An alternative import pathway of AIF to the mitochondria.

In eukaryotic cells, transport of the newly synthesized proteins and phospholipids to the appropriate subcellular target compartments is essential for maintaining organelle morphology and cell survival. In animal cells, mitochondria are major organelles containing DNA genome that encodes only for a small fraction of their proteins, which are required for the organelle function. Most mitochondrial proteins are encoded by the nuclear genes and imported to the mitochondria following protein synthesis. Apoptosis-inducing factor (AIF), an essential FAD-dependent NADH oxidase for the oxidative phosphorylation, is located in the intermembranous space and contains mitochondrial localization signals. However, the import mechanism of AIF to the mitochondria is not yet studied. Using sucrose gradient ultracentrifugation and immunoblotting, AIF was detected in fractions of the endoplasmic reticulum, mitochondria-associated membranes (MAM) and mitochondria, and AIF from these fractions was resistant to trypsin in the absence of digitonin, suggesting that AIF could be protected by phospholipids. Knockdown of dynamin-related protein 1 (DRP1kd) expression reduced AIF levels in the mitochondria, but increased AIF concentrations in the MAM. Knockdown of mitofusin-2 (Mfn-2kd) or ATPase family AAA domain containing 3A (ATAD3Akd) expression, however, reduced AIF levels in the mitochondria and increased the number of transport vesicles that contained AIF in the cytosol, indicating that ATAD3A and Mfn-2 were respectively essential for the import and fusion of transport vesicles into the mitochondria. Here we show that AIF is imported from the endoplasmic reticulum to the mitochondria via mitochondria-associated membranes and transport vesicles.

[1]  F. Sánchez‐Madrid,et al.  The mitochondrial fission factor dynamin‐related protein 1 modulates T‐cell receptor signalling at the immune synapse , 2011, The EMBO journal.

[2]  S. Sugano,et al.  ATPase family AAA domain-containing 3A is a novel anti-apoptotic factor in lung adenocarcinoma cells , 2010, Journal of Cell Science.

[3]  N. Pfanner,et al.  Distinct Forms of Mitochondrial TOM-TIM Supercomplexes Define Signal-Dependent States of Preprotein Sorting , 2009, Molecular and Cellular Biology.

[4]  T. Lithgow,et al.  Importing Mitochondrial Proteins: Machineries and Mechanisms , 2009, Cell.

[5]  G. Kroemer,et al.  AIF: Not Just an Apoptosis‐Inducing Factor , 2009, Annals of the New York Academy of Sciences.

[6]  V. Rangnekar,et al.  The Tumor Suppressor Par-4 Activates an Extrinsic Pathway for Apoptosis , 2009, Cell.

[7]  Yung-Yen Chiang,et al.  Nuclear expression of dynamin-related protein 1 in lung adenocarcinomas , 2009, Modern Pathology.

[8]  G. Hajnóczky,et al.  MAM: more than just a housekeeper. , 2009, Trends in cell biology.

[9]  T. Langer,et al.  Mitofusin 2 Builds a Bridge between ER and Mitochondria , 2008, Cell.

[10]  L. Scorrano,et al.  Mitofusin 2 tethers endoplasmic reticulum to mitochondria , 2008, Nature.

[11]  Miguel A. Andrade-Navarro,et al.  Cargo-Selected Transport from the Mitochondria to Peroxisomes Is Mediated by Vesicular Carriers , 2008, Current Biology.

[12]  Chad D Williamson,et al.  Colberg-poley Human Cells Mitochondrion-associated Membranes of Cytomegalovirus Ul37 Proteins Traffic into Mitochondrial and Secretory Human , 2007 .

[13]  L. Scorrano,et al.  Mitofusin 2: a mitochondria-shaping protein with signaling roles beyond fusion. , 2008, Antioxidants & redox signaling.

[14]  J. C. Hines,et al.  Identification of New Kinetoplast DNA Replication Proteins in Trypanosomatids Based on Predicted S-Phase Expression and Mitochondrial Targeting , 2007, Eukaryotic Cell.

[15]  C. Mannella,et al.  Structural and functional features and significance of the physical linkage between ER and mitochondria , 2006, The Journal of cell biology.

[16]  Y. Lai,et al.  Localization of GRP78 to mitochondria under the unfolded protein response. , 2006, The Biochemical journal.

[17]  M. Schrader.  Shared components of mitochondrial and peroxisomal division. , 2006, Biochimica et biophysica acta.

[18]  M. Wong-Riley,et al.  Is nuclear respiratory factor 2 a master transcriptional coordinator for all ten nuclear-encoded cytochrome c oxidase subunits in neurons? , 2005, Gene.

[19]  K. Mihara,et al.  Export of mitochondrial AIF in response to proapoptotic stimuli depends on processing at the intermembrane space , 2005, The EMBO journal.

[20]  B. de Kruijff,et al.  Continuous equilibration of phosphatidylcholine and its precursors between endoplasmic reticulum and mitochondria in yeast. , 2003, Molecular biology of the cell.

[21]  Nicholas J. Hoogenraad,et al.  Molecular Chaperones Hsp90 and Hsp70 Deliver Preproteins to the Mitochondrial Import Receptor Tom70 , 2003, Cell.

[22]  T. M. Lewin,et al.  Rat liver acyl-CoA synthetase 4 is a peripheral-membrane protein located in two distinct subcellular organelles, peroxisomes, and mitochondrial-associated membrane. , 2002, Archives of biochemistry and biophysics.

[23]  T. Dawson,et al.  Mediation of Poly(ADP-Ribose) Polymerase-1-Dependent Cell Death by Apoptosis-Inducing Factor , 2002, Science.

[24]  M. McNiven,et al.  The dynamin-like protein DLP1 is essential for normal distribution and morphology of the endoplasmic reticulum and mitochondria in mammalian cells. , 1999, Molecular biology of the cell.

[25]  Ruedi Aebersold,et al.  Molecular characterization of mitochondrial apoptosis-inducing factor , 1999, Nature.

[26]  T. Lithgow,et al.  The yeast nascent polypeptide-associated complex initiates protein targeting to mitochondria in vivo. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[27]  M. McNiven,et al.  A Novel Dynamin-like Protein Associates with Cytoplasmic Vesicles and Tubules of the Endoplasmic Reticulum in Mammalian Cells , 1998, The Journal of cell biology.

[28]  G. Lupo,et al.  Evidence that phosphatidylserine is imported into mitochondria via a mitochondria-associated membrane and that the majority of mitochondrial phosphatidylethanolamine is derived from decarboxylation of phosphatidylserine , 1995, The Journal of Biological Chemistry.

[29]  J. Vance.  Phospholipid synthesis in a membrane fraction associated with mitochondria. , 1990, The Journal of biological chemistry.

[30]  W. Ross,et al.  Topoisomerase-specific drug sensitivity in relation to cell cycle progression , 1987, Molecular and cellular biology.

[31]  P. Fisher,et al.  Import of nuclear-encoded mitochondrial proteins: a cotranslational perspective. , 2009, International review of cell and molecular biology.

[32]  M. Wong-Riley,et al.  NuclearRespiratoryFactor1RegulatesAllTenNuclear-encoded Subunits of Cytochrome c Oxidase in Neurons * , 2008 .

[33]  村上 薫.  Reconstitution of mitochondrial protein transport with purified ornithine carbamoyltransferase precursor expressed in Escherichia coli , 1990 .