**4. Methylation in DNA damage and repair**

DNA methylation is considered a post-replication modification by the methyl group (–CH3) addition at carbon 5 cytosine, known as 5-methylcytosine (5mC), primarily in dinucleotides of CpG [68]. This enzymatic reaction is catalyzed by three DNA methyltransferases (DNMTs) (DNMT1, DNMT3A, and DNMT3B). These modifications are commonly found in promoter regions, the CpG islands, as their main function is transcription silencing, decreasing, or repressing the gene function. Methylation is the epigenetic modification most studied in cancer, as it is well-known that DNA hypermethylation can transcriptionally silence tumor suppressors and DNA repair genes, giving neoplastic cells survival advantages [69]. Various genes related to cell cycle regulation, tumor cell invasion, cell signaling, apoptosis, and chromatin remodeling are hypermethylated and silenced in almost all tumors. As demonstrated by Pal et al. [70] for the first time, CpG sites in the promoter of H2AX, RNF8, and CYCS are methylated; they show the collaborative participation of hypermethylation of DR5, DCR1, DCR2, CASP8, CYCS, BRCA1, BRCA2 and H2AX and the hypomethylation of DR4, FLIP, and RNF8 in sporadic breast cancer; the authors proposed that promoter methylation of these apoptotic and DDR genes is not due to a random phenomenon, as the progressive modification of aberrant epigenetic alterations are associated with tumor advancement, which generates the dysregulation of the DDR-apoptotic pathway, promoting tumor development. Hinrichsen et al. [71] found that increased methylation in the promoters of MLH1, MSH2, PMS2, and p16 genes are correlated to an advanced stage in hepatocellular carcinoma. Epigenetic marks and DDR are crucial points to understand if changes in DNA methylation can contribute to resistance of cancer treatment, particularly by radiotherapy [72].

Various epigenetic marks have been associated with the response to damage, primarily the methylation of lysine 4 and 79 in H3 (H3K4me and H3K79me, respectively) that contribute to DDR and DNA repair [73]. The marks on H3K4me1/2/3 regulate the repair of DSBs through chromatin accessibility [74, 75]. Furthermore, H3K36 methylation is associated with "open" euchromatin and helps RNA pol II activate transcription [76]. Chang et al. [77] demonstrated that the tumor suppressor PHRF1 can move on the DSB due to H3K36me2/me3 and NBS1, then ubiquitinate PARP1, and trigger the subsequent repair by NHEJ. Yet, it is known that DOT1L methyltransferase catalyzes the H3K79 mono-di-trimethylation through its non-SET domain. Recently, in the study by Kari et al. [78], the depletion or inhibition of DOT1L activity was shown to result in altered DNA damage response, indicated by decreased levels of γH2AX, but with increased KAP1 phosphorylation. Loss

of DOT1L function leads to faulty HRR-mediated DSB repair without affecting NHEJ. Highlighting DOT1L-mediated H3K79me3 importance in the early response to DNA damage and DSB repair, its inhibition also increases radiation sensitivity and chemotherapeutic agents in colorectal cancer patients' treatment. Dot1L and H3K79 methylation was previously associated with the role of 53Bp1 in response to DNA damage [79]. FitzGerald et al. [80] corroborated that not only H3K79me recruits 53BP1 to DNA damage sites, but also H4K20me. This indicates that H4K20me concentration is essential for the repair pathway related to 53BP1 and BRCA1, in cooperation with the 53BP1-RIF1-MAD2L2 complex. The histone posttranslational modification of H3K27me3 and H4K20me is a diagnostic indicator in melanoma [81].

There is DNA methyltransferase, in which expression is regulated by epigenetic modification during DNA damage. MGMT is an O6 -methylguanine-DNA methyltransferase responsible for the repair of damaged guanine, without other cofactors transferring the methyl at 06-meG to cysteine residues, removing adducts in a single step, thus protecting chromosomes from mutations, carcinogenesis, and alkylating agents [82, 83]. MGMT can do this only once, after its DNA-binding domain change by alkylating, which is detached from DNA and targeted for degradation by ubiquitination [84]. Loss of MGMT expression is due to promoter methylation [85], used as an advantage for good prognosis in a glioma, which can predict whether alkylating agents can benefit treatment [86]. MGMT can interfere with TMZ response on tumor cells and is responsible for efficient repair of TMZ and induced toxic DNA adducts, reducing treatment efficacy. Targeting MGMT seems to overcome chemoresistance in gliomas, but the prognostic value of MGMT methylation is controversial, as genomic rearrangements result in MGMT overexpression, independent of its promoter methylation, contributing to resistance [87].

