EMBO/sd-character-level-ner huggingface dataset:
Figure 3(a-e) Cartoon (top) illustrates the genotypes of the larvae, eye-antennal disc (EAD, circle: tumor cells in green and microenvironment cell in black), systemic cells are illustrated as a square (wild type cells in light brown and atg13 mutant cells in light orange) at indicated ages. Cartoon (left) illustrates the structure of cephalic complex attached to mouth hook. Representative images of Larva (image of larva using the backlight of microscope), cephalic complex (green highlights the GFP-labelled tumor clones), muscle (Phalloidin in green stains actin and Hoechst in blue stains nucleus) and adipose tissue (Lipid Tox in red stains for lipid droplets, Hoechst stains nucleus) from top to bottom. (a) RasV12; ctrl tumor bearing animal at day 6. (b) RasV12; scrib-/- tumor bearing animal at day 6. (c) RasV12; scrib-/- tumor bearing animal at day 8. (d) RasV12, scrib-/-, atg13-/-//atg13-/- at day 8. (e) RasV12, scrib-/-, atg13-/-//atg13-/- animal complemented with transgenic atg13, rescuing the tumor growth, at day 8. (f) Quantification of area of space occupied with lobes of fat body within larval segments 4 to 8 (shown in yellow dashed line in 2a), of animals bearing tumors at day 6 RasV12; ctrl (n=45), RasV12; scrib-/- (n=30), RasV12, scrib-/-, atg13-/-//atg13-/- (n=30) and ey3.5-atg13; RasV12, scrib-/-, atg13-/-//atg13-/- (n=29) and at day 10 RasV12; scrib-/- (n=40), RasV12, scrib-/-, atg13-/-//atg13-/- (n=25) and ey3.5-atg13; RasV12, scrib-/-, atg13-/-//atg13-/- (n=28). (g) Quantification of tumor volumes day 6 RasV12; ctrl (n=7), RasV12; scrib-/- (n=11) and day 8 RasV12; scrib-/- (n=13), RasV12, scrib-/-, atg13-/-//atg13-/- (n=9) and ey3.5-atg13; RasV12, scrib-/-, atg13-/-//atg13-/- (n=9). (h) Quantification of Ventral Longitudinal muscle 4 (VL4) (shown in yellow dashed line in 2a) of third segment of larvae carrying at day 6 RasV12; ctrl (n=15), RasV12; scrib-/- (n=14), and at day8 RasV12; scrib-/- (n=11), RasV12, scrib-/-, atg13-/-//atg13-/- (n=12) and ey3.5-atg13; RasV12; scrib-/-, atg13-/-//atg13-/- (n=14). (i) Quantification of dry weight excluding tumor of RasV12; scrib- -, atg13-/-//atg13-/-at day 6 (n=8) and day 10 (n=7). (j) Quantification of larval motility measured by crawling distance and crawling pattern for RasV12; scrib-/-, atg13-/-//atg13-/- at day 6 (n=15) and at day 8 (n=15), each colored line represents a single larva.
Compartment-specific regulation of neuronal transcripts by PTX.Figure 2(A) Volcano plots representing transcript down- (red) or up-regulation (yellow) after 48 hours PTX in the somatic and process compartment. Representative transcripts are highlighted and labelled. Linear model was used to fit transcriptome data and subsequently contrasted using likelihood ratio testing for PTX-treatment effects across compartments. (B) Venn-Diagram of differentially expressed transcripts between somatic and process compartment (C) Spearman rank correlation between log2-fold changes of differentially expressed genes significant in either the somatic or process compartment color-coded for exclusive changes in the respective compartments. Representative genes are highlighted and labelled. The statistical significance of Spearman correlations was calculated in the standard way, using Spearman's rho statistic. Regression lines indicate fitted linear models, with the lightgrey shaded areas depicting the 95% confidence interval. (D) Top 6 Gene Ontology Pathways enriched in differentially expressed genes in either the somatic or process compartment color-coded for Cellular Component (blue), Molecular Function (yellow) or Biological Process (red).
