Activins Stimulate Human Follicle-Stimulating Hormone β (FSHB) Expression Via a FOXL2-Dependent Mechanism in Pituitaries of Transgenic Mice

Presentation Number: SUN 249
Date of Presentation: April 2nd, 2017

Luisina Ongaro Gambino*1, T. Rajendra Kumar2, Mathias Treier3 and Daniel J. Bernard4
1McGill University, 2University of Colorado Denver, Anschutz Medical Campus, 3Max Delbruck Center for Molecular Medicine, Berlin, Germany, 4McGill University, Montreal, QC, Canada

Abstract

Alterations in follicle-stimulating hormone (FSH) synthesis or action underlie many forms of infertility. FSH synthesis by pituitary gonadotrope cells is stimulated by GnRH from the hypothalamus and intra-pituitary activins. Activins bind complexes of type I/type II receptor serine/threonine kinases, which phosphorylate SMAD proteins. SMAD complexes then accumulate in the nucleus where they bind the promoter region of the FSHβ subunit (Fshb) encoding gene in combination with forkhead box L2 (FOXL2). Deletion of the genes encoding Smad4 and/or Foxl2 in gonadotrope cells causes profound FSH deficiency in mice. Mechanisms of activin-regulated human FSHB expression, in contrast, are relatively unknown. FOXL2 is expressed in human gonadotropes and can bind to three forkhead binding elements in the proximal human FSHB promoter in vitro. However, human FSHB promoter-reporters are poorly responsive to activins in immortalized cells, precluding an assessment of these cis-elements or the FOXL2 protein in transcription of the human gene. We therefore took an alternate approach to examine the role of FOXL2 in activin regulation of human FSHB expression. Mice harboring a 10 kb human FSHB transgene (hereafter hFSHB) express the corresponding mRNA and protein specifically in gonadotrope cells. We cultured pituitaries from these animals to assess the necessity and sufficiency of activin signaling for expression of FSHB. As we previously reported, murine Fshb mRNA expression was stimulated by exogenous activin A or B in cultures from male or female mice. Basal Fshb expression, which depends on endogenous activin-like signaling, was blocked by follistatin-288 or the type I receptor inhibitor SB431542. Similarly, in the female pituitary cultures, activins stimulated and the inhibitors attenuated human FSHB mRNA levels. In contrast, human FSHB mRNA expression was only modestly regulated by activins or the inhibitors in male cultures. Further, we crossed hFSHB transgenic mice with animals carrying floxed alleles for Foxl2. We cultured their pituitaries and infected them with control or Cre-expressing adenoviruses. FOXL2 ablation impaired basal and activin-stimulated murine Fshb and human FSHB mRNA levels. Thus, the human FSHB gene is activin responsive, in pituitaries of female mice, and is dependent on FOXL2 for its expression. These results suggest that mechanisms of Fshb/FSHB regulation by activins and FOXL2 are conserved across species.

 

Nothing to Disclose: LO, TRK, MT, DJB