The melanin-concentrating hormone receptor couples to multiple G proteins to activate diverse intracellular signaling pathways

• Published in: Endocrinology (Philadelphia) Vol. 141; no. 12; pp. 4524 - 4532
• Main Authors: Hawes, B E, Kil, E, Green, B, O'Neill, K, Fried, S, Graziano, M P
• Format: Journal Article
• Language: English
• Published: United States 01-12-2000
• Subjects: Adenylyl Cyclases - metabolism; Animals; Cell Membrane - metabolism; CHO Cells; Colforsin - pharmacology; Cricetinae; Cyclic AMP - biosynthesis; Gene Expression; GTP-Binding Protein alpha Subunits, Gi-Go - physiology; GTP-Binding Proteins - physiology; Heterotrimeric GTP-Binding Proteins - physiology; Humans; Hypothalamic Hormones - metabolism; Hypothalamic Hormones - pharmacology; Inositol Phosphates - biosynthesis; Lysophospholipids - pharmacology; Melanins - metabolism; Melanins - pharmacology; Mitogen-Activated Protein Kinases - metabolism; Pertussis Toxin; Pituitary Hormones - metabolism; Pituitary Hormones - pharmacology; Protein Kinase C - antagonists & inhibitors; Receptors, Pituitary Hormone - genetics; Receptors, Pituitary Hormone - physiology; Signal Transduction; Transfection; Virulence Factors, Bordetella - pharmacology
• ISSN: 0013-7227
• DOI: 10.1210/en.141.12.4524

The receptor for melanin-concentrating hormone (MCH) was recently identified as the orphan G protein-coupled receptor SLC-1. In this study, a CHO cell line expressing the MCH receptor (Kd = 1.3 nM; binding capacity, 3.6 pmol/mg protein) is used to assess the ability of the MCH receptor to couple to Gi, Go, and Gq proteins. The results demonstrate that MCH inhibits forskolin-stimulated cAMP production in a pertussis toxin- (PTX)-sensitive manner in CHO-MCHR cells (EC50 = 100 pM), indicating that the MCH receptor couples to one or more members of the Gi subfamily of G proteins. In addition, MCH stimulates increases in phosphoinositide metabolism (EC50 = 50 nM) and in intracellular free Ca2+ levels (EC50 = 10 nM). MCH-stimulated inositol phosphate production and increases in intracellular free Ca2+ are partially inhibited (60% and 40%, respectively) by PTX pretreatment, demonstrating that there are at least two components of each of these signaling pathways. One component is PTX sensitive and therefore mediated through a Gi/Go protein. A distinct G protein-coupled (probably Gq type) mediates the PTX-insensitive component. To distinguish Gi vs. Go coupling, MCH-stimulated mitogen-activated protein (MAP) kinase activity was examined. Gi and Go use separate signaling pathways to mediate MAP kinase activation in CHOcells. Protein kinase C (PKC) activity is essential in the Go-dependent MAP kinase signaling pathway, but is not required in the GC-dependent MAP kinase signaling pathway. MCH stimulated MAP kinase activity is decreased (50%), but not abolished, by inhibition of PKC activity or depletion of cellular PKC, indicating that MCH-stimulated MAP kinase activity is mediated through both Gi- and Go-dependent signaling mechanisms. The results of this study are the first to clearly demonstrate that the MCH receptor couples to multiple G proteins to mediate several diverse intracellular signaling pathways.