EMBO/sd-character-level-ner huggingface dataset:
Figure 6(A) Striped ptc.Gal4-driven expression of UAS.GFP in a control wing imaginal disc. Scale bars ~20µm. n > 6 independent biological replicates. (B) Striped ptc.Gal4-driven expression of UAS.GFP in an aurA homozygous mutant wing imaginal disc leads to increased numbers of pH3 positive mitotic cells. n > 7 independent biological replicates. (C) Striped ptc.Gal4-driven expression of UAS.GFP and UAS.aurB-RNAi in a control wing imaginal disc leads to enlarged cells with large nuclei (marked by DAPI; white arrowhead). n > 6 independent biological replicates. (D) Striped ptc.Gal4-driven expression of UAS.GFP and UAS.aurB -RNAi in an aurA homozygous mutant wing imaginal disc leads to extrusion and apoptosis of cells (marked by Dcp1 and pyknotic nuclei). n > 8 independent biological replicates. (E) Striped ptc.Gal4-driven expression of UAS.GFP and UAS.pbl-RNAi in the wing imaginal disc leads to extrusion and apoptosis of cells (marked by Dcp1 and pyknotic nuclei). n > 8 independent biological replicates. (F) ptc.Gal4-driven expression of UAS.aurB-RNAi in an aurA homozygous mutant endoreplicating salivary gland has no phenotypic consequence. n > 6 independent biological replicates. (G) MARCM clones (GFP-positive) expressing UAS.aurB-RNAi in an aurA homozygous mutant cells result in single cell clones (i.e.: no cell division), while individual loss of AurA or AurB allows some cell proliferation. Quantification shown on the right (n > 7 independent samples per genotype; Mean +/- 1 S.D. shown). Scale bar ~20µm.
Figure 9A. Schematic representation of RF bicistronic transcript is shown above the bar graphs. Rluc: Renilla luciferase (blank box); Fluc: Firefly luciferase (solid box). The luc activity of HEK 293T cells transfected with bicistronic mRNA, as indicated, in the presence (+) or absence (-) of endouc-Flag mRNA was determined. The relative luc activity of transfected cells shown on the right was normalized by the amount of the transfected bicistronic transcript shown on the left. The relative luc activity was represented by the fold increase of Fluc/Rluc ratio over that obtained from control group, normalized as 1. The luc activity was measured by dual-luciferase assay, and its corresponding bicistronic mRNAs were measured by RT-qPCR. The values were calculated from three independent experiments and presented as mean ± S.D. (n=3). Student's t test was used to determine significant differences between each group (***: P <0.001). B. The luc activity of Fluc (solid column) versus Rluc (blank column) in HEK 293T cells transfected with bicistronic mRNAs shown in Figures A was presented as an increase in fold over that obtained from control group of RF which was normalized as 1. These data suggested that the change of Fluc/Rluc ratio shown in Figures A was entirely dependent on the change of Fluc activity. The data were averaged from three independent experiments and presented as mean ± S.D. (n = 3). Student's t test was used to determine significant differences between each group (**: P<0.005). C. Schematic representation of hpRF bicistronic transcript. Rluc: Renilla luciferase (blank box); Fluc: Firefly luciferase (solid box). The luc activity of HEK 293T cells transfected with bicistronic mRNA, as indicated, in the presence (+) or absence (-) of endouc-Flag mRNA was determined. The relative luc activity of transfected cells shown on the right was normalized by the amount of the transfected bicistronic transcript shown on the left. The relative luc activity was represented by the fold increase of Fluc/Rluc ratio over that obtained from control group, normalized as 1. The luc activity was measured by dual-luciferase assay, and its corresponding bicistronic mRNAs were measured by RT-qPCR. The values were calculated from three independent experiments and presented as mean ± S.D. (n=3). Student's t test was used to determine significant differences between each group (***: P <0.001). D. The luc activity of Fluc (solid column) versus Rluc (blank column) in HEK 293T cells transfected with bicistronic mRNAs shown in Figures C was presented as an increase in fold over that obtained from control group of hpRF which was normalized as 1. These data suggested that the change of Fluc/Rluc ratio shown in Figures C was entirely dependent on the change of Fluc activity. The data were averaged from three independent experiments and presented as mean ± S.D. (n = 3). Student's t test was used to determine significant differences between each group (***: P <0.001).
