The anti-apoptotic molecule Aven was originally identified in a yeast two-hybrid screen for Bcl-x(L)-interacting proteins and has also been found to bind Apaf-1, thereby interfering with Apaf-1 self-association during apoptosome assembly. Aven is expressed in a wide variety of adult tissues and cell lines, and there is increasing evidence that its overexpression correlates with tumorigenesis, particularly in acute leukemias. The mechanism by which the anti-apoptotic activity of Aven is regulated remains poorly understood. Here we shed light on this issue by demonstrating that proteolytic removal of an inhibitory N-terminal Aven domain is necessary to activate the anti-apoptotic potential of the molecule. Furthermore, we identify Cathepsin D (CathD) as the protease responsible for Aven cleavage. On the basis of our results, we propose a model of Aven activation by which its N-terminal inhibitory domain is removed by CathD-mediated proteolysis, thereby unleashing its cytoprotective function.