FOXO3a and FOXO1a inhibit the transactivation activities of ER-α and ER-β. (a, b) 293T cells were co-transfected with estrogen receptor (ER)-responsive element (ERE)-luc (firefly luciferase [luc] reporter containing EREs), pRL-TK (renilla luc as a transfection control for normalization), ER-α (panel a) or ER-β (panel b), and forkhead box class O (FOXO)3a plus IκB kinase (IKK)-β or an empty vector (control) as indicated. Total lysates of the transfected cells were prepared and subjected to luc assays. (c, d) Total lysates of 293T cells were co-transfected with ERE-luc, pRL-TK, ER-α (panel c) or ER-β (panel d), and FOXO1a plus IKK-β or an empty vector as indicated and subjected to luc assays. All cells were cultured in the presence of 17β-estradiol (E2). The relative reporter luc activity was normalized with pRL-TK. Data are expressed means and standard deviations from three repeated experiments, which were performed independently. *P < 0.05 between FOXO (FOXO3a or FOXO1a) minus IKK-β (lane 3) versus FOXO plus IKK-β (lane 4).