EMBO/sd-character-level-ner huggingface dataset:
Figure 1A. Relative abundance of miR species in human naive CD4 T cells. Cell purity was >97%. B, C. Average miR expression levels in unstimulated naive CD4 T cells non-infected vs HIV-1NL4-3 (B) or HIV-2ROD (C) infected after 24h (n=2, three pooled samples per library). Mean normalized read counts across libraries are shown. Library normalization was performed using the mean normalization method available in the Deseq package of Bioconductor (Anders Huber, 2010). Lines indicate changes of +/- 1 log2 fold between samples.
Figure 5A Identification of DCAF12 substrates. Control (WT) or DCAF12 knockout (KO) A375 cells were subjected to quantitative TMT proteomics to identify potential DCAF12 targets. MAGE-A proteins (shown in blue) are stabilized in DCAF12 knockout cells. B Knockout of DCAF12 rescues degradation of MAGE-A proteins in A375 cells. DCAF12 KO A375 cells were treated with complete media or EBSS before quantitative TMT proteomics. Note MAGE-A genes are not significantly altered by EBSS in DCAF12 KO cells. C DCAF12 target proteins are differentially affected by EBSS compared to remainder of the proteome. DCAF12 targets (n=33) identified show significantly greater decrease upon EBSS treatment compared to non-DCAF12 altered genes (n=8243). Average with standard deviation is shown with statistical analysis by student t-test. Asterisks *** indicate p<0.001 (student t-test). D MAGE-A2, MAGE-A3, MAGE-A6, and MAGE-A12, but not other MAGE-A genes contain a DCAF12 degron motif (C-term -EE*). E Overexpression of DCAF12 decreases wild-type MAGE-A3, but not MAGE-A3 degron mutant. The indicated constructs were expressed for 48 hrs in HeLa cells before immunoblotting. The MAGE-A3 degron mutant was created ], in which DNYNEPKANQ* were added to the extreme C-term to obscure the DCAF12 degron motif. F Analysis of proteomics indicates that MAGE-A1, MAGE-A4, MAGE-A10, and MAGE-A11 are not regulated by nutrient deprivation or DCAF12. Average (n=3) with standard deviation is shown with statistical analysis by student t-test. Asterisks *** indicate p<0.001 (student t-test). G MAGE-A10 and MAGE-A11 are not altered upon nutrient deprivation or DCAF12 deletion. A375 parental or DCAF12 KO cells were treated with EBSS for the indicated times. Cell lysates were probed for the indicated proteins.
USP45 is recruited to sites of damage following UV‐C irradiationU2OS cells were transiently transfected with the GFP–USP45 or GFP alone.Figure 8A U2OS cells were transiently transfected with the GFP-USP45 or GFP alone. Thirty‐six hours post‐transfection cells were irradiated with UV‐C (150 J/m2) through a micropore filter (lower panels) or left untreated (upper panels). Five minutes post‐irradiation, cells were fixed and co‐immunostained with antibodies recognising cyclobutane pyrimidine dimer (CPD) DNA lesions and GFP. The white arrows indicate co‐localisation between GFP-USP45 and CPD. Scale bar, 10 μm. B U2OS wild‐type (WT) and USP45 knockout (KO) cells were irradiated with UV‐C (20 J/m2). At the indicated post‐irradiation times, genomic DNA was extracted and subjected to Southern dot blot analysis using antibodies recognising CPD and dsDNA total antibody. Data are shown from a triplicate experiment in which each blot is derived from independent cells. C Same as (B) but using a High Sensitivity CPD/Cyclobutane Pyrimidine Dimer Elisa kit (NM‐MA‐K001) from Cosmo Bio. For the DNA damage detection, the manufacturer protocol was followed. Absorbance at 492 was measured that represent amount of CPDs in each sample at indicated times. Calf thymus DNA, UV‐Cirradiated (10 J/m2) and not irradiated was used as positive and negative samples, respectively.
