EMBO/sd-character-level-ner huggingface dataset:
ELYS overexpression increases nuclear size, NPC density, and nuclear levels of lamin B2, Ran, and NTF2.Figure 3MCF-10AT1k.cl2 cells were co-transfected with control or ELYS siRNA and plasmids expressing eGFP alone or siRNA-resistant eGFP-ELYS. Cell lysates were analyzed by western blot and probed for ELYS and tubulin. ELYS band intensity was normalized to tubulin and then normalized to the eGFP/negative siRNA control. ELYS concentrations are indicated in the row labeled [ELYS] (average ± SD). Quantification was performed on two biological replicates. Cells were stained with an antibody against lamin B1. Representative images are shown. (B) For each experiment, nuclear cross-sectional areas were quantified for 68-298 nuclei per condition (162 nuclei on average) and averaged. Three biological replicates, data from one representative experiment shown. (C) For each experiment, nuclear lamin B2 staining intensities were quantified for 29-81 nuclei per condition (60 nuclei on average) and normalized to the eGFP/negative siRNA control. Two biological replicates, data from one representative experiment shown. Cells with and without cytoplasmic lamin B2 aggregates were counted and the percentage of cells with aggregates was calculated. For each experiment, 58-299 cells were examined per condition. Three biological replicates, data from one representative experiment shown. Cells were stained with antibodies against Ran and NTF2. For each experiment, 62-130 nuclei were quantified per condition (92 nuclei on average). Two biological replicates, data from one representative experiment shown. (E) Nuclear Ran and NTF2 staining intensities were quantified and normalized to the eGFP control. (F) To measure the N/C staining intensity ratio of a given cell, the average fluorescence intensity of a nuclear region was divided by the average fluorescence intensity of a cytoplasmic region. Transfections were performed as in (A) and cells were stained with an antibody against FG-Nups (mAb414) and Nup133. Representative images for mAb414 staining are shown. Confocal imaging was performed through nuclear cross-sections as well as on the surface of nuclei. (H) To measure NPC densities, confocal NE surface images were acquired for mAb414 and Nup133 stained nuclei. NPC numbers were counted per unit area for 25-116 nuclei per condition (53 nuclei on average) and normalized to the negative control. Two biological replicates, data from one representative experiment shown.
Silencing of PHD1 increases cell apoptosis after chemotherapy.Figure 2A Western blot for p53, cleaved caspase-3 (cleaved casp3), cleaved parp, and vinculin (vinc) in p53wt/wt and p53−/− HCT116 silenced for PHD1 upon exposure to 300 μM 5-FU for 24 h, showing an increased apoptotic response to 5-FU treatment in PHD1-silenced cells compared to the scrambled control. B Confirmation of these results by FACS analysis of propidium iodide-stained PHD1-silenced p53wt/wt and p53−/− HCT116 cells exposed for 26 h to 300-μM 5-FU treatment. *P = 0.002 toward p53wt/wt HCT116 shScr 5-FU-treated (two-tailed unpaired t-test) with n = 6 for non-treated p53wt/wt HCT116 and n = 3 in all other groups. C Detection of cleaved casp3, cleaved parp, and vinc in HCT116 silenced with a second siRNA for PHD1. D, E Apoptosis as detected by Western blot for cleaved casp3 and parp in PHD1-silenced HCT116 treated for 20 h with 200 nM SN-38 (D) or for 24 h with 20 μM oxaliplatin (E). F Western blot for cleaved casp3, parp, and vinc in LIM1215 silenced for PHD1 and treated for 24 h with 200 μM 5-FU. G Similar apoptosis levels were detected by Western blot for cleaved casp3 and parp with vinc as a loading control in p53S15A→p53−/− HCT116 silenced for PHD1 or scrambled (Scr) control and treated for 20 h with 300 μM 5-FU.
