title: Reconstitution of proapoptotic BAK function in liposomes reveals a dual role for mitochondrial lipids in the BAK-driven membrane permeabilization process.
abstract: BAK is a key effector of mitochondrial outer membrane permeabilization (MOMP) whose molecular mechanism of action remains to be fully dissected in intact cells, mainly due to the inherent complexity of the intracellular apoptotic machinery. Here we show that the core features of the BAK-driven MOMP pathway can be reproduced in a highly simplified in vitro system consisting of recombinant human BAK lacking the carboxyl-terminal 21 residues (BAKΔC) and tBID in combination with liposomes bearing an appropriate lipid environment. Using this minimalist reconstituted system we established that tBID suffices to trigger BAKΔC membrane insertion, oligomerization, and pore formation. Furthermore, we demonstrate that tBID-activated BAKΔC permeabilizes the membrane by forming structurally dynamic pores rather than a large proteinaceous channel of fixed size. We also identified two distinct roles played by mitochondrial lipids along the molecular pathway of BAKΔC-induced membrane permeabilization. First, using several independent approaches, we showed that cardiolipin directly interacts with BAKΔC, leading to a localized structural rearrangement in the protein that "primes" BAKΔC for interaction with tBID. Second, we provide evidence that selected curvature-inducing lipids present in mitochondrial membranes specifically modulate the energetic expenditure required to create the BAKΔC pore. Collectively, our results support the notion that BAK functions as a direct effector of MOMP akin to BAX and also adds significantly to the growing evidence indicating that mitochondrial membrane lipids are actively implicated in BCL-2 protein family function.
authors.affiliation: From the Unidad de Biofísica (Centro Mixto Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibertsitatea (UPV/EHU), P.O. Box 644, 48080 Bilbao, Spain.
authors.forename: Olatz
authors.initials: O
authors.lastname: Landeta
authors.affiliation: From the Unidad de Biofísica (Centro Mixto Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibertsitatea (UPV/EHU), P.O. Box 644, 48080 Bilbao, Spain.
authors.forename: Ane
authors.initials: A
authors.lastname: Landajuela
authors.affiliation: CIC-BIOGUNE Structural Biology Unit, Parque Tecnologico Zamudio, Bizkaia, 48160 Derio, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Gil
authors.affiliation: From the Unidad de Biofísica (Centro Mixto Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibertsitatea (UPV/EHU), P.O. Box 644, 48080 Bilbao, Spain.
authors.forename: Stefka
authors.initials: S
authors.lastname: Taneva
authors.affiliation: Université de Bordeaux, INSERM U869, Institut Européen de Chimie et de Biologie, Pessac F-33607, France, and.
authors.forename: Carmelo
authors.initials: C
authors.lastname: DiPrimo
authors.affiliation: the MRC Centre for Protein Engineering and MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 0QH, United Kingdom.
authors.forename: Begoña
authors.initials: B
authors.lastname: Sot
authors.affiliation: CIC-BIOGUNE Structural Biology Unit, Parque Tecnologico Zamudio, Bizkaia, 48160 Derio, Spain.
authors.forename: Mikel
authors.initials: M
authors.lastname: Valle
authors.affiliation: From the Unidad de Biofísica (Centro Mixto Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibertsitatea (UPV/EHU), P.O. Box 644, 48080 Bilbao, Spain,; the Departamento de Bioquímica y Biología Molecular, UPV/EHU, Leioa 48940, Spain,; Ikerbasque, Basque Foundation for Science, 48011 Bilbao, Spain.
authors.forename: Vadim A
authors.initials: VA
authors.lastname: Frolov
authors.affiliation: From the Unidad de Biofísica (Centro Mixto Consejo Superior de Investigaciones Cientificas-Universidad del Pais Vasco/Euskal Herriko Unibertsitatea (UPV/EHU), P.O. Box 644, 48080 Bilbao, Spain,. Electronic address: gbzbaasg@lg.ehu.es.
authors.forename: Gorka
authors.initials: G
authors.lastname: Basañez
mesh_terms: D000818:Animals; D051023:BH3 Interacting Domain Death Agonist Protein / Q000235:genetics / Q000378:metabolism; D002308:Cardiolipins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D008930:Mitochondria, Liver / Q000235:genetics / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000378:metabolism*; D010539:Permeability; D051381:Rats; D017207:Rats, Sprague-Dawley; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: C494772:BAK1 protein, human; C494826:BAX protein, human; D051023:BH3 Interacting Domain Death Agonist Protein; C494774:Bak1 protein, rat; C494828:Bax protein, rat; D002308:Cardiolipins; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein