Differential regulation of proteasome-dependent estrogen receptor α and β turnover in cultured human uterine artery endothelial cells

Estrogen-induced loss of estrogen receptor (ER) alpha expression limits estrogen responsiveness in many target cells. However, whether such a mechanism contributes to changes in vascular endothelial ERalpha and/or ERbeta levels is unclear. Using RT-PCR assays, we did not find any regulation of ERalpha or ERbeta mRNA expression in human uterine artery endothelial cell (HUAEC) nuclear extracts on stimulation with 17beta-estradiol for 1 or 2 h. By contrast, Western analysis on HUAEC extracts revealed that 17beta-estradiol was capable of down-regulating both ERalpha and ERbeta protein starting 1 h after treatment, an effect that can be blocked by pretreatment with tamoxifen as well as with the proteasome inhibitor lactacystin. The proteolysis inhibitors insulin, cycloheximide, and puromycin impede ERalpha, but not ERalpha, turnover. Ubiquitin, but not its competitive inhibitor methyl-ubiquitin, induces rapid turnover of both ERs in a cell-free system of MCF-7 and HUAEC extracts. We, thus, propose the existence of estrogen-induced ER degradation that serves to control physiological responses in an estrogen target tissue, i.e. human vascular endothelium, by down-regulating ERalpha as well as ERbeta through different proteasomal uptake mechanisms.