Structures of BIRC6-Client Complexes Provide Mechanism of Smac-Mediated Release of Caspases

Summary Apoptosis is tightly regulated and essential for metazoan development 1, 2 . Excessive apoptosis contributes to neurodegenerative disease, while diminished apoptosis can lead to inflammation and cancer 3 . Inhibitor of apoptosis (IAP) proteins are the principal actors that restrain apoptotic activity and are thus attractive therapeutic targets 4 . IAPs in turn are regulated by mitochondria-derived pro-apoptotic factors such as Smac and HtrA2 4 . Here, through a series of cryo-electron microscopy (cryo-EM) structures of full-length baculoviral IAP repeat-containing protein 6 (BIRC6) bound to Smac, caspase-3, caspase-7 and HtrA2, we provide the molecular basis for BIRC6-mediated caspase inhibition and its release by Smac. We demonstrate that BIRC6 cooperates preferentially with the non-canonical E1 enzyme UBA6 to ubiquitylate caspases and that caspase ubiquitylation is effectively inhibited by Smac through near-irreversible interactions. The dimeric arrangement of BIRC6 resolves the long-standing question of how the evolutionarily conserved single-BIR domain IAPs sequester caspases and how Smac binding antagonizes IAPs. This collection of BIRC6 structures provides critical insights into IAP-mediated apoptosis regulation, relevant for the development of current and future apoptosis-targeting cancer therapeutics.