BOD (BCL-2-RELATED OVARIAN DEATH GENE) IS AN OVARIAN BH3 DOMAIN-CONTAINING PROAPOPTOTIC BCL-2 PROTEIN CAPABLE OF DIMERIZATION WITH DIVERSE ANTIAPOPTOTIC BCL-2 MEMBERS

Using the yeast two-hybrid protein-protein interaction system to searc h for genes capable of forming dimers with the antiapoptotic protein M cl-1, we have isolated BOD (Bcl-2-related ovarian death agonist) from an ovarian fusion cDNA library. The three variants of BOD (long, mediu m, and short) have an open reading frame of 196, 110, and 93 amino aci ds, respectively; all of them contain a consensus Bcl-2 homology 3 (BH 3) domain but lack other BH domains found in channel-forming Bcl-2 fam ily proteins. In the yeast cell assay, BOD interacts with diverse anti apoptotic Bcl-2 proteins [Mcl-1, Bcl-2, Bcl-xL, Bcl-w, Bfl-1, and Epst ein-Barr virus (EBV) BHRF-1] but not with different proapoptotic Bcl-2 proteins (BAD, Bak, Bok, and Bar). After overexpression in mammalian Chinese hamster ovary (CHO) cells, BOD induces apoptosis that can be p revented by the baculoviral caspase inhibitor P35. The cell-killing ac tivity of BOD is also antagonized in cells cotransfected with the anti apoptotic Bcl-w protein, which showed high affinity for BOD in the two -hybrid assay. Furthermore, mutagenesis studies showed that BOD mutant s with alterations in the BH3 domain lose cell-killing ability, sugges ting that the BH3 domain is important for the mediation of cell killin g by BOD. BOD mRNA is ubiquitously expressed in ovary and multiple oth er tissues. The BOD gene is also conserved in diverse mammalian specie s. Identification of BOD expands the group of proapoptotic Bcl-2 prote ins that only contains the BH3 domain and allows future elucidation of the intracellular mechanism for apoptosis regulation in ovary and oth er tissues.