Differential roles of Akt, Rac, and Ral in R-Ras-mediated cellular transformation, adhesion, and survival

Multiple biological functions have been ascribed to the Ras-related G prote in R-Ras. These include the ability to transform NIH 3T3 fibroblasts, the p romotion of cell adhesion, and the regulation of apoptotic responses in hem atopoietic cells. To investigate the signaling mechanisms responsible for t hese biological phenotypes, we compared three R-Ras effector loop mutants ( S61, G63, and C66) for their relative biological and biochemical properties . While the S61 mutant retained the ability to cause transformation, both t he G63 and the C66 mutants were defective in this biological activity. On t he other hand, while both the S61 and the C66 mutants failed to promote cel l adhesion and survival in 32D cells, the G63 mutant retained the ability t o induce these biological activities. Thus, the ability of R-Ras to transfo rm cells could be dissociated from its propensity to promote cell adhesion and survival. Although the transformation-competent S61 mutant bound prefer entially to c-Raf, it only weakly stimulated the mitogen-activated protein kinase (MAPK) activity, and a dominant negative mutant of MEK did not signi ficantly perturb R-Ras oncogenicity. Instead, a dominant negative mutant of phosphatidylinositol 3-kinase (PI3-K) drastically inhibited the oncogenic potential of R-Ras. Interestingly, the ability of the G63 mutant to induce cell adhesion and survival was closely associated with the PI3-K-dependent signaling cascades. To further delineate R-Ras downstream signaling events, we observed that while a dominant negative mutant of Akt/protein kinase in hibited the ability of R-Ras to promote cell survival, both dominant negati ve mutants of Rac and Ral suppressed cell adhesion stimulated by R-Ras. Thu s, the biological actions of R-Ras are mediated by multiple effecters, with PI3-K-dependent signaling cascades being critical to its functions.