Fig. 4

APE1 positively regulates ZEB1 and E-cadherin promoter binding in cervical cancer cells. a Silencing of APE1 or inhibition of APE1 redox function rescued ZEB1-mediated inhibition of E-cadherin expression in HeLa cells. Cells were transfected with indicated plasmid or nucleotides and Western blotting was performed after 72 h of transfection. For APE1 inhibitor experiment, cells were treated with 5 μM APX3330 or DMSO for 24 h after 48 h of transfection. b Co-IP showing that APE1 binds to ZEB1 in HeLa cells. HeLa cells were transfected with the indicated plasmids. After 72 h of transfection, cells were subjected to Co-IP. c EMSA showing that silencing APE1 or inhibition of the redox function of APE1 decreased ZEB1 and E-cadherin promoter binding in HeLa cells. HeLa cells were transfected with nontargeting siRNA or siRNA targeting APE1 for 72 h and then subjected to total protein or nuclear protein extraction; HeLa cells were treated with DMSO or 5 μM APX3330 for 24 h and then subjected to total protein or nuclear protein extraction. Western blots were conducted using total protein. d Silencing of APE1 rescued the ZEB1-mediated loss of luciferase activity in HeLa cells transfected with luciferase under the control of an E-cadherin promoter (HeLa-Luc). Western blot and luciferase assays were performed after 72 h of transfection. Vector, cells transfected with empty vector; NC, cells transfected with nontargeting control siRNA; Ctrl, cells not treated with anything. **, P < 0.01; ***, P < 0.001; ns, no significant difference