title: Mitochondrial DNA repair and replication proteins revealed by targeted chemical probes.
abstract: Efficient and accurate replication and repair of mitochondrial DNA is essential for cellular viability, yet only a minimal complement of mitochondrial proteins with relevant activities have been identified. Here, we describe an approach to screen for new pathways involved in the maintenance of mitochondrial DNA (mtDNA) that leverages the activities of DNA-damaging probes exhibiting specific subcellular localization. By conducting a siRNA screen of known nuclear DNA maintenance factors, and monitoring synergistic effects of gene depletion on the activity of mitochondria-specific DNA-damaging agents, we identify a series of proteins not previously recognized to act within mitochondria. These include proteins that function in pathways of oxidative DNA damage repair and dsDNA break repair, along with a novel mitochondrial DNA polymerase, POLθ, that facilitates efficient DNA replication in an environment prone to oxidative stress. POLθ expression levels affect the mutational rate of mitochondrial DNA, but this protein also appears critical for efficient mtDNA replication.
authors.affiliation: Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Simon
authors.initials: S
authors.lastname: Wisnovsky
authors.affiliation: Department of Chemistry, Faculty of Arts and Science, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Sae Rin
authors.initials: SR
authors.lastname: Jean
authors.affiliation: Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada.|Department of Chemistry, Faculty of Arts and Science, University of Toronto, Toronto, Ontario, Canada.|Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Shana O
authors.identifier: 0000-0003-3360-5359
authors.initials: SO
authors.lastname: Kelley
mesh_terms: D004249:DNA Damage / Q000187:drug effects; D004260:DNA Repair / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects; D004272:DNA, Mitochondrial / Q000096:biosynthesis* / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D006801:Humans; D024101:Mitochondrial Proteins / Q000378:metabolism*; D015335:Molecular Probes / Q000737:chemistry / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects
chemical_list: D004272:DNA, Mitochondrial; D024101:Mitochondrial Proteins; D015335:Molecular Probes; D004259:DNA-Directed DNA Polymerase