Early growth response protein 1 binds to the luteinizing hormone-β promoter and mediates gonadotropin-releasing hormone-stimulated gene expression

Abstract

The hypothalamic neuropeptide, GnRH, regulates the synthesis and secretion of LH from pituitary gonadotropes. Furthermore, it has been shown that the LH β-subunit gene is regulated by the transcription factors steroidogenic factor-1 (SF-1) and early growth response protein I (Egr1) in vitro and in vivo. The present study investigated the roles played by Egr1 and SF-1 in regulating activity of the equine LHβ-subunit promoter in the gonadotrope cell line, αT3-1, and the importance of these factors and cis-acting elements in regulation of the promoter by GnRH. All four members of the Egr family were found to induce activity of the equine promoter. The region responsible for induction by Egr was localized to the proximal 185 bp of the promoter, which contained two Egr response elements. Coexpression of Egr1 and SF-1 led to a synergistic activation of the equine (e)LHβ promoter. Mutation of any of the Egr or SF-1 response elements attenuated this synergism. Endogenous expression of Egr1 in αT3-1 cells was not detectable under basal conditions, but was rapidly induced after GnRH stimulation. Reexamination of the promoter constructs harboring mutant Egr or SF-1 sites indicated that these sites were required for GnRH induction. In fact, mutation of both Egr sites within the eLHβ promoter completely attenuated its induction by GnRH. Thus, GnRH induces expression of Egr1, which subsequently activates the eLHβ promoter. Finally, GnRH not only induced expression of Egr1, but also its corepressor, NGFI-A (Egr1) binding protein (Nab1), which can repress Egr1- induced transcription of the eLHβ promoter.