Bcl-x S , a proapoptotic member of the Bcl-2 protein family, is localized in the mitochondria and induces apoptosis in a caspase-and BH3-dependent manner by a mechanism involving cytochrome c release. The way in which Bcl-x S induces caspase activation and cytochrome c release, as well as the relationship between Bcl-x S and other proapoptotic members of the Bcl-2 family, is not known. Here we used embryonic fibroblasts derived from mice deficient in the multidomain proapoptotic members of the Bcl-2 family (Bax and Bak) and the apoptotic components of the apoptosome (Apaf-1 and caspase-9) to unravel the cascade of events by which Bcl-x S promotes apoptosis. Our results show that Bak but not Bax is essential for Bcl-x S -induced apoptosis. Bcl-x S induced activation of Bak, which in turn promoted apoptosis by apoptosome-dependent and -independent pathways. These findings provide the first evidence that a proapoptotic Bcl-2 family protein induces apoptosis exclusively via Bak.