Fig. 5

EZH2 acts as a positive regulator of TOP2A by promoting H3K27me3-mediated epigenetic silencing of miR-139-5p. (A) Protein levels of TOP2A and H3K27me3 in cells treated with the EZH2 inhibitors UNC1999 and EPZ005687. (B and C) Protein levels (B) and mRNA levels (C) of EZH2 and TOP2A in cells transfected with EZH2-targeted siRNAs. (D and E) mRNA levels of miR-139-5p in cells treated with EZH2 inhibitors UNC1999 and EPZ005687 (D) and in cells transfected with EZH2 targeted siRNAs (E). (F) mRNA levels of miR-139-5p in cells transfected with EED- and SUZ12- targeted siRNAs. (G) ChIP‒qPCR analysis of the enrichment of EZH2 and H3K27me3 on the promoter region of miR-139-5p in HCC cell lines. IgG was used as a negative control and H3 was used as a positive control. (H) Western blot analysis of TOP2A and H3K27me3 in HCC cells transfected with inhibitor NC and miR139-5p inhibitor and treated with DMSO, UNC1999, or EPZ005687. (I) Western blot analysis of TOP2A, EZH2 and H3K27me3 in HCC cells transfected with inhibitor NC and miR139-5p inhibitor and transfected with NC or siEZH2. (J) Western blot analysis of TOP2A, Dicer, EZH2 and H3K27me3 in HCC cells transfected with NC, siEZH2, siDicer, and siEZH2 and siDicer. *p < 0.05, **p < 0.01, ***p < 0.001