ACTIVATION OF RAC1, RHOA, AND MITOGEN-ACTIVATED PROTEIN-KINASES IS REQUIRED FOR RAS TRANSFORMATION

Although substantial evidence supports a critical role for the activat ion of Raf-1 and mitogen-activated protein kinases (MAPKs) in oncogeni c Ras-mediated transformation, recent evidence suggests that Ras may a ctivate a second signaling pathway which involves the Ras-related prot eins Rad and RhoA Consequently, we used three complementary approaches to determine the contribution of Rad and RhoA function to oncogenic R as-mediated transformation. First, whereas constitutively activated mu tants of Rad and RhoA showed very weak transforming activity when tran sfected alone, their coexpression with a weakly transforming Raf-1 mut ant caused a greater than 35-fold enhancement of transforming activity . Second, we observed that coexpression of dominant negative mutants o f Rad and RhoA reduced oncogenic Ras transforming activity. Third, act ivated Rad and RhoA further enhanced oncogenic Ras-triggered morpholog ic transformation, as well as growth in soft agar and cell motility. F inally, we also observed that kinase-deficient MAPKs inhibited Ras tra nsformation. Taken together, these data support the possibility that o ncogenic Ras activation of Rad and RhoA, coupled with activation of th e Raf/MAPK pathway, is required to trigger the full morphogenic and mi togenic consequences of oncogenic Ras transformation.