EMBO/sd-character-level-ner huggingface dataset:
Inhibiting the SERCA pumps, activation of the IP3Rs and chelation of ER Ca2+ reduced ANO8-dependent current inactivation.Figure 5(A) Plot of the slope of inactivation as a function of current density in the presence (red) and absence of ANO8 (black). Increasing current density was obtained by varying external Ca2+ between 2-50 mM. The results are mean±s.e.m and difference were analyzed by unpaired t test. Data information: The first number in parenthesis indicates the number of similar experiments performed and the second number is the number of cells analyzed. All results are given as mean±s.e.m of the indicated number of experiments or cells analyzed. (B, C) CRAC current was measured in HEK cells expressing STIM1 and Orai1 and in pipette solution containing 3 mM EGTA (b) or 10 mM BAPTA (c). SERCA pump was inhibited with 25 µM CPA in the red traces where indicated and 100 µM IP3 was included in the pipette solution (green in c). Data information: The first number in parenthesis indicates the number of similar experiments performed and the second number is the number of cells analyzed. All results are given as mean±s.e.m of the indicated number of experiments or cells analyzed. (D, E) Experimental protocol and conditions except that cells were also transfected with ANO8. Data information: The first number in parenthesis indicates the number of similar experiments performed and the second number is the number of cells analyzed. All results are given as mean±s.e.m of the indicated number of experiments or cells analyzed. (F, G) HEK cells expressing STIM1, Orai1, and ANO8 were used to measure current in pipette solution containing 10 mM BAPTA and with (red) or without (black) 100 µM IP3 or 10 mM of the ER Ca2+ chelator TPEN (green). The current density (F) and the normalized current (G) are shown, illustrating the delay in current inactivation by IP3 and TPEN. Data information: The first number in parenthesis indicates the number of similar experiments performed and the second number is the number of cells analyzed. All results are given as mean±s.e.m of the indicated number of experiments or cells analyzed.
Figure 2(A) Schematic diagram of intraperitoneal administration of tamoxifen (TM) for continuous 5 days before intradermal injection of LL37 in mice (Raptor cKO and WT mice). Mice were sacrificed on day 2 to conduct subsequent experiments. The mouse experiments were repeated for three times, and 5-8 mice were included in each group for each time. The results of a representative mouse experiment were displayed. (B) Immunostaining of Raptor in skin sections from WT and Raptor cKO mice. Epi, epidermis. Der, dermis. Scale bar: 50 μm. (C) The back skins of WT and Raptor cKO mice were intradermally injected with LL37 or control vehicle (n=6 for each group). Images were taken 48 h after the first LL37 administration. Below panels, magnified pictures of yellow boxed areas. Scale bar: 2 mm. (D) HE staining of lesional skin sections from WT and Raptor cKO mice injected with LL37 or control vehicle (n=6 for each group). Scale bar: 50 μm. (E) Dermal infiltrating cells were quantified (n = 6 for each group) for each high power field (HPF). (F) The mRNA expression levels of Tnf-α, Il6, Mmp9 and Vegf in skin lesions (n = 6 for each group). (G) Schematic diagram of topical application of rapamycin (RAPA) after LL37 injection in mice (BALB/c). Mice were sacrificed on day 2 to conduct subsequent experiments. The mouse experiments were repeated for three times, and 4-6 mice were included in each group for each time. The results of a representative mouse experiment were showed. (H) The back skins of LL37 injected-mice topically applied with RAPA (n = 6 for each group). Images were taken 48 h after the first LL37 injection. Bottom panels, magnified images of yellow boxed areas. Scale bar: 2 mm. (I) HE staining of lesional skin sections from LL37 or control mice topically applied with RAPA or vehicle. Scale bar: 50 μm. (J) Dermal infiltrating cells were quantified (n = 6 for each group). Data represent the mean ± SEM. **P < 0.01. 1-way ANOVA with Bonferroni's post hoc test was used.
