ESTRADIOL MODULATES THYROID-HORMONE REGULATION OF THE HUMAN GLYCOPROTEIN HORMONE ALPHA-SUBUNIT GENE

We have examined mechanisms of regulation of the human glycoprotein ho rmone alpha subunit gene by thyroid hormone (T3) and estradiol. Pituit ary-derived GH3 cells were transiently transfected with chimeric const ructs comprising between 1,500 and 98 base pairs of human alpha subuni t gene 5'-flanking sequence fused to the bacterial gene encoding chlor amphenicol acetyltransferase (halphaCAT) and treated with T3 and estra diol, alone and in combination. In pituitary cells, 98 base pairs of a lpha gene 5'-flanking sequence were sufficient to mediate both inhibit ion of alpha gene promoter activity by T3 and stimulation by estradiol ; inhibition of the alpha promoter by T3 was antagonized by estradiol. Mutation of nucleotides essential for T3 receptor binding to the alph a gene thyroid hormone response element abolished the response of halp haCAT expression to estradiol as well as T3. In contrast to pituitary GH3 cells, estradiol treatment alone had no effect on expression of ei ther halphaCAT or the endogenous alpha gene in JEG-3 choriocarcinoma c ells cotransfected with a human thyroid hormone receptor expression ve ctor, but estradiol antagonized suppression of both endogenous and tra nsfected alpha promoter activity by T3. Gel mobility shift assays demo nstrated specific binding of in vitro synthesized human estrogen recep tor (ER) to the alpha gene thyroid hormone response element. These fin dings suggest that estradiol modulates expression of the human alpha s ubunit gene in pituitary and choriocarcinoma cells by direct binding o f ER to the alpha gene promoter, and that interaction of ER with the a lpha gene negative TRE accounts for the antagonistic effects of estrad iol and T3.