Phosphorylation of Tyr319 in ZAP-70 is required for T-cell antigen receptor-dependent phospholipase C-γ1 and Ras activation

Brandi L. Williams, Brenda J. Irvin, Shari L. Sutor, Claudia C.S. Chini, Elaine Yacyshyn, Juliane Bubeck Wardenburg, Mark Dalton, Andrew C. Chan, Robert T. Abraham

Research output: Contribution to journalArticlepeer-review

158 Scopus citations

Abstract

Accumulating evidence indicates that the interdomain B regions of ZAP-70 and Syk play pivotal roles in the coupling of T-cell antigen receptor (TCR) stimulation to the activation of downstream signaling pathways. The interdomain B region of ZAP-70 contains at least three candidate sites of tyrosine phosphorylation. In this report, we identify Tyr319 as a functionally important phosphorylation site in the ZAP-70 interdomain B region. TCR crosslinkage triggered a rapid increase in the phosphorylation of Tyr319 in Jurkat T cells. Although mutation of Tyr319 to Phe had no effect on the tyrosine kinase activity of ZAP-70, the resulting ZAP(Y319→F) mutant failed to reconstitute TCR-dependent Ca2+ mobilization, Ras activation, CD69 expression and NFAT-dependent transcription in ZAP-70-deficient Jurkat cells. These defects were correlated with reduced tyrosine phosphorylation of phospholipase C (PLC)-γ1 and the LAT adapter protein in the ZAP(Y319→F)-expressing cells. On the other hand, ZAP(Y319→F)-expressing cells displayed normal increases in SLP-76 phosphorylation and ERK activation during TCR stimulation. Phosphorylation of Tyr319 promoted the association of ZAP-70 with the SH2 domains of two key signaling molecules, Lck and PLC-γ1. These studies suggest that Tyr319 phosphorylation is required for the assembly of a ZAP-70-containing signaling complex that leads to the activation of the PLC-γ1- and Ras-dependent signaling cascades in antigen-stimulated T cells.

Original languageEnglish
Pages (from-to)1832-1844
Number of pages13
JournalEMBO Journal
Volume18
Issue number7
StatePublished - Apr 1 1999

Keywords

  • NFAT
  • Signal transduction
  • T-cell antigen receptor
  • ZAP-70

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