### **5. Genome damage repair via m6A participation**

Modified nucleotides on DNA create a vast list, shown by Fragou et al. [88], but methylation by methyltransferases is an epigenetic modification that occurs on DNA and RNA. In fact, RNA contains more modified bases than DNA, as these RNA modifications are present in rRNA, tRNA, snRNA, miRNA, or mRNA, giving rise to the emergence of a wide chemical diversity on its side chains. One of these modifications on mRNA has been studied since its discovery in 1974 [89, 90], the methylation of adenosine residues form N6-methyladenosine (m6A) is the most abundant post-transcriptional mRNA modification, and is detected in approximately 25% of mRNAs [91]. The m6A modification mechanism of action functions like methylation on DNA and histones, i.e., its presence or absence on mRNA determines the fate of the transcription. The addition of a methyl group to the N6 site of A, occurring in the RRACH sequence [92, 93], has a distinctive position in the vicinity of stop codons, internal exons, in 5'UTRs which could promote 5'capindependent translation when found on the first nucleotide adjacent to the 7-methylguanosine cap; this gives protection from decapping [7, 94, 95] and in 3'UTRs regulates the affinity of RNA binding proteins [96]. This is known as m6A modification, which is involved in a variety of cellular processes including gene expression through regulating RNA metabolism, such as mRNA translation, degradation, alternative splicing, export and folding [97, 98], control of protein translation [94], and others. All these effects are globally known as RNA epitranscriptomics.

For some time, m6A modification was considered static and unalterable, but is now well-known as reversible and dynamic. It is difficult to examine gene regulation at the RNA level without appropriate methods; genome-wide sequencing became a

**175**

factors [106].

*Epigenetics and DNA Repair in Cancer DOI: http://dx.doi.org/10.5772/intechopen.94030*

major tool for many years, but had limitations with the m6A assay. The sensibility of the methodologies used was a critical point, because until recently, techniques had a detection limit of about 0.01%, in contrast to the content of m6A, less than 0.001% is sufficient to regulate biological processes [99]. Antibody-based, highthroughput sequencing technology allowed us for the past few years to locate the specific m6A sites and explore their biological significance. Linder et al. [91] solved the problem of distinguishing between m6A and adenosine by using incubation of an m6A-antibody to induce a specific mutational signature of m6A residues after UV

This modification is added and eliminated by proteins called "writers" and "erasers," respectively; these proteins regulate the abundance, prevalence, and distribution of m6A; this exerts its biological function and is modulated by protein "readers." First, m6A is added to the nucleus by the methyltransferase complex, which is formed by two core proteins, METTL3 and METTL4 [92]. METTL3/ METTL4 form a functional heterodimer, METTL3, which is the catalytic subunit which uses S-adenosylmethionine (SAM) or S-adenosylhomocysteine (SAH) as donors for methyl transfer in adenosine within the consensus motif G(G/A) ACU [100]. METTL4 has the active site blocked, and lacks amino acid residues to form hydrogen bonds with the ribose hydroxyls of SAM to facilitate donor and acceptor substrate binding; this is important in maintaining the integrity of the complex, stabilizing METTL3, and enabling RNA substrate recognition [101]. Regarding m6A demethylases or "eraser" proteins, FTO and ALKBH5 are two well-known m6A demethylases located in nuclear speckles within the methyltransferase complex. They belong to the group of Fe(II)/2-oxoglutarate (2OG)-dependent dioxygenases [102] and act on both DNA and RNA. FTO demethylates m6A through oxidative reactions that generate two intermediate hydroxymethyl6A and formyl m6A [103]. ALKBH5 catalyzes the direct removal of m6A, oxidizing the N-methyl group of the m6A site to a hydroxymethyl group, as its m6A demethylation function affects total RNA synthesis and mRNA export [104]. The "reader" proteins belong to the YTH domain family proteins, and in humans include YTHDF1–3 and YTHDC1–2. YTHDF1 stimulates mRNA translation to interact with eIFs and ribosomes. YTHDF2 binds to m6A, located in the 3'UTR leading mRNA to become processing bodies for degradation in a methylation-dependent manner [105]. While YTHDC1 protein binds to m6A, as well as pre-mRNA splicing factor SRSF3 to its mRNA-binding elements, they are close to m6A sites but block SRSF10 mRNA binding, promoting exon exclusion, and modulating mRNA splicing by recruiting pre-mRNA splicing