Figure 5Representative images showing cleaved-caspase 3 staining in colonic section from WT or Ang-/- colitis mice (n = 3) at indicated time point; arrows indicate cleaved-caspase 3 positive cells; two fields per colon section are quantified. the number of apoptotic cells in colonic section from WT or Ang-/- colitis mice (n = 3) at indicated time point Cell apoptotic status of IECs from WT or Ang-/- colitis mice (n = 5) on day 4. Cell apoptotic status of IECs from WT or Ang-/- colitis mice (n = 5) on day 4. Representative images showing Ki67 staining in colonic section from WT or Ang-/- colitis mice (n = 3) at indicated time point; five representative crypts per colon section are quantified. the number of proliferating cells per crypt in colonic section from WT or Ang-/- colitis mice (n = 3) at indicated time point; five representative crypts per colon section are quantified. Cell-cycle status of IECs from WT or Ang-/- colitis mice (n = 5) on day 4. Representative images showing cleaved-caspase 3 staining in colonic section from Angfl/fl and LyzMCre;Angfl/fl, or AAV-shControl- and AAV-shPlxnb2- treated colitis mice on day 4 (n = 3); arrows indicate cleaved-caspase 3 positive cells; two fields per colon section are quantified. the number of apoptotic cells in colonic section from Angfl/fl and LyzMCre;Angfl/fl, or AAV-shControl- and AAV-shPlxnb2- treated colitis mice on day 4 (n = 3) Ki67 staining and (L) the number of proliferating cells per crypt in representative colonic section from Angfl/fl and LyzMCre;Angfl/fl, or AAV-shControl- and AAV-shPlxnb2- treated colitis mice on day 4 (n = 3); five representative crypts per colon section are quantified.
Ubiquitylation of nascent/newly synthesized proteins during acute heat stress or exposed to the proteasome inhibitor MG132A.Figure 4A. Lysates from MCF7 cells exposed to the proteasome inhibitor MG132 (20 μM, 1 h) or to heat shock (42 oC, 1 h) were subjected to sucrose density gradient centrifugation. Monosomal and polysomal fractions were quantified from the UV traces at 254 nm. Monosomes eluting in fractions 4- 6 and polysomes in fractions 7 - 14 were quantified and polysome to monosome (P/M) ratios were calculated. Average ratios and the individual datapoints (n = 5) are displayed in a bar graph (means +/- SD). Numbers above the bars represent p values calculated with the two-stage linear step-up procedure of Benjamini, Krieger and Yekutieli. Gradient fractions were assayed for the presence of lysine (K) 48-linked polyubiquitylated proteins by immunoblotting with K48 linkage-specific antibodies. The line graph on the right is a quantification of K48-linked polyubiquitin across the gradients relative to the levels obtained in MCF7 cells maintained at 37 oC (means +/- SEM, n = 4). B. The same experiment as described in (A) but with MCF7 cells exposed to MG132 (20 μM, 1 h) or puromycin (100 μg/ml, 10 min). Means +/- SEM, n = 2. C. Lysate from MCF7 cells exposed to MG132 (20 μM, 1 h) or puromycin (100 μg/ml, 10 min) as indicated were subjected to immunoprecipitation with anti-puromycin antibodies, followed by assaying for co-precipitation of K48-ubiquitylated proteins. Actin is shown as a reference.
Aβ generation in cells after autophagic modulation.Figure 5.Aβ generation in cells after autophagic modulation. Levels of Aβ40 (A), Aβ42 (B), βCTF (C), and APP (D) measured by sandwich ELISA after the incubation of L/APP cells (6 h) in conditions that block autophagy (+Leu, +His, +Leu/+His, and 5 mM 3MA), activate macroautophagy (−Leu, −His, −Leu/−His, and rapamycin), or do not affect autophagy (complete media and enrichment of the deprivation of Gly or Val). Values reported as percent difference of control ± SEM; *, P < 0.05 (at least). In similar experiments, Aβ40 and Aβ42 levels from SH-SY5Y (E and F) and N2a cells (G and H) after various treatments. Error bars represent SEM.
