Visualization of a functional Gαq-green fluorescent protein fusion in living cells -: Association with the plasma membrane is disrupted by mutational activation and by elimination of palmitoylation sites, but not by activation mediated by receptors or Alf4-*S

To investigate how G protein alpha subunit localization is regulated under basal and activated conditions, we inserted green fluorescent protein (GFP) into an internal loop of G alpha (q), ,alpha (q)-GFP stimulates phospholipase C in response to activated receptors and inhibits py dependent activation of basal G protein-gated inwardly rectifying K+ currents as effectively as alpha (q) does. Association of alpha (q)-GFP with the plasma membrane is reduced by mutational activation and eliminated by mutation of the alpha (q) pahnitoylation sites, suggesting that a, must be in the inactive, palmitoylated state to be targeted to this location. We tested the effects of activation by receptors and by Alf(4)(-) on the localization of alpha (q)-GFP in cells expressing both alpha (q)-GFP and a protein kinase C gamma -red fluorescent protein fusion that translocates to the plasma membrane in response to activation of G(q). In cells that clearly exhibit protein kinase C gamma -red fluorescent protein translocation responses, relocalization of (alpha (q)-GFP is not observed. Thus, under conditions associated with palmitate turnover and By dissociation, alpha (q)-GFP remains associated with the plasma membrane, These results suggest that upon reaching the plasma membrane alpha (q) receives an anchoring signal in addition to palmitoylation and association with By, or that during activation, one or both of these factors continues to retain alpha (q) in this location.