title: MitoRCA-seq reveals unbalanced cytocine to thymine transition in Polg mutant mice.
abstract: Mutations in mitochondrial DNA (mtDNA) can lead to a wide range of human diseases. We have developed a deep sequencing strategy, mitoRCA-seq, to detect low-frequency mtDNA point mutations starting with as little as 1 ng of total DNA. It employs rolling circle amplification, which enriches the full-length circular mtDNA by either custom mtDNA-specific primers or a commercial kit, and minimizes the contamination of nuclear encoded mitochondrial DNA (Numts). By analyzing the mutation profiles of wild-type and Polg (mitochondrial DNA polymerase γ) mutant mice, we found that mice with the proofreading deficient mtDNA polymerase have a significantly higher mutation load by expanding the number of mutation sites and to a lesser extent by elevating the mutation frequency at existing sites even before the premature aging phenotypes appear. Strikingly, cytocine (C) to thymine (T) transitions are found to be overrepresented in the mtDNA of Polg mutated mice. The C → T transition, compared to other types of mutations, tends to increase the hydrophobicity of the underlying amino acids, and may contribute to the impaired protein function of the Polg mutant mice. Taken together, our findings may provide clues to further investigate the molecular mechanism underlying premature aging phenotype in Polg mutant mice.
authors.affiliation: 1] State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China [2] Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Ting
authors.initials: T
authors.lastname: Ni
authors.affiliation: State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China.
authors.forename: Gang
authors.initials: G
authors.lastname: Wei
authors.affiliation: State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China.
authors.forename: Ting
authors.initials: T
authors.lastname: Shen
authors.affiliation: State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China.
authors.forename: Miao
authors.initials: M
authors.lastname: Han
authors.affiliation: State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China.
authors.forename: Yaru
authors.initials: Y
authors.lastname: Lian
authors.affiliation: State Key Laboratory of Genetic Engineering &Ministry of Education (MOE) Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center of Genetics and Development, School of Life Sciences, Fudan University, Shanghai, 200438, P.R. China.
authors.forename: Haihui
authors.initials: H
authors.lastname: Fu
authors.affiliation: Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Yanqin
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Laboratory of Molecular Biology, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Liu
authors.affiliation: Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Yoshi
authors.initials: Y
authors.lastname: Wakabayashi
authors.affiliation: Unit on Synapse Development and Plasticity, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Li
authors.affiliation: Laboratory of Molecular Biology, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Toren
authors.initials: T
authors.lastname: Finkel
authors.affiliation: Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Xu
authors.affiliation: Genetics and Development Biology Center, National Heart Lung Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D003596:Cytosine; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics*; D051379:Mice; D008817:Mice, Mutant Strains; D008969:Molecular Sequence Data; D017354:Point Mutation / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods*; D013941:Thymine
chemical_list: D004272:DNA, Mitochondrial; D003596:Cytosine; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; C498803:Polg protein, mouse; D013941:Thymine