UBTD1 is associated with β-catenin at the cell-cell adhesion site (A) Immunoblot of DU145 and A549 cells at various cell densities ranging from 20% to 100% confluence showing the increased expression of UBTD1 and β-catenin with cell confluency.Figure 2(A) Immunoblot of DU145 and A549 cells at various cell densities ranging from 20% to 100% confluence showing the increased expression of UBTD1 and β-catenin with cell confluency. E-cadherin was used as a positive control. Tubulin was used as a loading control. , immunofluorescent labelling of UBTD1 and E-cadherin (B) at the cell-cell contact in confluent DU145 cells and analyzed by confocal microscopy. Right, representative histogram of a co-localization profile prepared with RGB profiler plugin of ImageJ. immunofluorescent labelling of UBTD1 and β-catenin (C) at the cell-cell contact in confluent DU145 cells and analyzed by confocal microscopy. Right, representative histogram of a co-localization profile prepared with RGB profiler plugin of ImageJ. Proximal ligation assay monitoring (left) and quantification (right) of UBTD1 association with E-cadherin or β-catenin in DU145 cells transfected with the indicated siRNA (control, siCTRL; UBTD1: siUBTD1pool). Nuclei were stained with DAPI (Blue) on the MERGE image. Immunoblots of UBTD1, β-catenin and YAP after subcellular fractionation of confluent DU145 cells transfected with the indicated siRNA (control, siCTRL; UBTD1: siUBTD1pool). EGFR, RhoGDI and Lamin A/C were used as controls for the membrane (M), cytosol (C) and nucleus (N) compartments, respectively. Co-immunoprecipitation between endogenous β-catenin and UBTD1 was performed in confluent DU145 cells. IgG isotype was used as a negative control. (G) Representative confocal images of β-catenin and UBTD1 in DU145 under resting (Ctrl), after calcium chelator treatment (EGTA) and recovery (Rec) conditions. Nuclei were stained with DAPI (Blue) on the MERGE image. (H) Representative confocal images of β-catenin and UBTD1 in DU145 after cell scattering induced by HGF treatment (24 h). Nuclei were stained with DAPI (Blue) on the MERGE image.
Fig.Figure 3A X-ray cocrystal structure of XMU-MP-5 with wild-type ALK (PDB ID: 7BTT). Specific interactions are depicted as dash lines, hydrogen bonds are shown in yellow and salt bridge is colored red. B Structure analysis of XMU-MP-5 with ALK(L1196M) mutant (PDB ID: 2YFX) by molecular docking. Carbon chains of L1196M (yellow) model were overlaid on wild-type ALK (blue). C Comparison of XMU-MP-5 bound with wild-type ALK (blue) and G1202R (purple). ALK(G1202R) homology model was developed based on the published X-ray structure of ALK(L1196M) (PDB ID: 2YFX).
Figure 2A. Wide-type (RKIP+/+) and RKIP deficiency (RKIP-/-) chimera were intraperitoneally injected with VSV (1x107 pfu/g), 12 hours later, ELISA of IFN-β and IL-6 in serum (n=6 per group). B. Determination of VSV loads in organs by TCID50 assay 12 hours later after RKIP+/+ and RKIP-/- chimera mice (n = 6 per group) intraperitoneally injected with VSV (1x107 pfu/g). C. Real-time PCR analysis of IFN-β mRNA and VSV expression in liver from RKIP+/+ and RKIP-/- chimera mice 12 hours later intraperitoneally injected with VSV (1x107 pfu/g) (n = 6 per group). D. Pathology of wild type and RKIP deficient chimeric mice in response to VSV infection. Hematoxylin and eosin staining of liver and lung sections from mice in (A). Scale bar, 100 μm (left) or 50 μm (right). E. Real-time PCR analysis of IFN-β mRNA expression in peritoneal macrophages from RKIP+/+ and RKIP-/- chimera mice 12 hours after intraperitoneally injected with VSV (1x107 pfu/g) (n = 6 per group). F. Survival curve for 8 weeks-old wild type and RKIP deficient chimeras challenged with VSV (1x108 pfu/g, i.v.) (n=10 per group). *p<0.05 Wilcoxon Test .
Fig1.Figure 1, The model explicitly accounts for reactions of metabolism and gene expression processes. Metabolic reactions produce metabolites and energy for not only biomass formation but also gene expression processes. The reactions of gene expression, on the other hand, synthesize RNA and proteins, which catalyse reactions of metabolism and gene expression processes as machineries or enzymes. In addition, pcLactis accounts for degradation of mRNA and proteins as well as dilution of biomass constituents during cell division. Coupling constraints in pcLactis. The coupling constraint allows for relating the reaction rate to the synthesis rate of its catalyst based on the reaction rate equation and steady-state assumption, where turnover rates kcat of metabolic enzymes, catalytic rates k of machineries and degradation constants kdeg of the catalysts are needed. Simulated exchange fluxes compared with experimentally measured data (Goel et al, 2015). Simulated saturation of glucose transporter. , Simulated inactive enzyme. The inactive enzyme is the sum of the enzymes that are synthesized but do not carry fluxes. The production of the inactive enzyme indicates that total proteome is not constrained. The grey area represents where the proteome switch occurs in experiments (Goel et al, 2015), which is between 0.5 and 0.6 h-1. , Sensitivity analysis for glucose transporter and modelled proteome at different growth rates. Colour represents the sensitivity score. A higher score indicates a greater impact of a given increase in the constraint on growth rate.
