Increased palmitoylation of the Gs protein alpha subunit after activation by the beta-adrenergic receptor or cholera toxin.
The alpha subunit of the heterotrimeric Gs protein that couples the beta-adrenergic receptor to adenylyl cyclase undergoes post-translational palmitoylation. We examined the dynamics of this modification of alpha s by metabolic labeling of COS and S49 lymphoma cells under different conditions. The endogenous alpha 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 alpha s the palmitate turnover (t1/2 approximately 50 min) was significantly faster than the protein degradation. Treatment of cells with 10 microM isoproterenol, a beta-adrenergic receptor agonist, in the presence of [3H]palmitate led to a rapid 4-10-fold increase in the palmitoylation of alpha s. This increase in palmitoylation was concentration-dependent (EC50 approximately 0.9 microM) and blocked by the antagonist propranolol. The mutant alpha 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 alpha s subunits. These data indicate that palmitoylation of the alpha s subunit is dynamic and regulated by activation of the alpha s subunit.
Degtyarev
M Y
MY
Molecular Pathophysiology Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Spiegel
A M
AM
Jones
T L
TL
D000818:Animals; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D000882:Haplorhini; D008223:Lymphoma; D009154:Mutation; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011943:Receptors, Adrenergic, beta / Q000187:drug effects / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D010169:Palmitic Acids; D011943:Receptors, Adrenergic, beta; D019308:Palmitic Acid; D002772:Cholera Toxin; D019204:GTP-Binding Proteins