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X-ray Repair Cross Complementing Protein 1 (XRCC1) encodes a protein which is found to involve in the DNA base excision repair (BER) [1, b, 16] and it is the first isolated mammalian gene . BER gets involved in the removal of the damaged base pair and the repair synthesis of the DNA by making use of the opposite strands as the template . BER brings up the various constituents like DNA polymerase Î², DNA ligase III and polyadenosine diphosphate-ribose polymerases (PARP) of the BER to the location where the DNA has been damaged and improves the efficiency of the BER pathway [1, 3, 16, 20, 23]. XRCC1 is associated with numerous human cancers. It interacts with various DNA glycosylases such as hOGG1, MPG, hNTH1 and NEIL1 to participate in the first step of the BER. It has the N-terminal domain (NTD) and BRCT1 (amino acid 301-402) domains which have the affinity to form the covalent complex with the apurinic or apyrimidinic (AP) sites with the help of the Schiff base and it is the protein-protein interface which interacts with PARP [9, 15, 16, 17]. It is found to have higher affinity when it carries AP-lyase or APE1-incised AP site in its DNA and thus the overall repair process of damaged base pair is accelerated and it can be used as a substrate for the DNA polymerase beta (DNA pol Î²). The repaired DNA strand can be resealed with the help of its role in the multiple protein-protein interaction .
19q13.2 [1, 22]
DNA associated [h]
Structure of XRCC1
XRCC1 has 17 exons  and is found to encode a protein with 633 amino acids . It is a homodimer protein with the molecular weight of about 69526 Da [f]. The size of the DNA is about 32.27 kb [h, 22]. The NTD region of the gene binds with pol Î² very efficiently. It has been identified from the mutagenic study that the epitopic region of XRCC1 gets involved in the complex formation of pol Î². The XRCC1-NTD complexed with pol Î² creates an unexpected variation in the structure of the XRCC1-NTD. The oxidized form of the XRCC1-NTD has more affinity towards the poly Î² and the formation of the disulphide bond regulates the repair pathway. Thus XRCC1 has an indirect role in the DNA repair .
Functions of XRCC1
XRCC1 protein is essential for the integrity of the human genome . The protein product of the XRCC1 gene is used to repair the DNA strand break induced by the hydrolysis and the exchange of the sister-chromatids which is formed by the exposure of the ionizing radiation, alkylating agents and the chemicals present in the smoke [1, a, b, 3, 4, 12]. Due to the absence of the DNA repair process, adducts are formed which halts the DNA replication or the cytotoxic mutation and instability of the gene . It acts as the scaffold protein and gets involved in the BER pathway by interacting with DNA ligase III by means of COOH terminus in its BRCT domain, PARP 1 and 2, polynucleotide kinase, apurinic/apyrimidinic endonuclease 1 (APE1) and DNA pol Î² at its NH2 terminus [b, 2, 10, 12, 18, 21, 22, 26]. PARP is a zinc-finger containing enzyme which detects the single strand breaks in the DNA and it is also found to involve in the BER [14, 15]. It is found to have a minor function in the processing of the DNA during meiogenesis and recombination in germ cells [b, 16, 17]. It codes for an MÎ³ 70,000 protein . Absence of this gene leads to the decline in the level of ligase III . It is found to involve in the maintenance of the genome stability. It also stimulates the activity of the PKNP by transferring the PNKP from the phosphorylated DNA product. It recruits ARL6IP5 to the damaged sites of the DNA which mediates the DNA repair induced by the oxidative stress by means of stimulating the expression of XRCC1 through MAPK-E2F1 signaling pathway and protecting XRCC1 from ubiquitin-proteasome degradation. It is also necessary for the neurogenesis of the cerebellar interneurons and for the hippocampal homeostasis and the prevention of the neural damage of the DNA. It has the ability to activate PNKP by means of two independent mechanisms. The effect of simulation of the phosphorylated XRRC1 on PKNP is completely inhibited by the presence of the excess of FHA domain polypeptide whereas the XRCC1 which is non-phosphorylated is not at risk to the competition by FHA domain [h].