Figure 3Lymphocytes from the Peyer's patches of C57BL/6 and Vα14 mice were stained with anti-CD3 and CD1d (PBS57) tetramer. Percentage of lymphocytes that were CD3+CD1d-tetramer+ iNKT cells among the Peyer's patches of C57BL/6, Vα14, and Jα18-/- mice are shown. Mann-Whitney test. Error bars are SD. C57BL/6 n=22; Vα14 n=16; Jα18-/- n=8. (C-E)Mice were analyzed for total fecal IgA by ELISA. Mann-Whitney test. Error bars are SD. C57BL/6 n=14; Vα14 n=11; Jα18-/- n=13. (C-E)Mice were analyzed for total fecal IgG1 (D) or total serum IgG1 (E) by ELISA. Mann-Whitney test. Error bars are SD. C57BL/6 n=14; Vα14 n=11; Jα18-/- n=13. Percentages of total B cells that were IgG1+ in the spleen, mLN, and Peyer's patches of C57BL/6, Vα14, and Jα18-/- mice are shown. Mann-Whitney test. Error bars are SD. C57BL/6 n=15; Vα14 n=15; Jα18-/- n=5
Figure 5(A) Expression heatmap of ATAC-DEGs in TFAM knockdown and control SNU-368 cells (n=3 per group). Up- and down-regulated ATAC-DEGs in TFAM knockdown HCC cells were annotated. (B Top 20 GO terms enriched by ATAC-DEGs. Rich factor represents the ratio of the number of mapped genes to the number of all the annotated genes located in the pathway. C) enriched KEGG pathways enriched by ATAC-DEGs. Rich factor represents the ratio of the number of mapped genes to the number of all the annotated genes located in the pathway. (D) ATAC-seq signal tracks around the FN1, IL-6, TGFB1 and ITGB3 genes in HCC cells with treatment as indicated. Blue box indicates promoter region with increased accessibility. (E qRT-PCR for mRNA expression levels of metastasis related gene Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test. *P < 0.05; **P < 0.01. F) Western blot assays for protein expression levels of metastasis related gene in HCC cells (n = 3 independent experiments) and tumor tissues from mice (n =4 per group) with treatments as indicated. Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test. *P < 0.05; **P < 0.01. (G) ELISA analysis for the levels of IL-6 and TGFB1 in HCC cells and tumor tissues from mice with treatments as indicated (n = 3 independent experiments). Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test. *P < 0.05; **P < 0.01. (H Western blot assays levels of FN1, ITGB3 in HCC cells with treatment as indicated (n = 3 independent experiments). Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test. *P < 0.05; **P < 0.01. I) ELISA analysis for the levels of FN1, IL-6, TGFB1 and ITGB3 in HCC cells with treatment as indicated (n = 3 independent experiments). Data information: Graphs show mean ± s.e.m., two-tailed unpaired t-test. *P < 0.05; **P < 0.01.
