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 intracellular 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 cytoplasmic domain. To elucidate the role of PI3K during CD28-mediated interleukin-2 (IL-2) production, CD28 YMNM point and deletion mutants were expressed in Jurkat cells. We then measured IL-2 promoter activation after CD28 ligation. Our results showed that the Y189F mutant, which disrupts binding by PI3K, and the YMNM deletion mutant both demonstrated reduced but significant activity for IL-2 promoter activation. In contrast, the N191A mutant, which retains PI3K binding ability, resulted in a complete abrogation of activity, suggesting that PI3K mediates a negative effect upon transcriptional activation of the IL-2 gene. Consistent with this idea, we found that the addition 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 with the YMNM motif, may act as a negative mediator in CD28-mediated IL-2 gene transcription.
Harada
Y
Y
Research Institute for Biological Sciences, Science University of Tokyo, 2669 Yamazaki, Noda, Chiba 278-0022, Japan.
Tanabe
E
E
Watanabe
R
R
Weiss
B D
BD
Matsumoto
A
A
Ariga
H
H
Koiwai
O
O
Fukui
Y
Y
Kubo
M
M
June
C H
CH
Abe
R
R
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D018106:CD28 Antigens / Q000737:chemistry / Q000502:physiology*; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000502:physiology*; D008969:Molecular Sequence Data; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011401:Promoter Regions, Genetic; D017386:Sequence Homology, Amino Acid; D013601:T-Lymphocytes / Q000276:immunology*
D018106:CD28 Antigens; D002867:Chromones; D004791:Enzyme Inhibitors; D007376:Interleukin-2; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one