Figure 2A) Example genomic region of MSL2 MNase-ChIP-seq profiles in early (stage 5-8, orange) and late (stage 13-15, purple) mixed-sex embryos. DNA binding is defined as sub-nucleosomal fragments (10 - 130 bp), while chromatin binding as mono-nucleosomal fragments (130 - 220 bp). Called peaks/regions are represented as boxes above the tracks and HAS below. Late-appearing regions are marked by a plus (+) sign. Graph represents pooled IP/Input signal of 3 biological replicates. B) Average composite plots of sub-nucleosomal MSL2 MNase-ChIP-seq (DNA) centered at PionX (left) or HAS (right) in early and late mixed-sex embryos. Total number of PionX = 56 and HAS = 247. Graph represents average IP/Input signal of 3 biological replicates. Shaded area indicates the 95% confidence interval across sites. C) Average composite plots of mono-nucleosomal MSL2 ChIP-seq (Chromatin) centered at PionX (left) or HAS (right) in early and late mixed-sex embryos.
Effects of the deletion of APG5 or APG7 on Apg12p–Apg10p thioester formation.Figure 3Effects of the deletion of APG5 or APG7 on Apg12p-Apg10p thioester formation. Coimmunoprecipitation was performed with wild‐type (KA311B), Δapg5 (YNM122) or Δapg7 (YTS12) cells harboring HAAPG10 and MycAPG12 on 2μ plasmids. The resulting precipitates were analyzed by Western blotting. The DTT‐resistant Apg12p-Apg5p conjugate is indicated by ●.
Figure 6Structure derived from a Bd1971∆EAL construct (yellow/orange), in comparison to cAMP-bound full-length structure (dark/light blue, interdomain linker helices in red/pink). Residues from the opposing monomer labeled by a '. (A) The apo sensor overlaid with the cAMP-bound sensor indicates a relative swing out of the cNMP body relative to C-helix central stator (motion indicated by arrow); the apo form would now sterically clash with the linker helix in the full-length bound structure. (B and C) Comparison of P-loop environment in both apo and bound states. In the apo form, the P-loop (selected residues in green) migrates out of a hydrophobic pocket formed by the hinge and C-helices, breaking an interaction between R108' and D63. In the bound state, M59 stacks with the nucleotide base and the P-loop shifts upward to contact both sides of the pocket. (D) An alignment via the cNMP body demonstrates that swing out is licensed in part by P-loop twisting/migration - L61 in the bound state allows close packing of the hinge and C-helices, whereas L61 in the apo state would clash with V87 and L88, forcing a relative shift of the body and helical subdomains. (E and F) Engagement of the cAMP base and M59 with the C-helix interface. The two orthogonal views demonstrate that L105 and L106 form a hydrophobic plane at the C-helix interface, under which the M59 sidechain can pack against the cAMP base and allow the cNMP body to adopt the swung in conformation.
Figure 2A Muscle sections show variable labeling using an antibody against glycosylated α-dystroglycan (αDG-IIH6), whereas labeling using antibodies against α-dystroglycan core protein (αDG-Core), β-dystroglycan (βDG) and laminin-2 is similar to control (Scale bar, 100 μm). B Western blots and ligand-overlay (O/L) of wheat-germ agglutinin-enriched muscle and fibroblasts lysates from PII.2, PII.5, the healthy sibling (HII.3) and healthy controls (Ctr, Ctr1, and Ctr2) (Muscle: 250μg protein/lane, Fibroblasts: 800μg/lane). In muscle, expression of αDG-IIH6 is reduced but the αDG-Core shows similar expression as controls, with a slight reduction in molecular weight; laminin overlay assay detected some binding activity but was diminished compared to controls, whereas agrin-binding activity showed no difference with controls. In fibroblasts, both αDG-IIH6 and αDG-Core expression and laminin-binding activity were normal, suggesting that unlike other muscular dystrophies, α-dystroglycan glycosylation defect is muscle-specific in POGLUT1 patients. C No ultrastructural alterations are observed in muscle by electron microscopy (6-7 fields were captured from PII.1, PII.3, and PII.5). Note normal basement membrane compactation (arrowheads). Original magnification: 26,700.
