7916–7931 Nucleic Acids Research, 2012, Vol. 40, No. 16 doi:10.1093/nar/gks532 Published online 20 June 2012 Mitochondrial dynamics and autophagy aid in removal of persistent mitochondrial DNA damage in Caenorhabditis elegans Amanda S. Bess, Tracey L. Crocker, Ian T. Ryde and Joel N. Meyer* Duke University, Nicholas School of the Environment, Integrated Toxicology and Environmental Health Program, LSRC, PO Box 90328, Durham, NC 27708, USA Received March 12, 2012; Revised May 8, 2012; Accepted May 11, 2012 ABSTRACT Mitochondria lack the ability to repair certain helix-distorting lesions that are induced at high levels in mitochondrial DNA (mtDNA) by important environmental genotoxins and endogenous metab- olites. These lesions are irreparable and persistent in the short term, but their long-term fate is unknown. We report that removal of such mtDNA damage is detectable by 48 h in Caenorhabditis elegans, and requires mitochondrial fusion, fission and autophagy, providing genetic evidence for a novel mtDNA damage removal pathway. Furthermore, mutations in genes involved in these processes as well as pharmacological inhibition of autophagy exacerbated mtDNA damage-mediated larval arrest, illustrating the in vivo relevance of removal of persistent mtDNA damage. Mutations in genes in these pathways exist in the human population, demonstrating the potential for import- ant gene–environment interactions affecting mitochondrial health after genotoxin exposure. INTRODUCTION Mitochondria serve several critical cellular functions, including energy production via oxidative phosphoryl- ation (OXPHOS) and key roles in apoptosis. The majority of mitochondrial proteins involved in these processes are encoded by nuclear DNA (nDNA), but a small subset, 13 polypeptides that are incorporated into 4 of the 5 complexes of the OXPHOS system plus 2 rRNAs and 22 tRNAs, are encoded by mitochondrial DNA (mtDNA). A large and growing number of inherited mitochondrial diseases result from mutations in or deple- tion of mtDNA and likely affect >1 in 6000 people (1), emphasizing the importance of mtDNA integrity to human health. Furthermore, there is substantial evidence to support a role of mitochondrial dysfunction and mtDNA mutation in the pathogenesis of more common diseases such as neurodegenerative conditions (2,3), type 2 diabetes mellitus (4) and cancer (5). Compared with nDNA, mtDNA is particularly suscep- tible to damage from prevalent environmental contamin- ants. For instance, depending on exposure conditions, mtDNA is 2–100 times more susceptible to bulky adducts when exposed to the carcinogenic, polycyclic aromatic hydrocarbon (PAH) benzo(a)pyrene (6–11). This susceptibility is partially because of the fact that mitochondria lack nucleotide excision repair (NER), the repair pathway utilized to repair many helix-distorting or ‘bulky’ nDNA damage induced by common environmen- tal agents including PAHs, mycotoxins and ultraviolet C radiation (UVC) (12–15). Thus, helix-distorting mtDNA lesions resulting from exposure to these genotoxins are persistent and reduce mtDNA replication (8,16–18) and transcription (19). The resulting reduction of mtDNA copy number or protein synthesis could result in mitochondrial dysfunction, as suggested by cytochrome c release, reduced mitochondrial membrane potential (MP), decreased respiration and increased lipid peroxidation reported in studies with PAHs and the mycotoxin Aﬂatoxin B1 (20–22). Helix-distorting lesions induce nDNA mutations (23,24) and in vitro evidence supports the potential for these lesions to be mutagenic in mtDNA (25,26). However, exposures to benzo(a)pyrene diol epoxide and UVC in cell culture have resulted in little to no increase in mutation frequency above background (7,18,27), prompting the hypothesis that mtDNAs harboring helix-distorting lesions are degraded and mutations result- ing from these lesions are rare (28,29). Mitochondrial degradation is carried out by macroautophagy (30,31). Non-selective autophagy of mitochondria as well as other organelles and cytoplasmic materials (31) is triggered by starvation and nutrient deprivation. On the other hand, selective autophagy of mitochondria or *To whom correspondence should be addressed. Tel: +1 919 613 8109; Fax: +1 919 668 1799; Email: joel.meyer@duke.edu The Author(s) 2012. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.