Estrogen-induced proliferation of uterine epithelial cells is independent of estrogen receptor α binding to classical estrogen response elements

Acting via the estrogen receptor (ER), estradiol exerts pleomorphic effects on the uterus, producing cyclical waves of cellular proliferation and differentiation in preparation for embryo implantation. In the classical pathway, the ER binds directly to an estrogen response element to activate or repress gene expression. However, emerging evidence supports the existence of nonclassical pathways in which the activated ER alters gene expression through protein-protein tethering with transcription factors such as c-Fos/ c-Jun B (AP-1) and Sp1. In this report, we examined the relative roles of classical and nonclassical ER signaling in vivo by comparing the estrogen-dependent uterine response in mice that express wild-type ERα, a mutant ERα (E207A/G208A) that selectively lacks ERE binding, or ERα null. In the compound heterozygote (AA/-) female, the nonclassical allele (AA) was insufficient to mediate an acute uterotrophic response to 17β-estradiol (E₂). The uterine epithelial proliferative response to E₂ and 4-hydroxytamoxifen was retained in the AA/- females, and uterine luminal epithelial height increased commensurate with the extent of ERα signaling. This proliferative response was confirmed by 5-bromo-2′-deoxyuridine incorporation. Microarray experiments identified cyclin-dependent kinase inhibitor 1A as a nonclassical pathway-responsive gene, and transient expression experiments using the cyclin-dependent kinase inhibitor 1A promoter confirmed transcriptional responses to the ERα (E207A/G208A) mutant. These results indicate that nonclassical ERα signaling is sufficient to restore luminal epithelial proliferation but not other estrogen-responsive events, such as fluid accumulation and hyperemia. We conclude that nonclassical pathway signaling via ERα plays a critical physiologic role in the uterine response to estrogen.