BNIP3 heterodimerizes with Bcl-2/Bcl-X-L and induces cell death independent of a Bcl-2 homology 3 (BH3) domain at both mitochondrial and nonmitochondrial sites

BNIP3 (formerly NIP3) is a pro-apoptotic, mitochondrial protein classified in the Bcl-2 family based on limited sequence homology to the Bcl-2 homolog y 3 (BH3) domain and COOH-terminal transmembrane (TM) domain. BNIP3 express ed in yeast and mammalian cells interacts with survival promoting proteins Bcl-2, Bcl-X-L, and CED-9. Typically, the BH3 domain of pro-apoptotic Bcl-2 homologues mediates Bcl-2/Bcl-X-L heterodimerization and confers pro-apopt otic activity. Deletion mapping of BNIP3 excluded its BH3-like domain and i dentified the NH, terminus (residues 1-49) and TM domain as critical for Bc l-2 heterodimerization, and either region was sufficient for Bcl-X-L intera ction. Additionally, the removal of the BH3-like domain in BNIP3 did not di minish its killing activity. The TM domain of BNIP3 is critical for homodim erization, pro-apoptotic function, and mitochondrial targeting. Several TM domain mutants were found to disrupt SDS-resistant BNIP3 homodimerization b ut did not interfere with its killing activity or mitochondrial localizatio n. Substitution of the BNIP3 TM domain with that of cytochrome b(5) directe d protein expression to nonmitochondrial sites and still promoted apoptosis and heterodimerization with Bcl-2 and Bcl-X-L. We propose that BNIP3 repre sents a subfamily of Bcl-2-related proteins that functions without a typica l BH3 domain to regulate apoptosis from both mitochondrial and nonmitochond rial sites by selective Bcl-2/Bcl-X-L interactions.