Follicle-Stimulating Hormone Synthesis and Fertility Require the Independent and Complementary Activities of SMAD4 and FOXL2 in Gonadotropes

Presentation Number: OR30-6
Date of Presentation: June 23rd, 2014

Jerome Fortin*1, Ulrich Boehm2, Chuxia Deng3, Mathias Treier4 and Daniel J. Bernard1
1McGill University, Montreal, QC, Canada, 2University of Saarland School of Medicine, Homburg, Germany, 3NIH, Bethesda, MD, 4Max Delbruck Center for Molecular Medicine, Berlin, Germany

Abstract

Follicle-stimulating hormone (FSH) is an essential regulator of gonadal function and fertility in humans and mice. Gonadotropin-releasing hormone (GnRH) and activins stimulate FSH synthesis; yet, their relative roles and mechanisms of action in vivo are incompletely understood. In cell lines, activins signal through the receptor-regulated SMAD proteins, SMAD2/3, the obligatory co-SMAD, SMAD4, and the SMAD-interacting, cell-restricted factor, forkhead box L2 (FOXL2), to stimulate transcription of the FSHβ subunit gene (Fshb). Consistent with these observations, mice with gonadotrope-restricted deletions of Smad4 (S4cKO) or Foxl2 (F2cKO) are FSH-deficient and subfertile. Based on in vitro observations, we predict that SMAD4 can partially mediate activin-induced Fshb transcription in the absence of FOXL2 (and vice versa), potentially explaining the residual FSH synthesis and fertility in S4cKO or F2cKO mice. To test this idea, we generated mice lacking both factors in gonadotropes (S4F2cKO), by crossing mice carrying “floxed” alleles of Smad4 and Foxl2 with GnrhrIRES-Cre (GRIC) mice. In striking contrast with the effects of the single gene knockouts, S4F2cKO mice displayed severe hypogonadism. S4F2cKO females had small ovaries (devoid of antral follicles and corpora lutea), thread-like uteri, and were acyclic and sterile. Male S4F2cKO mice had small testes, but were fertile. Thus, S4F2cKO mice phenocopy Fshb-deficient animals. Indeed, male and female S4F2cKO mice were profoundly FSH- and Fshb-deficient. FSHβ immunoreactivity was absent in LHβ-positive pituitary gonadotropes of S4F2cKO mice, but retained in a small population of “FSHβ-only” cells. S4F2cKO animals had elevated pituitary expression of the luteinizing hormone β subunit gene (Lhb), possibly as a result of reduced negative feedback by gonadal sex steroids. In females, this resulted in elevated circulating LH levels. In contrast, serum LH was diminished in males, likely due to a sex-specific downregulation of the gonadotropin α subunit (Cga). Collectively, these results establish SMAD4 and FOXL2 as master regulators of Fshb expression in vivo, and suggest that activins may be more important regulators of FSH synthesis than GnRH.

 

Nothing to Disclose: JF, UB, CD, MT, DJB