ACTIVATION OF P56(LCK) BY P72(SYK) THROUGH PHYSICAL ASSOCIATION AND N-TERMINAL TYROSINE PHOSPHORYLATION

The p56(lck) and p59(fyn) protein tyrosine kinases are important signa l transmission elements in the activation of mature T lymphocytes by l igands to the T-cell antigen receptor (TCR)/CD3 complex. The lack of e ither kinase results in deficient early signaling events, and pharmaco logical agents that block tyrosine phosphorylation prevent T cell acti vation altogether. After triggering of the TCR/CD3 complex, both kinas es are moderately activated and begin to phosphorylate cellular substr ates, but the molecular mechanisms responsible for these changes have remained unclear. We recently found that the p72(syk) protein tyrosine kinase is physically associated with the TCR/CD3 complex and is rapid ly tyrosine phosphorylated and activated by receptor triggering also i n T cells lacking p56(lck). Here we examine the regulation of p72(syk) and its interaction with p56(lck) in transfected COS-1 cells. p72(syk ) Was catalytically active and heavily phosphorylated on its putative autophosphorylation site, Tyr-518/519. Mutation of these residues to p henylalanines abolished its activity in vitro and toward cellular subs trates in vivo and reduced its tyrosine phosphorylation in intact cell s by approximate to 90%. Coexpression of lck did not alter the catalyt ic activity of p72(syk), but the expressed p56(lck) was much more acti ve in the presence of p72(lck) than when expressed alone. This activat ion was also seen as increased phosphorylation of cellular proteins. C oncomitantly, p56(lck) was phosphorylated at Tyr-192 in its SH2 domain , and a Phe-192 mutant p56(lck) was no longer phosphorylated by p72(sy k). Phosphate was also detected in p56(lck) at Tyr-192 in lymphoid cel ls. These findings suggest that p56(lck) is positively regulated by th e p72(syk) kinase.