Figure 1(A 3rd instar larval fat bodies were stained by BODIPY (green) for lipid droplets and DAPI (blue) for nuclei. Scale bars represent 25 µm. (A) Top: schematic diagram of the TAG biosynthetic pathway. Bottom: overexpression of DGAT1 (ppl>DGAT1) does not affect lipid droplet size. (B) Relative TAG levels in different genetic backgrounds were measured by glyceride assay kit. For quantification, TAG levels were normalized to protein. Error bars represent ± SEM. ***: P < 0.001; NS: non-significant. C, 3rd instar larval fat bodies were stained by BODIPY (green) for lipid droplets and DAPI (blue) for nuclei. Scale bars represent 25 µm. (C) The lipid droplet size is not increased when larvae are reared on a 30% high-fat diet. ND: normal diet; HFD: high-fat diet. , D 3rd instar larval fat bodies were stained by BODIPY (green) for lipid droplets and DAPI (blue) for nuclei. Scale bars represent 25 µm. (D) CSN RNAi alone does not affect lipid droplet size, but CSN RNAi dramatically increases the lipid droplet size when DGAT1 is overexpressed. (E) Quantification of the lipid droplet size in (A) and (D). data were analyzed by one-way ANOVA with a post hoc Turkey's multiple-comparison test. Each point represents data from one fat body, and at least 30 cells were analyzed for each fat body. Error bars represent ±SEM. ***: P < 0.001; **: P < 0.01; NS: non-significant. F) 3rd instar larval fat bodies were stained by BODIPY (green) for lipid droplets and DAPI (blue) for nuclei. Scale bars represent 25 µm. (F) Lipid droplet size is increased in CSN7 RNAi larvae fed on a 30% high-fat diet. ND: normal diet; HFD: high-fat diet. (G) Quantification of the lipid droplet size in (C and F). ND: normal diet; HFD: high-fat diet. data were analyzed by one-way ANOVA with a post hoc Turkey's multiple-comparison test. Each point represents data from one fat body, and at least 30 cells were analyzed for each fat body. Error bars represent ±SEM. ***: P < 0.001; **: P < 0.01; NS: non-significant.
Figure 4A. The Pearson correlation analysis of RNA-seq results of oocyte samples from unidentified control patients and MOSAsn95Lys patient 1. B, C. Scatter plots showing transcriptional changes between human GV and MII stage oocytes from unidentified control patients (B) and between control and MOSAsn95Lys MII oocytes (C), separately. Genes that were increased or decreased more than two-folds are indicated in red and blue, respectively. RPKM, reads per kilobase per million mapped reads. D. Upregulated gene numbers based on different expression level in control and MOSAsn95Lys MII oocytes. E. Venn diagram showing the MOS-regulated decay genes, overlapped between the maturation-directed decay genes (downregulated transcripts from GV to MII oocytes from control individuals) and the upregulated genes in MOSAsn95Lys oocytes. F. Heatmap showing the expression dynamics of MOS-regulated decay genes in GV and MII oocytes from control or MOSAsn95Lys patient. G. The enriched biological process of upregulated MOS-regulated decay genes in MOSAsn95Lys oocytes, obtained via Gene Ontology (GO) analysis. The transcripts numbers of each biological process were labeled on the right. H. The bar graphs show the expression levels of selected MOS-regulated decay genes (n=2 or 3 biological replicates).