Figure 8 Figure 8. The multiple substrates of PARN in telomere biology and rRNA biogenesis. PARN regulates the expression of telomere-related gene transcripts, including shelterin transcripts and the hTR RNA component of telomerase. How PARN stabilizes hTR has been well described and involves its 3' end deadenylation. However, at present, the mechanisms underlying PARN-dependent regulation of telomere-related gene transcripts are still unknown and may involve direct or indirect regulations through 3' end deadenylation of either the shelterin mRNAs or some miRNAs that bind to these mRNAs. Our data also show that PARN regulates the level of DKC1 mRNA through the control of p53 levels. Taken together, PARN impacts on both telomere stability and telomere length. An additional role of PARN is to regulate rRNA maturation in both human and mouse cells. Our data suggest that PARN-dependent regulation of rRNA maturation occurs independently of p53. The involvement of PARN in various aspects of cell biology likely explains the severity of the phenotype in Høyeraal-Hreidarsson syndrome patients carrying biallelic PARN mutations.
Figure 5(A) Experimental design to treat HET pups from P2 to P8 with Miconazole. DMSO (controls) and Miconazole-treated pups were sacrificed at P8 or at P28 to evaluate the ON myelination. (B-E) Sox10 (red, oligodendrocyte precursors) and MBP (green, fully differentiated oligodendrocytes) IF on cross-sections of P8 HET ONs after 6 days of treatment with DMSO (D) or Miconazole (E). Miconazole increased the number of Sox10+ cells compared to HETs treated with DMSO (quantified in B) and rescued MBP staining pixel intensity of HET pups to level comparable to those of WT pups (quantified in C). Nuclei (blue) were stained with DAPI. , the error bars represent the SEM of the means; N=3 Statistical significance was obtained by ANOVA (*P<0.05; **P<0.01; ***P<0.001). Scale bars: 50µm (F-I) Sox10 (oligodendrocyte marker, red) and Phospho-ERK (dually phosphorylated forms of active ERK1 and ERK2, green) IF in WT and HET ONs, treated with DMSO (F) or Miconazole (G) between P2 and P8. As previously reported, Miconazole strongly activate ERK signaling pathway (G). Signal intensity quantification in whole ON area is shown in (H), while specific evaluation of pixel intensity around Sox10+ nuclei is shown in (I). Nuclei (blue) were stained with DAPI. the error bars represent the SEM of the means N=2 Statistical significance was obtained by ANOVA (*P<0.05; **P<0.01; ***P<0.001). Scale bars: 50µm (J-M) EM thin sections of P28 WT and HET ONs, treated with DMSO (J,K) or Miconazole (L) between P2 and P8. Different degrees of myelin compaction can be appreciated at high magnification, with HET ONs showing non compacted sheaths (white arrows in K) and vacuoles (yellow arrows). Miconazole treatment rescues almost normal myelin g-ratio, as quantified in M. Nuclei (blue) were stained with DAPI. the error bars represent the SEM of the means N=2 Statistical significance was obtained by ANOVA (*P<0.05; **P<0.01; ***P<0.001). Scale bars: 50µm, except (J-L) (500nm). (N-P') S100β (green, astrocytes) and Sox2 (red, reactive/proliferative astrocytes) IF in the ONs of WT (N,N') and HET (O-P') animals after 6 days of treatment with DMSO (N-O') or Miconazole (P,P'), then sacrificed at P28. Astrocytes undergo remodeling and express Sox2 in Nr2f1 HET animals (O',P'), even after Miconazole treatment. Histogram showing the average number of S100β+/Sox2+ astrocytes per P28 ON section in WT or HET animals treated with DMSO or Miconazole as indicated, with Sox2 high (magenta columns) and low levels (pink columns). Nuclei (blue) were stained with DAPI. the error bars represent the SEM of the means; ; N=2 Statistical significance was obtained by ANOVA (*P<0.05; **P<0.01; ***P<0.001). Scale bars: 50µm