Damage to DNA is derived from replication stress, telomere shortening, UV light, chemical toxins, and ROS - but RNA can suffer alterations and respond to damage agents. This variety of DNA lesions is removed in cells by protein complexes in specific repair systems (**Figure 2**). Until recently, response to damage was unique to DNA, while analyzing the response of DNA to UV, with Xiang et al. [107] changing that view. They found an accumulation of m6A on poly(A) + RNA two minutes after UV irradiation in response to DNA damage. They observed that the methyltransferase complex (METTL3, METTL4, WTAP) and Poly(ADP-ribose) polymerase (PARP) is localized to sites of UV-induced damage, with FTO demethylase recruiting. DNA polymerase translesion (Pol κ) is necessary for METTL3 and METTL4 recruitment. The authors suggested that PARP, METTL3, m6A RNA, and Pol κ could be alternative repair pathways to respond to UV-induced damage, with m6A in the main role for rapid recruitment of Pol κ to damaged sites. Colocalization of Pol κ with m6A to sites with a high content of cyclobutene pyrimidine dimers (CPDs) corroborate that m6A RNAs have a regulatory role in the NER pathway. Svobodová et al. [108] found that m6A RNAs are diffuse to damaged DNA, but

light-induced antibody-RNA crosslinking and reverse transcription.

#### *Epigenetics and DNA Repair in Cancer DOI: http://dx.doi.org/10.5772/intechopen.94030*

*DNA - Damages and Repair Mechanisms*

modification during DNA damage. MGMT is an O6

**5. Genome damage repair via m6A participation**

melanoma [81].

of DOT1L function leads to faulty HRR-mediated DSB repair without affecting NHEJ. Highlighting DOT1L-mediated H3K79me3 importance in the early response to DNA damage and DSB repair, its inhibition also increases radiation sensitivity and chemotherapeutic agents in colorectal cancer patients' treatment. Dot1L and H3K79 methylation was previously associated with the role of 53Bp1 in response to DNA damage [79]. FitzGerald et al. [80] corroborated that not only H3K79me recruits 53BP1 to DNA damage sites, but also H4K20me. This indicates that H4K20me concentration is essential for the repair pathway related to 53BP1 and BRCA1, in cooperation with the 53BP1-RIF1-MAD2L2 complex. The histone posttranslational modification of H3K27me3 and H4K20me is a diagnostic indicator in

There is DNA methyltransferase, in which expression is regulated by epigenetic

transferase responsible for the repair of damaged guanine, without other cofactors transferring the methyl at 06-meG to cysteine residues, removing adducts in a single step, thus protecting chromosomes from mutations, carcinogenesis, and alkylating agents [82, 83]. MGMT can do this only once, after its DNA-binding domain change by alkylating, which is detached from DNA and targeted for degradation by ubiquitination [84]. Loss of MGMT expression is due to promoter methylation [85], used as an advantage for good prognosis in a glioma, which can predict whether alkylating agents can benefit treatment [86]. MGMT can interfere with TMZ response on tumor cells and is responsible for efficient repair of TMZ and induced toxic DNA adducts, reducing treatment efficacy. Targeting MGMT seems to overcome chemoresistance in gliomas, but the prognostic value of MGMT methylation is controversial, as genomic rearrangements result in MGMT overexpression, independent of its promoter methylation, contributing to resistance [87].

Modified nucleotides on DNA create a vast list, shown by Fragou et al. [88], but methylation by methyltransferases is an epigenetic modification that occurs on DNA and RNA. In fact, RNA contains more modified bases than DNA, as these RNA modifications are present in rRNA, tRNA, snRNA, miRNA, or mRNA, giving rise to the emergence of a wide chemical diversity on its side chains. One of these modifications on mRNA has been studied since its discovery in 1974 [89, 90], the methylation of adenosine residues form N6-methyladenosine (m6A) is the most abundant post-transcriptional mRNA modification, and is detected in approximately 25% of mRNAs [91]. The m6A modification mechanism of action functions like methylation on DNA and histones, i.e., its presence or absence on mRNA determines the fate of the transcription. The addition of a methyl group to the N6 site of A, occurring in the RRACH sequence [92, 93], has a distinctive position in the vicinity of stop codons, internal exons, in 5'UTRs which could promote 5'capindependent translation when found on the first nucleotide adjacent to the 7-methylguanosine cap; this gives protection from decapping [7, 94, 95] and in 3'UTRs regulates the affinity of RNA binding proteins [96]. This is known as m6A modification, which is involved in a variety of cellular processes including gene expression through regulating RNA metabolism, such as mRNA translation, degradation, alternative splicing, export and folding [97, 98], control of protein translation [94],

and others. All these effects are globally known as RNA epitranscriptomics.

