RAC REGULATION OF TRANSFORMATION, GENE-EXPRESSION, AND ACTIN ORGANIZATION BY MULTIPLE, PAK-INDEPENDENT PATHWAYS

yRad and RhoA are members of the Rho family of Ras-related proteins an d function as regulators of actin cytoskeletal organization, gene expr ession, and cell cycle progression, Constitutive activation of Rad and RhoA causes tumorigenic transformation of NIH 3T3 cells, and their fu nctions may be required for full Ras transformation. The effecters by which Rad and RhoA mediate these diverse activities, as well as the in terrelationship between these events, remain poorly understood, Rad is distinct from RhoA in its ability to bind and activate the p65 PAK se rine/threonine kinase, to induce lamellipodia and membrane ruffling, a nd to activate the c-Jun NH2-terminal kinase (JNK). To assess the role of PAK in Rad function, we identified effector domain mutants of Rad and Rac1-RhoA chimeric proteins that no longer bound PAK. Surprisingly , PAK binding was dispensable for Rad-induced transformation and lamel lipodium formation, as well as activation of JNK, p38, and serum respo nse factor (SRF). However, the ability of Rad to bind to and activate PAK correlated with its ability to stimulate transcription from the cy clin DI promoter, Furthermore, Rad activation of JNK or SRF, or induct ion of lamellipodia, was neither necessary nor sufficient for Rad tran sforming activity, Finally, the signaling pathways that mediate Rad ac tivation of SRF or JNK were distinct from those that mediate Rad induc tion of lamellipodia, Taken together, these observations suggest that Rad regulates at least four distinct effector-mediated functions and t hat multiple pathways may contribute to Rad-induced cellular transform ation.