**3. Mechanisms of DNA repair**

Maintainance of genomic integrity is complex due to great diversity of damage that can occur in DNA. In contrast to other biomolecules, DNA cannot be replaced, only repaired. To avoid the deleterious consequences of damage accumulation, cells have a variety of DNA repair pathways, each recognize and repair specific types of DNA damage. Base excision repair (BER), nucleotide excision repair (NER), mismatch repair (MMR), homologous recombination (HR), non-homologous end-joining repair (NHEJ) and direct damage reversal repair are some of the most important pathways used by cells to repair oxidative and alkylating DNA damage (Table I). These cellular repair pathways are not completely independent from one another. Some studies have show physical interactions between some

DNA Damage Protection and Induction of Repair

2008).

by Dietary Phytochemicals and Cancer Prevention: What Do We Know? 241

(MPG). MPG is responsible for enzymatic hydrolysis of the N-glycosylic bond resulting an abasic site in the DNA that is repaired by other enzymes of BER pathway. Over expression of MPG may produce an imbalance between abasic sites formation and repair in favor of abasic sites formation leading an increase of alkylating agents cytotoxicity (Doak et al.,

It has been estimated that a large number of AP sites are generated per cell per day. AP sites are unstable and are highly mutagenic because they result in non-template DNA and RNA synthesis. However, the number of mutations is extremely low, which demonstrate the efficient repair of this damage by the repair mechanisms (Jaiswal and Narayan, 2008). The ability of one glycosylase to recognize more than one type of damage, and the fact that each damage may be recognized by more than one type of glycosylases, give a degree of redundancy in the DNA repair processes which contribute to efficient damage repair (Maynard et al., 2009). Several studies have been showed that post-translational modifications, such as phosphorylation, acetylation and sumoylation may modulate repair

Decrease on BER activity can predispose humans to development of certain cancers, such as colon cancer (Jaiswal and Narayan, 2008; Wilson and Bohr, 2007). Otherwise, an increase of BER activity has been associated with resistance to certain cancer treatments (Liu and Gerson, 2006; Marchesi *et al.*, 2007). Nevertheless, the functional significance of BER in

**DNA Repair pathway DNA damage Reviews** 

Hegde, M. L., et al., 2008 Robertson, A.B., et al., 2009

Kaina, B., et al., 2007 and

Helleday et al., 2007

Hanawalt, P.C., 2002 Nouspikel, T., 2009

Jiricny J., 2006

2010

Double strand breaks Dudas and Chovanec, 2004

alkylated bases; AP sites; SSBs

insertion/deletion loops (IDLs)

Direct damage reversal repair O6MeG lesions Kondo N., et al., 2010

Table 1. The main DNA repair pathways and types of DNA damage repaired.

DNA adducts (e.g. thymidine dimers and 6–4 photoproducts) induced by UV radiation or electrophilic chemicals

NER is another important repair pathway involved in DNA adduct repair (e.g. thymidine dimers and 6–4 photoproducts) that are induced by ultraviolet radiation, chemicals or ROS (Benhamou and Sarasin, 2000). These adducts change the normal structure of the DNA helix, breaking transcription and replication processes. Two distinct NER sub-pathways, transcription coupled repair (TCR) and global genomic repair (GGR), have been described. Briefly, TCR repair transcription-blocking lesions present in transcribed DNA strands; and GGR pathway repairs lesions over the bulk genome including non-transcribed strands of active genes (Knudsen et al., 2009). Lesions like thymidine dimmers are usually repaired by

activity of BER enzymes, influencing repair efficiencies (Tudek, 2007).

prevention, development and treatment of disease remains unclear.

Base Excision Repair (BER) Base modifications (e.g. oxidized and

Mismatch Repair (MMR) Mispaired nucleotides and

Nucleotide Excision Repair

Homologous recombination (HR) and non-homologous end-joining repair (NHEJ)

**3.2 Nucleotide Excision Repair** 

(NER)

repair proteins from different repair pathways, suggesting that regulation of DNA repair involves protein cross-talk (Knudsen *et al.*, 2009). Protein expression, pos-translational modifications and nuclear translocation of DNA repair proteins have been referred as essential in the regulation of DNA repair activity and to maintain genomic stability. However, these subjects need to be clarified in the future (Knudsen *et al.*, 2009; Tudek, 2007). When DNA damage is not repaired, the cell by a complex network that collectively forms the DNA damage response (DDR) machinery delays cell-cycle progression, acting on cell cycle checkpoints. This delay gives the DNA repair machinery more time, allowing correct repair of the damage. DDR machinery can induce other mechanisms such as apoptosis or necrosis to avoid that altered cells continue to proliferate and result in disease (Bartek *et al.*, 2007; Maynard *et al.*, 2009).