USP45 recruitment to sites of DNA damage is rapid and transientLive‐cell fluorescence analysis of U2OS cells stably expressing either GFPNLS empty, wild‐type (WT) USP45 or catalytically inactive USP45 [Cys199Ala].Figure 9A Live‐cell fluorescence microscopy analysis of U2OS cells stably expressing either GFPNLS empty, wild‐type (WT) USP45 or catalytically inactive USP45 [Cys199Ala]. Cells were treated with BrdU (10 μM, 24 h), psoralen or angelicin (25 μM, 3 h) and images captured at the indicated times after laser micro‐irradiation. The location of the micro‐irradiation stripe in each cell is indicated with a yellow line. Similar results were obtained in at least two independent experiments. Scale bar, 10 μm.
Figure 2(A-C) qRT-PCR measurement of Ifnb1 transcripts in mouse peritoneal macrophages (A), THP1 cells (B) and human peripheral blood mononuclear cells (PBMC) (C) that were left unstimulated or stimulated with 2'3' cGAMP (0.1μg/mL) for 4 h in the presence of DMSO or indicated concentrations of Palbociclib (0.1, 1, 10 μM). The data are presented as fold induction of Ifnb1/Gapdh transcripts. The data shown are mean±SEM of n=3 biological replicates. Statistical significance was determined by one-way ANOVA followed by Dunnett's post hoc test. *, P<0.05; ****, P<0.0001. (D) MTS detection of the viability of mouse peritoneal macrophages (PM), THP1 and PBMC cells left untreated (DMSO) or treated with Palbociclib (1 μM, 10 μM) for 4 h. The data shown are mean ± SEM from n=3 biological replicates. Statistical significance was determined by one-way ANOVA. ns, not significant. (E-F) Immunoblotting of indicated proteins in the lysates of mouse peritoneal macrophages (E) and THP1 cells (F) stimulated with 2'3' cGAMP (0.1 μg/mL) in the presence of DMSO or Palbociclib (10 μM) for indicated times. The data shown are representative of n=3 biological replicates. (G-H) qRT-PCR measurement of Il1b (G) and Il6 (H) transcripts in mouse peritoneal macrophages left unstimulated or stimulated with 2'3' cGAMP (0.1 μg/mL) in the presence of DMSO or Palbociclib (10 μM) for indicated times. The data shown are mean ± SEM of n=3 biological replicates. Statistical significance was determined by two-way ANOVA followed by Tukey's post hoc test. *, P<0.05; ****, P<0.0001 ; ns, not significant. (I-J) qRT-PCR measurement of Il1b (I) and Il6 (J) transcripts in THP1 cells left unstimulated or stimulated with 2'3' cGAMP (0.1μg/mL) in the presence of DMSO or Palbociclib (10 μM) for indicated times. The data shown are mean ± SEM of n=3 biological replicates. Statistical significance was determined by two-way ANOVA followed by Tukey's post hoc test. *, P<0.05; ****, P<0.0001 ; ns, not significant. (K-L) Immunoblotting of indicated proteins in the lysates of mouse peritoneal macrophages (K) and THP1 (L) cells stimulated with 2'3' cGAMP (0.1μg/mL) in the presence of DMSO or Palbociclib (10 μM) for indicated times. The data shown are representative of n=3 biological replicates. (M-N) Immunoblotting of indicated proteins in the lysates of mouse peritoneal macrophages (M) and THP1 (N) cells stimulated with 2'3' cGAMP (0.1 μg/mL) in the presence of DMSO or Palbociclib (10 μM). The data shown are representative of n=3 biological replicates.
Function and expression of AREG in cervical cancer cells.Figure 7Western blot analysis showing that AREG treatment induced dephosphorylation of LATS1, MOB1, and YAP. ME180 cells were starved for 6 h after reaching cell confluence, then cells were treated with AREG (50 ng/ml) for 0, 30, 60 min. AREG (50 ng/ml) treatment induced appearance of elongated cells in ME180 cells (top and middle panels) and significantly stimulated ME180 cell proliferation (lower panel). Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (P = 0.003). Scale bar: 100 μm. Wound-healing assay showing that AREG stimulates migration of ME180 cervical cancer cells within 10 h in serum-free medium. Scale bar: 200 μm. Real-time PCR showing that treatment of ME180 cells with AREG for 24 h significantly increased AREG mRNA expression. Each bar represents mean ± SEM (n = 9). Bars with different letters are significantly different from each other (P < 0.0001). Real-time PCR showing that knockdown of YAP in ME180 cells with YAP siRNA (siYAP) significantly suppressed AREG-induced AREG mRNA expression. siCTRL (non-target siRNA) was used as a siRNA control. Each bar represents mean ± SEM (n = 4). Bars with different letters are significantly different from each other (siCTRL vs. siCTRL+AREG, P < 0.0001; siYAP+CTRL vs. siYAP+AREG, P < 0.0001; siCTRL+AREG vs. siYAP+AREG, P = 0.0037).
