has been provided by the fact that DNA polymerase g is capable of mutagenic bypass through DNA lesions, introducing a deoxyadenosine opposite an AP site or an 8-oxodG [133] and inserting deoxynucleotides opposite bulky DNA adducts derived from benzo[a]pyrene diol epoxide [134]. Although much remains to be learned about the possibility of TLS in mitochondria, it seems reasonable that a damage tolerance mechanism might be operative in mitochondria to prevent replication and repair stalling following genotoxic injury.

10.4.6.4Nucleotide Excision Repair (NER)

Nucleotide excision repair (NER) is the major pathway active against UV irradiation and involves removal and resynthesis of a short fragment on the damaged DNA strand [135]. To date, no evidence for the NER pathway has been presented in mammalian mitochondria [136]. However, in yeast, some established NER substrates i.e., UV-induced photoproducts are repaired, but by alternative mechanisms. For instance, in Schizosaccharomyces pombe mitochondrial cyclobutane pyrimidine dimers (CPDs) and (6-4) pyrimidineÐpyrimidone photoproducts [(6-4)PPs] can be removed by UV-damaged DNA endonuclease-dependent excision repair (UVER) [137]. In such a pathway, UVDE cleaves the phosphodiester bond 5¢-adjacent to the CPD or (6-4)PP, and the repair is thought to be completed by a BER-like process [138].

10.4.7Intramitochondrial Localization of DNA Repair Proteins

mtDNA is organized in nucleoprotein particles called nucleoids, containing several copies of the mitochondrial genome and multiple proteins [139, 140]. Nucleoids from most organisms have been demonstrated to contain at least 25 proteins, most of which have not been fully characterized. Among those proteins that have been characterized, a wide array play important functions in DNA replication and repair, as well as in organization including its DNA packaging, Some nucleoid-associated proteins, i.e., high mobility group (HMG) box proteins, have roles in mtDNA maintenance and packaging; others, such as Aco1 and IIv5, are multifunctional, having structural roles in addition to metabolic functions [141, 142]. Nucleoids are associated with the inner mitochondrial membrane and are spaced evenly along the mitochondrial reticulum [143]. Factors proposed to control mammalian mitochondrial nucleoid organization include prohibitin 1 (PHB1) [142] and the ATPase family AAA domain-containing protein 3 (ATAD3) [144]. Multiple DNA repair associated proteins have also been identiÞed and include mitochondrial ssDNA-binding protein (mtSSB), mitochondrial RNA polymerase, breast cancer 1 protein (BRCA1), TWINKLE helicase, and polymerase g [145]. Consistent with a membrane localization of the mtDNA, most of the mitochondrial BER activities are membrane-bound [98]. With the exception of AP endonuclease, the entire BER pathway interacts with

the inner membrane in mammalian organelles and is generally accepted to associate with nucleoids [98]. Interestingly, p53 was recovered in a mitochondrial inner membrane subfraction containing BER components [146]. In addition, components of NER and BER have been shown to interact in proximity of mitochondrial nucleoids. SpeciÞcally, proteins encoded the Cockayne syndrome gene CSA and CSB associate with mtSSB and OGG1 [147]. Furthermore, CSB seems to play a role in recruiting BER proteins to DNA repair complexes at the mitochondrial inner membrane [148].

10.4.8mtDNA Damage Sensing and Signaling

Under physiological conditions, DNA repair proteins (mtDNA and nDNA proteins) are maintained in the cytosol and are rapidly translocated to either the mitochondria or nucleus in response to DNA damaging conditions. Recently, the yeast N-glycosylases NTG1 and NTG2 were shown to be dynamically relocalized to the mitochondria following mitochondrial DNA damage [149]. Furthermore, it was shown NTG1 localization to mitochondria was triggered by mtDNA oxidative damage and not by the ROS level. This suggests that there is a sensing mechanism for mtDNA damage which would be mediated by speciÞc mitochondrial signals. Posttranslational modiÞcations via sumoylation have been shown to be involved in NTG1 trafÞcking to the damaged genome [149]. In mammalian cells, OGG1, AP endonuclease, CSA and CSB proteins are rapidly relocalized to the damaged DNA in stress conditions [147, 150].

10.4.9Import of Nuclear Encoded DNA Repair Enzymes into the Mitochondria

As noted above, the majority of mitochondrial proteins are encoded by the nuclear genome as precursors with an N-terminal extension, containing a mitochondrial targeting sequence [105, 151]. After recognition of the targeting sequence by membrane receptors, the proteins are transported across the mitochondrial membrane by the translocase complexes, TOM and TIM, which are present in the outer and inner membrane, respectively. Cleavage of the mitochondrial targeting sequence is carried out by a peptidase [152]. Oxidative stress inhibits the import and processing of mitochondrial matrix proteins in vitro and in vivo [153, 154] and impaired mitochondrial import of DNA repair proteins has been implicated in certain neurological diseases [155]. Intriguingly, there appears to be a general deÞciency in the import of base excision repair proteins into the mitochondria matrix during aging, namely, OGG1 and uracil DNA-glycosylase [92]. It appears that these enzymes become trapped in the outer membrane in an age-dependent manner, which may contribute to the inefÞcient