**4.1 Base excision repair (BER)**

Base excision repair (BER) is the commissioned pathway to removed nonbulky lesions like alkylated, deaminated, and oxidized bases from the DNA. This pathway has been well studied in the nucleus and was the first repair mechanism reported in mitochondria and to date the best characterized in this last organelle [33, 34]. BER mechanism is highly conserved from bacteria to humans and basically comprises five stages: (1) recognition and excision of the damaged base, (2) removal of the abasic site formed, (3) DNA end processing, (4) repair synthesis, and (5) ligation [35], and at the same time, BER can be divided into two branches (both founded in mitochondria): short patch and long patch, where the difference lies in the procedure used to repair, while short patch forms a single nucleotide gap, long patch forms a bigger one of 2–10 nucleotides [36]. One of the main elements involved in BER processes are the DNA glycosylases, enzymes that catalyze the excision of the N-glycosidic bond between the altered base and its corresponding carbohydrate, thus creating an abasic site, and activity that was first observed by Lindahl et al. [37]. These glycosylases can be subdivided into monofunctional or bifunctional, depending on whether they have lyase activity or not, ability that determines the type of damage they can repair, while monofunctional glycosylases focus on nonoxidative damage, bifunctional exert their action against oxidized DNA bases [35]. In mammalian mitochondria, seven glycosylases have been reported, three monofunctional: uracil-DNA glycosylase (UNG1), *E. coli*  MutY homolog (MYH), and N-methylpurine DNA glycosylase (AAG/MPG) and four bifunctional: 8-oxoguanina-DNA glycosylase (OGG1), *E. coli* endonuclease III homolog (NTH1) endonuclease VIII-like glycosylase 1 and 2 (NEIL1 and 2) [38–41]. After the action of glycosylases, the next step, DNA end processing, is catalyzed by nuclear BER enzymes that are also found in mitochondria, as apurinic-apyrimidinic endonuclease 1 (APE1) and polynucleotide kinase-3′-phosphatase (PNKP) [35], further the

#### **Figure 1.**

*Mitochondrial import and mtDNA repair pathways. The elements that participate in mitochondrial genome maintenance are nuclear encoded; therefore, mitochondria need to import these effector proteins, which are involved in different repair pathways.* 

synthesis is carried out by the main mitochondrial polymerase (Polγ). It was recently reported that polymerase beta (Polβ) also localizes within mitochondria, whereas similar to the nucleus, it has a relevant role in mtDNA maintenance and mitochondrial homeostasis through its participation in BER [42]. It has also been observed that FEN1, DNA2, and EXOG are involved in the removal of DNA flap generated by polymerase strand displacement synthesis [43]. Finally, the ligation activity is made by mitochondrial ligase III [44]. In conclusion, despite that BER is the most characterized pathway in mitochondria, it is still unknown whether all the nuclear elements are conserved in mitochondrial BER, or on the contrary, if this pathway have specific proteins that do not participate in nuclear version of BER, it is not well understood either which are the regulatory signals that controls the import of these elements to mitochondria.

#### **4.2 Nucleotide excision repair (NER)**

 Nucleotide excision repair (NER) pathway is a complex machinery involved in recognition of lesions, adducts or structures that modifies the DNA double helix structure, having the possibility of blocking transcription, replication, and thus affecting DNA stability. One of the most important features of NER is its versatility on a wide kind of lesions since they are detected and repaired. Roughly, the process starts with damage recognition, next, the damaged strand is cleaved at both sides of the lesion to be removed later, then the missing sequence is synthesized using the complementary strand as a template, and finally the ends are ligated, thus restoring DNA sequence and integrity [45].

 There is a large body of evidence reporting the absence of nucleotide excision repair activity in mitochondria. The first observation about this lack was made by Clayton et al. in 1974 [2], who reported that mitochondria were unable to remove UV-induced pyrimidine dimers. As a result, lesions that are normally repaired in nucleus by NER could persist in mitochondrial genome; this damage is not only restricted to photodimers, since NER also participates in repair of other bulky lesions and a subset of oxidative DNA damage [1]. In the nucleus, bulky lesions can also be overcome through the use of specialized translesion synthesis (TLS) polymerases; however, in mitochondria, the major polymerase (Polγ) presents a weakly thymine dimer bypass activity in vitro and at present, it is unknown whether this activity is conserved in vivo [46]. On the other hand, it was recently reported that polymerase theta (Polθ), an enzyme that acts as a translesion bypass polymerase, thus promoting the pass of replicative stalling lesions in the nucleus [47], is localized to mitochondria. Additionally, it was observed that there is an increase of Polθ localization to mitochondria after treating the cells with an oxidative agent, suggesting that the enzyme is recruited to the organelle when this kind of damage is inflicted, where it could facilitate translesion bypass synthesis. Interestingly, in POLQ KO cells, the rate of point mutation in mtDNA was significantly reduced after oxidative treatment; this observation indicates that Polθ is involved in a pathway of error-prone DNA synthesis that may facilitate replication in mitochondria [48].

 Despite that the nucleotide excision repair pathway has not been clearly recognized in mitochondria, there are several proteins involved in the nuclear version of this pathway that not only localize to the organelle but also they accumulate upon oxidative damage, and when they are depleted from cells, the number of point mutation is increased, observations that strongly suggest their participation in mtDNA maintenance. Some of these proteins are: Cockayne syndrome A (CSA), Cockayne syndrome B (CSB), and Rad23A [48–50]. Therefore, further investigation is needed to completely rule out NER pathway from mitochondria, or to elucidate whether these mentioned proteins are involved in other pathways different to nucleus.
