title: Nucleotide pools dictate the identity and frequency of ribonucleotide incorporation in mitochondrial DNA.
abstract: Previous work has demonstrated the presence of ribonucleotides in human mitochondrial DNA (mtDNA) and in the present study we use a genome-wide approach to precisely map the location of these. We find that ribonucleotides are distributed evenly between the heavy- and light-strand of mtDNA. The relative levels of incorporated ribonucleotides reflect that DNA polymerase γ discriminates the four ribonucleotides differentially during DNA synthesis. The observed pattern is also dependent on the mitochondrial deoxyribonucleotide (dNTP) pools and disease-causing mutations that change these pools alter both the absolute and relative levels of incorporated ribonucleotides. Our analyses strongly suggest that DNA polymerase γ-dependent incorporation is the main source of ribonucleotides in mtDNA and argues against the existence of a mitochondrial ribonucleotide excision repair pathway in human cells. Furthermore, we clearly demonstrate that when dNTP pools are limiting, ribonucleotides serve as a source of building blocks to maintain DNA replication. Increased levels of embedded ribonucleotides in patient cells with disturbed nucleotide pools may contribute to a pathogenic mechanism that affects mtDNA stability and impair new rounds of mtDNA replication.
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Anna-Karin
authors.initials: AK
authors.lastname: Berglund
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Clara
authors.initials: C
authors.lastname: Navarrete
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Martin K M
authors.identifier: 0000-0003-2174-2225
authors.initials: MK
authors.lastname: Engqvist
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Emily
authors.initials: E
authors.lastname: Hoberg
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Szilagyi
authors.affiliation: Wellcome Trust Centre for Mitochondrial Research, Institute of Neuroscience, The Medical School, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Robert W
authors.initials: RW
authors.lastname: Taylor
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Claes M
authors.initials: CM
authors.lastname: Gustafsson
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Maria
authors.initials: M
authors.lastname: Falkenberg
authors.affiliation: Institute of Biomedicine, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Anders R
authors.identifier: 0000-0002-5069-0930
authors.initials: AR
authors.lastname: Clausen
mesh_terms: D004247:DNA / Q000096:biosynthesis; D000074002:DNA Polymerase gamma; D004260:DNA Repair / Q000235:genetics*; D004261:DNA Replication / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics*; D005347:Fibroblasts; D054629:Genome, Mitochondrial; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000235:genetics / Q000473:pathology; D012313:RNA / Q000096:biosynthesis; D012260:Ribonucleases / Q000235:genetics; D012265:Ribonucleotides / Q000235:genetics*
chemical_list: D004272:DNA, Mitochondrial; D012265:Ribonucleotides; D012313:RNA; D004247:DNA; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; D012260:Ribonucleases