G(i)-dependent activation of c-Jun N-terminal kinase in human embryonal kidney 293 cells

Heterotrimeric G proteins stimulate the activities of two stress-activated protein kinases, c-dun N-terminal kinase (JNK) and p38 mitogen-activated pr otein kinase in mammalian cells. In this study, we examined whether alpha s ubunits of G(i) family activate JNK using transient expression system in hu man embryonal kidney 293 cells. Constitutively activated mutants of G alpha (il), G alpha(i2), and G alpha(i3) increased JNK activity. In contrast, con stitutively activated C alpha(o) and G alpha(z) mutants did not stimulate J NK activity. To examine the mechanism of JNK activation by G alpha(i), kina se-deficient mutants of mitogen-activated protein kinase kinase 4 (MKK4) an d 7 (MKK7), which are, known to be JNK activators, were transfected into th e cells. However, G alpha(i)-induced JNK activation was not blocked effecti vely by kinase-deficient MKK4 and MKK7. In addition, activated G alpha(i) m utant failed to stimulate MKK4 and MKK7 activities. Furthermore, JNK activa tion by Gai was inhibited by dominant-negative Rho and Cdc42 and tyrosine k inase inhibitors, but not dominant-negative Rac and phosphatidylinositol 3- kinase inhibitors. These results indicate that G alpha(i) regulates JNK act ivity dependent on small GTPases Rho and Cdc42 and on tyrosine kinase but n ot on MKK4 and MKK7.