Figure 3A Colonization patterns of GFP-Pst DC3000 and Pst DC3000 (AvrRpt2) in wild-type plants. Data information: Leaves were inoculated with GFP-labeled bacteria at 108 cfu/ml for 2 days. IS, infected site; UIS, uninfected site. White dash lines indicate the boundary between IS and UIS. Scale bars, 100 μm B Time-lapse images of GFP-Pst DC3000 and Pst DC3000 (AvrRpt2) captured at 0 (magenta), 15, (yellow), and 30 (green) sec Time in seconds in the movies is shown at the top of images. Blue color was used for chlorophyll autofluorescence. Data information: Leaves were inoculated with GFP-labeled bacteria at 108 cfu/ml for 2 days. Scale bars, 10 μm C Colonization patterns of GFP-Pst DC3000 (AvrRpt2) in wild-type and palQ plants after PA and CA pretreatments. Data information: Leaves were inoculated with GFP-labeled bacteria at 108 cfu/ml for 2 days. M, mock; PA, piperonylic acid; CA, coniferyl alcohol; IS, infected site; UIS, uninfected site. White dash lines indicate the boundary between IS and UIS. Scale bars, 100 μm
Inhibition of endocytosis restores DHHC5 to the plasma membrane in the presence of mutant Golga7b.Figure 530 minute DMSO treatment of HeLa cells co-expressing WT DHHC5 (FLAG tagged) and mutant Golga7b (HA tagged). 30 minute Dynasore treatment (25µM) of HeLa cells co-expressing WT DHHC5 (FLAG tagged) and mutant Golga7b (HA tagged). Non-targeting control siRNA treatment (10nM) of HeLa cells expressing mutant Golga7b (HA tagged). Images are detecting endogenous DHHC5 and the HA tagged Golga7b AP2µ siRNA treatment (10nM) of HeLa cells that were subsequently transfected with mutant Golga7b (HA tagged). Images are detecting endogenous DHHC5 and the HA tagged Golga7b. Quantification of plasma membrane signal of DHHC5 in (A and B). ****=p<0.001, unpaired t-test. N=37, +Dynasore. N=28, +DMSO. Quantification of plasma membrane signal of DHHC5 in (C and D). ****=p<0.001, unpaired t-test. All scale bars 10μm. Biotin endocytosis assays of HeLa cells with endogenous DHHC5, over-expression of DHHC5 alone, co-expression of DHHC5 and WT Golga7b, co-expression of DHHC5 and mutant Golga7b or expression of the DHHC5 C-terminal mutant. Biotinylated cell surface proteins endocytosed within 15 minutes were purified using streptavidin resin and blots probed with an antibody to DHHC5. The lack of signal in the pull-down of the DHHC5 C-terminal mutant or when DHHC5 and WT Golga7b are co-expressed indicates that DHHC5 is stabilised at the plasma membrane in these conditions. The robust signal in the pull-down of endocytosed proteins from cells co-expressing DHHC5 and mutant Golga7b shows a high rate of endocytosis of DHHC5 in this condition. N=3, *=p<0.05, paired t test"
Figure 2Expression of HIF1A-AS2 was suppressed in HUVECs by oligo phosphodiester nucleotides (ODN)-based knockdown. Cells were then treated with CoCl2 for 5 hrs to mimic hypoxic condition, and subjected to gene expression analyses: The RNA abundance of HIF-1A and HIF1A-AS2 was analyzed by real-time PCR and normalized by 18s rRNA (A). HIF-1α protein level was monitored by western blot, with β-actin as the internal control (B). Nuclear run-on assays were performed to assess the extent of nascent transcription of target genes under mock vs. CoCl2 treatment (C) Nuclear run-on assays were performed to assess the extent of nascent transcription of target genes in the HIF1A-AS2 knockdown cells (D). HIF-1A mRNA expression was detected in a region-specific manner (5' vs. 3') by primers corresponding to exons 1-2 (E1-2) and exon 14-15 (E14-15). To further monitor transcription activity, ChIP was performed to characterize RNA polymerase II binding on chromatin isolated from MCF7 cells under CoCl2 treatment (E), The precipitated DNA fragments were quantitatively analyzed by real-time PCR using primers for the HIF1A-AS2 and HIF-1A promoters, and normalized to the values of IgG. HIF1A-AS2 corresponds to the transcribed region. HIF-1A mRNA expression was detected in a region-specific manner (5' vs. 3') by primers corresponding to exons 1-2 (E1-2) and exon 14-15 (E14-15). To further monitor transcription activity, ChIP was performed to characterize RNA polymerase II binding on chromatin isolated from MCF7 cells in HIF1A-AS2-knockdown vs. control cells (F). The precipitated DNA fragments were quantitatively analyzed by real-time PCR using primers for the HIF1A-AS2 and HIF-1A promoters, and normalized to the values of IgG. HIF1A-AS2 corresponds to the transcribed region. Schematic of the convergent mode of transcriptional regulation of the HIF-1A sense and antisense gene pair. For graphs shown in this figure, means ± S.D. calculated from three independent experiments are shown.