a–f, Electron micrographs of MCF7.figf2a-f, Electron micrographs of MCF7.control (clone 38) (a, b) and MCF7.beclin1 (clone 17) cells (c- f) grown in nutrient-rich media (a, c) or subjected to 4 h of serum and amino-acid deprivation (b, d-f). Asterisks in e and f denote autophagic vacuoles that would be counted in the experiment shown in g. Autophagic vacuoles were defined as double-membrane vacuolar structures containing recognizable cytoplasmic contents. Scale bars, 1 µm. g, Quantitative effects of beclin 1 on basal and nutrient-deprivation-induced autophagy of MCF7 cells. Bars indicate mean (±s.e.m.) number of autophagic vacuoles per cell for cells growing in normal media (black), for cells subjected to 4 h of serum and amino-acid deprivation (grey), and for cells pre-treated for 30 min with 10 mM 3-MA and subjected to 4 h of serum and amino-acid deprivation in the presence of 3-MA (open). Mean was determined by counting the total number of autophagic vacuoles in each cell for 100 cells per clone per treatment. h, Comparison of the rates of degradation of long-lived proteins in MCF7.control cells (squares, clone 38), MCF7.beclin1 cells (circles, clone 17) and MCF7.beclin1stop cells (triangles, clone 70), in EBSS + 10% serum and complete amino acids (open symbols, solid lines) in EBSS alone (closed symbols, solid lines), and in EBSS + 10 mM 3-MA (closed symbols, dotted lines). Results are mean (±s.e.m.) of triplicate wells. Similar results were obtained in three independent experiments. Results shown with these clones are representative of results obtained with other clones analysed by electron microscopy in g.
Figure 1Immunoblotting of SPATC1L in testicular cells (TC), testicular sperm (TS), and epididymal sperm (S). An anti-ADAM2 antibody was used as a control. Expression of SPATC1L in human testis lysates (TL) and mature sperm (S). An anti-α-tubulin antibody was used as a control. Developmental expression pattern of SPATC1L in testis, examined by immunoblotting using total lysates obtained from prepubertal and adult male mice (days 7, 14-21, 28, 35, 42, 49, and 56). An anti-α-tubulin antibody was used as a control. Localization of SPATC1L in germ cells during spermatogenesis. Isolated spermatogenic cells were immunostained for SPATC1L (red) and γ-tubulin (green). γ-Tubulin was used as a centrosome marker, and cells were counterstained with the nuclear dye Hoechst 33342 (blue). Fluorescence images of individual staining are shown at the top and merged images are shown in the middle. Insets are magnified iamges. Sc, spermatocyte; RS, round spermatid; ES, elongating spermatid; TS, testicular sperm; MS, epididymal mature sperm; BF, overlay with bright field image.
Cardiomyocyte specific modulation of pyruvate metabolism cell-autonomously affects their dedifferentiation and proliferation following cardiac injury.Figure 3(a) Transgenic animals and experimental time course for data shown in (b-e). Immunostaining of heart sections for PCNA and MEF2 in 5 dpci animals (Tg(hsp70l:LOXP-STOP-LOXP-pdha1aSTA-T2A-mCherry) and Tg(hsp70l:LOXP-STOP-LOXP-pdk3b-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); magnified view of area in white boxes shown below; arrowheads point to PCNA+ CMs; white dashed lines mark the wound border; percentage of PCNA+ CMs in the border zone shown on the right (n = 4-5 ventricles of each genotype). Immunostaining of heart sections for PCNA and MEF2 in 5 dpci animals (Tg(hsp70l:LOXP-STOP-LOXP-pdha1aSTA-T2A-mCherry) and Tg(hsp70l:LOXP-STOP-LOXP-pdk3b-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); magnified view of area in white boxes shown below; arrowheads point to PCNA+ CMs; white dashed lines mark the wound border; percentage of PCNA+ CMs in the border zone shown on the right (n = 4-5 ventricles of each genotype). Immunostaining of heart sections for N2.261 and MEF2 in 5 dpci animals (Tg(hsp70l:LOXP-STOP-LOXP-pdha1aSTA-T2A-mCherry) and Tg(hsp70l:LOXP-STOP-LOXP-pdk3b-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); magnified view of area in white boxes shown below; arrowheads point to N2.261+ CMs; white dashed lines mark the wound border; percentage of N2.261+ CMs in the border zone shown on the right (n = 4-5 ventricles of each genotype). Immunostaining of heart sections for N2.261 and MEF2 in 5 dpci animals (Tg(hsp70l:LOXP-STOP-LOXP-pdha1aSTA-T2A-mCherry) and Tg(hsp70l:LOXP-STOP-LOXP-pdk3b-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); magnified view of area in white boxes shown below; arrowheads point to N2.261+ CMs; white dashed lines mark the wound border; percentage of N2.261+ CMs in the border zone shown on the right (n = 4-5 ventricles of each genotype). (f AFOG staining of heart sections from 60 dpci animals (Tg(hsp70l:LOXP-STOP-LOXP-pdha1aSTA-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); black dashed lines outline the scar area; scar area measured on the right (n = 4-5 ventricles of each genotype). g) AFOG staining of heart sections from 60 dpci animals and Tg(hsp70l:LOXP-STOP-LOXP-pdk3b-T2A-mCherry) alone (control) or in combination with Tg(myl7:Cre-ERT2), all after tamoxifen and heat shock treatments); black dashed lines outline the scar area; scar area measured on the right (n = 4-5 ventricles of each genotype).
