THE SRC HOMOLOGY 2 DOMAIN OF THE PROTEIN-TYROSINE KINASE P56(LCK) MEDIATES BOTH INTERMOLECULAR AND INTRAMOLECULAR INTERACTIONS

A key event in signaling by many cell surface receptors is the activat ion of Src-like protein-tyrosine kinases and the assembly of protein c omplexes at the plasma membrane mediated by Src homology 2 and 3 (SH2 and SH3) domains. p56lck is a Src-related protein-tyrosine kinase whic h has SH2 and SH3 domains and is involved in T-cell signaling and onco genic transformation. Here we demonstrate that purified recombinant SH 2 and HSH3/SH2 domains of p56lck can mediate intermolecular interactio ns with a number of tyrosine-phosphorylated proteins present in lysate s of NIH 3T3 cells transformed by a constitutively activated form of p 56lck (p56lckF505). Two of the interacting tyrosine-phosphorylated pro teins were identified as the p85 subunit of phosphatidylinositol 3-kin ase and the GTPase-activating protein of p21ras Using a synthetic phos phopeptide corresponding to the tyrosine-phosphorylated carboxyl termi nus of p56lck (amino acids 494-509), purified recombinant Lck SH2 doma in, and differentially phosphorylated forms of p56lck we provide evide nce that the SH2 domain of p56lck can also mediate intramolecular inte ractions with the phosphorylated carboxyl terminus. Together these res ults suggest that the SH2 domain of p56lck has a dual function: (i) it can mediate intermolecular interactions with cellular proteins phosph orylated on tyrosine and thus might be involved in building up signali ng complexes at the plasma membrane and (ii) it can bind to the tyrosi ne-phosphorylated carboxyl terminus of p56lck in an intramolecular fas hion and thereby might be involved in the regulation of its intrinsic protein-tyrosine kinase activity. Phosphorylation/dephosphorylation of the regulatory tyrosine residue 505 might serve as a switch between t hese two functions.