Figure 5Effects of COUP-TFII inhibition and lactate treatment on tumor growth in vivo. (A) Growth curve in tumors derived from xenografted shScramble or shCOUP-TFII-HCT116 cells and treated with sodium chloride or sodium L-lactate (60 l x g-1 body weight; 150 mM). (B) tumor volume at the experimental endpoint, in tumors derived from xenografted shScramble or shCOUP-TFII-HCT116 cells and treated with sodium chloride or sodium L-lactate (60 l x g-1 body weight; 150 mM). Effects of COUP-TFII inhibition and lactate treatment on tumor growth in vivo. (C) Western blot analysis of the indicated proteins in tumors derived from xenografted shScramble or shCOUP-TFII-HCT116 cells and treated with sodium chloride or sodium L-lactate (60 l x g-1 body weight; 150 mM). (D) Effects of trametinib treatment and overexpression of COUP-TFII on tumor growth in vivo. Tumors were derived from xenografted HCT116 expressing HA-COUP-TFII or carrying a control vector and treated with trametinib (3mg/kg). (E) Western blot analysis of indicated proteins in tumors derived from xenografted HCT116 treated with trametinib with or without COUP-TFII overexpression. Asterisk indicates exogenous HA-COUP-TFII. (F) Schematic showing COUP-TFII-mediated growth pathway in KRAS activated cancer cells.
ERK7 overexpression inhibits lipid biosynthesisA.Figure 2A. Fatty acid composition of total lipids in CG>control and CG>ERK7 larvae, measured by gas chromatography (N=3 replicates of 15 larvae/replicate for each genotype). B. Schematic presentation of biosynthesis of fatty acids and their likely chain shortening by peroxisomal partial β-oxidation. C. Amount of glucose-derived [13C]-labeled carbons incorporated into the TAG pool of control and CG>ERK7 fat bodies (N=3 replicates of 15 fat bodies/replicate for each genotype and diet). D. Ectopic ERK7 expression in the fat body results in elevated hemolymph glucose levels (N=3 replicates of 15 larvae/replicate for each genotype). E. Circulating glucose levels are decreased in ERK71 mutants (N=4 replicates of 15 larvae/replicate for each genotype).
Figure 2Schematic of the H-1400 assay. A DSB is induced at MATa and repaired by recombination with HMLα-inc. There is ~1400 bp homology between MATa and HMLα-inc sequences on the second end (green box). Representative Southern blots showing DSB repair products in WT, rad59∆, rad52-R70A and rad59∆ rad52-R70A cells. DNA was digested with XhoI and EcoRI and probed with a Z sequence (yellow box). Percentage of BIR among all repair products by 6 hr (mean ± SD; n = 3). Data information: Welch's unpaired t-test was used to determine the p-value Viability of indicated strains (mean ± SD; n = 3). Data information: Welch's unpaired t-test was used to determine the p-value Graphs show analysis of repair efficiency compared to uncut parental band (left) (mean ± SD; n = 3) and repair efficiency compared to WT (right) at 6 hrs after DSB induction (right). Data information: Welch's unpaired t-test was used to determine the p-value Schematic of the H-2100 no-gap assay. A DSB is induced at MATα and repaired by recombination with HMLα-inc. There is ~2100 bp homology between MATα and HMLα-inc sequences on the second end (green box). Representative Southern blots showing DSB repair products in WT, rad59∆, and rad52-R70A. DNA was digested with XhoI and EcoRI and probed with a Z sequence (yellow box). Percentage of BIR product among repair products by 6 hr are shown. (mean ± SD; n = 3). Data information: Welch's unpaired t-test was used to determine the p-value Viability of indicated strains (mean ± SD; n = 3). Data information: Welch's unpaired t-test was used to determine the p-value Graphs show analysis of repair efficiency compared to uncut parental band (left) (mean ± SD; n = 3) and repair efficiency compared to WT (right) at 6 hrs after DSB induction (right). Data information: Welch's unpaired t-test was used to determine the p-value
Figure 4A. Oligo(dT) FISH and IF analysis of nucleolar protein RPF1 and Cajal body protein coilin. B. Oligo(dT) FISH and IF analysis of nucleolar protein RPF1 and nuclear speckle protein SRRM2. C. Oligo(dT) FISH and IF analysis of Cajal body protein coilin and nuclear speckle protein SRRM2.
