G Protein and Beta-arrestin Signaling Bias at the Ghrelin Receptor [Signal Transduction]

September 26th, 2014 by Evron, T., Peterson, S. M., Urs, N. M., Bai, Y., Rochelle, L. K., Caron, M. G., Barak, L. S.

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 GH release, appetite, and dopamine signaling through Gq/11, Gi/o and G12/13 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 if signaling bias occurs for the GHSR1a, we investigated ghrelin signaling in a number of cell-based assays, including Ca2+ mobilization, SRF-RE, stress fiber formation, ERK1/2 phosphorylation and β-arrestin translocation, utilizing intracellular second loop (ICL2) and C-tail mutants of GHSR1a. We observed that GHSR1a and β-arrestin rapidly form metastable plasma-membrane complexes following exposure to 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 P148 and L149 in GHSR1a ICL2 generate receptors with 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.