Analysis of G protein betagamma dimer formation in live cells using multicolor bimolecular fluorescence complementation demonstrates preferences of beta1 for particular gamma subunits

Molecular Pharmacology
Stacy M MervineCatherine H Berlot

Abstract

The specificity of G protein betagamma signaling demonstrated by in vivo knockouts is greater than expected based on in vitro assays of betagamma function. In this study, we investigated the basis for this discrepancy by comparing the abilities of seven beta1gamma complexes containing gamma1, gamma2, gamma5, gamma7, gamma10, gamma11, or gamma12 to interact with alphas and of these gamma subunits to compete for interaction with beta1 in live human embryonic kidney (HEK) 293 cells. betagamma complexes were imaged using bimolecular fluorescence complementation, in which fluorescence is produced by two nonfluorescent fragments (N and C) of cyan fluorescent protein (CFP) or yellow fluorescent protein (YFP) when brought together by proteins fused to each fragment. Plasma membrane targeting of alphas-CFP varied inversely with its expression level, and the abilities of YFP-N-beta1YFP-C-gamma complexes to increase this targeting varied by 2-fold or less. However, there were larger differences in the abilities of the CFP-N-gamma subunits to compete for association with CFP-C-beta1. When the intensities of coexpressed CFP-C-beta1CFP-N-gamma (cyan) and CFP-C-beta1YFP-N-gamma2 (yellow) complexes were compared under conditions in which CFP-C...Continue Reading

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Citations

Jul 26, 2011·Nature Chemical Biology·Eneko UrizarJonathan A Javitch
Oct 7, 2008·Annual Review of Pharmacology and Toxicology·Denis J DupréTerence E Hébert
Jun 14, 2013·The Journal of Neuroscience : the Official Journal of the Society for Neuroscience·Jenna M RamakerPhilip F Copenhaver
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Sep 30, 2009·Cellular Signalling·Timothy MulliganSteven A Farber

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