Cooperative transformation of NIH3T3 cells by G alpha 12 and Rac1

The heterotrimeric G-protein, G alpha 12, together with the closely-related G alpha 13, are members of the G12 class of alpha-subunits important in mediating the signaling from seven transmembrane domain-spanning receptors, Recent evidence implicating both G alpha 12 and G alpha 13 in the activation of signaling pathways involving members of the RHO gene family led us to examine the role of Rad, RhoA and Cdc42Hs in the transforming properties of G alpha 12. Asparagine 17 (Asn 17) dominant inhibitory mutants of Rad, and to a lesser extent RhoA, block focus forming ability of the GTPase-deficient mutant of G alpha 12 (G alpha 12 Leu 229) in NIH3T3 cells, In turn, wild-type G alpha 12 cooperates well with Rad Val 12 but not with RhoA Leu 63 mutant in transforming NIH3T3 cells, Interestingly, the morphology of foci induced by G alpha 12 and RhoA mutants are strikingly similar and is distinct from those displayed by Rad Val 12 mutant, The fact that G alpha 12's ability to induce mitogenesis in NIH3T3 cells is not significantly perturbed by C3 ribosyltransferase suggested that RhoA does not play a major role in G alpha 12-induced mitogenic events, Activated mutant of Rad has previously been demonstrated to stimulate the activity of the stress-induced c-Jun IV-terminal kinase/stress-activated protein kinases (JNK/SAPKs), Transient co-transfection of Rad Val 12 mutant with the wild-type G alpha 12 in COS7 cells leads to the further activation of an exogenously expressed hemagglutinin(HA)-tagged JNK. Furthermore, the cooperation between G alpha 12 and Rad in cellular transformation is correlated with their ability to stimulate transcription from c-fos serum response element (SRE).