title: Oxidants and not alkylating agents induce rapid mtDNA loss and mitochondrial dysfunction.
abstract: Mitochondrial DNA (mtDNA) is essential for proper mitochondrial function and encodes 22 tRNAs, 2 rRNAs and 13 polypeptides that make up subunits of complex I, III, IV, in the electron transport chain and complex V, the ATP synthase. Although mitochondrial dysfunction has been implicated in processes such as premature aging, neurodegeneration, and cancer, it has not been shown whether persistent mtDNA damage causes a loss of oxidative phosphorylation. We addressed this question by treating mouse embryonic fibroblasts with either hydrogen peroxide (H(2)O(2)) or the alkylating agent methyl methanesulfonate (MMS) and measuring several endpoints, including mtDNA damage and repair rates using QPCR, levels of mitochondrial- and nuclear-encoded proteins using antibody analysis, and a pharmacologic profile of mitochondria using the Seahorse Extracellular Flux Analyzer. We show that a 60min treatment with H(2)O(2) causes persistent mtDNA lesions, mtDNA loss, decreased levels of a nuclear-encoded mitochondrial subunit, a loss of ATP-linked oxidative phosphorylation and a loss of total reserve capacity. Conversely, a 60min treatment with 2mM MMS causes persistent mtDNA lesions but no mtDNA loss, no decrease in levels of a nuclear-encoded mitochondrial subunit, and no mitochondrial dysfunction. These results suggest that persistent mtDNA damage is not sufficient to cause mitochondrial dysfunction.
authors: Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
authors: Amy M
authors: AM
authors: Furda
authors: Adele M
authors: AM
authors: Marrangoni
authors: Anna
authors: A
authors: Lokshin
authors: Bennett
authors: B
authors: Van Houten
mesh_terms: D000477:Alkylating Agents / Q000633:toxicity*; D000818:Animals; D002460:Cell Line; D004249:DNA Damage; D004272:DNA, Mitochondrial / Q000187:drug effects*; D006019:Glycolysis / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000633:toxicity; D008741:Methyl Methanesulfonate / Q000633:toxicity; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000502:physiology; D016877:Oxidants / Q000633:toxicity*; D010085:Oxidative Phosphorylation / Q000187:drug effects
chemical_list: D000477:Alkylating Agents; D004272:DNA, Mitochondrial; D016877:Oxidants; D008741:Methyl Methanesulfonate; D006861:Hydrogen Peroxide