Domains of estrogen receptor α (ERα) required for ERα/Sp1-mediated activation of GC-rich promoters by estrogens and antiestrogens in breast cancer cells

Estrogen receptor α (ERα)/Sp1 activation of GC-rich gene promoters in breast cancer cells is dependent, in part, on activation function 1 (AF1) of ERα, and this study investigates contributions of the DNA binding domain (C) and AF2 (DEF) regions of ERα on activation of ERα/Sp1. 17β-Estradiol (E2) and the antiestrogens 4-hydroxytamoxifen and ICI 182,780 induced reporter gene activity in MCF-7 and MDA-MB-231 cells co-transfected with human or mouse ERα (hERα or MOR), but not ERβ and GC-rich constructs containing three tandem Sp1 binding sites (pSp1₃) or other E2-responsive GC-rich promoters. Estrogen and antiestrogen activation of hERα/Sp1 was dependent on overlapping and different regions of the C, D, E, and F domains of ERα. Antiestrogen-induced activation of hERα/Sp1 was lost using hERα mutants deleted in zinc finger 1 [amino acids (aa) 185-205], zinc finger 2 (aa 218-245), and the hinge/helix 1 (aa 265-330) domains. In contrast with antiestrogens, E2-dependent activation of hERα/Sp1 required the C-terminal F domain (aa 579-595), which contains a β-strand structural motif. Moreover, in peptide competition experiments overexpression of a C-terminal (aa 575-595) F domain peptide specifically blocked E2-dependent activation of hERα/Sp1, suggesting that F domain interactions with nuclear cofactors are required for ERα/Sp1 action.