Intracellular Calcium Signaling Is Required for GnRH Mediated Regulation of T-Cell Factor Dependent Transcription in Gonadotropes.

Abstract

Gonadotropes of the anterior pituitary synthesize and secrete the gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH). LH and FSH are essential for normal reproductive function and act on the ovary and testes to regulate steriodogenesis and gametogenesis. The genes that encode these two hormones are regulated by gonadotropin-releasing hormone (GnRH), which is secreted from the hypothalamus and binds to its receptor on gonadotropes. Upon binding to its receptor, GnRH activates Gαq that in turn activates multiple members of the mitogen-activated protein kinase (MAPK) signaling family, including extracellular regulated kinase (ERK), JUN N-terminal kinase (JNK) and p38 MAPK as well as an increase Ca++ mobilization. Recent work from our laboratory has shown that the co-activator β-catenin is a downstream target of GnRH and required for hormonal regulation of several genes including Jun. GnRH regulation of Jun also requires T-cell factor (TCF) in addition to β-catenin. In this study we test the hypothesis that GnRH regulates β-catenin dependent genes and that this regulation requires activation of c-src tyrosine kinase (SRC), JNK as well as an increase in intracellular Ca++. Pretreatment of LβT2 cells with SU6656, a SRC specific inhibitor reduces GnRH mediated activation of TOPflash, an artificial β-catenin dependent reporter, as well as expression of Jun. Similar results were also observed with a JNK specific inhibitor (SP600125). Pretreatment of LβT2 cells with BAPTA-AM, an intracellular Ca++ chelator, reduces GnRH mediated activation of TOPflash as well as expression of Jun. In contrast, inhibition of ERK and p38 MAPK had no effect on GnRH regulated activity of TOPflash or Jun, suggesting that the JNK pathway acts uniquely to mediate the effects of the neurohormone. In summary, GnRH appears to signal through SRC, JNK and calcium to regulate TCF/β-catenin dependent transcription of Jun, an immediate early gene that ultimately confers hormonal responsiveness to several key genes required for gonadotropin synthesis and secretion. This research was supported by NIH RO1 HD055776 to J.H.N. (platform)