Osteoblast-specific EGFR deletion reduces c-Fos driven osteosarcoma formation.Figure 2X-ray analysis of 7-months-old H2-c-fosLTR/Egfrwt and H2-c-fosLTR/Egfr∆Ob littermates. Scale bars: 1cm Bone tumor number per mouse at 6-7 months of age (n=22 wt, 11 ∆Ob) Quantification of tumor size in tibiae (n=22 wt, 11 ∆Ob) ALP levels in the serum at 7-month endpoint (n=17 wt, 8 ∆Ob) µPET/CT analysis of 7-months-old H2-c-fosLTR/Egfrwt and H2-c-fosLTR/Egfr∆Ob littermates Standardized uptake values (SUV) of the µPET-tracer Na[18F]F in the pelvic osteosarcoma of 4- and 7-months-old H2-c-fosLTR/Egfrwt and H2-c-fosLTR/Egfr∆Ob mice. n=6 wt, 4 ∆Ob for 4 month time-point, n=6 wt, 3 ∆Ob for 7 month time-point
a) Huh7 cells expressing GFP-LC3 treated with 0.f5(a) Huh7 cells expressing GFP-LC3 treated with 0.25 μM Torin1 and 0.4 μM bafilomycin A1 for 1 h. The autophagosome was identified as the IMP-poor double-membrane structure labelled with anti-GFP antibody (large gold, 10 nm). Both PtdIns(3) P (small gold, 6 nm) and GFP-LC3 were seen exclusively in the PF in both convex and concave structures. The small and large gold particles are shown as green and red dots, respectively, in the figures at right. Scale bars, 0.1 μm. Figures displaying wider areas, as well as additional examples, are presented in Supplementary Fig. 4c. ( (b) NIH3T3 cells expressing Atg4C78A were incubated in Earle's balanced salt solution for 1 h. The isolation membrane was observed as parallel membranes with few IMPs. PtdIns(3) P was labelled only in the PF. Scale bar, 0.1 μm. Other examples are presented in Supplementary Fig. 5. (c) Labelling density of PtdIns(3) P in mammalian autophagosomal membrane (mean±s.e.m.). The experiment was repeated three times and the number of colloidal goldparticles per μm2 was measured in more than ten randomly selected images. Some labelling was observed in the EF but the majority of gold particles in the EF occurred near the EF-PF boundary (white circles; see Supplementary Fig. 6c for more examples), suggesting that many of them actually represent PtdIns(3) P in the PF.
