Rapid T-cell receptor-mediated tyrosine phosphorylation of p120, an Fyn/Lck Src homology 3 domain-binding protein

Kris A. Reedquist, Toru Fukazawa, Brian Druker, Govindaswamy Panchamoorthy, Steven E. Shoelson, Hamid Band

Research output: Contribution to journalArticlepeer-review

74 Scopus citations

Abstract

Tyrosine phosphorylation of cellular proteins is the earliest identifiable event following T-cell antigen receptor (TCR) stimulation and is essential for activating down-stream signaling machinery. Two Src-family protein- tyrosine kinases, the TCR-associated p59(fyn) (Fyn) and the CD4/8-associated p56(lck) (Lck), have emerged as the likely mediators of early tyrosine phosphorylation in T cells. Here, we show direct binding of a 120-kDa TCR- induced phosphotyrosyl polypeptide, p120, to glutathione S-transferase fusion proteins of the Src homology 3 (SH3) domains of Fyn, Lck, and p60(src) (Src) but not other proteins. While binding of p120 to Fyn SH2 domain was phosphotyrosine-dependent as expected, its binding to the SH3 domain was independent of tyrosine phosphorylation, as shown by lack of competition with a phosphotyrosyl competitor peptide. In contrast, an SH3-specific proline- rich peptide completely abolished p120 binding to SH3. p120 was tyrosine- phosphorylated within 10 sec following stimulation of Jurkat cells with anti- CD3 monoclonal antibody, with maximal phosphorylation at 30 sec. Importantly, p120 was found associated with Fyn and Lck proteins in unstimulated Jurkat cells and served as an in vitro substrate for these kinases. These results provide evidence for a role of the SH3 domains of Fyn and Lck in the recruitment of early tyrosine-phosphorylation substrates to the TCR- associated tyrosine kinases.

Original languageEnglish (US)
Pages (from-to)4135-4139
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number10
DOIs
StatePublished - May 10 1994

Keywords

  • Src- family kinases
  • T-cell activation
  • protein-tyrosine kinase
  • signal transduction

ASJC Scopus subject areas

  • General

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