Figure 7Freezing phenotype (A) of wild type, ost1-3, egr2-3, and ost1 egr2 mutants. Two-week-old seedlings grown on MS medium were treated at −5°C (NA) or −8°C for 0.5 h (CA). survival rate and ion leakage (B) of wild type, ost1-3, egr2-3, and ost1 egr2 mutants. Two-week-old seedlings grown on MS medium were treated at −5°C (NA) or −8°C for 0.5 h (CA). Data information: each bar represents the mean ± SEM of three independent experiments, each of which has three technical repeats. Asterisks indicate significant differences compared to the wild type with the same treatment (*P < 0.05, **P < 0.01, two-tailed t-test). C Expression of CBFs in wild type, ost1-3, egr2-3, and ost1 egr2-3 plants under cold stress. Two-week-old seedlings grown at 22°C were treated at 4°C for 0 h and 3 h. Data information: each bar represents the mean ± SEM of three independent experiments, each of which has three technical repeats. Asterisks indicate significant differences compared to the wild type with the same treatment (*P < 0.05, **P < 0.01, two-tailed t-test). D Model of EGR2-OST1 modulation of cold stress responses in Arabidopsis. At warm temperature (22°C), EGR2 can be modified by myristoylation, which allows it to recognize OST1 and inhibit its activity to ensure plant growth. Under cold stress (4°C), the interaction of NMT1 and EGR2 is decreased, thereby causing the accumulation of unmyristoylated EGR2, which shows reduced interaction with OST1 and disrupts the interaction of EGR2-OST1 as well. This ultimately leads to activating OST1 protein kinases and enhancing plant freezing tolerance. Clade A PP2Cs, such as ABI1, also partially contributes to regulating OST1 activity in response to cold stress. EGR2-M and u-EGR2 represent myristoylated and unmyristoylated EGR2, respectively.
Schematic model showing how Hsp90 regulates SG dynamics via DYRK3.Figure 6Upper part: in resting cells (no stress), Hsp90 interacts with DYRK3 and regulates its stability and activity. Upon stress, DYRK3 dissociates from Hsp90 and partitions inside SGs, where it is protected from irreversible aggregation and degradation. During the recovery phase after stress, soluble DYRK3 associates with Hsp90 outside or at the boundary of SGs to be stabilized. Once stabilized by Hsp90, DYRK3 could autophosphorylate its activation loop to achieve full activation and trigger the disassembly of SGs, by phosphorylating yet unknown molecular targets. Active DYRK3 would then promote translation restoration. Lower part: upon treatment with GA or 17AAG, DYRK3 dissociates from Hsp90, becomes inactive and is either targeted to SC35 speckles or degraded by the 26S proteasomes. During stress, blocking the loading of DYRK3 onto Hsp90 with GA or 17AAG redirects DYRK3 to SGs or to 26S proteasomes. During the recovery phase after stress, inhibition of Hsp90 prevents the stabilization and activation of DYRK3. As a consequence, SGs do not disassemble and translation is not restored. The model only depicts the interplay between Hsp90 and DYRK3 in the regulation of SG dynamics, although other Hsp90-dependent/DYRK3-independent mechanisms may also occur.
