Molecular Control of Mitochondrial Apoptosis by the BCL-2 Family.

Abstract SCI-13

The BCL-2 family proteins constitute a crucial checkpoint in apoptosis. They can be divided into 3 subfamilies based on their death regulatory activity and domain composition: 1) multidomain anti-apoptotic members such as BCL-2 and BCL-X_L; 2) multidomain pro-apoptotic BAX and BAK; 3) pro-apoptotic BH3-only molecules (BH3s). Our prior study has established a hierarchical regulation of mitochondrion-dependent apoptosis by BCL-2 subfamilies. Upon apoptosis, the “activator” BH3s, including BID, BIM, and PUMA, trigger homo-oligomerization of BAX and BAK to mediate cytochrome c efflux, leading to caspase activation. Conversely, the anti-apoptotic BCL-2/BCL-X_L/MCL-1 sequesters “activator” BH3s into insert complexes, thus preventing BAX/BAK activation. The remaining BH3s including BAD and NOXA do not activate BAX/BAK directly, but instead prevent the anti-apoptotic BCL-2 members from sequestering the “activator” BH3s. How the death effectors BAX and BAK are activated to trigger the mitochondria-dependent death program remains as one of the most vigorously debated topics in apoptosis research. Recently, we propose a stepwise activation model of BAX and BAK driven by activator BH3s. We demonstrate that the α1 helix of BAX keeps the α9 helix engaged in the dimerization pocket, rendering BAX as a monomer in cytosol. tBID, BIM, and PUMA bind transiently to the α1 helix of BAX to unleash auto-inhibition which allows for the structural reorganization by exposing both N- and C- termini. The C-terminal transmembrane domain hence becomes available for insertion into the mitochondrial outer membrane. tBID/BIM/PUMA remains associated with the N-terminally exposed BAX to drive the homo-oligomerization. BAK, an integral mitochondrial membrane protein, constitutively exposes its α1 helix and requires tBID/BIM/PUMA to trigger its homo-oligomerization. The homo-oligomerization of BAX/BAK appears to involve the interaction between the BH3 domain of one molecule and the canonical dimerization pocket of the other. Our study provides novel mechanistic insights regarding the spatiotemporal execution of BAX/BAK-governed cell death. As BAX and BAK are present in viable cells, their activity must be tightly regulated. Using a biochemical approach, we have previously identified VDAC2 as a gatekeeper of BAK. In viable cells, VDAC2 associates with BAK to keep BAK in check. Following death stimuli, BAK is released from VDAC2 by “activator” BH3s, thus undergoing homo-oligomerization. Our recent study provides in vivo evidence demonstrating a critical role of the VADC2-BAK complex in gauging the thymocyte survival. Genetic depletion of Vdac2 in thymus resulted in excessive cell death and hypersensitivity to diverse death stimuli including the T cell receptor engagement. Remarkably, these phenotypes were completely rescued by the concurrent deletion of Bak but not Bax. Thus, the VDAC2-BAK axis serves as a novel mechanism that governs the homeostasis of thymocytes. Impairment of apoptosis is not only central to cancer development but also renders tumors refractory to cytotoxic therapy. Accordingly, inhibitors of the anti-apoptotic BCL-2 proteins have been developed as promising anti-cancer therapeutics. Further characterization of the differential regulation between various BCL-2 subfamilies should help refine the strategies targeted on BCL-2-regulated apoptosis.