INDUCTION OF THE INCREASED FYN KINASE-ACTIVITY IN ANERGIC T-HELPER TYPE-1 CLONES REQUIRES CALCIUM AND PROTEIN-SYNTHESIS AND IS SENSITIVE TO CYCLOSPORINE-A

Several alterations in T cell receptor-associated signal transduction have been observed following induction of anergy of T helper type 1 (Th1) clones, including a modified intracellular free calcium ([Ca2+](i)) response and increased kinase activity associated with the protein tyrosine kinase p59(fyn). In the current study, we demonstrate that, although the kinetics of acquisition of both of these signaling alterations correlated with the generation of anergy, a normal calcium response returned within 48 h after removal from the anergizing stimulus, whereas the increased p59(fyn) activity persisted and the cells remained hyporesponsive. Generation of both the anergic state and the increased p59(fyn) activity was prevented in the presence of calcium-free medium, cycloheximide (CHX), or cyclosporin A (CsA), and could be mimicked by the calcium ionophore ionomycin. In contrast, the altered calcium response was inhibited by stimulation in the presence of calcium-free medium or CsA, but not CHX. Thus, surprisingly, these data suggest that a chronic elevation of [Ca2+](i) is proximal to and necessary for the increase in p59(fyn)-associated kinase activity observed in anergic Th1 clones. Increased p59(fyn) activity, but not the altered calcium response, correlates with maintenance of the anergic state.