Phosphatidylinositol 3‐kinase activity is not essential for CD28 costimulatory activity in Jurkat T cells: studies with a selective inhibitor, wortmannin

The interaction of CD28 with its counter‐receptor, B7‐1 (CD 80), on antigenpresenting cells induces a co‐signal in T cells required to promote antigendependent interleukin‐2 (IL‐2) production and to prevent clonal anergy. CD28 stimulation causes both protein‐tyrosine kinase and phosphatidylinositol3‐kinase (PI3‐K) activation, suggesting a possible role for these enzyme activities in CD28 co‐signal transduction. Here, we investigate the effect of wortmannin, a selective and irreversible PI3‐K inhibitor on CD28 co‐signaling events in the Jurkat T cell line. Wortmannin added to cell cultures partially inhibits CD28‐induced tyrosine phosphorylation of the putative p110 catalytic subunit of PI3‐K, but does not block CD28‐induced association of the p85 PI3‐K regulatory subunit with the CD28 receptor. Wortmannin inhibits in a dose‐dependent manner both total cellular PI3‐K activity and CD28‐induced receptor‐associated PI3‐K activity. Wortmannin (1 μM) inhibits cellular PI3‐K activity by 90% with complete inhibition achieved at 10 μM. The inhibitory effect of wortmannin on cellular PI3‐K activity is prolonged (> 18 h), suggesting that the drug is not readily metabolized by Jurkat T cells. Wortmannin, at concentrations that blocked PI3‐K activity, fails to inhibit the synergistic effect of CD28 on IL‐2 secretion in the presence of phorbol 12‐myristate 13‐acetate and ionomycin. These data demonstrate that CD28‐induced signaling events other than the activation of PI3‐K catalytic activity contribute to the control of IL‐2 secretion.