CD28 signal transduction: tyrosine phosphorylation and receptor association of phosphoinositide-3 kinase correlate with Ca(2+)-independent costimulatory activity

Eur J Immunol. 1994 Nov;24(11):2732-9. doi: 10.1002/eji.1830241124.

Abstract

The interaction of CD28 with its counter-receptor, B7, induces a cosignal in T cells required to prevent clonal anergy and to promote antigen-dependent interleukin-2 production. The molecular basis of the CD28 cosignal is not well understood but involves the activation of protein tyrosine kinase(s) (PTK). In this report we demonstrate that CD28 cross-linking on Jurkat T leukemic cells causes the activation of at least two PTK pathways. A CD28-induced, p56lck kinase-independent pathway causes tyrosine-phosphorylation of a 110-kDa substrate while recruitment of p56lck kinase activity is apparently required for CD28-induced tyrosine-phosphorylation of 97- and 68-kDa substrates as well as CD28-induced increases in intracellular calcium. The tyrosine phosphorylation of p110, but not p97 or p68, correlated with CD28 calcium-independent costimulatory activity. The pp110 molecule was tentatively identified as the catalytic subunit of phosphoinositide (PI)-3 kinase based upon its coimmunoprecipitation with the p85 regulatory subunit of PI-3 kinase. PI-3 kinase protein and catalytic activity were found complexed with the CD28 receptor if the receptor was "activated" by cross-linking on the surface of intact cells prior to detergent solubilization. The kinetics of association of PI-3 kinase with the "activated" CD28 receptor was rapid, occurring within 30 s of receptor cross-linking and was stable for at least 30 min. Analysis of the CD28 cytoplasmic peptide sequence revealed a putative PI-3 kinase src homology 2 binding motif and CD28 tyrosine phosphorylation site, DYMNM. Tyrosine phosphorylation of CD28 was detected in pervanadate-treated Jurkat B2.7 cells, but not untreated cells. Pervanadate-induced tyrosine phosphorylation of CD28 correlated with receptor association of PI-3 kinase in the absence of CD28 cross-linking, suggesting that CD28 association with PI-3 kinase uses a tyrosine phosphorylation-dependent mechanism. These data provide a model for CD28 signal transduction and support a role for PI-3 kinase in mediating the CD28 calcium-independent, cyclosporin A-insensitive costimulatory signal.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • CD28 Antigens / physiology*
  • Calcium / physiology*
  • Humans
  • Lymphocyte Specific Protein Tyrosine Kinase p56(lck)
  • Molecular Sequence Data
  • Phosphatidylinositol 3-Kinases
  • Phosphorylation
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Protein-Tyrosine Kinases / physiology
  • Signal Transduction*
  • Tetradecanoylphorbol Acetate / pharmacology
  • Tumor Cells, Cultured
  • Tyrosine / metabolism*

Substances

  • CD28 Antigens
  • Tyrosine
  • Phosphatidylinositol 3-Kinases
  • Phosphotransferases (Alcohol Group Acceptor)
  • Protein-Tyrosine Kinases
  • Lymphocyte Specific Protein Tyrosine Kinase p56(lck)
  • Tetradecanoylphorbol Acetate
  • Calcium