Figure 5(a) Schematic of repressive interactions via Notch-Delta between neighbouring cells whereby the effects of HES5 protein in Cell 1 (marked as P1) on transcription in Cell 2 and vice-versa are represented using an inter-cellular Hill function J(P1, 2(t−τND)) where t denotes time and τND represents the intercellular time delay, the time interval required to synthesise the intermediate molecular species (detailed in Figure EV4a); HES5 auto-repression is represented using an intra-cellular Hill function G(P1, 2(t−τH)) where τH represents the inter-cellular time delay, the time interval required for protein to be produced and repress its own transcription. (b) Mathematical description of the inter-cellular Hill function and its parameters: time delay (τND), repression threshold (P0) and Hill coefficient (n); (bottom left panel) higher P0 corresponds to reduced inter-cellular repression (i.e. decreased coupling strength) and conversely lower P0 corresponds to higher coupling strength; (bottom right panel) increasing values of n correspond to increased steepness of the inter-cellular response. (c) Multiscale coupled mathematical model of the tissue environment consisting of a 2D hexagonal grid of cells expressing HES5 protein with corresponding auto-repression (described in (a)) coupled together by repressive interactions between its 6 immediate neighbours (see Methods); single cell inter-cellular repression is a Hill function (with parameters described in (b)) dependent on mean protein abundance in the neighbouring cells. (d) Parameter exploration of single cell temporal period emerging from the model at different repression threshold and time delay values. (e) Parameter exploration of phase synchronisation quantified using the Kuramoto Order Parameter (see Materials and Methods) where 1 indicates global in-phase activity and 0 indicates no co-ordination of phase between cells. (f) Parameter selection strategy combining experimentally determined temporal phase (insert left panel) and KOP (insert right panel) values in spinal cord tissue (see (Manning et al., 2019) and Materials and Methods) to indicate areas where model statistics (i.e. mean temporal period and KOP of synthetic data) resemble real tissue; values within ±1SD and 2±SD from the mean of the tissue are identified and values found outside of ±2.4SD from the mean of tissue are excluded. (g) Representative examples of synthetic kymograph data obtained at specific levels of repression threshold: Alternating high-low (P0 = 400), Global in phase (P0 = 15000) and Local in phase (P0 = 21000) and corresponding KOP values; the presence of microclusters at weak coupling is indicated with red arrowheads; time delay 150min, n = 4. (h) Kymograph data obtained in the absence of coupling between cells; phase relationships are un-coordinated resulting in a KOP≈0. (i-j) Synthetic data timeseries corresponding to simulations in (g).