CLU is required for rapid mitotic exit in cabazitaxel-resistant PC3 cells.Figure 9(A) Cell cycle distribution of syncronized PC3 and PC3-R-caba cells 60 min after nocodazole release at indicated time points. (B) Western blot for CLU, Cdc25C, Cdc25C-T48, Cdk1, Cdk1-Y15 and Wee1 in IGRCaP-1 and PC3 parental (WT) or cabazitaxel-resistant (R-caba) cells as well as for PC3-R-caba cells transfected with siSCR or siCLU. Vinculin was used as loading control. (C) Cell survival measured using WST1 in PC3-R-caba cells after SCR or OGX-011 transfection and treatment with 10 nM cabazitaxel for 48h. Error bar represent mean ± SEM, n=3, *** p<0.001 by one way ANOVA followed by Bonferroni's post hoc correction. (D) Cell survival measured using WST1 in PC3 resistant cells after transfection with siSCR or siCLU followed by treatment with cabazitaxel (10 nM; C10), MK-1775 (10 nM; MK10) or both for 72 hours. .Error bar represent the mean ± SEM, n=3, *** p<0.001 by one way ANOVA followed by Bonferroni's post hoc correction. (E) PARP cleavage was assessed by western blot in cells treated as in D and its relative expression was quantified by densitometry using ImageJ. Vinculin was used as loading control (F) Left panel: Quantification of tumour volume over 52 days in nude male mice xenotransplanted with PC3-docetaxel resistant cells and treated with either MK-1775 (M, black line), MK-1775 and micellar taxane (MT, blue line), micellar taxane and OGX-011 (TO, red line) or MK-1775, micellar taxane and OGX-011 (MTO, green line). The error bars indicate mean ± SEM, n=8. Right panel: Slopes of tumor growth between groups, were compared by ANOVA followed by Dunnett's post hoc analysis..*p<0.05, ***p<0.001 .p=0.0488 (M to TO); p=0.0007 (M to MTO).
Figure 2(A) 786-0 cells were subjected to aa starvation for the indicated times. The protein level of GDH1 was determined by WB using the indicated antibody. (B) The mRNA level of GDH1 in the cells in (A) was determined by qPCR using specific primers against GDH1 mRNA. Data information: The data are represented as mean ± SEM of three independent experiments. The one-way ANOVA test were used (N.S.: not significant; ***p < 0.001). (C) 786-0 cells were treated with or without aa starvation for 6 hours and then treated with CHX (1 μM) for the indicated times. The protein level of GDH1 was determined by western blotting (left), and the relative GDH1 protein levels were calculated (right). CHX: cycloheximide. Unpaired t-test was used. Data information: The data are represented as mean ± SEM of three independent experiments. The unpaired t-test were used (N.S.: not significant; ***p < 0.001). (D) Immunofluorescence (IF) assay was performed to determine the subcellular location of GDH1 with or without aa starvation. GDH1 localization was indicated by GDH1 antibody and MitoTracker (left). Signal intensities and distances were quantified (right). Scale bars: 20 μm. (E) Cytoplasm-mitochondrial fractionation assay was performed to determine the subcellular location of GDH1 with or without aa starvation. The fractionation efficiency and GDH1 location were determined by WB using antibodies against VDAC1 (mitochondrial membrane marker) and GDH1. Tubulin was used as loading control. (F) The total GDH1 protein in 786-0 and 769-P cells with or without aa starvation for 12 hours was measured by WB. (G) Immunofluorescence (IF) assay was performed to determine the subcellular location of WT or mitochondrial signal peptide deleted (dMSP) GDH1. GDH1 localization was indicated by GDH1 antibody and MitoTracker (left). Signal intensities and distances were quantified (right). Scale bars: 20 μm. (H) Cytoplasm-mitochondrial fractionation assay was performed to determine the subcellular location of WT or mitochondrial signal peptide deleted (dMSP) GDH1. The fractionation efficiency and GDH1 location were determined by WB using antibodies against VDAC1 (mitochondrial membrane marker) and GDH1. Tubulin was used as loading control. (I) 786-0 cells with FLAG-tagged WT or dMSP GDH1 were treated with CHX (1 μM) for the indicated times. The protein level of FLAG-tagged WT or dMSP GDH1 was determined by western blotting (left). The relative WT or dMSP GDH1 protein level was calculated (right). Unpaired two-tailed t-test was used. Data information: The data are represented as mean ± SEM of three independent experiments. The unpaired t-test were used (N.S.: not significant; ***p < 0.001).
