title: The exonuclease activity of DNA polymerase γ is required for ligation during mitochondrial DNA replication.
abstract: Mitochondrial DNA (mtDNA) polymerase γ (POLγ) harbours a 3'-5' exonuclease proofreading activity. Here we demonstrate that this activity is required for the creation of ligatable ends during mtDNA replication. Exonuclease-deficient POLγ fails to pause on reaching a downstream 5'-end. Instead, the enzyme continues to polymerize into double-stranded DNA, creating an unligatable 5'-flap. Disease-associated mutations can both increase and decrease exonuclease activity and consequently impair DNA ligation. In mice, inactivation of the exonuclease activity causes an increase in mtDNA mutations and premature ageing phenotypes. These mutator mice also contain high levels of truncated, linear fragments of mtDNA. We demonstrate that the formation of these fragments is due to impaired ligation, causing nicks near the origin of heavy-strand DNA replication. In the subsequent round of replication, the nicks lead to double-strand breaks and linear fragment formation.
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Bertil
authors.initials: B
authors.lastname: Macao
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Jay P
authors.initials: JP
authors.lastname: Uhler
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Triinu
authors.initials: T
authors.lastname: Siibak
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Zhu
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Yonghong
authors.initials: Y
authors.lastname: Shi
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Wenwen
authors.initials: W
authors.lastname: Sheng
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Monica
authors.initials: M
authors.lastname: Olsson
authors.affiliation: Department of Mitochondrial Biology, Max Planck Institute for Biology of Ageing, D-50931 Cologne, Germany.
authors.forename: James B
authors.identifier: http://orcid.org/0000-0002-2902-4968
authors.initials: JB
authors.lastname: Stewart
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Claes M
authors.initials: CM
authors.lastname: Gustafsson
authors.affiliation: Department of Medical Biochemistry and Cell Biology, University of Gothenburg, Medicinaregatan 9 A, P.O. Box 440, SE-40530 Gothenburg, Sweden.
authors.forename: Maria
authors.initials: M
authors.lastname: Falkenberg
mesh_terms: D000818:Animals; D015139:Blotting, Southern; D000074002:DNA Polymerase gamma; D004261:DNA Replication; D004272:DNA, Mitochondrial / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism; D006801:Humans; D051379:Mice; D016133:Polymerase Chain Reaction; D061987:Sf9 Cells; D018411:Spodoptera
chemical_list: D004272:DNA, Mitochondrial; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; C413027:POLG protein, human; C498803:Polg protein, mouse; D005090:Exodeoxyribonucleases