title: G Protein and β-arrestin signaling bias at the ghrelin receptor.
abstract: The G protein-coupled ghrelin receptor GHSR1a is a potential pharmacological target for treating obesity and addiction because of the critical role ghrelin plays in energy homeostasis and dopamine-dependent reward. GHSR1a enhances growth hormone release, appetite, and dopamine signaling through G(q/11), G(i/o), and G(12/13) as well as β-arrestin-based scaffolds. However, the contribution of individual G protein and β-arrestin pathways to the diverse physiological responses mediated by ghrelin remains unknown. To characterize whether a signaling bias occurs for GHSR1a, we investigated ghrelin signaling in a number of cell-based assays, including Ca(2+) mobilization, serum response factor response element, stress fiber formation, ERK1/2 phosphorylation, and β-arrestin translocation, utilizing intracellular second loop and C-tail mutants of GHSR1a. We observed that GHSR1a and β-arrestin rapidly form metastable plasma membrane complexes following exposure to an agonist, but replacement of the GHSR1a C-tail by the tail of the vasopressin 2 receptor greatly stabilizes them, producing complexes observable on the plasma membrane and also in endocytic vesicles. Mutations of the contiguous conserved amino acids Pro-148 and Leu-149 in the GHSR1a intracellular second loop generate receptors with a strong bias to G protein and β-arrestin, respectively, supporting a role for conformation-dependent signaling bias in the wild-type receptor. Our results demonstrate more balance in GHSR1a-mediated ERK signaling from G proteins and β-arrestin but uncover an important role for β-arrestin in RhoA activation and stress fiber formation. These findings suggest an avenue for modulating drug abuse-associated changes in synaptic plasticity via GHSR1a and indicate the development of GHSR1a-biased ligands as a promising strategy for selectively targeting downstream signaling events.
authors.affiliation: From the Departments of Cell Biology.
authors.forename: Tama
authors.initials: T
authors.lastname: Evron
authors.affiliation: From the Departments of Cell Biology.
authors.forename: Sean M
authors.initials: SM
authors.lastname: Peterson
authors.affiliation: From the Departments of Cell Biology.
authors.forename: Nikhil M
authors.initials: NM
authors.lastname: Urs
authors.affiliation: From the Departments of Cell Biology.
authors.forename: Yushi
authors.initials: Y
authors.lastname: Bai
authors.affiliation: From the Departments of Cell Biology.
authors.forename: Lauren K
authors.initials: LK
authors.lastname: Rochelle
authors.affiliation: From the Departments of Cell Biology, Neurobiology, and Medicine, Duke University, Medical Center, Durham, North Carolina 27710 marc.caron@dm.duke.edu.
authors.forename: Marc G
authors.initials: MG
authors.lastname: Caron
authors.affiliation: From the Departments of Cell Biology, lawrence.barak@dm.duke.edu.
authors.forename: Larry S
authors.initials: LS
authors.lastname: Barak
mesh_terms: D019393:Arrestin / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D020935:MAP Kinase Signaling System / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D009473:Neuronal Plasticity / Q000502:physiology; D055550:Protein Stability; D017433:Protein Structure, Secondary; D021381:Protein Transport / Q000502:physiology; D054440:Receptors, Ghrelin / Q000235:genetics / Q000378:metabolism*; D017483:Receptors, Vasopressin / Q000235:genetics / Q000378:metabolism
chemical_list: C585768:AVPR2 protein, human; D019393:Arrestin; D054440:Receptors, Ghrelin; D017483:Receptors, Vasopressin; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D019204:GTP-Binding Proteins
keywords: Actin; Arrestin; G Protein; G Protein-coupled Receptor (GPCR); Ghrelin; Rho (Rho GTPase); Signaling Bias