SDHB and IDH2 protein levels are associated with Gleason grade A.Figure 5A. SDHB protein expression levels in a TMA (n = 83) detected by immunohistochemistry (IHC). Box-plot shows median, 1st and 3rd quartiles, and whiskers extend to ± 1.5 interquartile range. Jitter represents single values in groups. Kruskal-Wallis test and Dunn's all pairs test were applied. Q = adj. p.-value, n.s. = not significant. TMA = Tissue Micro Array. B. Representative IHC- staining of SDHB in normal prostate glands and Gleason grade (GL) 3 - 5 PCa glands. Scale bar = 100µm. C. IDH2 protein expression levels in a TMA (n = 83) detected by IHC. Box-plot shows median, 1st and 3rd quartiles, and whiskers extend to ± 1.5 interquartile range. Jitter represents single values in groups. Kruskal-Wallis test and Dunn's all pairs test were applied. Q = adj. p.-value, n.s. = not significant. D. Representative IHC staining of IDH2 in normal prostate glands and GL 3 - 5 PCa glands. Scale bar = 100µm. E. Nuclear (N) and cytoplasmatic (C) STAT3 protein expression levels in a TMA (n = 83) detected by IHC. Box-plot shows median, 1st and 3rd quartiles, and whiskers extend to ± 1.5 interquartile range. Jitter represents single values in groups. Kruskal-Wallis test and Dunn's all pairs test were applied. Q = adj. p.-value, n.s. = not significant.
Phylogeographic analysis revealed global transmission patterns of human-associated S.Figure 5 Figure 5 Phylogeographic analysis revealed global transmission patterns of human-associated S. suis. Countries of relevance in the map are colored. Geographic presence, inferred arrivals (arrows) and principal long-distance transmission events (line) are based on phylogeographic analysis. The date are the inferred values for the most recent common ancestor (taken from BEAST2). The origin of most transmission waves was Europe. One of these transmission waves was spread to China leading to two major outbreaks (Jiangsu and Sichuan) (red line).
mtON activation increases the PMF.Figure 2Representative TMRE fluorescence traces in arbitrary units (a.u.) before and after Δψm dissipation with FCCP. Dashed lines indicate where FCCP was added. +/- ATR traces were performed in the absence of succinate. mtON activation was continuous throughout the traces. Light green trace is from mitochondria with ATR, gray traces are without. Quantification of change in fluorescence (ΔF) normalized to the maximum change in fluorescence given by succinate respiration (ΔFmax) for each mitochondrial preparation. Data are from the maximum light dose in Fig. EV 3A. Two-way ANOVA with Sidak's test for multiple comparisons, *p = 0.0469, +ATR succinate vs. +ATR +light p = 0.9978, n = 6 mitochondrial isolations. Data show mean ± SEM. Representative BCECF-AM 490/440 nm ratio trace. Ratio of 545 nm fluorescence intensity at either 440 or 490 nm excitation. Dashed lines are where light or FCCP treatment occurred. Light green trace is from mitochondria with ATR, gray traces are without. Quantification of change in BCECF-AM fluorescence ratio normalized to maximum change given by succinate matrix alkalization. Two-way ANOVA with Sidak's test for multiple comparisons, -ATR succinate vs. -ATR, +light *p = 0.0212, +ATR succinate vs. +ATR light p = 0.999, +ATR succinate vs. +ATR,-light p = 0.0237, +ATR, +light vs, +ATR, -light p = 0.0474, n = 3 mitochondrial isolations. Some individual points overlap. Data show mean ± SEM. ATP levels normalized to total ATP synthesis given by succinate respiration. Data from Fig. EV 3B. Succinate data shown for comparison after normalization. Two-way ANOVA with Sidak's test for multiple comparisons, *p = 0.0011, +ATR succinate vs +ATR light p = 0.5680, n = 3,7,3,7, biological replicates for each bar from left to right. Some individual points overlap. Data show mean + SEM. O2 required to consume 50 nmoles of ADP after mtON activation. Data are the maximum illumination from Fig. EV 4D using time-matched dark conditions, One-way ANOVA *p = 0.013 (-ATR, -light vs. +ATR, +light p = 0.010. -ATR, +light vs. +ATR, +light p = 0.013.) -ATR, -light n = 5, rest n = 6 mitochondrial preparations. Data show mean + SEM.