DUAL MYRISTYLATION AND PALMITYLATION OF SRC FAMILY MEMBER P59(FYN) AFFECTS SUBCELLULAR-LOCALIZATION

The Src family consists of nine related tyrosine protein kinases with a common domain structure, including a myristylated N-terminal glycine residue. In this report, we identify cysteine residues within the N-t erminal region of the Src family member Fyn which serve as sites for p almitylation. To facilitate detection of protein fatty acylation, p59( fyn) was overexpressed in COS cells and incubated with radioiodinated fatty acid analogs of myristate (IC13) or palmitate (IC16), Incorporat ion of both fatty acids into p59(fyn) was readily observed. Acylation with the palmitate analog was prevented when Gly-2 was mutated to alan ine, implying that N-myristylation is required for palmitylation, and when either Cys-3 or Cys-6 was mutated to serine. Palmitylation was sh own to alter the distribution of p59(fyn) between mem brane-bound and soluble fractions. In contrast, no incorporation of the palmitate anal og into pp60(v-src), which lacks N-terminal cysteine residues, was obs erved. Mutation of Ser-3 of Src to cysteine, but not Ser-6, resulted i n incorporation of the palmitate analog. These results serve to deline ate sequence elements important for dual acylation of proteins, and fu rther illustrate the utility of radioiodinated fatty acid analogs for studies of protein fatty acid acylation.