Red indicates a higher expression and blue indicates a lower expression. cell and innate immune response elucidate immunopathology of COVID-19.
Figure 1Green line: Normalized population expression level averaged over all OxyR, and SoxS-regulated promoters; red line: average over all LexA-regulated promoters as measured with a promoter-GFP plasmid library in a plate reader. Oxidative stress promoters clearly precede SOS response promoters in response to NIT stress, when measured at the population level Error bars show standard deviation over all detected oxidative stress and SOS promoters, respectively It is not clear if this temporal order correctly reflects the temporal order in single cells. B. Schematic showing response of two different genes (green and red) in three different cells (solid, dashed, dotted line). The temporal order observed at the population level correctly reflects the temporal order in each individual cell. C. As B. Here the temporal order at the population level is not the same in every single cell: Although in most cells, the green gene responds before the red one, one cell expresses the green gene before the red one (dashed lines).
Promoter enrichment‐quantitative mass spectrometry (PE‐QMS) identifies transcriptional regulators of Abca1 expressionRT–qPCR of Abca1 transcripts following 1 μM T0901317 stimulation of LXR+/+ and LXR−/− primary BMMs.Figure 1RT-qPCR of Abca1 transcripts following 1 μM T0901317 stimulation of LXR+/+ and LXR−/− primary BMMs. Fold changes are shown relative to LXR+/+ 0 h. Error bars represent ± SEM for n = 4-8 (**P = 0.001 at 8 h and **P = 0.00004 at 16 h versus LXR−/−; **P = 0.0003 at 0 h versus LXR+/+); bottom, proposed model explaining the ligand‐dependent induction of Abca1. Following ligand stimulation, LXR-corepressor complexes are exchanged for LXR-coactivator complexes, which promote gene expression. The LXR-cofactor interactions responsible for the attenuation of expression beginning after 16 h remain unknown. Overview of the PE‐QMS experimental strategy to identify LXR ligand‐stimulated and LXR binding‐dependent interactions with the Abca1 promoter. Nuclear extracts were prepared from RAW 264.7 macrophages treated with control or 1 μM T0901317 (depicted as a black circle) for 18 h. The Abca1 promoter region was used to isolate gene regulatory complexes under the indicated conditions. Immobilized templates were washed, and bound proteins eluted and digested. Peptides were purified, then identified, and quantified by mass spectrometry. Heat map of the PE‐QMS experiment. Ligand‐dependent subsets were quantified using label‐free and isotope labeling approaches and are indicated on the left. Colors represent Log2 relative abundance ratios (T0901317/vehicle) of identified proteins. LXR binding‐dependent subsets were quantified using an isotope labeling approach and are indicated on the right. Changes in binding > 1.5‐fold were employed as cutoffs for depicted subsets. ND = not determined. Full dataset is available as Supplementary Table S1.
Figure 2(A) Immunofluorescence staining against Iba-1 in Cb of ASMko and wt mice treated or not with PLX for 2 months. DAPI staining shows cell nuclei. Scale bar, 100 μm. (B) Magnified images (from A) showing ramified (left) versus amoeboid (right) morphology in wt and ASMko microglia treated or not with PLX. Scale bars, 30 μm. (C) Mean ± SEM number of Iba-1 positive cells in the Cb of the different mouse groups (n = 7 mice per group, Two-way Anova, Bonferroni post hoc). (D, E) Performance in the Rotarod test of ASMko and wt mice treated or not with PLX for 2 months expressed as the mean ± SEM time spent in the rotating rod on each of four trials (D) or the mean ± SEM time of the four trials (E) (n = 7 mice per group, Two-way Anova, Games-Howell (D) and Bonferroni (E) post hoc). (F) Mean ± SEM body weight of ASMko and wt mice treated or not with PLX at the indicated time during long-term treatment (n= 7 mice per group, Two-way Anova, Bonferroni post hoc). (G) Survival curve of ASMko and wt mice with or without long-term PLX treatment (n = 7 mice per group). (H) Immunofluorescence staining against the PC marker Calbindin in mid and posterior lobules of Cb in ASMko and wt mice treated or not with PLX for 2 months. DAPI staining shows cell nuclei. Scale bar, 500 μm. (I) Mean ± SEM number of Calbindin positive cells in the anterior (lobules I-V), middle (lobules VI-VIII) and posterior zone (lobules IX-X) of the Cb from the different mouse groups (n = 7 mouse per group, Two-way Anova, Bonferroni post hoc).
Figure 5A) HEK293 cells were transfected to co-express KDF1 with USP7. IP and WCL were blotted with different antibodies as indicated. B) Immunoprecipitation was carried out with α-KDF1 antibody in keratinocyte cultured in medium with low or high concentration of calcium (Ca), same as in Fig. 1D. IPs were blotted with different antibodies as indicated. The star denotes KDF1. C) WCL from WT and USP7 CRISPR KO cells were subjected to immunoblotting with different antibodies as indicated. D) IKKα protein level in WT and USP7 KO keratinocyte were examined by immunoblotting before and after calcium shift. E) WT and USP7 KO keratinocyte were treated with MG132 at 10μM for 6 hours, then subjected to immunoprecipitation using anti-ubiquitin antibody. Precipitated product was analyzed by immunoblotting with IKKα antibody. Band intensity was determined by densitometry and shown as bar graphs. Statistical analysis is conducted using unpaired Student's t-test. Error bar represents S.D. (standard deviation). N=3 (biological replicates). **, p<0.01. Kd: kilodalton for molecular weight markers. F-G) Immunoblot of Krt10 and Loricrin (Lor) with WT and USP7 KO cells before and after calcium shift (F). Star denotes an unspecific band in α-Loricrin blots. Band intensity was determined by densitometry and shown as bar graphs Statistical analysis is conducted using unpaired Student's t-test. Error bar represents S.D. (standard deviation). N=3 (biological replicates). ****, p<0.0001; **, p<0.01. H) Sections of engrafted skin developed from USP7 KO or control WT cells were immunostained with different antibodies as indicated. Dotted lines denote dermal-epidermal boundaries. Epi: epidermis, Der: dermis. Scale bar=100μm. I) A working model of epidermal differentiation regulated by KDF1. Upon differentiation signals, KDF1 associates with IKKα and recruits deubiquitination enzyme USP7 to the protein complex. USP7 can deubiquitinate IKKα and promote its protein stability, which will in turn promote epidermal differentiation.