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. 2019 Oct 15;10(24):5964–5974. doi: 10.7150/jca.35704

Figure 2.

Figure 2

Cip2a induces miR-301a expression through E2F1. (A-B) BT549 and MDA-MB-231 cells were transfected with Cip2a shRNA (A) or Cip2a-expressing vector (B), the expression of SKA2, c-myc and E2F1 were measured by Western blot. (C) BT549 and MDA-MB-231 cells were transfected with Cip2a shRNA in combination with pCMV-c-myc, or in combination with pCMV-E2F1, the expression of miR-301a was measures by qRT-PCR. (D) BT549 and MDA-MB-231 cells were transfected with Cip2a shRNA in combination with pCMV-c-myc, or in combination with pCMV-E2F1, the expression of Cip2a, E2F1, c-myc and SKA2 were measured by Western blot. (E) Sequence analysis of the promoter region revealed two conserved E2F1-binding sites at the core promoter region of SKA2/miR-301a (E2F1-1, E2F1-2). (F) The binding of E2F1 on SKA2 promoter region were measure by ChIP analysis. (G) BT549 and MDA-MB-231 cells were transfected with Cip2a-expressing vector in combination with pCMV-E2F1, together with a luciferase reporter construct containing the indicated promoter regions or mutant promoter regions. Relative luciferase activities were measured 48 h after transfection. *P <0.05, **P <0.01.