Quantitative analysis of estrogen receptor proteins in rat ovary

The mRNAs of estrogen receptor β (ERβ), and its splice variant, ERβ2, are abundant in granulosa cells in the ovary. With the use of antibodies, ERβ protein has also been shown to be abundantly expressed, but to date no ERβ2 protein has been demonstrated in the ovary. ERβ2 has a peptide, 18 amino acids in length, inserted into its ligand-binding domain, resulting in a reported 35-fold reduction in its affinity for estrogen (E2). ERα, ERβ1 and ERβ2 were quantified by Western blotting and by RT-PCR and their cellular localization in the ovary was examined by immunohistochemistry. In 3- and 5-week-old virgin, pregnant, lactating and post-lactating rats, the level of ERα protein ranged between 1.6 and 3.8 fmol/μg total protein. That of ERβ was 8.8-11.2 and of ERβ2, in the same samples, 4.1-5.9 fmol/μg total protein. ERβ2 and ERβ1 proteins were, therefore, present in approximately equal amounts in the ovary throughout the various reproductive stages. The major ERβ proteins in rat ovary, detected by their molecular weights on Western blots, were ERβ1-530 and ERβ2-548 (530 + 18 amino acids (aa)). Immunohistochemical staining revealed that ERβ and ERβ2 were expressed predominantly in granulosa cells of growing follicles, while ERα was found only in theca cells. In some theca cells, both ERα, ERβ2 were expressed. The data suggests that in theca cells, where it is co expressed with ERα, ERβ2 could function as a repressor of ERα. However, in granulosa cells where no ERα is detectable, and where E2 levels are high, ERβ2, with its low affinity for E2, could be an important sensor through which E2 exerts regulatory control.