Effects of Estradici on Estrogen Receptor, Progesterone Receptor, and Tyrosinase in Hamster Melanoma Transplanted into Athymic Mice1

Nuclear estrogen binding was characterized in IIM-1, a malignant hamster melanoma cell line transplanted into male and female athymic mice following acute, subchronic, and chronic injection of estradiol. Nuclear binding was saturable, of high affinity ( IO'" M ') and readily soluble in low salt buffer. Saturation analyses revealed that (3H]estradiol in excess of 5.0 IIMapparently bound to a second class of lower affinity ¡III'1M~'), higher capacity cytosol sites. Enzyme-linked immunoassay with a specific monoclonal antibody (H222 SPY) directed against the human estrogen receptor protein was in excellent agreement (r —0.93) with values obtained using hydroxyapatite to separate bound from free ligand. Nuclear estrogen receptor content in 11M-1 cells was increased maxi mally l h after acute s.c. injection of a low dose (0.1 UK}of estradiol. The increase in nuclear receptor content was accompanied by an apparent rapid reduction in cytosol binding. Subchronic (3 days) and chronic exposure (35 days) to estradiol also produced a significant, dose-related increase in tumor nuclear estrogen receptor content. Cytosol binding for progestin was low (<2 fmol) to absent in IIM-1 xenografts not exposed to estradiol. Subchronic and chronic exposure to estradiol induced a dose-related, specific, high affinity (10'' M~') cytosol binding protein for progestin(s) in l IM-1 xenografts carried in male and female athymic mice. In contrast, progestin binding to nuclear receptor was not increased in estrogen-primed animals, nor did acute injection of progesterone (100 ftg s.c.) increase the amount of saturable, high affinity (10* M ') nuclear progestin receptor in control or estradiol-primed athymic mice. In contrast to the induction of progestin binding, tyrosinase activity was not altered by a similar exposure to estradiol when assayed at a saturating concentration of tyrosine. These observations suggest that the estrogen receptor in HM-1 cells may be functional but that pigmentary changes observed in mammals following chronic exposure to estradiol may not be mediated by a direct effect on the rate limiting enzyme of melanin synthesis.