Figure 1(A) Determination of differentiation states of skin cutaneous melanoma (SKCM) biopsies from the TCGA cohort (n=458) The vast majority of these tumors exhibited a differentiated profile (89%; melanocytic and transitory states). The others display a dedifferentiated profile (11%; neural crest-like cells and undifferentiated states). Human melanoma 501Mel cell line is classified as differentiated melanoma cells (melanocytic cells), according to the gene expression profile (four categories were defined by (Tsoi et al, 2018); melanocytic, transitory, neural crest-like and undifferentiated cells). 501mel cell line was selected for the CRISPR screens. (B) Workflow depicting the in vivo CRISPR-SAM screen to identify tumor-promoting genes. Parental cells and CRISPR-SAM cell library were xenografted on nude mice (3x106 cells/mouse, n=6 and n=10, respectively) and tumor growth was monitored during 5 months. Seven tumors were collected and analyzed by DNA-Seq to identify the sgRNAs (C) sgRNAs distribution in the cell library. sgRNA mean reaches 550+/-2. (D) Tumor growth curves for the 7 tumors arising from the CRISPR-SAM-engineered cells xenografted in nude mice (E) Distribution of sgRNAs in cell library and in the 7 tumors (log10(sgRNAs counts)). Blue and red lines indicated the enrichment ≥10 fold or ≥100 fold (tumors vs cell library (in vitro)) (F) From the seven tumors, the top hundred genes (enriched) have been selected and common genes are SMAD3, BIRC3 and SLC9A5 (G) sgRNA counts in tumors versus in CRISPR-SAM cell library (respectively, black and orange points). Each black point corresponds to one tumor. Two replicates have been shown for CRISPR-SAM cell library (orange points). (H) Workflow depicting the validation step: 501Mel cells overexpressing SMAD3, BIRC3 or SLC9A5 (obtained by CRISPR-SAM) were xenografted on nude mice and tumor volume was monitored using caliper. 3x106 cells/mouse. n= 7, 6 and 6 mice, respectively. (I) SMAD3 expression levels in 501Mel cells overexpressing the cofactors for CRISPR-SAM approach and the control sgRNA (501Mel 3+ backbone) or the SMAD3 sgRNA. SLC9A5 and BIRC3 sgRNAs are used as controls to show the specificity of the SMAD3 overexpression. HSC70 serves as loading control. (J) SMAD3, BIRC3 and SLC9A5 mRNA expression levels in melanoma cell lines described in H and I. n=7 independent biological experiments. Dashed lines for medians. (K) Tumor growth curves from 501Mel cells overexpressing SMAD3, BIRC3 or SLC9A5.
Morphological studies of proAPI in the cytosol.Figure 1(A) Immunostaining of API in the vacuole of SEY6210 (wild-type) cell showing a dispersed pattern. (B-D) Immunostaining of TVY1 (pep4) cells showing API in the Cvt complex. The arrow points to a Cvt complex. Higher magnification images are shown in C and D. E) Freeze-substitution fixation image of Cvt complex in TVY1 cell. (F) Freeze-substitution fixation image of a Cvt complex in strain SEY6210 transformed with a 2μ plasmid encoding API grown in SD medium. (G) Immunostaining of strain SEY6210 containing a 2μ APE1 plasmid grown in SD medium. N, Nucleus; V, vacuole.
Figure 6A. SecB fails to restore proOmpA translocation ATPase activity stimulation of SecA(noC-tail) in the presence of TF. proOmpA translocation ATPase activity stimulation (as in Fig. 1C; SecB molar excess over proOmpA indicated). T/M ratio of SecA(noC-tail) in the absence of TF or SecB= 100%; lane 1; all other values were expressed as a % of this value. n= 4 biological replicates; mean values ± SEM. Unpaired parametric t-test, 95% confidence interval: **: p= 0.0048; ****: p < 0.0001. B. Excess of SecB inhibits the proOmpA-stimulated translocation ATPase activity of SecA(noCtail). ATPase measured as in Fig. 1C; SecB concentration range: 0-6 µM. Values were normalized (effectmax=100%; effectmin=0%) and plotted (n= 3 biological replicates; mean values ± SEM) versus the log10[SecB4]. The Ki was determined as in Fig. 1E. C. TF and SecB-regulated, SecA-dependent in vitro translocation of proOmpA into SecYEG/A-IMVs, analyzed after proteolytic treatment. Translocated, protease-resistant proOmpA was visualized using immunostaining (α-His) in the absence or presence of TF2 or/and SecB4 (as indicated). Translocation in the absence of TF or SecB was considered 100% (lane 3); all other values were expressed as a percentage of this. A summary of all repeats is presented (mean ± SEM; n= 6 biological replicates; ns: not significant (p = 0.1568 for lane 3 vs 4; p = 0.4835 for lane 3 vs 6); Unpaired parametric t-test, 95% confidence interval: ****: p < 0.0001) with a representative western blot (bottom).
