Fig. 2

E2F1 and EIF4A3 enhance the expression of circSEPT9 through the binding to the SEPT9 promoter and pre-mRNA, respectively. a The expression of SEPT9 was detected in TNBC cells transfected with E2F1 overexpression plasmids or siRNAs targeting E2F1 by qRT-PCR. b Schematic illustration of wild type (WT) and mutant (Mut) sequences of three putative binding sites of E2F1 on SEPT9 promoter are shown. c The relative luciferase activities were detected in MDA-MB-231 cells co-transfected with luciferase reporter plasmids containing wild type or mutant SEPT9 promoter sequence and overexpression plasmids of E2F1. d ChIP-qPCR assays were performed to determine which putative E2F1 binding site the SEPT9 promoter was bound to in MDA-MB-231 cells, IgG was used as a negative control. e The circSEPT9 expression was valued in TNBC cells transfected with E2F1 overexpression plasmids or si-E2F1 by qRT-PCR. f and g The putative binding sites of EIF4A3 in the upstream and downstream region of the circSEPT9 pre-mRNA were predicted with circInteractome database, and the RIP assay confirmed that EIF4A3 could directly bind to the SEPT9 pre-mRNA in MDA-MB-231 cells. The intron 10 of the SEPT9 pre-mRNA and IgG were used as the negative controls. h circSEPT9 expression was detected in TNBC cells after EIF4A3 up-regulation or down-regulation by qRT-PCR. i The expression of cell cycle-related proteins in TNBC cells co-transfected with EIF4A3 expression vector and si-circSEPT9 was higher than that in TNBC cells transfected with si-circSEPT9 alone by western blot. The data are presented as the mean ± SD, *P < 0.05, **P < 0.01, ***P < 0.001