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

Estrogenreceptors(ERs),ERaandERb,belongtoagroupoftranscriptionfactorsthat,uponligandbinding,regulategene 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 ERa and ERb on various DNA-binding regionsinintactchromatin.ERa/bheterodimerswereisolatedbyprecipitationwithanti-ERbantibodyfollowedbyanti-ERa antibody from a stable MCF-7-derived cell line that expresses endogenous ERa and an inducible version of ERb. The Re-ChIP method was first validated based on the detection of ERa/b heterodimers bound to a promoter region of the pS2 gene known to bind both ERa and ERb. We next examined 12 ER-binding sites using Re-ChIP assays for ERa/b heterodimer recruitment. Our results confirmed the recruitment of ERa/b heterodimers to all these regions. This study represents the first demonstration of binding of ERa/b heterodimers to various DNA-binding regions in intact chromatin.