For some time, m6A modification was considered static and unalterable, but is now well-known as reversible and dynamic. It is difficult to examine gene regulation at the RNA level without appropriate methods; genome-wide sequencing became a


**174**

major tool for many years, but had limitations with the m6A assay. The sensibility of the methodologies used was a critical point, because until recently, techniques had a detection limit of about 0.01%, in contrast to the content of m6A, less than 0.001% is sufficient to regulate biological processes [99]. Antibody-based, highthroughput sequencing technology allowed us for the past few years to locate the specific m6A sites and explore their biological significance. Linder et al. [91] solved the problem of distinguishing between m6A and adenosine by using incubation of an m6A-antibody to induce a specific mutational signature of m6A residues after UV light-induced antibody-RNA crosslinking and reverse transcription.

This modification is added and eliminated by proteins called "writers" and "erasers," respectively; these proteins regulate the abundance, prevalence, and distribution of m6A; this exerts its biological function and is modulated by protein "readers." First, m6A is added to the nucleus by the methyltransferase complex, which is formed by two core proteins, METTL3 and METTL4 [92]. METTL3/ METTL4 form a functional heterodimer, METTL3, which is the catalytic subunit which uses S-adenosylmethionine (SAM) or S-adenosylhomocysteine (SAH) as donors for methyl transfer in adenosine within the consensus motif G(G/A) ACU [100]. METTL4 has the active site blocked, and lacks amino acid residues to form hydrogen bonds with the ribose hydroxyls of SAM to facilitate donor and acceptor substrate binding; this is important in maintaining the integrity of the complex, stabilizing METTL3, and enabling RNA substrate recognition [101]. Regarding m6A demethylases or "eraser" proteins, FTO and ALKBH5 are two well-known m6A demethylases located in nuclear speckles within the methyltransferase complex. They belong to the group of Fe(II)/2-oxoglutarate (2OG)-dependent dioxygenases [102] and act on both DNA and RNA. FTO demethylates m6A through oxidative reactions that generate two intermediate hydroxymethyl6A and formyl m6A [103]. ALKBH5 catalyzes the direct removal of m6A, oxidizing the N-methyl group of the m6A site to a hydroxymethyl group, as its m6A demethylation function affects total RNA synthesis and mRNA export [104]. The "reader" proteins belong to the YTH domain family proteins, and in humans include YTHDF1–3 and YTHDC1–2. YTHDF1 stimulates mRNA translation to interact with eIFs and ribosomes. YTHDF2 binds to m6A, located in the 3'UTR leading mRNA to become processing bodies for degradation in a methylation-dependent manner [105]. While YTHDC1 protein binds to m6A, as well as pre-mRNA splicing factor SRSF3 to its mRNA-binding elements, they are close to m6A sites but block SRSF10 mRNA binding, promoting exon exclusion, and modulating mRNA splicing by recruiting pre-mRNA splicing factors [106].

Damage to DNA is derived from replication stress, telomere shortening, UV light, chemical toxins, and ROS - but RNA can suffer alterations and respond to damage agents. This variety of DNA lesions is removed in cells by protein complexes in specific repair systems (**Figure 2**). Until recently, response to damage was unique to DNA, while analyzing the response of DNA to UV, with Xiang et al. [107] changing that view. They found an accumulation of m6A on poly(A) + RNA two minutes after UV irradiation in response to DNA damage. They observed that the methyltransferase complex (METTL3, METTL4, WTAP) and Poly(ADP-ribose) polymerase (PARP) is localized to sites of UV-induced damage, with FTO demethylase recruiting. DNA polymerase translesion (Pol κ) is necessary for METTL3 and METTL4 recruitment. The authors suggested that PARP, METTL3, m6A RNA, and Pol κ could be alternative repair pathways to respond to UV-induced damage, with m6A in the main role for rapid recruitment of Pol κ to damaged sites. Colocalization of Pol κ with m6A to sites with a high content of cyclobutene pyrimidine dimers (CPDs) corroborate that m6A RNAs have a regulatory role in the NER pathway. Svobodová et al. [108] found that m6A RNAs are diffuse to damaged DNA, but