#### **3.1 Base Excision Repair**

BER is the major pathway involved in the repair of oxidation and alkylation DNA damage and occurs in both the nucleus and mitochondria (D'Errico *et al.*, 2008). BER recognizes and repairs AP sites, DNA SSBs and different types of base modifications, such as oxidized/reduced bases (e.g. 8-oxoG or formamidopyrimidines), alkylated bases, deaminated bases (e.g. uracil) or base mismatches (Maynard et al., 2009). BER pathway involves steps as recognition, excision, filling and ligation that are carried out by four or five enzymes. Briefly, the repair process is initiated by one of several DNA glycosylases, each recognizing a specific DNA lesion (e.g. OGG1, NTH, NEIL and MYH recognize oxidation damage; deamination damages are recognized by UDG, MED1, UNG, and TDG, and MPG initiate alkylation repair) (Knudsen et al., 2009). These DNA glycosylases excise the damaged base (by cleave of N-glycosidic bond between the sugar and the base), generating an AP site. An AP endonuclease, as APE1, cleaves the AP site, to generate 3′ OH and 5′ deoxyribose phosphate (dRP) terminus. The third step is carried through DNA polymerase that fills the nucleotide gap generated due to lesion base removal. Finally, DNA ligase seals the nick on DNA (Hegde *et al.*, 2008; Evans *et al.*, 2004). Several other protein factors have been identified as interacting with the essential BER proteins and/or the DNA to modulate BER activity. BER pathway is described in detail in the several reviews (Fortini *et al.*, 2003; Hegde *et al.*, 2008; Robertson *et al.*, 2009; Fortini *et al.*, 2003).

Hydroxylation of guanine at C-8 position, 8-oxoGua (8-Oxo-7,8-dihydroguanine) is one of the most abundant forms of DNA oxidation and the most studied because of its mutagenic potential. If not properly repaired, 8-oxoG can pair with cytosine or adenine. Replication of 8-oxoG paired with C by DNA polymerases is a non-mutagenic process. However, replication of 8-oxoG paired with A results in GC to TA or TA to GC transversions that are strong mutagenic DNA lesions (Pastoriza Gallego and Sarasin, 2003). In mammalian cells 8 oxoGua is predominantly recognized and excised by a specific DNA glycosylases. hOGG1 removes 8-oxoG when paired with a cytosine and the glycosylase hMYH removes adenine when mispaired with 8-oxoG, both through the BER pathway. Activity of OGG1 is enhanced by cofactors such as human apurinic/apyrimidinic endonuclease (APE1), Xeroderma Pigmentosum complementation group C (XPC) and human endonuclease VIIIlike (NEIL1) (D'Errico et al., 2008).

N7alkylG adducts can be repaired by spontaneous depurination resulting in abasic sites that are often correctly repaired by BER. If not repaired, abasic sites may result in singlestranded gaps that can stall replication forks resulting in double stranded breaks. N7alkylG adducts are also recognized and removed from DNA by N-methylpurine-DNAglycoslase

repair proteins from different repair pathways, suggesting that regulation of DNA repair involves protein cross-talk (Knudsen *et al.*, 2009). Protein expression, pos-translational modifications and nuclear translocation of DNA repair proteins have been referred as essential in the regulation of DNA repair activity and to maintain genomic stability. However, these subjects need to be clarified in the future (Knudsen *et al.*, 2009; Tudek, 2007). When DNA damage is not repaired, the cell by a complex network that collectively forms the DNA damage response (DDR) machinery delays cell-cycle progression, acting on cell cycle checkpoints. This delay gives the DNA repair machinery more time, allowing correct repair of the damage. DDR machinery can induce other mechanisms such as apoptosis or necrosis to avoid that altered cells continue to proliferate and result in disease (Bartek *et al.*,