Figure 1A, Western blots in HCT116 cells treated for 24 hrs with increasing concentrations of MPA, or 5 nM Actinomycin D (ActD). GAPDH served as a loading control. B, HCT116 cells were treated for 24 hrs with vehicle alone (-), 1 µM MPA, 10 µM MPA, or the combination of 10 µM MPA with 400 µM guanosine. Guanylate nucleotides levels were measured by LC-MS, and normalized to protein content. Mean ± SEM is representative of 3 independent experiments carried out in triplicate. Data information data are presented as relative to control. *p<0.05, **p<0.01, ***p<0.001 by two tail T-Student's test. Parental HCT116 cells expressing a stable tetracycline-inducible shRNA against IMPDH2 (IM2iKD) and either a stable inducible non-targeting (NT) shRNA (IM2iKD-shNT) or an shRNA targeting IMPDH1 (IM2iKD - shIM1), were grown for 7 days in the absence or presence of doxycycline (2 µg/mL) (- dox or + dox, respectively). mRNAs of IMPDH1 and IMPDH2 were quantified by real-time qPCR in 2 independent experiments carried out in triplicate and normalized to β-actin mRNA (C). Data information: data are presented as relative to control. *p<0.05, **p<0.01, ***p<0.001 by two tail T-Student's test. Parental HCT116 cells expressing a stable tetracycline-inducible shRNA against IMPDH2 (IM2iKD) and either a stable inducible non-targeting (NT) shRNA (IM2iKD-shNT) or an shRNA targeting IMPDH1 (IM2iKD - shIM1), were grown for 7 days in the absence or presence of doxycycline (2 µg/mL) (- dox or + dox, respectively). Whole cell extracts were analyzed on Western blots with the indicated antibodies. GAPDH was used as a loading control. Quantification of band intensity of p53 and p21 is shown. Mean ± SD is representative of four independent experiments. (right panel) (D). Data information: data are presented as relative to control. *p<0.05, **p<0.01, ***p<0.001 by two tail T-Student's test. E, Western blots in HCT116 cells treated for 24 hrs with the indicated concentration of MPA, in presence of dimethyl sulfoxide (DMSO) or 400 µM guanosine. GAPDH was used as a loading control.
Figure 6Morphology of four-week-old soil-grown plants of Col-0 and two independent transgenic lines of 35S-SNIPER1H129Y. Scale bar = 1 cm. PR1 and PR2 gene expression in the indicated plants as determined by RT-PCR. Error bars represent mean ± SD (n = 3). Two independent experiments were carried out with similar results. Morphology of four-week-old soil-grown plants of Col-0, sniper1-1, sniper2-1, eds1-2, SNIPER1H129Y, SNIPER1H129Y eds1-2, sniper1-1 sniper2-1 eds1-2 and sniper1-c1 sniper2-1 eds1-2. sniper1-1 is an exonic T-DNA insertional mutant. sniper1-c1 is a SNIPER1 knockout mutant containing 807bp deletion that was generated by CRISPR-Cas9 system. Scale bar = 1 cm. Quantification of H.a. Noco2 sporulation in the indicated plants 7 days post inoculation (dpi) with 105 spores per ml water. Error bars represent mean ± SD (n=4). Two independent experiments were carried out with similar results. Immunoblot of RPS4HA (E), SNC1 (F), RPS2HA (G), and RPM1myc (H) in SNIPER1H129Y background. Equal loading is shown by Ponceau S staining of a non-specific band. The numbers below represent the normalized ratio between the intensity of the protein band and the Ponceau S band ± SD (n=3). Molecular mass marker in kilo Daltons is indicated on the left. Three independent experiments were carried out with similar results.
