Novel role of phosphatidylinositol 3-kinase in CD28-mediated costimulation

Ligation of the CD28 surface receptor provides a major costimulatory signal for full scale T cell activation. Despite extensive studies, the intracell ular signaling pathways delivered by CD28 ligation are not fully understood . A particularly controversial matter is the role of phosphatidylinositol 3 -kinase (PI3K) in CD28-mediated costimulation. It is known that the binding site for PI3K and Grb-2 lies nested within the YMNM motif of the CD28 cyto plasmic domain. To elucidate the role of PI3K during CD28-mediated interleu kin-2 (IL-2) production, CD28 YMNM point and deletion mutants were expresse d in Jurkat cells. We then measured IL-2 promoter activation after CD28 lig ation. Our results showed that the Y189F mutant, which disrupts binding by PI3K, and the YMNM deletion mutant both demonstrated reduced but significan t activity for IL-2 promoter activation. In contrast, the N191A mutant, whi ch retains PI3K binding ability, resulted in a complete abrogation of activ ity, suggesting that PI3K mediates a negative effect upon transcriptional a ctivation of the IL-2 gene. Consistent with this idea, we found that the ad dition of a PI3K pharmacological inhibitor augmented IL-2 promoter activity , whereas coexpression of a constitutively active form of PI3K reduced this activity. Taken together, these data indicate that PI3K, when associated w ith the YMNM motif, may act as a negative mediator in CD28-mediated IL-2 ge ne transcription.