Fig. 3

CRNDE upregulates EGFR through mediating chromatin relaxation in HCC. A Schematic representation of the EGFR promoter. Full-length EGFR promoter regions 600 bp and 1000 bp upstream of the transcriptional start site from the 5'-end of pGl3TK-EGFR. B, C Relative luciferase activities of each promoter were determined in Hep3B under conditions of CRNDE depletion and Huh7 under conditions of CRNDE overexpression using the reporter assay. D, E Western blot analysis of H3K9Ac, H3K27Ac, and histone H3 protein levels in Hep3B (control and CRNDE depletion groups) and Huh7 cells (pcDNA3 and CRNDE overexpression groups). F, G Western blot and qRT-PCR analysis of p300 and EGFR protein and EGFR mRNA in Huh7 cells under CRNDE overexpression and shRNA-induced silencing of p300, respectively. H Relative luciferase activity of the EGFR promoter in Huh7 cells with CRNDE overexpression or shRNA-silenced p300, determined using the reporter assay. I Huh7 cells transfected with pcDNA3-control or CRNDE overexpression plasmids were incubated with DMSO or C646 and EGFR, H3K9Ac, H3K27Ac, and histone H3 protein levels were assessed by western blot. J qRT-PCR analysis of EGFR mRNA levels. K Huh7 cells transfected with pcDNA3-control or CRNDE overexpression plasmids were incubated with DMSO or C646 and relative luciferase activities of the promoter were assessed by the reporter assay. Statistical significance (p value) was calculated with the two-tailed Student's t-test for a single comparison between two groups. Data are presented as mean ± SD (*p < 0.05; **p < 0.01; ***p < 0.001)