Figure 7(A) siRNA-based knockdown of Pex3, Pex16, and Pex19. HeLa cell lysates treated with the indicated plasmids and siRNAs were immunoblotted with the indicated antibodies. (B) A comparable amount of MITOL was detected in control, PEX3-, PEX16-, and PEX19-knockdown cells. HeLa cell lysates stably expressing HA-Parkin and 3Flag-MITOL treated with the corresponding siRNAs were immunoblotted with the indicated antibodies. (C) MITOL translocation from the mitochondria to peroxisomes is highly dependent on Pex3. HeLa cells stably expressing 3Flag-MITOL and HA-Parkin were transfected with control, PEX3, PEX16, or PEX19 siRNA, treated with 15 µM CCCP for 3 hours, and then subjected to immunocytochemistry with anti-Flag and anti-catalase antibodies. Scale bars, 10 µm. (D) Correlation statistics for the localization of 3Flag-MITOL and catalase in cells transfected with the indicated siRNAs. Dots indicate individual Pearson correlation coefficient data points. In the box-plots, the center lines indicate the medians, the box limits indicate the 25th and 75th percentiles as determined in the R software package, and the whiskers extend 1.5 times the interquartile range from the 25th and 75th percentiles. Means and the number of samples are shown on the box and X-axis, respectively. Statistical significance was calculated using a one-tailed Welch's t-test.
Myosin VI contributes to OFD1 removal from the centriolar satellites required for ciliogenesis.Figure 6(A) A scheme of the IF analysis performed to calculate the total intensity of OFD1 staining at the centriolar satellites. (B) IF analysis of OFD1 signal at the centriolar satellites upon serum starvation. hTERT-RPE1 p53 KO cells were transfected with siRNA against myosin VI. After four days, cells were fixed (growing) or serum starved for 24 hours (SS). Cells were immunostained with anti-OFD1 and anti-PCM1 antibodies. The intensity of OFD1 signal in the area covered by PCM1 was quantified and normalised against the intensity of PCM1 staining in the same area. Left, representative images, scale bar, 2 μm. Right, results are expressed as fold change with respect to mock average intensity. Bars represent mean ±SD. Mock_growing, n=150 cells; Mock_SS, n=149 cells; MyoVI KD_growing, n=149 cells; MyoVI KD_SS, n=150 cells, from three independent experiments. ns, not significant; * P<0,05; **** P<0,0001 by Kruskal-Wallis test. (C) IB analysis of OFD1 after serum starvation in control and myosin VI-depleted cells. hTERT-RPE1 p53 KO cells were transfected with siRNA against myosin VI. After four days, cells were serum starved for 24 hours (SS). Lysates were analysed by IB with anti-OFD1, anti-myosin VI, anti-p62, and anti-LC3 antibodies. Anti-GAPDH was used as loading control. The amount of OFD1 protein was normalised against GAPDH signal and is expressed as percentage of OFD1 levels in cells grown in serum-starved conditions compared to cells grown in media containing serum. Bars represent mean ±SD. n=5 independent experiments. * P<0,05 by Kruskal-Wallis test. (D) IF analysis of primary cilium upon serum starvation. hTERT-RPE1 p53 KO cells were transfected with siRNA against myosin VI. After four days, cells were fixed (growing) or serum starved for 24 hours (SS). Cells were immunostained with anti-acetylated tubulin (to identify the cilia), anti-Cep135 (to identify the centrioles) and DAPI. Left, representative images, scale bar, 2 μm. Right, results are expressed as fold change with respect to mock average intensity. Bars represent mean ±SD. n=3 independent experiments. 100-200 cells/condition were counted for each experiment. ***P < 0.001 by two-way ANOVA test.
