Regulation of the rat follicle-stimulating hormone beta-subunit promoter by activin

Abstract

FSH is controlled by a variety of positive and negative stimuli, and the unique FSHbeta-subunit is a major target for this regulation. Activin is a key modulator of FSHbeta transcription and hormone secretion. The signal transduction pathway leading to FSH expression was previously unknown. Here, we show that the transcription factors Smad3 and Smad4 mediate activin-stimulated activity of the rat FSHbeta promoter in a pituitary-derived cell line, LbetaT2. Cells were transiently transfected with the rat FSHbeta promoter fused to a luciferase reporter gene (-338rFSHbeta-Luc), and a minimal activin-responsive region was identified. Transfection of Smad3, but not the highly related Smad2, led to a ligand-independent stimulation of the FSHbeta promoter activity. As expected, activin caused an additional increase of luciferase expression, which was blocked by cotreatment with follistatin. Although Smad4 alone had no effect on FSHbeta transcription, it significantly augmented Smad3 and activin-mediated stimulation of the promoter. A palindromic consensus Smad-binding element in the proximal promoter was found to bind Smad4, and elimination of the region resulted in a loss of activin-mediated FSHbeta transcription. The activin signaling pathway is conserved in a number of cells, but FSHbeta expression is restricted to gonadotropes. A pituitary-specific transcription factor necessary for activin-dependent induction of the FSHbeta promoter has been identified that permits FSHbeta expression in nongonadotrope cells. Pitx2 is a member of Pitx subfamily of bicoid-related homeodomain factors that is required for pituitary development and is present in the adult pituitary. This factor was transfected into LbetaT2 cells, where it caused up-regulation of basal and activin-mediated FSHbeta promoter activity. Furthermore, cotransfection of Pitx2c with Smad3 in kidney-derived TSA cells resulted in activin-regulated FSHbeta response, suggesting its important role in tissue-restricted regulation of FSHbeta by activin. A Pitx2c binding site was identified within the proximal promoter, and elimination of this region also resulted in a loss of activin-regulated FSHbeta promoter activity. Taken together, these studies suggest that the regulation of FSHbeta is dependent on activin-mediated signaling factors in concert with pituitary-derived nuclear regulatory proteins.