PHYSICAL AND FUNCTIONAL INTERACTIONS OF PROTEIN-TYROSINE KINASES, P59(FYN) AND ZAP-70, IN T-CELL SIGNALING

The src family protein tyrosine kinases participate in signaling throu gh cell surface receptors that lack intrinsic tyrosine kinase domains, One of the src family kinases, p59(fyn)(Fyn), plays an important role in the TCR-mediated signaling, Here we report that Fyn becomes associ ated with the zeta-associated tyrosine kinase, ZAP-70, in a T cell hyb ridoma upon stimulation, The association was transient; it occurred as early as 10 s after stimulation and disappeared after 10 min. The two proteins were also associated with each other when coexpressed in COS cells, Coexpression of the zeta-chain was not required for their inte raction, Mutational analysis of Fyn and ZAP-70 revealed that their kin ase activities were relevant to the association, Deletion of both the SH2 and SH3 domains of Fyn resulted in the decrease of the association with ZAP-70, Consistently, Fyn-SH2 and Fyn-SH3 fused to glutathione S -transferase were able to bind to ZAP-70, These data suggest that mult iple sites of Fyn and ZAP-70 are involved in the association, Furtherm ore, coexpression of the wild-type of both kinases in COS cells enhanc ed tyrosine phosphorylation of the helix-turn-helix-containing protein , HS1, HS1 was also tyrosine phosphorylated upon TCR stimulation, Thus , we propose that Fyn phosphorylates and activates ZAP-70 and that bot h kinases cooperate in TCR signaling.