FIG. 5.

Rac1 is essential to the CD5 costimulatory signal pathway leading to an upregulation of the IL-2 promoter activity, while Rho and Cdc42 play no role in this pathway. The CD5-induced signaling pathway in T lymphocytes expressing constitutively active Rac1 is insensitive to wortmannin but still sensitive to KN-62 or the expression of a dominant negative CaM kinase IV mutant. T cells, prestimulated as described in Materials and Methods, were transfected with 15 μg of pCAT3e-IL-2(−319/+47) together with 15 μg of either an empty control expression plasmid (control) (A) or the expression plasmid(s) for dominant negative or constitutively active mutants (B to H): dominant negative Rac1 (Rac1 · N17) (B), dominant negative Rho (Rho · N19) (C), dominant negative Cdc42 (Cdc42 · N17) (D), constitutively active Rac1 (Rac1 · V12) (E to G), and constitutively active Rac1 (Rac1 · V12) in combination with dominant negative CaM kinase IV (H). Transfected cells were left alone for 1 h, divided into three groups, and subsequently left unstimulated (□) or stimulated with PHA plus anti-CD28 (▪) or PHA plus anti-CD28 and anti-CD5 (▤) for 24 h in the presence of 100 nM wortmannin (F) or 10 μM KN-62 (G). CAT expression was measured as described in Materials and Methods. The results are expressed as the relative CAT expression compared to the PHA- plus anti-CD28-induced CAT expression in the transfected control T cells, which was set at 1. The mean values ± SEMs found for the relative CAT expression in three to six independent experiments are shown.