Molecular analysis of estrogen induction of preproenkephalin gene expression and its modulation by thyroid hormones

Estrogen receptors (ER) and thyroid hormone receptors (TR) are ligand-depen dent nuclear transcription factors. Estrogen-induced preproenkephalin (PPE) gene expression in the hypothalamus is directly related to estrogen-induce d lordosis behavior in the rat. In the present study, we showed that the PP E mRNA level in the ventromedial hypothalamus of female rats was significan tly decreased by ovariectomy. This decrease was reversed by estrogen replac ement in a dose- and time-dependent manner. Using transient transfection an d electrophoretic mobility shift assays (EMSA), functional estrogen respons e elements (ERE) were identified between -437 and -145 base pairs (bp) of t he rat PPE gene promoter region. Two ERE-like elements are present between -405 and -364 of the rat PPE gene promoter, which bind ER alpha as demonstr ated by EMSA. Estrogen produced a dose-dependent increase in CAT activity i n cotransfection assays with ER alpha expression vector and a 437PPE-CAT re porter construct containing 437 bp of the: rat PPE gene promoter and the CA T reporter gene. This estrogen-induced PPE promoter activity was inhibited by liganded-TR in transient cotransfection assays. Analysis of DNA-protein interactions by EMSA revealed that both ER alpha and TR (alpha1 and beta1) could bind to the EREs in the rat PPE gene promoter. Furthermore, estrogen induction of PPE mRNA in the ventromedial hypothalamus of the ovariectomize d female rat was significantly attenuated by concomitant administration of triiodothyronine. These results suggest that estrogen regulation of the hyp othalamic PPE gene expression is mediated through an estrogen-receptor comp lex directly interacting with the functional EREs in its promoter region; a nd that this estrogen effect can be modified by thyroid hormones. (C) 2001 Elsevier Science B.V. All rights reserved.