The src protein contains multiple domains for specific attachment to membranes.

Abstract

The proteins encoded by the oncogene v-src and its cellular counterpart c-src (designated generically here as ppWsrc) are tightly associated with both plasma membranes and intracellular membranes. This association is due in part to the amino-terminal myristylation of pp6OsrC, but several lines of evidence suggest that amino-terminal portions of the protein itself are also involved. We now report that pp60src contains at least three domains which, in conjunction with myristylation, are capable of mediating attachment to membranes and determining subcellular localization. We identified these domains by fusing various portions of pp6fsrc to pyruvate kinase, which is normally a cytoplasmic protein. Amino acids 1 to 14 of pp6Osc are sufficient to mediate both myristylation and the attachment of pyruvate kinase to cytoplasmic granules. In contrast, amino acids 38 to 111 mediate association with the plasma membrane and perinuclear membranes, whereas amino acids 204 to 259 mediate association primarily with perinuclear membranes. We conclude that pp60Osc contains independent domains that target the protein to distinctive subcellular locations and thus may facilitate diverse biological functions of the protein.The products of the retroviral oncogene v-src and its cellular counterpart c-src (designated generically here as pp6Osrc) are protein tyrosine kinases that are associated with membranes. Association of pp60 src with membranes is essential for transformation by v-src (19). In various cell types, the viral and cellular src proteins are associated with plasma membranes (7,8,21,44), with perinuclear membranes (36), with secretory organelles in both chromaffin cells and platelets (30, 33), and with growth cones in developing neurons (27,41). Little is known about how pp6Osrc is specifically targeted to these subcellular locations.The amino terminus of pp6Osrc is covalently coupled to a 14-carbon fatty acid, myristic acid (4, 39). The myristyl moiety promotes the association of pp6Osrc with membranes (2, 11). On the other hand, not all myristylated src proteins are associated with membranes (3, 11), and some nonmyristylated src proteins are membrane associated, albeit weakly (12,20). These results suggest that pp6Osrc contains amino acid sequences which, in conjunction with amino-terminal myristylation, mediate the association of the protein with membranes.Previous work has implicated the amino-terminal 10 kilodaltons (kDa) of pp6Osrc in attachment to membranes (22,35

Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations

“…Myristylated proteins can be separated into two categories, those that are soluble and those that associate with membranes (Towler et al, 1988). In the case of the transforming protein of Rous sarcoma virus (pp60""c), the myristyl group is clearly necessary, although perhaps not sufficient, to convey membrane-associating properties (Kaplan et al, 1990). In the case of the poliovirus capsid protein, VP4, the myristyl group appears to provide a hydrophobic anchor between subunits on the coat surface and plays a structural role in capsid assembly (Chow et al, 1987).…”

Immunolocalization of the cellular src protein in interphase and mitotic NIH c-src overexpresser cells.

“…1 a); (c) no staining was observed when irrelevant mAbs are substituted for mAb 327 (not shown). It is equally unlikely that the pp60 c'~ location patterns obtained with the mAb 327 are peculiar to the particular cell line used here since Kaplan et al (1990;Kaplan, K. B., H~ E. Varmus, and D. O. Morgan, personal communication) have described similar pp60 ..... distributions in COS7 cells and in RAT-1 fibroblasts overexpressing the chicken pp60 c-'~. The possibility also must be considered that the localization of an overexpressed chicken pp60 .... in mouse cells does not reflect the true location of the endogeneous mouse pp60 .... .…”

“…10). pp60 .... has been shown recently (Kaplan et al, 1990) to contain independent amino-terminal domains that are specific for attachment to different cellular targets. The viral and cellular src proteins differ in their amino acid sequence, including regions in their amino-terminal domain.…”

Selected glimpses into the activation and function of Src kinase

“…However, myristoylation is not su cient for membrane anchorage (Kaplan et al, 1990b). Instead, other residues of Src in conjunction with myristoylation are required for mediating membrane association (Kaplan et al, 1990b).…”