Gonadotropin-Releasing Hormone Receptor Couples to Multiple G Proteins in Rat Gonadotrophs and in GGH3 Cells: Evidence from Palmitoylation and

There is evidence in several cell systems suggesting that the GnRH receptor couples to multiple G proteins. Presently there are no published studies showing GnRH receptor coupling to Gia, Gsa, and Gq/11a in a single cell type. To examine this pos-sibility we measured palmitoylation of G proteins in response to GnRH receptor occupancy, since this event is a measure of G-protein activation by cognate receptors. GnRH stimulated time (0–120 min)- and dose (10212–1026 g/ml)-dependent palmitoy-lation of both Gia and Gsa. Palmitoylation is G-protein activa-tion dependent; accordingly, pertussis toxin (100 ng/ml; PTX), phorbol myristic acid (100 ng/ml), and Antide (50 nM; a GnRH antagonist) did not stimulate palmitoylation of Gia or Gsa above basal levels. However, cholera toxin (5 mg/ml), an activator of Gsa, stimulated palmitoylation of Gsa but not Gia. We used a lactotrope-derived cell line expressing the GnRH receptor (GGH3) to examine whether the ability of the receptor to couple multiple G proteins is gonadotroph specific. GGH3 cells were transfected with specific cDNA coding for different G proteins, and agonist-stimulated second messenger production was as-sessed. Buserelin (a GnRH agonist) stimulated increased cAMP release in Gsa cDNA-transfected GGH3 cells, whereas in Gia cDNA-transfected cells, both inositol phosphate (IP) production and cAMP release were decreased in response to buserelin. Transfection of Gqa, G11a, G14a, and G15a cDNA into GGH3 cells resulted in an increased IP production in response to bus-erelin, indicating that GnRH receptor couples to this PTX-insen-sitive G-protein family. The observations presented in this study provide evidence for GnRH receptor coupling to multiple G pro-teins in a single cell type