2 radical theory of aging, the accumulation of reactive oxy- gen species-(ROS-)-induced mtDNA damage and mtDNA mutations over time are the main contributors to deleterious changes leading to cellular dysfunction and aging [17, 18]. In eukaryotic cells, the integrity of mtDNA is constantly challenged by the endogenous production of mitochondrial ROS, which are generated during mitochondrial respiration. ROS are particularly important genotoxic agents and can generate a large number of DNA lesions, including oxidized DNA bases, abasic sites, and single- and double-strand breaks. Because many of the ROS-induced DNA lesions show mutagenic or cytotoxic eﬀects, mitochondrial-free radicals are thought to be an important source of DNA mutations and DNA instability, particularly in mitochon- drial DNA. The proximity of mtDNA to the main sites of mitochondrial ROS generation, located within complexes I and III of the mitochondrial electron transport chain in the inner mitochondrial membrane, is the major reason for the higher steady-state levels of oxidative lesions and the higher instability observed in mtDNA when compared to nuclear DNA [19, 20]. For many years, such high level of oxidative lesions in mtDNA was considered to be a consequence, at least in part, of (i) the absence of eﬃcient DNA repair mechanisms in mitochondria and (ii) the lack of protective histones in mtDNA. However, diﬀerent studies have shown that mtDNA is organized into protein-DNA complexes called nucleoids, where DNA has been described to be associated with diﬀerent proteins and anchored to the inner mitochondrial membrane [21–23]. These membrane- associated structures may provide more protection to the mtDNA against oxidative attack than was formerly thought. On the other hand, it is now well established that various DNA repair pathways actively take place in mitochondria. The initial studies on mitochondrial DNA repair capacity focused on UV light-induced DNA damage repair, and they provided the basis for the notion that mitochondria lacked functional DNA repair mechanisms [24]. Thirty years later, it is known that various DNA repair pathways do take place in mitochondria [25]. Most of the studies on mitochondrial DNA repair have focused on the BER mechanisms, which are the DNA repair pathway coping with oxidative lesions and it is the best-characterized DNA repair pathway in mitochondria [26, 27]. 2. DNA Repair and Mitochondria As mentioned above, DNA repair research has focused mainly on the nuclear compartment; however, the investiga- tions performed in the last two decades have conﬁrmed that mitochondria do possess eﬀective DNA repair mechanisms, and the understanding of how these mechanisms function has signiﬁcantly increased in the last few years. The ﬁrst DNA repair pathway that was described to actively take place in mammalian mitochondria was the BER pathway. Today, other DNA repair mechanisms that were thought to occur exclusively in the nucleus have been described to take place in mammalian mitochondria such as MMR [28, 29] and the long-patch BER [30–32]. Moreover, classical nuclear DNA repair enzymes have been identiﬁed in mitochondria Journal of Aging Research [28, 31, 33–35], with similar function than the one observed in the nucleus [31, 35], or with novel functions according to its mitochondrial location [33]. However, and despite the increased knowledge in mtDNA repair, important questions still remain opened, such as the precise mechanisms for the repair of some speciﬁc lesions in mtDNA such as double-strand breaks or DNA adducts derived from lipid peroxidation. The latter are likely generated in mtDNA, since mitochondrial phospholipids are main targets of mitochondrial-free radicals [36] and mtDNA is associated with the inner mitochondrial membrane; DNA adducts derived from lipid peroxidation are known to be processed by NER [37, 38], but whether mitochondria possess the capacity to repair DNA lesions others than UV light-induced lesions via the NER pathway remains to be elucidated. 3. Mitochondrial Base Excision Repair BER is the primary pathway known for repair of small DNA modiﬁcations caused by alkylation, deamination, or oxidation. BER takes place both in the nucleus and in mitochondria, and although the mechanisms are similar, mitochondria possess an independent BER machinery. Both nuclear BER and mtBER are based on a cascade of reactions starting with the recognition of the damage followed by enzymatic processing steps that aim to remove the lesion and restore genomic integrity. All the components of the mtBER pathway are nuclear encoded and imported into mitochondria [39]. The BER pathway includes 4 distinct steps: (i) recognition and removal of the modiﬁed base, (ii) processing of the generated apurinic/apyrimidinic (AP) site, (iii) incorporation of the correct nucleotide(s), and (iv) strand ligation. 3.1. Mitochondrial BER: Early Steps, Recognition, and AP Site Processing. The ﬁrst step of BER is catalyzed by DNA glycosylases, which recognize the modiﬁed base and cleave the N-glycosidic bond, creating an abasic site. In addition to the cleavage activity towards the N-glycosidic bond, some DNA glycosylases also have AP lyase activity, which allows the cleavage of the DNA backbone. These DNA glycosylases are called bi-functional glycosylases while those possessing only cleavage activity are named monofunctional. Oxidized bases are generally removed by bifunctional DNA glycosylases. Among all the diﬀerent DNA glycosylases that have been described to be present in the nucleus, only some of them have been detected in mitochondria [26, 40]. As the rest of enzymes involved in mtBER, nuclear genes encode mitochondrial DNA glycosylases. Mitochondrial and nuclear isoforms of the same DNA glycosylases are encoded by the same gene and are generated by alternative transcription initiation sites and alternative splicing [41]. That is the case of two of the main mitochondrial DNA glycosylases, 8-oxoguanine DNA glycosylase-1 (Ogg1) and Uracil-DNA glycosylase (UNG). UNG was the ﬁrst DNA-glycosylase to be identiﬁed [42]. Uracil in DNA is generated by deamination of cytosine or by misincorporation of dUMP. The removal of uracil