A) The effects of GAPDH on degradation of Htt-552.Figure 6(A) The effects of GAPDH on degradation of Htt-552. The isolated lysosomes were incubated with 10 µg GAPDH and Htt-552 for 30 min at 37°C. At the end of the incubation, lysosomes were collected by centrifugation and the levels Htt-552 were determined with immunoblotting. Values are the mean±SE of three experiments. **P<0.01, *P<0.05 (compared with Htt-552-18Q without lysosomes); ##P<0.01, #P<0.05 (compared with Htt-552-100Q without lysosomes); $$P<0.01 (compared with Htt-552-18Q without GAPDH); P0.01 (compared with Htt-552-100Q without GAPDH). (B) The effects of LAMP-2A neutralizing antibody on degradation of Htt-552 by isolated lysosomes. The incubation of lysosomal preparation with Htt-552 was described as above. Values are the mean±SE of three independent experiments **P<0.01 (compared with Htt-552-18Q without lysosomes); #P<0.05, ##P<0.01 (compared with Htt-552-100Q without lysosomes); $$P<0.01 (compared with Htt-552-18Q without LAMP-2A antibody); P0.01 (compared with Htt-552-100Q without LAMP-2A antibody). (C) The incubation of lysosomal preparation with Htt-552 was described as above. The incubation of lysosomal preparation with Htt-552 was described as above. The effects of Hsc70 neutralizing antibody on degradation of Htt-552 by isolated lysosomes. Values are the mean±SE of three independent experiments. **P<0.01 (compared with Htt-552-18Q without lysosomes); ##P<0.01, #P<0.05 (compared with Htt-552-100Q without lysosomes); $$P<0.01 (compared with Htt-552-18Q without Hsc70 antibody); P0.05 (compared with Htt-552-100Q without Hsc70 antibody).
Figure 7A Graphic illustration of two human liver cancer PDXs mouse model based sorafenib (SOR) treatment regimen. In vivo analyses of tumor (B) in mice that were subcutaneously implanted with tumor tissues from two human liver cancer patients and divided into four groups. In vivo analyses of tumor growth (C) in mice that were subcutaneously implanted with tumor tissues from two human liver cancer patients and divided into four groups. In vivo analyses of tumor weight (D) in mice that were subcutaneously implanted with tumor tissues from two human liver cancer patients and divided into four groups. Data information: In all relevant panels, *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; Two-tailed t-test. Data are presented as mean ± SD. E H&E and immunohistochemical images of Ki67, LC3-B, VEGF-A in randomly selected PDX#1 and PDX#2 tumors. Quantification of Ki67 (F) expression in immunohistochemical images from PDX tumors by Image Pro Plus (IPP) analysis. Data information: In all relevant panels, *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; Two-tailed t-test. Data are presented as mean ± SD. Quantification of LC3-B (G), and VEGF-A (H) expression in immunohistochemical images from PDX tumors by Image Pro Plus (IPP) analysis. Data information: In all relevant panels, *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; Two-tailed t-test. Data are presented as mean ± SD. I Knockdown of METTL3 by siRNA in PDX#1 and PDX#2 tissues were confirmed by Western blot. J Working model. The molecular mechanisms of m6A-dependent sorafenib-resistance in liver cancer. The expression of METTL3 was down-regulated in sorafenib-resistant liver cancer, and caused decrease of FOXO3 by the m6A modification in an YTHDF1-dependent manner. Degradation of FOXO3 thus promoted autophagy-associated sorafenib-resistance in liver cancer.
Figure 1(A) Left quadriceps muscle biopsy performed at age 10 months (i-iii) demonstrating a dystrophic process with variation in fiber size (black arrow), increased connective tissue (blue arrow) and rare internalized nuclei (white arrow) on hematoxylin and eosin (H&E) staining (i) and Gömöritrichrome staining (ii) and type 1 fiber atrophy on nicotinamide adenine dinucleotide (NADH) staining (iii). Muscle biopsy in an unaffected (control) individual (iv-vi). Scale bar: 100μm. (B) Western blot data (n=1) with quantification demonstrated reduced α-Dystroglycan (α-DG) in P1's muscle compared to control. (C) Western blot data with quantification demonstrated reduced α-Dystroglycan in fibroblasts from P1 and P2 compared to control (c), although not to the extent of the reduction observed in fibroblasts from a patient with a confirmed α-dystroglycanopathy (α-DG pt) due to biallelic pathogenic variants in LARGE (n=1). (D) Brain MRI findings in P1 at 28 months of age (top) revealing normal appearing cortex, cerebellum and pons on T1-weighted images and T2-weighted image. There is evidence of mild thinning of the corpus callosum (arrow). Brain MRI in an unaffected (control) individual at 30 months of age (bottom) demonstrating normal sized corpus callosum.
