Steroid hormone regulation and tissue-specific expression of the human GnRH gene in cell culture and transgenic animals.

In order to study the molecular mechanisms involved in the control of GnRH gene expression, the human GnRH gene was cloned and characterized. The gene was expressed in cells obtained from CNS tumors in transgenic mice generated utilizing 1131 bp of 5' flanking GnRH DNA fused to the simian virus 40 large T antigen. We have shown a stimulatory estrogen response element in the human GnRH gene by transient transfection studies. DNase I footprinting and an avidinbiotin DNA binding assay demonstrated that the human GnRH gene bound ER. The GN cell line was found to have nuclear ERs utilizing an 125I estradiol binding study and by in situ hybridization histochemistry. In order to study GnRH expression in vivo, either 5000 or 484 bp of GnRH flanking DNA was fused to the luciferase (Luc) reporter gene, and transgenic mice generated. Expression in the transgenic animals was found in the hypothalamus of animals bearing the -5000Luc transgene, but not in animals bearing the -484Luc transgene. The transgenic mice expressing the -5000Luc gene were gonadectomized resulting in a 20-30% increase in hypothalamic Luc expression in the males and a 65% increase in females, while mice who were gonadectomized and replaced with testosterone (males) or E2 (females) showed a 50% decrease in Luc expression over control levels. Thus, these studies present in vitro evidence of E2 modulation of GnRH gene expression and an in vivo model in which sensitive studies of GnRH regulation and expression can be performed.