The small GTP-binding protein Rac is a downstream effector of the oncogene product p21-ras. Rac is involved in actin polymerization, Jun kinase activation, and intracellular superoxide anion production, through distinct pathways in tumor cells. Here we investigated the role of activated Rac in the response of tumor cells to apoptosis triggered by anti-cancer drugs or the cell surface death receptor CD95. Using M14 melanoma cells stably transfected with a constitutively active form of Rac1, we show that activated Rac inhibits tumor cell response to apoptosis. The inhibitory effect of activated Rac on apoptotic signaling is mediated by the interaction of Rac with intracellular oxidase and the subsequent production of superoxide, which is supported by experiments performed with M14 and NIH3T3 cells transiently transfected with the loss-of-function mutants of Rac in an activated RacV12 background. Consistent with these findings, we also demonstrate that inhibition of the Rac pathway in the HaRas-expressing T24 bladder carcinoma cell line induces a decrease in superoxide anion concentration, and results in a significant increase in tumor cell sensitivity to apoptosis. These findings demonstrate the existence of a novel Rac-dependent survival pathway mediated by intracellular superoxide in tumor cells.