Fig.Figure 1A) Upper panels: Representative single z-planes from 3D-SIM image stacks of the DAPI-stained nuclei of AC16 cells kept at 37°C or exposed to different cold temperatures for 24h. Lower panels: DAPI signal segmented into chromatin and interchromatin regions according to relative intensity. Scale bar: 5 μm. Arrows mark chromatin concentrated at the nuclear periphery (P) and around nucleoli (N). B) Boxplots of the ratio of chromatin volume to nuclear volume for cells exposed to different temperature conditions. Median (central line), interquartile range (IQR, box edges), most extreme data point no more than 1.5x IQR from box edges (whiskers). Number of nuclei: 37°C (31), 28°C 24h (26), 18°C 5h (19), 18°C 24h (22), 18°C 24h then 37°C 2h (25), 8°C 24h (23). C) Left and central panels: representative single z-planes from 3D-SIM image stacks of the DAPI-stained nuclei of cells kept at 37°C or exposed to 18°C for 24h showing the spatial distribution of the immunofluorescence (IF) signal of hnRNPC (green) and nuclear pore complexes (NPCs, red). Right panel: DAPI signal overlaid with dots indicating IF signal centroid coordinates for centroids located in the z-plane shown. Scale bar whole nuclei: 5 μm. Scale bar enlarged section: 1 μm. D) Bar graph of the mean NPC centroid to nearest hnRNPC centroid distance for each temperature condition. Number of distances: 37°C (112511), 28°C 24h (94521), 18°C 5h (72340), 18°C 24h (74998), 18°C 24h then 37°C 2h (92896), 8°C 24h (80570). E) Schematic illustrating how chromatin concentrated at the periphery after exposure to 18°C could widen the average NPC to nearest mRNP distance and thus hinder mRNA nuclear export. Data information: Actual p-values are presented in the source data file.
Figure 6(A) Human lung epithelial A549 cells and primary bronchial epithelial cells (hAECB) lack FcγR (CD16, CD64 and CD32) and FcαR (CD89) cell surface expression as analyzed by flow cytometry. Human THP1 cells serve as positive control. (B) The neonatal FcR (FcRn) is expressed in both A549 cells and human THP1 cells as determined by Western Blotting. Data are representative of at least two independent experiments. (C) Absolute bacterial counts (CFUs) in human THP1 macrophages 1 h after infection with MTB pre-incubated with the LAM-reactive IgA1 (left) or IgG1 (right) antibody TB24PB037 or the respective isotype control antibody as indicated. Mean and SEM are indicated.
Figure 8OsPRR73 binds the promoter of OsHKT2;1, by recruiting HDAC10, to negatively regulate the expression of OsHKT2;1 to reduce the import of cellular sodium ion, which subsequently reduced ROS accumulation, thus conferring salinity stress (left panel). In the absence of OsPRR73, the transcription inhibition of OsHKT2;1 was released, hence resulted in sodium ion accumulation. The excessive sodium ion subsequently accumulation caused ROS accumulation, to forming a Na+-ROS stress cascade, leading to decreased survival.
Figure 3 A MA-plot of RNA expression profiles from female Lin-Sca1+ progenitors 2 days after induction of differentiation with 100 nM Rosi, displaying the log2-ratio of Cited4-/- to Cited4+/+ intensities against the average log2-intensities for all microarray probe sets (n=3)  B Comparison of the lists of genes significantly changed by Rosi in wild type cells (Cited4+/+: Rosi vs. Ctrl, P<0.01) or by Cited4-knockout under Rosi treatment (Rosi: Cited4-/- vs. Cited4+/+, P<0.01) in expression profiles from female Lin-Sca1+ progenitors 2 days after induction of differentiation with 100 nM Rosi or vehicl C Comparison of the lists of gene sets significantly enriched by Rosi in wild type cells (Cited4+/+: Rosi vs. Ctrl, false discovery rate (FDR) <0.1) or by Cited4-knockout under Rosi treatment (Rosi: Cited4-/- vs. Cited4+/+, FDR<0.1) in GSEA (KEGG) (n=3)  D-G Enrichment plots from GSEA with the KEGG (D-F) or TFT (G) gene set collection (Cited4-/- vs. Cited4+/+), performed on RNA expression profiles 2 days after induction of differentiation with 100 nM Rosi (n=3). Vertical bars represent the individual genes of the gene set  H Comparison of the lists of gene sets affected by Cited4-knockout under Rosi or under vehicle treatment (Cited4-/- vs. Cited4+/+, FDR<0.1) in GSEA as in (C)
Figure 2A, Treatment of PRL2 with a synthetic substrate leads to complete phosphorylation of the catalytic residue, cysteine 101. Recombinant PRL2 was treated with 3-O-methylfluorescein phosphate (OMFP) and analyzed by SDS-PAGE with the Phos-tag reagent. Phosphocysteine is hydrolyzed by boiling. B, Detection of phosphorylated by electrospray mass spectrometry. OMFP-treatment increased the mass of the intact protein by 79.99 amu but had no effect on the C101A mutant. C, 31P NMR spectrum of the thioPO3 signal from phosphocysteine in PRL2. The broad peak is TCEP in the buffer. D, Cysteine phosphorylated PRL2 does not bind to CNNM3. Recombinant PRL2 was phosphorylated by treatment with OMFP and subjected to pull-down by the CBS-pair domain of CNNM3 as a GST fusion. Only unphosphorylated PRL2 was observed in the pull-down lanes while phosphorylated PRL2 was in the flow-through lanes. E, ITC measurement of the affinity of PRL2 for the CNNM3 GST-CBS-pair domain. Phosphorylation by OMFP completely blocked binding while oxidization of PRL2 by diamide decreased the affinity 200-fold. F, The cysteine residues in the PRL2 active site are not required for binding the CNNM3 CBS-pair domain.