#### **Figure 2.**

*Regulatory complexes of m6A in DNA damage repair. The m6A mark on mRNAs is involved in some repair pathways in response to DNA damage. It has been observed that the interaction of writing, eraser and reader proteins with other proteins that are part of canonical repair pathways is essential for the response to damage. SAM: S-adenosylmethionine; UV: ultraviolet light, NER: nucleotide excision repair, HRR: homologous recombination repair, eIFS: eukaryotic initiation factors, phosphate group: red circle.*

a new participant, METTL16, accumulated 20–30 min after induced damage in a subset of irradiated cells. This response was specific to CPDs, as the authors observed that m6A RNAs' accumulation pattern was specific to repair of CPDs' sites, which do not accumulate in other lesions, such as NHEJ.

Apyrimidinic or apuric (AP) sites can arise by spontaneous hydrolysis, cleaving the N-glycosidic bond through elimination of an incorrect or damaged base by DNA glycosylases or ionizing radiation. Considered one of the most predominant lesions in the genome, it is repaired by the BER pathway, which has PARP-1 as the first sensor and responder, activated by poly(ADP-ribose) polymerase [1]. Recently, Xiang et al. [107] found it was associated with m6A in RNA. AP lyases cleave 5′ or 3′ to AP sites to further processing by DNA polymerase and ligase. ALKBH1 is another member of the AlkB family that participates in demethylation of histone H2A in mouse stem cells. It is capable of cleavage to DNA at AP sites, using a lyase mechanism to produce a DNA nick on the 3′ side of a basic site, leaving a product that is missing a 3′-phosphate and an adduct in the 5'-DNA product. ALKBH1 acts on both ss-DNA and ds-DNA and can produce DSBs related to AP lyase activity. Human ALKBH1 possesses m6A demethylation activity, although this is not its main function. It cannot be ruled out this has a role in epigenetic gene silencing. Due to its abundance in mitochondria, it could play a primary role in mitochondrial DNA repair and function [109–111]. Other AlkB, such as ALKBH3, function as RNA repair enzyme; it is equally distributed in cytosol, the nucleus, and is active on RNA and ssDNA [112]. Zhang et al. [113] showed how METTL3 is phosphorylated by ATM protein for its localization to DSBs, where it catalyzes m6A in RNAs. METTL3 stimulates the recruitment of RAD51 to DSBs in a DNA-RNA

**177**

*Epigenetics and DNA Repair in Cancer DOI: http://dx.doi.org/10.5772/intechopen.94030*

**6. Cancer drugs: resistance and epigenetics**

Wnt/β-catenin pathway in BRCA-deficient EOC cells.

immune-related adverse side effects, vs. with ipilimumab.

Xiang et al. [107] showed that FTO could be recruited to damaged γH2AX chromatin after irradiation, strengthening FTO participation in DNA damage repair. In melanoma, FTO is upregulated and promotes cell proliferation, cell migration, invasion, and cell viability. Yang et al. [121] demonstrated that FTO regulates PD-1 expression that also promotes mTOR signaling. FTO can promote resistance to anti PD-1 blockade in melanoma through m6A, mediating PD-1 (PD-1 or PDCD1 is a negative regulator of T-cell activity). This supports the anti PD-1 blockade with an anti-tumor response in advanced cancers and reduces

Glioblastoma is a common and aggressive primary brain tumor in adults, and is highly resistant to treatment such as surgery, irradiation, and adjuvant TMZ chemotherapy, which failed to improve the outcome. One cause of poor response to TMZ, as suggested by Visvanathan et al. [8], is that GSCs show high levels of m6A and METTL3, supporting the proposal that METTL3 is key in GSC maintenance, making those cells resistant to therapy and refractory to radiotherapy by efficient repair of DNA. Here, METTL3 alters the DNA repair efficiency and radiation sensitivity through m6A sites in SOX2–3'UTR, stabilizing it in GSC, as the recruitment of human antigen R (HuR) to m6A modified mRNA is crucial for SOX2 stabilization by METTL3. This supports a role for METTL3, shared with SOX2 in another repair pathway, mediated HR: SOX2 protects GSCs from radiation-induced cytotoxicity

chemo- and radiotherapy.

hybrid-dependent manner, promoting efficient homologous recombinationmediated DSB repair. The upregulation of METTL3 may contribute to resistance to