Figure 1(A) NET-seq data of wild type (WT) yeast at the GCG1/SUT098 and ORC2/SUT014 loci. NET-seq reads in black and grey represent the Watson (+) and Crick (-) strands, respectively. The NET-seq track combines remapped data from previous publications (see Material and Methods for more details). (B) Schematic representation of a shared promoter region initiating a coding (blue) and a DNC (purple) transcript in the opposite orientation. Fluorescent protein reporter (FPR) construction replaces the endogenous coding and DNC genomic regions with sequences encoding mCherry and YFP, respectively. (C) SGA and flow cytometry analysis in budding yeast. The array involves the crossing of MATα query strain harboring the FPR construct with the MATa non-essential gene deletion mutant library. The systematic selection as described in Baryshnikova et al, 2010 results in haploids containing the gene deletion and FPR. Quantification of the resultant haploids by high-throughput flow cytometry records fluorescent signals in up to 50,000 cells for each gene deletion mutant. (D, E) Scatter plot visualization of YFP vs mCherry fluorescence signals in the GCG1 and ORC2 promoter FPR screens. Each data point represents the median signal of a deletion mutant normalized to individual plate median. Highlighted data points represent the mutants of Hda1C (purple), SAGA acetylation module (blue), SAGA deubiquitination module (orange), and cac2∆ (yellow). The regression line is marked in black. The mutants favoring DNC transcription are found above the regression line. The mutants favoring coding transcription are below the regression line. Technical repeat = 1, biological observations = 50,000 individual cells before filtering.
Metabolomics of mutant DHFR strains in minimal media without or with added amino acids.Figure 2(A) and (B) shows abundance of selected nucleotides and amino acids for mutants I91L+W133V and W133V as well as WT strain treated with 0.5μg/ml of Tmp (Trimethoprim) after 4 hours of growth at 40/42°C in M9 minimal medium (condition of no filamentation), Concentration of all metabolites were normalized to WT levels at 4 hours when grown under similar conditions. In minimal medium (B) Methionine levels are extremely low Levels of purines (IMP, AMP) are also largely rescued with amino acid supplementation, however dTMP, dTDP and dTTP levels remain extremely low. Error bars represent SEM of at least three biological replicates (see Methods). The dashed line in each plot represents value of 1 for WT. (C) and (D) represents nucleotide and amino acid abundances after 4 hours of growth in amino acid supplemented M9 medium at 40/42°C (condition of filamentation). Concentration of all metabolites were normalized to WT levels at 4 hours when grown under similar conditions. Methionine levels are extremely low, which recover largely in panel (D). Levels of purines (IMP, AMP) are also largely rescued with amino acid supplementation, however dTMP, dTDP and dTTP levels remain extremely low. Error bars represent SEM of at least three biological replicates (see Methods). The dashed line in each plot represents value of 1 for WT.
Figure 7Nicotine- and acetylcholine-induced whole-cell current traces of α7-nAChRs expressed in Xenopus oocytes recorded in the absence or presence of CTα16 or CTα16scr, respectively. Note specific robust potentiation of nicotine- and acetylcholine-induced currents by CTα16, but no direct activation of α7-nAChRs by either peptide in the absence of agonist (not shown). Bars indicate application of EC50 agonist concentrations and of the peptide indicated (green: CTα16: grey: CTα16scr). Quantification of potentiation shows that CTα16 enhances nicotine- and acetylcholine-induced currents to a similar extend whereas CTα16scr had no effect. CTα16 dose-response curve obtained with submaximal nicotine concentrations (100 µM) with a corresponding EC50 value of 10.7 ± 4.6 nM and a hill-coefficient of 2.1 ± 0.3 (n=4). Nicotine-induced whole-cell current traces of α7-nAChRs in the presence of 20 nM recAPPsα (green bar) or 20 nM recAPPsβ (blue bar). Note that only recAPPsα efficiently potentiates α7-nAChRs, which is blocked by BTX (orange bar). Quantification of potentiation shows that recAPPsα enhances nicotine-induced currents to a similar extend as CTα16 (see for comparison Figure 7B) while recAPPsβ had no effect. Nicotine dose-response curve obtained either in the presence of 20 nM recAPPsα (green circles, left) or without treatment (black circles, right). Note that recAPPsα treatment shifts the curve to the left, significantly reducing the apparent agonist affinity by about 2-fold (EC50 of 40.3 ± 6.0 µM, green curve vs. EC50 of 80.5 ± 12.7 µM, black curve; n=5; *p<0.05). For statistical evaluation of potentiation we performed a paired two-tailed Students t-test of the agonist-induced currents in the absence and presence of the indicated peptide. n=number of oocytes. Data represent mean ± SEM. nsp>0.05, *p<0.05.
Model describing the photoperiod response of flowering and hypocotyl elongation in WT plants.Figure 5Simulated levels of FT and ATHB2 mRNA (as calculated by taking the area under the curve) across multiple photoperiods. Using FTAREA values, the number of days to flower was calculated and compared to data from Corbesier et al (1996) (see Supplementary Information). Error bars represent standard deviation. Simulated flowering: solid black line, filled squares; data: dashed black lines, empty squares. Same as in (C) using ATHB2AREA to calculate hypocotyl length. Data taken from Kunihiro et al (2011) (error bars were unavailable).