Increased palmitoylation of the G_s protein α subunit after activation by the β-adrenergic receptor or cholera toxin

The α subunit of the heterotrimeric G_s protein that couples the β-adrenergic receptor to adenylyl cyclase undergoes post-translational palmitoylation. We examined the dynamics of this modification of α_s by metabolic labeling of COS and S49 lymphoma cells under different conditions. The endogenous α_s proteins were immunoprecipitated with a peptide-specific antibody, separated by SDS-polyacrylamide gel electrophoresis, and analyzed by fluorography and densitometry. A pulse-chase study of COS cells incubated with [³H]palmitate or [³⁵S]methionine showed that for α_s the palmitate turnover (t_1/2 ≈ 50 min) was significantly faster than the protein degradation. Treatment of cells with 10 μM isoproterenol, a β-adrenergic receptor agonist, in the presence of [³H]palmitate led to a rapid 4-10-fold increase in the palmitoylation of α_s. This increase in palmitoylation was concentration-dependent (EC₅₀ = 0.9 μM) and blocked by the antagonist propranolol. The mutant α_s proteins in the unc and H21a S49 cell lines did not show an increase in [³H]palmitate incorporation with isoproterenol treatment. Cholera toxin treatment of COS cells increased the [³H]palmitate incorporation into the α_s subunits. These data indicate that palmitoylation of the α_s subunit is dynamic and regulated by activation of the α_s subunit.