Tpl2[D270A] mutation impairs maturation of Staphylococcus aureus phagosomesA-K.Figure 5BMDMs of the indicated genotypes were infected with YFP-S. aureus (MOI 10) for 1 h. (A) Phagocytic uptake (CFU) of YFP-labelled S. aureus into BMDMs of indicated genotypes 1h post-infection (n = 12; 2 biological and 6 technical replicates per condition per experiment). Data information: One representative experiment out of three shown. Error bars represent SEM. P < 0.01, **** P < 0.0001. Student's unpaired t-test. BMDMs of the indicated genotypes were infected with YFP-S. aureus (MOI 10) for 1 h. (B) Representative images of YFP+ BMDMs labelled with Magic Red cathepsin L substrate (red) and DAPI (nuclear stain) 1h post-infection. (C) Quantification of Magic Red cathepsin L staining from panel (A). Average fluorescence intensity of Magic Red cathepsin L substrate per cell relative to fluorescence intensity of YFP-S.aureus (n = 26-29 cells). (D) Quantification of Magic Red cathepsin L staining co-localised with YFP-S. aureus from panel (A) (n = 26-29 cells). Data information: One representative experiment out of three shown. Error bars represent SEM. P < 0.01, **** P < 0.0001. Student's unpaired t-test. BMDMs of the indicated genotypes were infected with YFP-S. aureus (MOI 10) for 1 h. (E) Representative images of YFP+ BMDMs labelled with LysoTracker Red DND-99 dye (red) and DAPI, 1h post-infection with YFP-S. aureus (green).(F) Quantification of LysoTracker Red signal from panel (D). Average fluorescence intensity of LysoTracker Red per cell relative to fluorescence intensity of YFP-S. aureus (n = 66-98 cells). (G) Quantification of LysoTracker Red co-localisation with YFP-S. aureus from panel (D) (n = 66-98 cells). Data information: One representative experiment out of three shown. Error bars represent SEM. P < 0.01, **** P < 0.0001. Student's unpaired t-test. BMDMs of the indicated genotypes were infected with YFP-S. aureus (MOI 10) for 1 h. (H) Representative images of BMDMs stained with an anti-EEA1 antibody (red) and DAPI, 1h post-infection with YFP-S. aureus (green). (I) Quantification of EEA1+ S. aureus-containing phagosomes from panel (G) (n = 27 cells). (J) Representative images of BMDMs stained with an anti-LAMP-1 antibody (red) and DAPI, 1h post-infection with YFP-S. aureus (green). (K) Quantification of LAMP-1+ S. aureus phagosomes from panel (I) (n = 25 cells). Data information: One representative experiment out of three shown. Error bars represent SEM. P < 0.01, **** P < 0.0001. Student's unpaired t-test.
Figure 2A. Representative images showing hTERT-RPE1 grown in the presence of serum and stained with anti acetylated α-tubulin antibody. There were more ciliated cells after treatment with siRNA against KV10.1 (KV10.1 KD), as compared to scrambled siRNA-transfected cells, which were devoid of cilia (control). Scale bar: 10 µm.B. Quantification of the effect depicted in A for the number of images indicated at the base of the columns. C. Time course of ciliary disassembly. hTERT-RPE1 cells were serum starved for 24 hours, fixed at the times indicated after serum reintroduction, and stained with anti acetylated α-tubulin to reveal cilia. While in the presence of scrambled siRNA the abundance of cilia decreased rapidly (black line), it remained virtually unchanged in cells where KV10.1 was knocked down (red). N=8-13.D. Time course of ciliary shortening after serum reintroduction. In the same experiments as in C, the length of the remaining cilia was measured automatically using ImageJ (FIJI). While ciliary length decreased with a similar time course in the siRNA-treated cells as in controls, cilia were longer in KV10.1-knockdown cells. A re-elongation of cilia was evident in both populations after 6 hours. Values are mean ± SEM for a number of cilia between 62 (control, 4.5 h) and 308 (both groups, 1.5 h). E. Blockade of permeation through the K+ channel altered the speed of deciliation. After serum starvation, medium containing serum and Astemizole (10 µM) was added, and cells were fixed and stained at he indicated time points (N=12).