Y244F mSTING mutant translocates to autophagosomes but not late endosomes.Figure 7(A) STING-/- MEF cells were transfected with mSTING WT-GFP and Y244F-GFP (Green) for 24h, pretreated with gefitinib for 1h and transfected with cGAMP for the indicated time; cells were fixed and labeled with LC3 (autophagosome marker, Red) antibody. Co-localization (Col) is indicated by white dots. (A) STING-/- MEF cells were transfected with mSTING WT-GFP and Y244F-GFP (Green) for 24h, pretreated with gefitinib for 1h and transfected with cGAMP for the indicated time; cells were fixed and labeled with CD63 (late endosome marker). (C) A model for STING's trafficking routes in stimulated cells in the presence or the absence of EGFR.
Figure 1A) Differential interference contrast (DIC) image of cortical neurons in culture. The arrowhead points to a neuron expressing Ub-R-GFP. B) GFP fluorescence before and after exposure to lactacystin (10µM). C) A neuron coexpressing Ub-R-GFP and CFP before (left) and 10 hours after exposure to lactacystin (right). Top row - GFP fluorescence; bottom row - CFP fluorescence. Bars: 50µm (A,B), 20 µm (C). D) Changes in GFP and CFP fluorescence following exposure to lactacystin at t=1 hour (vertical dashed line). Changes normalized to initial fluorescence in same cells. 9 neurons from 5 experiments; averages and SEM. E) Same as in D, except that here neurons were exposed only to carrier solution. 11 neurons from 5 experiments; averages and SEM (barely observable). F) Comparison of GFP fluorescence accumulation rates. Linear fits shown as black lines; fit parameters shown next to fits. Same data as in D, and E.
A minority of CAR T cell-tumors interactions results in target cell apoptosis in vitroEµ-myc B cells expressing a FRET-based reporter for caspase 3 activity were labeled with the calcium-sensitive dye Indo-1, incubated with GFP-expressing anti-CD19 CAR T cells and subjected to live imaging.Figure 6(A-B) Calcium responses were recorded over time in (A) unconjugated tumor cells or in (B) tumor cells forming a conjugate with a CAR T cell. Each line corresponds to an individual tumor cell and each square to a time point. Squares are colored in magenta upon calcium elevation or in blue otherwise. Apoptotic events (detected by caspase 3 activity) are indicated by an overlaid green dot. Data is representative of three independent experiment. (C) The number of calcium fluxes in individual tumor cells is plotted for unconjugated tumors or conjugated tumors. Values were normalized to 100 min. Each dot represents one tumor cell. Bars represent mean values. **p<0.01, unpaired t‑test. (D) The cumulated period of calcium signals was recorded for individual unconjugated or conjugated target cells and expressed as percentage of total time. Each dot represents one tumor cell. Bars represent mean values. **p<0.01, unpaired t‑test. (E) Distinct tumor cell outcomes in response to CAR T cell attack. Three categories were defined as previously described: no hit, sublethal hit, lethal hit. Pie chart representing the percentage of each category. Data were pooled from three independent experiments with 3 distinct image acquisitions in each experiment.