Only membrane-associated RSV src proteins have amino-terminally bound lipid

Abstract

The product of the Rous sarcoma virus (RSV) transforming gene, src, is a phosphoprotein of molecular weight (M_r) 60,000 (pp60^src)¹ that interacts with cellular membranes via an 8,000 M_r amino-terminal hydrophobic domain^2–6, possibly after post-translational modification^5,7,8. Recently, it has been demonstrated that fatty acid is tightly associated with some membrane-bound proteins^9–15, including pp60^src (ref. 16). The fatty acid attachment site is located within the membrane-embedded region of the vesicular stomatitis and Sindbis virus glycoproteins^11,13–15. To test for a similar modification of the ammo-terminal domain of pp60^src, we measured the incorporation of ³H-palmitic acid into pp60^src of cells that had been transformed by wild-type RSV or recovered avian sarcoma viruses (rASVs)^6,17, some of which have variant forms of pp60^src. We report here that membrane-associated pp60^src proteins encoded by either wild-type RSV or rAS V1441^6,17 contain bound lipid. However, pp60^src proteins with amino-terminal size variations encoded by two different rASV isolates, rASV 157 and rASV 1702¹⁷, that behave as soluble, cytoplasmic proteins⁶, contain no detectable lipid. Furthermore, we observed that lipid association is temperature sensitive in cells infected with RSV mutant tsNY68¹⁸, whose pp60^src membrane association is temperature sensitive^19–21. These data support the role of lipid binding in the membrane association of pp60^src and suggest that the amino-terminal 8,000 M_r of pp60^src contains the lipid attachment site.