title: Diverting CERT-mediated ceramide transport to mitochondria triggers Bax-dependent apoptosis.
abstract: A deregulation of ceramide biosynthesis in the endoplasmic reticulum (ER) is frequently linked to induction of mitochondrial apoptosis. Although in vitro studies suggest that ceramides might initiate cell death by acting directly on mitochondria, their actual contribution to the apoptotic response in living cells is unclear. Here, we have analyzed the consequences of targeting the biosynthetic flow of ceramides to mitochondria using a ceramide transfer protein (encoded by COL4A3BP) equipped with an OMM anchor, mitoCERT. Cells expressing mitoCERT import ceramides into mitochondria and undergo Bax-dependent apoptosis. Apoptosis induction by mitoCERT was abolished through (i) removal of its ceramide transfer domain, (ii) disruption of its interaction with VAMP-associated proteins (VAPs) in the ER, (iii) addition of antagonistic CERT inhibitor HPA12, (iv) blocking de novo ceramide synthesis and (v) targeting of a bacterial ceramidase to mitochondria. Our data provide the first demonstration that translocation of ER ceramides to mitochondria specifically commits cells to death and establish mitoCERT as a valuable new tool to unravel the molecular principles underlying ceramide-mediated apoptosis.
authors.affiliation: Molecular Cell Biology Division, Department of Biology/Chemistry, University of Osnabrück, Osnabrück D-49076, Germany.
authors.forename: Amrita
authors.initials: A
authors.lastname: Jain
authors.affiliation: Molecular Cell Biology Division, Department of Biology/Chemistry, University of Osnabrück, Osnabrück D-49076, Germany.|Max-Planck-Institute for Molecular Cell Biology and Genetics, Dresden D-01307, Germany.
authors.forename: Oliver
authors.initials: O
authors.lastname: Beutel
authors.affiliation: Molecular Cell Biology Division, Department of Biology/Chemistry, University of Osnabrück, Osnabrück D-49076, Germany.
authors.forename: Katharina
authors.identifier: 0000-0001-9800-2781
authors.initials: K
authors.lastname: Ebell
authors.affiliation: Molecular Cell Biology Division, Department of Biology/Chemistry, University of Osnabrück, Osnabrück D-49076, Germany.
authors.forename: Sergey
authors.initials: S
authors.lastname: Korneev
authors.affiliation: Molecular Cell Biology Division, Department of Biology/Chemistry, University of Osnabrück, Osnabrück D-49076, Germany holthuis@uos.de.|Membrane Biochemistry & Biophysics, Bijvoet Center and Institute of Biomembranes, Utrecht University, Utrecht 3584 CH, The Netherlands.
authors.forename: Joost C M
authors.identifier: 0000-0001-8912-1586
authors.initials: JC
authors.lastname: Holthuis
mesh_terms: D017209:Apoptosis; D055162:Biocatalysis; D001692:Biological Transport; D002518:Ceramides / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D033921:Vesicular Transport Proteins / Q000378:metabolism; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: D002518:Ceramides; C114051:VAPA protein, human; D033921:Vesicular Transport Proteins; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; C119090:CERT1 protein, human; D017346:Protein Serine-Threonine Kinases
keywords: Bcl-2 proteins; Ceramide transfer protein; Cytochrome c; Endoplasmic reticulum; Membrane contact sites; Mitochondrial apoptosis; VAP receptor