%0 Journal Article %A Sundeep Malik %A Mousumi Ghosh %A Tabetha M. Bonacci %A Gregory G. Tall %A Alan V. Smrcka %T Ric-8 Enhances G Protein βγ-Dependent Signaling in Response to βγ-Binding Peptides in Intact Cells %D 2005 %R 10.1124/mol.104.010116 %J Molecular Pharmacology %P 129-136 %V 68 %N 1 %X Peptides derived from a random-peptide phage display screen with purified Gβ1γ2 subunits as the target promote the dissociation of G protein heterotrimers in vitro and activate G protein signaling in intact cells. In vitro, one of these peptides (SIRKALNILGYPDYD; SIRK) promotes subunit dissociation by binding directly to Gβγ subunits and accelerating the dissociation of GαGDP without catalyzing nucleotide exchange. The experiments described here were designed to test whether the mechanism of SIRK action in vitro is in fact the mechanism of action in intact cells. We created a mutant of Gβ1 subunits (β1W332A) that does not bind SIRK in vitro. Transfection of Gβ1W332A mutant into Chinese hamster ovary cells blocked peptide-mediated activation of extracellular signal-regulated kinase (ERK), but it did not affect receptor-mediated Gβγ subunit-dependent ERK activation, indicating that Gβγ subunits are in fact the direct target in cells responsible for ERK activation. To determine whether free Gα subunits were released from G protein heterotrimers upon peptide treatment, cells were transfected with Ric-8A, a guanine nucleotide exchange factor for free GαGDP, but not heterotrimeric G proteins. Ric-8A-transfected cells displayed enhanced myristoyl-SIRKALNILGYPDYD (mSIRK)-dependent inositol phosphate (IP) release and ERK activation. Ric-8A also enhanced ERK activation by the Gi-linked G protein coupled receptor agonist lysophosphatidic acid. Inhibitors of Gβγ subunit function blocked Ric-8-enhanced activation of ERK and IP release. These results suggest that one potential function of Ric-8 in cells is to enhance G protein Gβγ subunit signaling. Overall, these experiments provide further support for the hypothesis that mSIRK promotes G protein subunit dissociation to release free βγ subunits in intact cells. %U https://molpharm.aspetjournals.org/content/molpharm/68/1/129.full.pdf