Anti-estrogen activity in the yeast transcription system: Estrogen receptor mediated agonist response

Abstract

The mouse estrogen receptor was expressed in yeast cells to study the mechanism of action of anti-estrogens. Tamoxifen and hydroxytamoxifen, estrogen antagonists in mammalian tissues, faled to antagonize estradiol-induced expression of a Vit A2-ERE-CTC1-lacZ reporter gene construct and exhibited full agonist activity, while nafoxidine exhibited partial antagonism as well as partial agonism. ICI 164,384 is a potent anti-estrogen in both mouse and human estrogen receptor system. Our previous studies in the mouse uterus indicated that rapid degradation of the estrogen receptor accounted for the loss of estrogen responsiveness. In yeast however, ICI 164,384 or an isomer ICI 182,780 were unable to antagonize estradiol at concentration of 200 μM. On the contrary, both ICI compounds exhibited partial agonist activity by stimulating β-galactosidase activity to 50% that of estradiol. We examined the level of estrogen receptor in the yeast after treatment with estradiol, ICI 164,384 or vehicle by Western blot and found no ICI-induced erduction of estrogen receptor levels, but observed an increase in estrogen receptor following estradiol treatment. This indicates that the proteolytic activity responsible for degrading estrogen receptor in ICI 164,384-treated uteri or eukaryotic cells is not present in yeast. The agonist activity seen with ICI indicated that ICI-bound estrogen receptor is able of induce expression of an estrogen-responsive reporter gene. In support of this, estrogen receptor from ICI 164,384-treated yeast was able to bind an estrogen-responsive element in a gel-shift assay. The inability of ICI 164,384 to antagonize estradiol in the yeast gene-expression system will allow us to investigate mechanisms relevant to hormonal antagonism which are present in mammalian ssytems, but lacking in yeast. Agonistic activity of ICI 164,384 or ICI 182,780 and the lack of antagonism indicates that an active ICI receptor complex can be formed due to the deficient proteolysis in the yeast compared to mammalian systems where ICI stimulates increased receptor turnover and loss