PHOSPHORYLATION OF SLP-76 BY THE ZAP-70 PROTEIN-TYROSINE KINASE IS REQUIRED FOR T-CELL RECEPTOR FUNCTION

Two families of tyrosine kinases, the Src and Syk families, are requir ed for T-cell receptor activation. While the Src kinases are responsib le for phosphorylation of receptor-encoded signaling motifs and for up -regulation of ZAP-70 activity, the downstream substrates of ZAP-70 ar e unknown. Evidence is presented herein that the Src homology 2 (SH2) domain-containing leukocyte protein of 76 kDa (SLP-76) is a substrate of ZAP-70. Phosphorylation of SLP-76 is diminished in T cells that exp ress a catalytically inactive ZAP-70. Moreover, SLP-76 is preferential ly phosphorylated by ZAP-70 in vitro and in heterologous cellular syst ems. In T cells, overexpression of wild-type SLP-76 results in a hyper active receptor, while expression of a SLP-76 molecule that is unable to be tyrosine-phosphorylated attenuates receptor function. In additio n, the SH2 domain of SLP-76 is required for T-cell receptor function, although its role is independent of the ability of SLP-76 to undergo t yrosine phosphorylation. As SLP-76 interacts with both Grb2 and phosph olipase C-gamma 1, these data indicate that phosphorylation of SLP-76 by ZAP-70 provides an important functional link between the T-cell rec eptor and activation of ras and calcium pathways.