BER is the major pathway involved in the repair of oxidation and alkylation DNA damage and occurs in both the nucleus and mitochondria (D'Errico *et al.*, 2008). BER recognizes and repairs AP sites, DNA SSBs and different types of base modifications, such as oxidized/reduced bases (e.g. 8-oxoG or formamidopyrimidines), alkylated bases, deaminated bases (e.g. uracil) or base mismatches (Maynard et al., 2009). BER pathway involves steps as recognition, excision, filling and ligation that are carried out by four or five enzymes. Briefly, the repair process is initiated by one of several DNA glycosylases, each recognizing a specific DNA lesion (e.g. OGG1, NTH, NEIL and MYH recognize oxidation damage; deamination damages are recognized by UDG, MED1, UNG, and TDG, and MPG initiate alkylation repair) (Knudsen et al., 2009). These DNA glycosylases excise the damaged base (by cleave of N-glycosidic bond between the sugar and the base), generating an AP site. An AP endonuclease, as APE1, cleaves the AP site, to generate 3′ OH and 5′ deoxyribose phosphate (dRP) terminus. The third step is carried through DNA polymerase that fills the nucleotide gap generated due to lesion base removal. Finally, DNA ligase seals the nick on DNA (Hegde *et al.*, 2008; Evans *et al.*, 2004). Several other protein factors have been identified as interacting with the essential BER proteins and/or the DNA to modulate BER activity. BER pathway is described in detail in the several reviews (Fortini *et al.*, 2003;

Hydroxylation of guanine at C-8 position, 8-oxoGua (8-Oxo-7,8-dihydroguanine) is one of the most abundant forms of DNA oxidation and the most studied because of its mutagenic potential. If not properly repaired, 8-oxoG can pair with cytosine or adenine. Replication of 8-oxoG paired with C by DNA polymerases is a non-mutagenic process. However, replication of 8-oxoG paired with A results in GC to TA or TA to GC transversions that are strong mutagenic DNA lesions (Pastoriza Gallego and Sarasin, 2003). In mammalian cells 8 oxoGua is predominantly recognized and excised by a specific DNA glycosylases. hOGG1 removes 8-oxoG when paired with a cytosine and the glycosylase hMYH removes adenine when mispaired with 8-oxoG, both through the BER pathway. Activity of OGG1 is enhanced by cofactors such as human apurinic/apyrimidinic endonuclease (APE1), Xeroderma Pigmentosum complementation group C (XPC) and human endonuclease VIII-

N7alkylG adducts can be repaired by spontaneous depurination resulting in abasic sites that are often correctly repaired by BER. If not repaired, abasic sites may result in singlestranded gaps that can stall replication forks resulting in double stranded breaks. N7alkylG adducts are also recognized and removed from DNA by N-methylpurine-DNAglycoslase

2007; Maynard *et al.*, 2009).

**3.1 Base Excision Repair** 

Hegde *et al.*, 2008; Robertson *et al.*, 2009; Fortini *et al.*, 2003).

like (NEIL1) (D'Errico et al., 2008).

(MPG). MPG is responsible for enzymatic hydrolysis of the N-glycosylic bond resulting an abasic site in the DNA that is repaired by other enzymes of BER pathway. Over expression of MPG may produce an imbalance between abasic sites formation and repair in favor of abasic sites formation leading an increase of alkylating agents cytotoxicity (Doak et al., 2008).

It has been estimated that a large number of AP sites are generated per cell per day. AP sites are unstable and are highly mutagenic because they result in non-template DNA and RNA synthesis. However, the number of mutations is extremely low, which demonstrate the efficient repair of this damage by the repair mechanisms (Jaiswal and Narayan, 2008). The ability of one glycosylase to recognize more than one type of damage, and the fact that each damage may be recognized by more than one type of glycosylases, give a degree of redundancy in the DNA repair processes which contribute to efficient damage repair (Maynard et al., 2009). Several studies have been showed that post-translational modifications, such as phosphorylation, acetylation and sumoylation may modulate repair activity of BER enzymes, influencing repair efficiencies (Tudek, 2007).

Decrease on BER activity can predispose humans to development of certain cancers, such as colon cancer (Jaiswal and Narayan, 2008; Wilson and Bohr, 2007). Otherwise, an increase of BER activity has been associated with resistance to certain cancer treatments (Liu and Gerson, 2006; Marchesi *et al.*, 2007). Nevertheless, the functional significance of BER in prevention, development and treatment of disease remains unclear.


Table 1. The main DNA repair pathways and types of DNA damage repaired.
