RGS14 regulates the lifetime of Gα-GTP signaling but does not prolong Gβγ signaling following receptor activation in live cells.

• Type of Material: Article
• Authors: Nicole E. Brown, Emory University; Nevin A. Lambert, Augusta University; John Hepler, Emory University
• Language: English
• Date: 2016-08-18
• Publisher: Wiley Open Access: Various Creative Commons Licenses
• Publication Version: Final Published Version
• Copyright Statement: © 2016 The Authors.
• License: Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1002/prp2.249
• Title of Journal or Parent Work: Pharmacology Research and Perspectives
• ISSN: 2052-1707
• Volume: 4
• Issue: 5
• Start Page: e00249
• End Page: e00249
• Grant/Funding Information: American Heart Association Predoctoral. Grant Number: 14PRE18850017; National Institutes of Health. Grant Numbers: 5R01NS037112, 5R21NS074975, R01GM078319, R01GM109879; National Institutes of Health Pharmacological Sciences Predoctoral Research Training. Grant Number: T32M008602
• Keywords: Gβγ; RGS protein; Gαi1; RGS14.; G protein; GPCR
• Research Categories: Health Sciences, Pharmacology; Health Sciences, Toxicology

Abstract

RGS14 is a multifunctional scaffolding protein possessing two distinct G protein interaction sites including a regulator of G protein signaling (RGS) domain that acts as a GTPase activating protein (GAP) to deactivate Gαi/o-GTP proteins, and a G protein regulatory (GPR) motif that binds inactive Gαi1/3-GDP proteins independent of Gβγ. GPR interactions with Gαi recruit RGS14 to the plasma membrane to interact with Gαi-linked GPCRs and regulate Gαi signaling. While RGS14 actions on Gα proteins are well characterized, consequent effects on Gβγ signaling remain unknown. Conventional RGS proteins act as dedicated GAPs to deactivate Gα and Gβγ signaling following receptor activation. RGS14 may do the same or, alternatively, may coordinate its actions to deactivate Gα-GTP with the RGS domain and then capture the same Gα-GDP via its GPR motif to prevent heterotrimer reassociation and prolong Gβγ signaling. To test this idea, we compared the regulation of G protein activation and deactivation kinetics by a conventional RGS protein, RGS4, and RGS14 in response to GPCR agonist/antagonist treatment utilizing bioluminescence resonance energy transfer (BRET). Co-expression of either RGS4 or RGS14 inhibited the release of free Gβγ after agonist stimulation and increased the deactivation rate of Gα, consistent with their roles as GTPase activating proteins (GAPs). Overexpression of inactive Gαi1 to recruit RGS14 to the plasma membrane did not alter RGS14's capacity to act as a GAP for a second Gαo protein. These results demonstrate the role of RGS14 as a dedicated GAP and suggest that the G protein regulatory (GPR) motif functions independently of the RGS domain and is silent in regulating GAP activity in a cellular context.