BER pathway is a multi-step process which is used to remove the damaged base pairs in the DNA caused by the oxidation, methylation, deamination or the loss of the DNA base pair [10, 26]. These alterations are known to be mutagenic. It has two subpathways which are initiated by the action of the DNA glycosylase. Barnes and Lindhal reported that eleven different glycosylases were found in the mammalian cells . The DNA glycosylase such as the 8-oxoguanine glycosylase (OGG1) is used to cleave the N-glycosidic bond between the damaged base pair and the sugar phosphate backbone of the DNA and this cleavage creates an AP or abasic sites in the DNA. In humans, eight DNA glycosylases has been found with to have the partially overlapped bases. The AP sites can also be formed by the hydrolysis of the N-glycosidic bond. In both the cases, AP Endonuclease 1 (APE1) cleaves the phosphodiester bond at 5' end of the AP site which results in the 3' hydroxyl group and a transient 5' abasic deoxyribose phosphate (dRP). The dRP can be removed by the action of the DNA pol Î² which acts by removing the dRP moiety by means of its AP lyase activity and by adding one nucleotide to the 3' end of the nick [d, 19]. Finally, the strand nicks are sealed by the activity of the DNA ligase that restores the integrity of the DNA. The above process of replacing the damaged base pair with the new base pair is known as "short-patch" repair and it represents approximately 80-90% of all BER. The other subpathway of BER is known as the "long-patch" repair pathway which is used only when the modified basepair is resistant to the AP lyase activity of the DNA pol Î² present in the DNA. It leads to the replacement of 2-10 damaged bases. It also requires some of the factors such as DNA glycosylase, APE1 and DNA pol Î² which has involved in the "short-patch" repair pathway but the long-patch repair is a PCNA-dependent pathway in which several base pairs can be added to the repair gap by displacing the dRP as a part of a "flap" oligonucleotide with the help of the DNA polymerase. The overhanging oligonucleotides are sealed by the Flap endonuclease FEN-1 whose nick has been formerly sealed by the DNA ligase [d]. Short-patch DNA repair is used when the gap is only one nucleotide whereas the long-patch DNA repair is used when the gap is about two to eight nucleotides long [19, 25].
BER pathway [d]
Short and long patch pathway
Cigarette smoke mainly contains the carcinogens and the Reactive Oxygen Species (ROS) . The smoke contains the free radicals which is present in both the mainstream and sidestream smoke. It is present in both the gas and the tar phase. The tar phase contains more reductants like quinone when compared to that of the gas phase . Polycyclic aromatic hydrocarbons (PAH), aromatic amines and N-nitroso compounds are some of the carcinogens present in the cigarette smoke which damages the DNA by forming the adducts. ROS which is a component of the cigarette smoke is found in both the gas-phase and the particulate matter. ROS includes radicals of the oxygen such as the superoxide and the hydroxyl radicals and some of the oxygen derivatives which lack unpaired electrons like hydrogen peroxide and hypochlorous acid. The enzymatic reactions carried out by the bacteria and inflammatory cells revealed that N-nitroso compounds present in cigarette smoke produces nitric oxide radicals which causes the oxidative damage and the accumulation of ROS leads to the oxidative stress which results in the cancer. ROS damages the DNA directly or indirectly by inducing lipid peroxidation and the oxidation of the protein. The indirect damage of the DNA includes the inactivation of the target enzymes which gets involved in the synthesis of the DNA whereas the direct damage of the DNA includes the strand breaks of the DNA, formation of the abasic sites and base adduct formation like thymine glycol, 5-hydrozymethyluracil and 8-hydroxy-2-deoxyguanosine. Damages in the base of the DNA and the single strand breaks in the DNA can be repaired by the BER pathway .
General Mechanism of Benzo[a]pyrene
Benzo[a]Pyrene Diol Epoxide (BPDE) is a bioactivated form of the B[a]P and is the polycyclic Aromatic Hydrocarbon (PAH) that damages the DNA irreversibly by means of covalent binding or by the process oxidation which leads to the formation of the BPDE-DNA adducts. The adducts are formed with the DNA by binding to the amino group of adenine or guanine [24, 27]. The formation of adduct obstructs transcription of the gene which are not repaired by the NER pathway and this leads to the carcinogenesis. Thus, the reduced DNA repair capacity of the gene is related to the increased risk of the environmentally induced cancer .