Cell type specificity of chromatin interactions.Figure 4A) The normalized interaction frequency (as above) in MCF-7 cells cultured in hypoxia (vertical axis) was plotted against that in VHL defective 786-Ocells in normoxia (horizontal axis) for each site that interacted significantly with a HIF-binding site in either cell line or both. Highlighted in red are sites that had a significantly different interaction frequency in either cell type. B) The log2 (fold difference) between MCF-7 cells in normoxia and hypoxia (left) and VHL defective 786-O cells in normoxia and MCF-7 cells in hypoxia (right) from Figure 3B and Figure 4A respectively. Highlighted in red are sites that had a significantly different interaction frequency in either condition/cell type. C) Capture-C (MCF-7hypoxia - blue; 786-O - green) at the CCND1 locus from an enhancer (red highlight) bound by HIF almost exclusively in VHL defective 786-O and showing cell-specific cis-interactions, blue highlighted region indicates region of interaction at the TSS of CCND1, green highlighted region indicates region of interaction at an enhancer 3' to the MYEOV gene. D) Capture-C (MCF-7hypoxia - blue; 786-O - green) at the TGFA locus from an enhancer bound by HIF exclusively in VHL defective 786-O cells but showing constitutive interaction. Blue highlighted region indicates interaction with the TSS of TGFA. Green highlighted region indicates interaction with an enhancer immediately 3' to MYEOV.
Figure 2Quantitative phosphoproteomic analysis defining the role of Ddc1 in the set of Mec1-dependent phosphorylation in rad9∆ cells. Ddc1 dependency (y axis) is plotted against Mec1 dependency (x axis). These phosphoproteomic analyses were conducted in the presence of 0.02% MMS for 2 hours. Quantitative phosphoproteomic analysis defining the role of Dna2 (WY-AA mutation impairs Mec1-activating function of Dna2) in the set of Mec1-dependent phosphorylation in rad9∆ cells. Dna2 dependency (y axis) is plotted against Mec1 dependency (x axis). These phosphoproteomic analyses were conducted in the presence of 0.02% MMS for 2 hours. Plot comparing Dna2 and Ddc1 dependency of selected Mec1 substrates. Error bars represent standard deviation of two or more independent peptide-spectrum matches (PSMs) of the indicated site. Quantitative phosphoproteomic analysis defining the role of long-range resection factors Exo1 and Dna2 in the set of Mec1-dependent phosphorylation events in rad9∆ cells. MEC1 dependency (x axis) is plotted against long range resection dependency (y axis). These phosphoproteomic analyses were conducted in the presence of 0.02% MMS for 2 hours. For the DNA2-AID cell line, cells were pre-treated for 15 minutes with 100μM IAA and then treated for 2 hours with 0.02% MMS.
a–d, Electron micrographs of etoposide-treated Atg5-/- MEFs.figf3a-d, Electron micrographs of etoposide-treated Atg5-/- MEFs. a, AVs were observed near the Golgi apparatus (G). Inset, the isolation membrane (I) was extended from the Golgi stack. b, Developing autophagosome. rER, rough endoplasmic reticulum. c, d, Isolation membrane fusing with vesicles containing thick membrane (d) and a complete autophagosome (c) is generated. e, f, Co-localization of Lamp2 with GFP-M6PR, GFP-Stx7 or GFP-Rab9 in etoposide-treated Atg5-/- MEFs. Arrows indicate co-localized dots. Original magnification, × 200. g-i, Atg5-/- MEFs were transfected with the indicated siRNAs for 24 h and treated with etoposide. Induction of macroautophagy was assessed by Lamp2 IF (g) and by EM (h), and the number of isolation membranes was calculated (i). In g, error bars indicate s.d. (n = 4). In h and i, red and blue lines indicate means and s.e.m., respectively; n = 35 cells each. j, No effect of silencing of Rab9 (siRab9) on the production of LC3-II in etoposide-treated WT MEFs.