Development of unique antibodies directed against each of the six different phosphotyrosine residues within the T cell receptor CD3ζ chain

Sigal Gelkop, Batya Weisman, Ranjan Nath Pulak, Dorit Zharhary, Noah Isakov

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

Signal transduction from the T cell antigen receptor (TCR)/CD3 complex involves six different immunoreceptor tyrosine-based activation motifs (ITAM) located within the cytoplasmic tails of the CD3 chains. Each ITAM possesses two conserved tyrosine residues that can undergo phosphorylation upon TCR/CD3 crosslinking and become a docking site for SH2-containing effector molecules. Specificity of the SH2 domains is determined by their ability to bind a phosphorylated tyrosine in the context of a longer peptide motif within the target protein. As a result, phosphorylation of different tyrosines within the CD3 cytoplasmic tails creates docking sites for distinct SH2-containing signaling proteins that differentially impact on the quality of the T cell response. In the present study, we prepared antibodies specific for each of the six different phosphotyrosines of the mouse CD3ζ chain. The antibodies were characterized with respect to their cross-reactivity, ability to recognize the phosphorylated versus non-phosphorylated forms of tyrosine-containing motifs, and cross-reactivity with the homologous phospho-motifs on the human CD3ζ protein. The antibodies were found to be specific and selective for phospho-CD3ζ. They can serve as useful tools for distinguishing between the six potential tyrosine phosphorylation sites on the CD3ζ chain, and for correlating the phosphorylation of specific CD3ζ tyrosine residues with activation of signaling pathways that dictate T cell differentiation into responding, anergic, or apoptotic cells.

Original languageEnglish
Pages (from-to)129-137
Number of pages9
JournalJournal of Immunological Methods
Volume375
Issue number1-2
DOIs
StatePublished - 31 Jan 2012

Keywords

  • CD3ζ chain
  • ITAM
  • Immunoreceptor
  • Phosphopeptides
  • Tyrosine phosphorylation

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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