Regulation of mitochondrial dysfunction and apoptosis by BCL-2 family members BID, BAX, and BAK

BCL-2 family proteins control a critical event during apoptosis, the release of cytochrome c from mitochondria to cytosol responsible for the activation of effector caspases and orderly demise of the cell. BCL-2 family members are regulated by a complex series of interactions that modify their conformation, subcellular localization, and function. I have studied interactions between the BH3 domain-only family member BID and the fully conserved multiple domain, "multi-domain" pro-apoptotic members BAX and BAK, and their role in cytochrome c release and apoptosis.;Following engagement of cell surface death receptors, cytosolic p22 BID is cleaved to p15 tBID, which targets mitochondria and causes cytochrome c release.;Mechanistically BID might act as an upstream ligand to activate other pro-apoptotic molecules such as BAX, or could function as a downstream pore-forming effector. To begin to test these hypotheses an in vitro mitochondrial targeting/cytochrome c release system was established, and the effects of a series of BID mutants on wild-type mitochondria were assessed. I noted that of the "multi-domain" members BAK was prominent on the surface of the mitochondria. Comparison with Bak -/- mitochondria provided evidence for a model in which tBID functions as a ligand that causes the intramembranous oligomerization of BAK into a pore for cytochrome c efflux in a permeability transition-independent manner. Characterization of Fas-activated Bid -/- hepatocytes indicated that BID is essential for both BAK oligomerization and the translocation of monomeric BAX from cytosol to mitochondria.;Bax, Bak doubly-deficient animals and primary cells revealed that tBID activates and requires either BAX or BAK to induce cytochrome c release and apoptosis in vivo. Bax, Bak doubly-deficient, but not singly-deficient, mice show a similar resistance as Bid-deficient mice to Fas-induced hepatocellular apoptosis. Bax, Bak doubly-deficient murine embryonic fibroblasts are completely resistant to retrovirus-mediated tBID expression. These cells are also quite resistant to multiple death stimuli that initiate damage at distinct subcellular compartments including the nucleus, plasma membrane, and endoplasmic reticulum. Taken together these results suggest a model where BH3 domain-only death ligands such as BID activate "multi-domain" proapoptotic members BAX and BAK to unlock the essential gateway to mitochondrial dysfunction and apoptosis.