INHIBITION OF TCR CD3-MEDIATED SIGNALING BY A MUTANT OF THE HEMATOPOIETICALLY EXPRESSED G16 GTP-BINDING PROTEIN/

We have investigated the role of the hematopoietically expressed G16 G TP-binding protein on T cell activation. We constructed transfectants of Jurkat T cells that express a function-deficient mutant of G alpha 16 predicted to prevent activation of this G protein. Upon stimulation with anti-CD3 epsilon antibodies, mutant G alpha 16 transfectants dis play a profound defect in the production of IL-2 and IL-10, as well as in the expression of CD69. In contrast, the phorbol 12-myristate 13-a cetate (PMA)-induced IL-10 production and CD69 expression, and the ion omycin plus PMA-induced IL-2 production are not affected. Consistent w ith the reduction in cytokine production is the inhibition of early si gnaling events in the mutant G alpha 16-expressing cells. There are si gnificant reductions in anti-epsilon-induced tyrosine phosphorylation of zeta, epsilon, ZAP-70, and phospholipase C gamma 1, as well as in i ntracellular Ca2+; mobilization. In accordance with the effects on tyr osine phosphorylation is the reduction of TCR/CD3-mediated Fyn and Lck activities in G alpha 16 mutant cells. Even though the mechanism thro ugh which the G alpha 16 mutant mediates inhibition of T cell activati on is not known, the data suggest a model where G proteins become acti vated upon TCR/CD3 engagement and regulate the activation of tyrosine kinases and subsequent downstream signaling events that lead to the ac tivation of cytokine genes.