PSAP Induces A Unique Mitochondrial Apoptotic Pathway Independent Of Bcl-2Family Proteins

Presenillin-associated protein (PSAP) was identified as a mitochondrialproapoptotic protein. However, the mechanism by which PSAP induces apoptosisremains unknown. To this end, we have established an inducible expression system.Using this system, we have examined the roles of caspases, Bcl-2family proteins,cytochrome c, Smac (Smac/Diablo, second mitochondria-derived activator ofcaspases/direct IAP binding protein with low PI), and Apaf-1(apoptotic protease-activating factor) in PSAP-induced apoptosis. Our results demonstrated that caspaseactivation is required for the execution of PSAP-induced apoptosis. Knockdown ofcaspase-9, but not caspase-8, abolished PSAP-induced PARP cleavage and theactivation of other caspases, indicating that caspase-9is the initiator caspase in theapoptotic cascades induced by PSAP. Knockdown of Apaf-1abolished PSAP-inducedcaspase activation and PARP cleavage, indicating that the apoptosome formationtriggered by cytochrome c is crucial for PSAP-induced apoptosis. Knockdown ofSmac abolished PSAP-induced caspase activation and PARP cleavage, indicating that,in addition to Apaf-1or apoptosome formation, Smac is also essential for PSAP-induced apoptosis; overexpression of Bcl-2and Bcl-xL did not protect cells fromPSAP-induced apoptosis, and knockdown of Bid, Bax, and Bak had no effect onPSAP-induced cytochrome c and Smac release, indicating that PSAP-inducedapoptosis is not regulated by Bcl-2family proteins. These results strongly suggest thatPSAP evokes mitochondrial apoptotic cascades via a novel mechanism that is notregulated by Bcl-2family proteins, but both the formation of cytochrome c-Apaf-1apoptosome and the presence of Smac are absolutely required for PSAP-inducedapoptosis.