Mitochondrial Dysfunction Due to Oxidative Mitochondrial DNA Damage Is Reduced through Cooperative Actions of Diverse Proteins

Abstract

The mitochondrial genome is a significant target of exogenous and endogenous genotoxic agents; however, the determinants that govern this susceptibility and the pathways available to resist mitochondrial DNA (mtDNA) damage are not well characterized. Here we report that oxidative mtDNA damage is elevated in strains lacking Ntg1p, providing the first direct functional evidence that this mitochondrion-localized, base excision repair enzyme functions to protect mtDNA. However, ntg1 null strains did not exhibit a mitochondrial respiration-deficient (petite) phenotype, suggesting that mtDNA damage is negotiated by the cooperative actions of multiple damage resistance pathways. Null mutations in ABF2 or PIF1, two genes implicated in mtDNA maintenance and recombination, exhibit a synthetic-petite phenotype in combination with ntg1 null mutations that is accompanied by enhanced mtDNA point mutagenesis in the corresponding double-mutant strains. This phenotype was partially rescued by malonic acid, indicating that reactive oxygen species generated by the electron transport chain contribute to mitochondrial dysfunction in abf2⌬ strains. In contrast, when two other genes involved in mtDNA recombination, CCE1 and NUC1, were inactivated a strong synthetic-petite phenotype was not observed, suggesting that the effects mediated by Abf2p and Pif1p are due to novel activities of these proteins other than recombination. These results document the existence of recombination-independent mechanisms in addition to base excision repair to cope with oxidative mtDNA damage in Saccharomyces cerevisiae. Such systems are likely relevant to those operating in human cells where mtDNA recombination is less prevalent, validating yeast as a model system in which to study these important issues.Mitochondria are important cellular targets for spontaneous and induced DNA damage (3,7,36,38,43), and mutations in mitochondrial DNA (mtDNA) cause diseases and likely contribute to late-onset neurodegenerative disorders and the aging process in humans (25, 51). Oxidative damage persists longer in mtDNA than it does in nuclear DNA (52) and appears to be a major contributor to mtDNA mutagenesis in vivo. In addition, decreased mitochondrial oxidative phosphorylation capacity is linked to oxidative stress pathways that perturb other cellular components and functions (52). The susceptibility of mtDNA to oxidative damage has been hypothesized to arise from a combination of biological determinants (36), including the proximity of mtDNA to reactive oxygen species that are by-products of normal respiration, the lack of a compact nucleosome structure to protect mtDNA from damage, and a paucity of mtDNA damage-processing pathways relative to those known to exist in the nucleus (6). However, it is largely unknown if and how these factors ultimately contribute to the susceptibility of mtDNA to damage.DNA damage caused by reactive oxygen species is removed by the base excision repair pathway (32), which is initiated by the action of glycosylases that excise specific d...

Murine Pif1 Interacts with Telomerase and Is Dispensable for Telomere Function In Vivo

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Spontaneous DNA Damage in Saccharomyces cerevisiae Elicits Phenotypic Properties Similar to Cancer Cells

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