Journal Article

Agonist-induced formation of unproductive receptor-G 12 complexes

Inoue, Asuka, Bouvier, Michel, Lambert, Nevin A., Lu, Sumin, Mathiasen, Signe, Javitch, Jonathan A., Okashah, Najeah, Zhou, Joris, Wright, Shane C., Kawakami, Kouki

2020

Overview

G protein-coupled receptors (GPCRs) are targeted by a large fraction of approved drugs and regulate many important cellular processes. Association of GPCRs with heterotrimeric G proteins in response to agonist activation is thought to invariably lead to G protein activation. We find instead that G 12 heterotrimers can associate with agonist-bound receptors in a manner that does not lead to activation. These unproductive agonist–receptor-G protein ternary complexes sequester G 12 heterotrimers and thus inhibit rather than support G 12 signaling. These findings reveal a mechanism whereby agonist activation of GPCRs can inhibit as well as promote G protein signaling. G proteins are activated when they associate with G protein-coupled receptors (GPCRs), often in response to agonist-mediated receptor activation. It is generally thought that agonist-induced receptor-G protein association necessarily promotes G protein activation and, conversely, that activated GPCRs do not interact with G proteins that they do not activate. Here we show that GPCRs can form agonist-dependent complexes with G proteins that they do not activate. Using cell-based bioluminescence resonance energy transfer (BRET) and luminescence assays we find that vasopressin V 2 receptors (V 2 R) associate with both G s and G 12 heterotrimers when stimulated with the agonist arginine vasopressin (AVP). However, unlike V 2 R-G s complexes, V 2 R-G 12 complexes are not destabilized by guanine nucleotides and do not promote G 12 activation. Activating V 2 R does not lead to signaling responses downstream of G 12 activation, but instead inhibits basal G 12 -mediated signaling, presumably by sequestering G 12 heterotrimers. Overexpressing G 12 inhibits G protein receptor kinase (GRK) and arrestin recruitment to V 2 R and receptor internalization. Formyl peptide (FPR1 and FPR2) and Smoothened (Smo) receptors also form complexes with G 12 that are insensitive to nucleotides, suggesting that unproductive GPCR-G 12 complexes are not unique to V 2 R. These results indicate that agonist-dependent receptor-G protein association does not always lead to G protein activation and may in fact inhibit G protein activation.