title: Complementation between polymerase- and exonuclease-deficient mitochondrial DNA polymerase mutants in genomically engineered flies.
abstract: Replication errors are the main cause of mitochondrial DNA (mtDNA) mutations and a compelling approach to decrease mutation levels would therefore be to increase the fidelity of the catalytic subunit (POLγA) of the mtDNA polymerase. Here we genomically engineer the tamas locus, encoding fly POLγA, and introduce alleles expressing exonuclease- (exo(-)) and polymerase-deficient (pol(-)) POLγA versions. The exo(-) mutant leads to accumulation of point mutations and linear deletions of mtDNA, whereas pol(-) mutants cause mtDNA depletion. The mutant tamas alleles are developmentally lethal but can complement each other in trans resulting in viable flies with clonally expanded mtDNA mutations. Reconstitution of human mtDNA replication in vitro confirms that replication is a highly dynamic process where POLγA goes on and off the template to allow complementation during proofreading and elongation. The created fly models are valuable tools to study germ line transmission of mtDNA and the pathophysiology of POLγA mutation disease.
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Ana
authors.initials: A
authors.lastname: Bratic
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Timo E S
authors.initials: TE
authors.lastname: Kauppila
authors.affiliation: Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Medicinaregatan 9A, Gothenburg SE-40530, Sweden.
authors.forename: Bertil
authors.initials: B
authors.lastname: Macao
authors.affiliation: Department of Biological Mechanisms of Ageing, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Grönke
authors.affiliation: Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Medicinaregatan 9A, Gothenburg SE-40530, Sweden.
authors.forename: Triinu
authors.initials: T
authors.lastname: Siibak
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: James B
authors.identifier: http://orcid.org/0000-0002-2902-4968
authors.initials: JB
authors.lastname: Stewart
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Francesca
authors.initials: F
authors.lastname: Baggio
authors.affiliation: Department of Biological Mechanisms of Ageing, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Dols
authors.affiliation: Department of Biological Mechanisms of Ageing, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.
authors.forename: Linda
authors.initials: L
authors.lastname: Partridge
authors.affiliation: Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Medicinaregatan 9A, Gothenburg SE-40530, Sweden.
authors.forename: Maria
authors.initials: M
authors.lastname: Falkenberg
authors.affiliation: Department of Laboratory Medicine, Karolinska Institutet, Stockholm SE-17177, Sweden.
authors.forename: Anna
authors.initials: A
authors.lastname: Wredenberg
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Strasse 9b, Cologne D-50931, Germany.|Department of Laboratory Medicine, Karolinska Institutet, Stockholm SE-17177, Sweden.
authors.forename: Nils-Göran
authors.initials: NG
authors.lastname: Larsson
mesh_terms: D000818:Animals; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D004330:Drosophila / Q000235:genetics*; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D005818:Genetic Engineering; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D021122:Protein Subunits
chemical_list: D004272:DNA, Mitochondrial; D021122:Protein Subunits; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; C413027:POLG protein, human; D005090:Exodeoxyribonucleases