Lipid-dependent targeting of G proteins into rafts

Domains rich in sphingolipids and cholesterol, or rafts, may organize signa l transduction complexes at the plasma membrane. Raft lipids are believed t o exist in a state similar to the liquid-ordered phase. It has been propose d that proteins with a high affinity for an ordered lipid environment will preferentially partition into rafts (Melkonian, K. A., Ostermeyer, A. G., C hen, J. Z., Roth, M. G., and Brown, D. A. (1999) J. Biol. Chem. 274, 3910-3 917), We investigated the possibility that lipid-lipid interactions between lipid-modified proteins and raft lipids mediate targeting of proteins to t hese domains. G protein monomers or trimers were reconstituted in liposomes , engineered to mimic raft domains. Assay for partitioning of G proteins in to rafts was based on Triton X-100 insolubility. Myristoylation and palmito ylation of G alpha(i) were necessary and sufficient for association with li posomes and partitioning into rafts. Strikingly, the amount of fatty-acylat ed G alpha(i) in rafts was significantly reduced when myristoylated G alpha (i) was thioacylated with cis-unsaturated fatty acids instead of saturated fatty acids such as palmitate. Prenylated beta gamma subunits were excluded from rafts, whether reconstituted alone or with fatty-acylated cy subunits , These results suggest that the structural difference between lipids that modify proteins is one basis for the selectivity of protein targeting to ra fts.