Ligand-specific regulation of proteasome-mediated proteolysis of estrogen receptor-␣. Am J Physiol Endocrinol Metab 282: E891-E898, 2002. First published December 18, 2001 10.1152/ajpendo.00353.2001.-Proteasome-mediated proteolysis modulates the cellular concentration of estrogen receptor-␣ (ER␣) and is induced by treatment of cells with 17␤-estradiol. Herein, we show that multiple receptor agonists, including 17␣-estradiol and estriol as well as the antagonist ICI-182780, stimulate proteasome-dependent proteolysis of ER␣ in a process that requires ligand binding to the receptor. Proteolysis of receptor depends on ligand concentration, and there exists a direct correlation between ligand-binding affinity and the half-maximal dose of ligand required to stimulate receptor degradation. Furthermore, introduction of a point mutation into the receptor ligand-binding pocket yields a stable receptor resistant to proteolysis. Interestingly, although all ligands stimulate receptor degradation, the extent to which overall ER levels are affected varies with each ligand and is not related to ligand-binding affinity or activation of transcription. These results demonstrate ligand-specific regulation of ER␣ proteolysis, and they introduce the concept that cellular receptor concentration is governed not only at the level of induction of proteolysis but also by the efficiency with which the receptor is degraded. steroid; nuclear receptor; antagonist; estriol; pituitary CELLULAR ESTROGEN RECEPTOR LEVELS are dynamic and are particularly sensitive to changes in circulating levels of 17␤-estradiol. It has been demonstrated through a number of studies that the decline in estrogen receptor-␣ (ER␣) upon exposure to 17␤-estradiol results from a combination of mechanisms that control both receptor synthesis and degradation through transcriptional, posttranscriptional, and posttranslational mechanisms (20,27,32,33,35,36). The most rapid of these regulatory mechanisms is the direct loss of ER␣ protein brought about by the induction of proteasome-mediated proteolysis (1,21,28).Regulated proteolysis by proteasomes accounts for the turnover of most short-lived proteins, including many nuclear receptors (9,16,23,29,41,43,45). Through a series of three enzymatic reactions, ubiquitin moieties are attached to a protein substrate, which targets it to the 26S proteasome. The molecular events that direct ER␣ into this pathway have not been clearly established. However, earlier studies that examine changes in receptor-binding capacity have shown that receptor levels can be controlled by both receptor agonists and antagonists, suggesting the possibility that receptor occupation by ligand may provide specificity (3,4,15,18,22,33).In our original report of ER␣ protein regulation by proteolysis, we utilized a pituitary lactotrope model system, the PR1 cell line. The lactotrope cell population of the anterior pituitary is a major target of estrogen action. Animals that lack ER␣ show a decrease in lactotrope cell density and prolactin expression (37). In ...