Free radicals in Skin cancer
Tobacco smoking which includes the cigarette and the pipe smoking increases the risk of the cutaneous squamous cell carcinoma . When the human skin is exposed to the cigarette smoke, it creates an oxidative stress on the skin and the exposure increases the level of the hydroperoxide which has been derived from the cigarette smoke. It has various effects on the skin like wrinkling and also includes some of the disorders of the skin like impairment in the barrier function of the skin. The cigarette smoke contains the free radicals which help in the lipid peroxidation. It mainly participates in the oxidation of Squalene (SQ), which is one of the important constituent of the human skin surface lipid in to the squalene monohydroperoxide (SQHPO). The oxidation of the human skin by the cigarette smoke is a very tedious process. It involves two types of oxidation. The first type is the oxidation of lipids to the corresponding hydroperoxides and the second type involves the oxidation of the hydroperoxides. It is also found that the hydroperoxides present on the human skin which can be detected by the chemiluminescence (CL) detector is oxidized more when compared to the one which is not exposed. The oxidation can be prevented by treating it with the antioxidants like glutathione, thiotaurine, hypotaurine and ascorbic acid .
XRCC1 is one of the genes which get involved in the BER pathways. BER is the main pathway which guards the DNA from the damages caused by UV, ionizing radiation, cellular metabolism, including the reactive oxygen species, methylation, deamination and hydroxylation. XRCC1 is used in the BER by interacting with the DNA repair proteins like PARP, DNA ligase III and DNA pol Î². It is found to have eight nonsynonymous SNPs. Among which three of the polymorphism is found to be common and it includes the amino acid substitutions of Arg194 to Trp (C to T) of exon 6, Arg280 to His (G to A) of exon 9 and Arg399 to Gln (G to A)  of exon 10 [17, 22]. The Arg399 to Gln polymorphism is found in the BRCT-1 interaction domain of XRCC1 within the region of PARP and this polymorphism is related to the cancer risk [4, 17]. Variation in the Gln399 allele decreases the DNA repair capacity of XRCC1 which is due to the formation of the DNA adducts, high level of the sister chromatid exchange, increased RBC glycophorin A, p53 mutations and prolonged delay in the cell cycle [4, 16, 23]. The variations in the residue Arg194 and 280 is known to occur in the proliferating cell nuclear antigen binding region which contains polar Pro, Ser and Arg/Lys rich regions. The function of XRCC1 is altered by the transition of the positively charged amino acid Arg to the hydrophobic Trp within the conserved region. It was reported by Wang et al that the variations in the Trp194 allele had fewer belomycinor benzo[a]pyrene diol epoxides induced chromosomal breaks than that of the wild genotype. Polymorphisms at the codon 288 and 399 didn't have any association with the altered DNA levels and the G2 cell cycle delay.
Substitution of G28152 with A at codon 399 of exon 10 increases the risk of cancer. Lunn reported that the XRCC1 28152 polymorphism in codon 399 is related to the increased level of aflatoxin B1-DNA adducts and glycophorin A variants in the normal population [4, 16]. BER repairs the PAH-DNA adducts by metabolizing them by means of radical cation intermediates to electrophiles which binds to DNA and destabilizes the N-glycosyl bond by quick depurination or depyrimidation of the bases that are adducted .
