Reciprocal regulation of Gs alpha by palmitate and the beta gamma subunit.
Hormonal activation of Gs, the stimulatory regulator of adenylyl cyclase, promotes dissociation of alpha s from G beta gamma, accelerates removal of covalently attached palmitate from the G alpha subunit, and triggers release of a fraction of alpha s from the plasma membrane into the cytosol. To elucidate relations among these three events, we assessed biochemical effects in vitro of attached palmitate on recombinant alpha s prepared from Sf9 cells. In comparison to the unpalmitoylated protein (obtained from cytosol of Sf9 cells, treated with a palmitoyl esterase, or expressed as a mutant protein lacking the site for palmitoylation), palmitoylated alpha s (from Sf9 membranes, 50% palmitoylated) was more hydrophobic, as indicated by partitioning into TX-114, and bound beta gamma with 5-fold higher affinity. beta gamma protected GDP-bound alpha s, but not alpha s-GTP[gamma S], from depalmitoylation by a recombinant esterase. We conclude that beta gamma binding and palmitoylation reciprocally potentiate each other in promoting membrane attachment of alpha s and that dissociation of alpha s.GTP from beta gamma is likely to mediate receptor-induced alpha s depalmitoylation and translocation of the protein to cytosol in intact cells.
Iiri
T
T
Department of Cellular and Molecular Pharmacology, University of California, San Francisco 94143, USA.
Backlund
P S
PS
Jones
T L
TL
Wedegaertner
P B
PB
Bourne
H R
HR
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010168:Palmitates / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
D010168:Palmitates; D011994:Recombinant Proteins; D019205:GTP-Binding Protein alpha Subunits, Gs