Binding of estrogen receptor beta to estrogen response element in situ is independent of estradiol and impaired by its amino terminus.

The functions of 17beta-estradiol (E2) are mediated by estrogen receptor (ER) alpha and beta. ERs display similar DNA- and ligand-binding properties in vitro. However, ERbeta shows lower transcriptional activity than ERalpha from the estrogen response element (ERE)-dependent signaling. We predicted that distinct amino termini contribute to differences in transcription efficacies of ERs by affecting in situ ER-ERE interactions. We used chromatin immunoprecipitation and a novel in situ ERE competition assay, which is based on the ability of ER to compete for ERE binding with a designer activator that constitutively induces transcription from an ERE-driven reporter construct. Interference of activator-mediated transcription by unliganded or liganded ERs was taken as an indication of ER-ERE interaction. Results revealed that ERs interacted with ERE similarly in the absence of E2. However, E2 enhanced the ERE binding of ERalpha but not that of ERbeta. The removal of the amino terminus increased the ERbeta-ERE interaction independent of E2. The ERbeta amino terminus also prevented E2-mediated enhancement of the chimeric ERalpha-ERE interaction. Thus, the amino terminus of ERbeta impairs the binding of ERbeta to ERE. The abrogation of ligand-dependent activation function 2 of the amino-terminally truncated ERbeta resulted in the manifestation of E2 effect on ERbeta-ERE interaction. This implies that E2-mediated enhancement of ERbeta-ERE interaction is masked by the activation function 2, whereas the intact amino terminus is a dominant region that decreases the binding of ERbeta to ERE. Thus, ERbeta-ERE interaction is independent of E2 and is impaired by its amino terminus. These findings provide an additional explanation for differences between ERalpha and ERbeta functions that could differentially affect the physiology and pathophysiology of E2 signaling.