Progestin Regulation of Estrogen Receptor Messenger RNA in Human Breast Cancer Cells

Abstract

Progestin antagonism of estrogen action is thought to be due, at least in part, to progestin down-regulation of the estrogen receptor (ER). The molecular mechanisms subserving this effect, and the functional consequences in terms of target cell sensitivity to estrogens, are poorly understood. The present study was undertaken to address these issues with particular emphasis on progestin regulation of ER gene expression at the mRNA level. The T-47D human breast cancer cell line was treated with the synthetic progestin, ORG 2058, and the resultant changes in ER mRNA and ER levels determined by Northern analysis and radioligand binding, respectively. Treatment of T-47D cells with ORG 2058 resulted in rapid down-regulation of ER mRNA levels to a nadir of 35–40% of control by 6 h. This fall in ER mRNA levels was accompanied by a slower but more sustained fall in ER binding to a nadir of 20% of control at 24 h. Between 12 and 24 h ER mRNA levels recovered partially while ER ligand binding continued to fall. At 48 h both ER mRNA and ER concentrations remained depressed, although the latter to a greater extent. ER mRNA half-life was determined by [³H]uridine incorporation to be approximately 60 min and was unaffected by progestin treatment during the early rapid phase of ER mRNA down-regulation. These data demonstrate that progestins cause rapid down-regulation of the ER mRNA and suggest that during the early rapid phase of this effect, reduced transcription of the ER gene rather than altered ER mRNA half-life mediate this effect. In an attempt to relate these decreased levels of ER to estrogen sensitivity the bacterial chloramphenicol acetyl transferase gene driven by an estrogen responsive promoter was transiently transfected into T-47D cells. Progestin treatment of transfected cells partially antagonized estrogen induction of reporter gene activity in a concentration dependent fashion. Furthermore the extent and concentration dependence of this effect correlated with the degree of down-regulation of the ER by progestins supporting a role for ER down-regulation in the reduced sensitivity of target cells to estrogen afte progestin treatment.