The cross-regulation of the regulatory circuit in induced HepG2 cells. (A) As illustrated, HNF-4α, HNF-1α, HNF-6 and USF-1 were cross-regulated to form a positive feedback circuit. (B) The predicted binding sites of HNF-4α, HNF-1α, HNF-6 and USF-1 within their target promoters were identified. (C) Binding of HNF-4α, HNF-1α, HNF-6 and USF-1 to their target sites was evaluated by ChIP. After HepG2 cells were cocultured with 13.5-d mouse embryonic hepatocytes for 48 h, the binding of HNF-4α, HNF-1α, HNF-6 and USF-1 to their target elements was significantly promoted. (D) The contents of HNF-4α bound in hnf-1α and usf-1 promoters were detected by DNA pull-down assay. After HepG2 cells were treated with 13.5-d mouse embryonic hepatocyte medium for 48 h, the bound HNF-4α increased significantly. *P < 0.05 and **P < 0.01 versus control. The experiments were repeated three times. Con, control.