Mutation of XRCC1 related to lung cancer
The substitution Arg280 with His in exon 9 is the polymorphism which is related to that of the lung cancer [5, 19]. The gene product of XRCC1 acts as a scaffold protein which coordinates the action of polymerase Î², DNA ligase III and poly (ADP-ribose) polymerase in short-patch BER. Arg399 to Gln polymorphism of the gene which is located in the conserved region alters the function of the gene . The polymorphism of the non-conservative amino acid Arg399 to Gln is accompanied by the transversion of the nucleotide from G 28152 to A. The polymorphism occurs within the BRCA1 COOH terminus of the PARP which might have an effect on complex assembly or efficiency of the DNA repair [g, 16, 19, 20]. Polymorphism of XRCC1 with the base pair substitution of C26304 to T in exon 6 is accompanied by the nonconservative amino acid change from Arg194 to Trp [16, 19]. It has been reported by Lunn et al that 399Gln allele is related to the higher levels of the aflatoxin B1-DNA adducts and glycophorin A somatic mutations . The polymorphism also had some remarkable variation among the Caucasians and Taiwanese [13, 14]. It was also reported by Duell et al that 399Gln allele has the higher frequencies of the sister chromatid exchange when compared to that of the 399Arg homozygous allele . PAHs such as benzo[a]pyrene are the most important carcinogens which forms the DNA adducts which increases the risk of the lung cancer . During the enzymatic oxidation of the PAHs, a variety of the reactive oxygen species such as the hydroxyl radical and hydrogen peroxides are generated and the production of the reactive oxygen species leads to the oxidative damage of the DNA which is removed by means of the BER that includes XRCC1. BER is also used to target the depurinating DNA adducts such as the N7-methylguanine and N3-methyladenine which has been derived from the radical cations formed by one-electron oxidation of PAHs . The variation in the Arg399 to Gln is found to be related to the decreased sensitivity to belomycin and benzo[a]pyrene-diol-epoxide (BPDE) and the variations in Arg280His and Arg194Trp is found to have decreased risk to cancer [18, 19, 23].
Mutation of XRCC1 related to Oral Cancer
XRCC1 plays a vital role in the BER pathway by interacting with DNA pol Î², PARP and DNA ligase III. It has the BRCT domain which is responsible for its function in cell cycle checkpoint and is responsible for the DNA damage [7, 12]. XRCC1 has a central role in the carcinogenic pathway of oral cancer in Taiwanese. Shen et al reported five types of polymorphism in the XRCC1 gene . Among the five types of polymorphism, three types of polymorphisms are found in the codon 194, 280 and 399 with the substitution of Arg to Trp, Arg to His on exon 9 and Arg to Gln respectively [7, 8]. Polymorphism in codon 399 of XRCC1 is related to the p53 mutation of the oral cancer caused by the chemical carcinogens .
Head and neck cancer
Head and Neck Squamous Cell Carcinoma (HNSCC) encompass 6% of all the malignant neoplasm. Smoking or the consumption of the alcohol is found to be the major risk factor of HNSCC. Two types of the XRCC1 polymorphism have been identified at the codon 194 and 399 which includes the substitution of Arg to Trp (C26304T) of exon 6 and Arg to Gln (G28152A) of exon 10 respectively [4, 9]. Polymorphism of Gln399 is located in the COOH terminus of the PARP which interacts with the BRCT domain [9, 17, 26]. PARP detects the strand breaks in the DNA and it is the enzyme which contains the zinc-finger. The higher level of the DNA adducts are found in the individuals who carry polymorphism in the residue Gln399 of XRCC1 [9, 17]. Single Nucleotide Polymorphism (SNP) of the residue Arg194 is mainly related to the lower belomycin and benzo[a]pyrene diol epoxides sensitivity. The residue 280His which is found to be situated in the proliferating cell nuclear antigen binding region is sensitive to the higher level of the belomycin. The 399Gln allele which is found at the carboxylic acid terminal side of the polyadenosine diphosphate-ribose polymerase-interacting domain is linked to the higher levels of the aflatoxin B1-DNA adducts and higher belomycin sensitivity . XRCC1 protein gets interacted with the DNA ligase III and DNA pol Î². DNA ligase III is used to re-join the strand breaks in the DNA whereas DNA pol Î² is used in the base excision repair . The variation in Arg 194 and 399 increased the risk to the HNSCC in the smoking individuals [e]. Polymorphism of Gln399/Gln399 has an increased risk of the cancer only among the light smokers and Arg280Gln is related to the risk of the Head and Neck Cancer .
Polymorphism at the codons Arg 194 and 399 increased the risk of the laryngeal cancer in the individuals who smoke [e].