Diva, a Bcl-2 Homologue that Binds Directly to Apaf-1 and Induces BH3-independent Cell Death

We have identified and characterized Diva, which is a novel regulator of apoptosis. Sequence analysis revealed that Diva is a member of the Bcl-2 family of proteins containing Bcl-2 homology domain 1, 2, 3, and 4 (BH1, BH2, BH3, and BH4) regions and a carboxyl-terminal hydrophobic domain. The expression of Diva mRNA was detected in multiple embryonic tissues but was restricted to the ovary and testis in adult mice. The expression of Diva promoted the death of 293T, Ramsey, and T47D cells as well as that of primary sensory neurons, indicating that Diva is a proapoptotic protein. Significantly, Diva lacks critical residues in the conserved BH3 region that mediate the interaction between BH3-containing proapoptotic Bcl-2 homologues and their prosurvival binding partners. Consistent with this, Diva did not bind to cellular Bcl-2 family members including Bcl-2, Bcl-XL, Bcl-w, Mcl-1, and A1/Bfl-1. Furthermore, mutants of Diva lacking the BH3 region fully retained their proapoptotic activity, confirming that Diva promotes apoptosis in a BH3-independent manner. Significantly, Diva interacted with a viral Bcl-2 homologue (vBcl-2) encoded by the Kaposi's sarcoma-associated herpesvirus. Consistent with these associations, apoptosis induced by Diva was inhibited by vBcl-2 but not by Bcl-XL. Importantly, Diva interacted with Apaf-1, an adapter molecule that activates caspase-9, a central death protease of the apoptotic pathway. The expression of Diva inhibited the binding of Bcl-XL to Apaf-1, as determined by immunoprecipitation assays. Thus, Diva represents a novel type of proapoptotic Bcl-2 homologue that promotes apoptosis independently of the BH3 region through direct binding to Apaf-1, thus preventing Bcl-XL from binding to the caspase-9 regulator Apaf-1.