Research ArticleGPCR SIGNALING

Distinct profiles of functional discrimination among G proteins determine the actions of G protein–coupled receptors

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Sci. Signal.  01 Dec 2015:
Vol. 8, Issue 405, pp. ra123
DOI: 10.1126/scisignal.aab4068

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Fingerprinting GPCRs

G protein–coupled receptors (GPCRs) influence most aspects of physiology and are targeted by many clinically used drugs. The physiological functions of this large family of proteins are thought to be mediated by highly specific G protein interactions (see the Focus by Smrcka). Masuho et al. devised a bioluminescence-based assay in transfected cells to examine the G protein–coupling specificities of different GPCRs for 13 different G proteins, generating fingerprint-like profiles for each receptor. These assays revealed unexpected use of G proteins by certain GPCRs, biased G protein usage by a given GPCR in response to different agonists, and G protein activation by a ligand thought to be an antagonist, findings that were verified in relevant cells. Given the clinical importance of this family of receptors, assays such as this one aid in understanding the physiological effects of currently used drugs, as well as in designing better therapeutics and limiting their potential side effects.

Abstract

Members of the heterotrimeric guanine nucleotide–binding protein (G protein)–coupled receptor (GPCR) family play key roles in many physiological functions and are extensively exploited pharmacologically to treat diseases. Many of the diverse effects of individual GPCRs on cellular physiology are transduced by heterotrimeric G proteins, which are composed of α, β, and γ subunits. GPCRs interact with and stimulate the binding of guanosine triphosphate (GTP) to the α subunit to initiate signaling. Mammalian genomes encode 16 different G protein α subunits, each one of which has distinct properties. We developed a single-platform, optical strategy to monitor G protein activation in live cells. With this system, we profiled the coupling ability of individual GPCRs for different α subunits, simultaneously quantifying the magnitude of the signal and the rates at which the receptors activated the G proteins. We found that individual receptors engaged multiple G proteins with varying efficacy and kinetics, generating fingerprint-like profiles. Different classes of GPCR ligands, including full and partial agonists, allosteric modulators, and antagonists, distinctly affected these fingerprints to functionally bias GPCR signaling. Finally, we showed that intracellular signaling modulators further altered the G protein–coupling profiles of GPCRs, which suggests that their differential abundance may alter signaling outcomes in a cell-specific manner. These observations suggest that the diversity of the effects of GPCRs on cellular physiology may be determined by their differential engagement of multiple G proteins, coupling to which produces signals with varying signal magnitudes and activation kinetics, properties that may be exploited pharmacologically.

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