GENETIC-EVIDENCE FOR DIFFERENTIAL COUPLING OF SYK FAMILY KINASES TO THE T-CELL RECEPTOR - RECONSTITUTION STUDIES IN A ZAP-70-DEFICIENT JURKAT T-CELL LINE

T-cell antigen receptor (TCR) engagement activates multiple protein ty rosine kinases (PTKs), including the Src family member, Lck, and the S yk-related PTK, ZAP-70. Studies in ZAP-70-deficient humans have demons trated that ZAP-70 plays crucial roles in T-cell activation and develo pment, However, progress toward a detailed understanding of the regula tion and function of ZAP-70 during TCR signaling has been hampered by the lack of a suitable T-cell model for biochemical and genetic analys es, In this report, we describe the isolation and phenotypic character ization of a Syk- and ZAP-70-negative somatic mutant derived from the Jurkat T-cell line. The P116 cell line displays severe defects in TCR- induced signaling functions, including protein tyrosine phosphorylatio n, intracellular Ca2+ mobilization, and interleukin-2 promoter-driven transcription, These signaling defects were fully reversed by reintrod uction of catalytically active versions of either Syk or ZAP-70 into t he P116 cells, However, in contrast to ZAP-70 expression, Syk expressi on triggered a significant degree of cellular activation in the absenc e of TCR ligation, Transfection experiments with ZAP-70-Syk chimeric p roteins indicated that both the amino-terminal regulatory regions and the carboxy-terminal catalytic domains of Syk and ZAP-70 contribute to the distinctive functional properties of these PTKs. These studies un derscore the crucial role of ZAP-70 in TCR signaling and offer a power ful genetic model for further analyses of ZAP-70 regulation and functi on in T cells.