title: Both CD28 ligands CD80 (B7-1) and CD86 (B7-2) activate phosphatidylinositol 3-kinase, and wortmannin reveals heterogeneity in the regulation of T cell IL-2 secretion.
abstract: In this report, the co-stimulatory signals provided by CD80 (B7-1) or CD86 (B7-2) were compared to CD28 ligation by mAb. We demonstrate that while both anti-CD3 and anti-CD28 antibodies induced activation of phosphoinositide (PI) 3-kinase, the kinetics of activation differed. Anti-CD28 produced a sustained activation of PI 3-kinase while anti-CD3 induced activation was transient. Both B7-1 and B7-2 could induce prolonged activation of PI 3-kinase. The co-stimulatory effects of B7-1 and B7-2 were dependent on CD28 cross-linking, based on complete inhibition of PI 3-kinase activation by CD28 antibody Fab fragments. While Jurkat T cells co-stimulated with anti-CD3 and B7-1 or B7-2 secreted high levels of IL-2, there were distinct effects of anti-CD28 mAb and B7-1 or B7-2 on IL-2 secretion in conjunction with protein kinase C activation. To assess functional effects of CD28 ligation, pharmacologic inhibitors of PI 3-kinase were evaluated. In Jurkat cells, efficient inhibition of PI 3-kinase activation after B7-2 stimulation was achieved using wortmannin; however, we observed a surprising increase in IL-2 secretion after B7 or anti-CD28 stimulation. The effect of wortmannin was concentration dependent. Moreover, the effect was specific for receptor-mediated activation as wortmannin did not enhance phorbol ester plus ionomycin-induced IL-2 secretion. Another inhibitor of PI 3-kinase, LY294002, also resulted in augmentation of anti-CD28-induced IL-2 secretion by Jurkat cells. The effects of wortmannin on IL-2 secretion were also examined in primary T cells. In marked contrast, wortmannin resulted in a potent inhibition of anti-CD3 plus B7-1 or anti-CD28-induced IL-2 secretion while phorbol ester plus ionomycin-induced IL-2 secretion was wortmannin resistant. Together these observations demonstrate that signal transduction by both B7-1 and B7-2 involves PI 3-kinase, and that PI 3-kinase or other wortmannin-sensitive targets are important for IL-2 secretion. Finally, treatment of Jurkat cells with PI 3-kinase inhibitors alone was sufficient to induce low levels of IL-2 secretion. This is consistent with the notion that a wortmannin-sensitive target such as PI 3-kinase may down-regulate IL-2 secretion in Jurkat cells.
authors.affiliation: Immune Cell Biology Program, Naval Medical Research Institute, Bethesda, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Ueda
authors.forename: B L
authors.initials: BL
authors.lastname: Levine
authors.forename: M L
authors.initials: ML
authors.lastname: Huang
authors.forename: G J
authors.initials: GJ
authors.lastname: Freeman
authors.forename: L M
authors.initials: LM
authors.lastname: Nadler
authors.forename: C H
authors.initials: CH
authors.lastname: June
authors.forename: S G
authors.initials: SG
authors.lastname: Ward
mesh_terms: D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015703:Antigens, CD / Q000494:pharmacology*; D018122:B7-1 Antigen / Q000494:pharmacology*; D051940:B7-2 Antigen; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D016466:CHO Cells; D006224:Cricetinae; D004790:Enzyme Induction / Q000187:drug effects / Q000276:immunology; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates / Q000096:biosynthesis; D010716:Phosphatidylinositols / Q000096:biosynthesis; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D015398:Signal Transduction / Q000276:immunology; D013601:T-Lymphocytes / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018122:B7-1 Antigen; D051940:B7-2 Antigen; D018106:CD28 Antigens; D017252:CD3 Complex; C497964:CD86 protein, human; D007376:Interleukin-2; D008562:Membrane Glycoproteins; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D000077191:Wortmannin