Fig. 6

E2F5 activates the transcription of CDK13 gene and upregulates circCDK13 expression. a and b, PC3 cells were transfected with oeE2F5, shE2F5 or their corresponding control vectors, and then RT-qPCR detected CDK13 mRNA (A) or circCDK13 (B). *P < 0.05, **P < 0.01 vs. control vector. c, The schematic diagram shows 3 putative E2F binding sites within the 1-kb promoter region of CDK13. d, ChIP-qPCR detected the binding of E2F5 to the CDK13 promoter region in PC3 cells. Proximal: − 263 ~ − 459; Distal: − 444 ~ − 704. Arrowheads indicate the position of primers used for qPCR. *P < 0.05 vs. IgG. e, PC3 cells were transfected with the indicated constructs. Luciferase activity was measured by dual-luciferase reporter assays. *P < 0.05 vs. CDK13-Luc or CDK13-Luc + oeE2F5, #p<0.05 vs. oeE2F5 + circCDK13. f and g, Western blot analysis examined the E2F5, CDK13 and p21 expression in PC3 and 22RV1 cells transfected with circCDK13 and oeE2F5 either alone or together (F), or in PC3 and 22RV1 cells transfected with shcircCDK13 and miR-449a/212-5p mimic either alone or together (G). h, PC3 and 22RV1 cells were transfected as in (G), and cell proliferation was measured by colony formation assay. Upper panel shows the quantitative analysis of colony numbers from three independent experiments. *P < 0.05 vs. control constructs or shcircCDK13 and miR-449a/212-5p alone