Binding of phosphatidylinositol-3-OH kinase to CD28 is required for T-cell signalling.
The engagement of CD28 with its ligand B7.1/CD80 results in potent costimulation of T-cell activation initiated through the CD3/T-cell receptor complex. The biochemical basis of CD28 costimulatory function is poorly understood. The signalling pathways used by CD28 are unlike those used by the CD3/T-cell receptor in that they are resistant to cyclosporin A and independent of changes in cyclic AMP concentrations. These differences suggest that each pathway provides unique biochemical information which is required for T-cell activation. We report here that CD28 becomes tyrosine-phosphorylated following interaction with B7.1/CD80, which induces formation of a complex with phosphatidylinositol-3-OH kinase, mediated by the SH2 domains of the p85 subunit of the kinase. Phosphatidylinositol-3-OH kinase is a heterodimer of this 85K regulatory subunit and a 110K catalytic subunit, and is a common substrate for most receptor tyrosine kinases and some cytokine receptors, binding through its SH2 domain to phosphotyrosine in the motif Tyr-X-X-Met in the CD28 sequence, which is highly conserved between human, mouse and rat and lies in the intracellular domain. We show that CD28 mutants that have their kinase-binding site deleted or the tyrosine at position 173 substituted by phenylalanine do not associate with the kinase after CD28 stimulation and cannot stimulate production of interleukin-2. Our results suggest that phosphatidylinositol-3-OH kinase is critical for signalling by CD28.
Pagès
F
F
INSERM U119, Marseille, France.
Ragueneau
M
M
Rottapel
R
R
Truneh
A
A
Nunes
J
J
Imbert
J
J
Olive
D
D
D000818:Animals; D018106:CD28 Antigens / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D006825:Hybridomas; D015151:Immunoblotting; D007739:L Cells; D051379:Mice; D009154:Mutation; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins; D015398:Signal Transduction / Q000276:immunology*; D013601:T-Lymphocytes / Q000502:physiology*; D014162:Transfection
D018106:CD28 Antigens; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D011993:Recombinant Fusion Proteins