Alterations in m6A and proteins responsible for its regulation on the RNA level were shown to interfere with the response to cancer treatment. Cyclophosphamide (CTX) is an alkylating chemotherapy drug used in cancer treatment; it forms adducts at the N7-guanine position, which are unstable and therefore undergo spontaneous dissociation. These features are exploited in rapidly dividing cells, which are likely to be disrupted before repair takes place [114]. Little is known about CTX effect on RNA epigenetic complexes, yet it was shown to increase m6A levels, inhibiting the gene and protein expression of FTO, YTHDF1, YTHDF2, YTHDC1, and YTHDF3 in a time- and concentration-dependent manner [115]. Cervical squamous cell carcinoma (CSCC) is an example of increased chemotherapy resistance; Zhou and colleagues [116] found that β-catenin is an FTO target: they observed FTO overexpression and reduced m6A β-catenin levels, with the effect of this change upregulation of β-catenin protein and the subsequent activation of ERCC1, a critical player in NER, which contributes to chemotherapy resistance and a poor prognosis. In case of BRCA-mutated epithelial ovarian cancers (EOC), the use of Olaparib has been clinically beneficial: it is a poly(ADP-ribose) polymerase inhibitor (PARPi), which detects and binds DNA SSBs and DSBs, using the N-terminal DNA binding domain [117, 118]. When DNA is damaged, PARP-1 can recognize damaged sites and their formation; the binding exposes the enzymatic site of PARP-1, resulting in its activation and the recruitment of XRCC1, the first protein for assembly and activation of DNA bases excision repair machinery [119]. The resistance of tumor cells to PARPi is not well-known, but in the study of Fukumoto and colleagues [120], FZD10 was observed as a receptor in Wnt/β-catenin signaling, increasing m6A modification of mRNA in resistant cells, thus stabilizing. This increase contributes to PARPi resistance by upregulating the

*DNA - Damages and Repair Mechanisms*

a new participant, METTL16, accumulated 20–30 min after induced damage in a subset of irradiated cells. This response was specific to CPDs, as the authors observed that m6A RNAs' accumulation pattern was specific to repair of CPDs'

*Regulatory complexes of m6A in DNA damage repair. The m6A mark on mRNAs is involved in some repair pathways in response to DNA damage. It has been observed that the interaction of writing, eraser and reader proteins with other proteins that are part of canonical repair pathways is essential for the response to damage. SAM: S-adenosylmethionine; UV: ultraviolet light, NER: nucleotide excision repair, HRR: homologous* 

Apyrimidinic or apuric (AP) sites can arise by spontaneous hydrolysis, cleaving the N-glycosidic bond through elimination of an incorrect or damaged base by DNA glycosylases or ionizing radiation. Considered one of the most predominant lesions in the genome, it is repaired by the BER pathway, which has PARP-1 as the first sensor and responder, activated by poly(ADP-ribose) polymerase [1]. Recently, Xiang et al. [107] found it was associated with m6A in RNA. AP lyases cleave 5′ or 3′ to AP sites to further processing by DNA polymerase and ligase. ALKBH1 is another member of the AlkB family that participates in demethylation of histone H2A in mouse stem cells. It is capable of cleavage to DNA at AP sites, using a lyase mechanism to produce a DNA nick on the 3′ side of a basic site, leaving a product that is missing a 3′-phosphate and an adduct in the 5'-DNA product. ALKBH1 acts on both ss-DNA and ds-DNA and can produce DSBs related to AP lyase activity. Human ALKBH1 possesses m6A demethylation activity, although this is not its main function. It cannot be ruled out this has a role in epigenetic gene silencing. Due to its abundance in mitochondria, it could play a primary role in mitochondrial DNA repair and function [109–111]. Other AlkB, such as ALKBH3, function as RNA repair enzyme; it is equally distributed in cytosol, the nucleus, and is active on RNA and ssDNA [112]. Zhang et al. [113] showed how METTL3 is phosphorylated by ATM protein for its localization to DSBs, where it catalyzes m6A in RNAs. METTL3 stimulates the recruitment of RAD51 to DSBs in a DNA-RNA

sites, which do not accumulate in other lesions, such as NHEJ.

*recombination repair, eIFS: eukaryotic initiation factors, phosphate group: red circle.*

**176**

**Figure 2.**

hybrid-dependent manner, promoting efficient homologous recombinationmediated DSB repair. The upregulation of METTL3 may contribute to resistance to chemo- and radiotherapy.
