Binding of estrogen receptor α/β heterodimers to chromatin in MCF-7 cells

Estrogen receptors (ERs), ER alpha and ER beta, belong to a group of transcription factors that, upon ligand binding, regulate gene expression by binding to specific DNA regions in chromatin as dimers. In this article, we applied the sequential chromatin immunoprecipitation assay (Re-ChIP) to study the simultaneous presence of ER alpha and ER beta on various DNA-binding regions in intact chromatin. ER alpha/beta heteroclimers were isolated by precipitation with anti-ER beta antibody followed byanti-ER alpha antibody from a stable MCF-7-derived cell line that expresses enclogenous ER alpha and an inducible version of ER beta. The Re-ChIP method was first validated based on the detection of ER alpha/beta heterodimers bound to a promoter region of the pS2 gene known to bind both ER alpha and ER beta. We next examined 12 ER-binding sites using Re-ChIP assays for ER alpha/beta heterodimer recruitment. Our results confirmed the recruitment of ER alpha/beta eterodimers to all these regions. This study represents the first demonstration of binding of ER alpha/beta heterodimers to various DNA-binding regions in intact chromatin.