Causes of DNA Damage and Genomic Instability: A Review

ABSTRACT


INTRODUCTION 1.1 Telomeric DNA (tDNA)
Telomeric DNA (tDNA) is a segment of DNA that occurs at the ends of linear eukaryotic chromosomes in eukaryotic cells (Chow et al., 2018). They are made up of repeated segments of DNA (tandem repeats) that consist of the sequence 5′-TTAGGG-3′ [in which T, A, and G are the bases thymine, adenine, and guanine, respectively] (Rodrigues and Lydall 2018a). It plays an essential role in maintaining the integrity and stability of the genome (Dewhurst et al., 2021 andSixtus A. Okafor, et al., 2022), as they cap the end sequences of the chromosomes (Maciejowski et al., 2021). It is non linear and serves to protect the vulnerable ends of the chromosomes from degradation (Markiewicz-Potoczny et al., 2021), functions of the DNA repair system and are susceptible to oxidative DNA damage (Ergünm, andSahin 2010 andYang et al., 2020). tDNA was discovered by Elizabeth Blackburn and colleagues In 1975In -1977. They are non coding, but act as buffer in protecting the coding sequences further behind (Rodrigues, Banks and Lydall 2018), by differentiating them from the DNA double-strand breaks (DSBs) (Schmutz et al., 2020) and also protecting them against homologous recombination (HR) and non-homologous end joining (NHEJ) (Guo et al., 2011 andTorrance and.). (Nature Education)

Genomic instability
Genomic instability is the term used by geneticists to refer to a high frequency or probability of inheritable changes known as mutations occurring within the genome of an organism (Betlem et al., 2018). These changes may involve rearrangement of base pairs, changes in nucleic acid sequences, chromosomal rearrangements or aneuploidy (Wojcicki et al., 2018 (Takai et al, 2003). Since each protein or enzyme in the replisome must perform its function well so as to result in a perfect copy of the DNA, then mutations of proteins such as DNA polymerase, ligase, can lead to impairment of replication and lead to spontaneous chromosomal exchanges (Rodrigues and Lydall 2018b). Proteins such as TEL1, MEC1 (ATR, ATM in humans) can detect single and doublestrand breaks and recruit factors such as Rmr3 helicase to stabilize the replication fork in order to prevent its collapse. Therefore, mutations in tel1, mec1, and rmr3 helicase is known to result in a significant increase in DNA damage occasioned by chromosomal recombination (Betlem et al., 2018 and Rodrigues and Lydall 2018b).

Fragile sites
There are locations in the genome where DNA sequences are prone to gaps and breaks after inhibition of DNA synthesis such as in the checkpoint arrest (Takai et al, 2003 andDemin et al., 2021). These sites are called fragile sites, and can occur commonly and naturally. It is present in most mammalian genomes or occurs rarely as a result of mutations, such as DNA-repeat expansion (Garcia and Sanchez-Puerta 2021). These rare fragile sites can lead to genetic disease such as fragile X mental retardation syndrome (

The 'end replication problem'
During cell division cycle, the DNA is copied and the chromosomes duplicated (Sixtus et al., 2022). If a cell's chromosome lacks telomere, the cell will loose its chromosomal end and genomic integrity in a phenomenon called "end replication problem". This end replication problem occurs because the end of linear DNA cannot be replicated completely during replication of the lagging strand at DNA synthesis (Ohki et al., 2001 andRusso et al., 2021) leading to telomere attrition (Olovnikov, 1973 andCharifi et al., 2021). During DNA synthesis, the leading strand is synthesize completely, while the lagging strand is gradually truncated at the ̴ 500-bp with the 3' overhang left behind (Ohki et al., 2001and Errichiello et al., 2020. Although, the extent of the end replication problem during DNA synthesis is poorly understood, it is thought, that the telomere shortening in telomerase-negative cells are resulted from the end replication problem. The end replication problem has been suggested to occur as a result of the priming failure of the okazaki fragments at the extreme end, and/or the failure of the most distal RNA primer to be replaced by DNA (Ohki et al., 2001 andRusso et al., 2021). Unlike prokaryotes, eukaryotes possess linear chromosome, and their DNA is replicated bi-directionally. The inability to replicate completely the terminal region of the lagging strand occupied by the Okazaki fragments (Sixtus et al., 2022), could lead to loss of terminal sequences and genetic information, following each cell division, and can induce loss of cell viability (Ohki et al., 2001 andSixtus et al., 2022). The end replication problem is also thought to explain the reason why somatic cells stop replicating after a number of cell replication (Olovnikov, 1973), observe the hayflick effect (Russo et al., 2021); undergo senescence and subsequently apoptosis (Olovnikov, 1973 andJamieson et al., 2021). There are various theoretical models to the end replication problem. The first model suggests that the synthesis of DNA by polymerase from 5' to 3' should have not only a catalytic site, but also a binding site in front of the catalytic site (Russo et al., 2021). This site will enable the attachment of enzyme to the parent DNA strand, such that DNA polymerase moving in front of the biding site during DNA replication will dissociate, as it has nowhere to bind, creating the end problem (Olovnikov, 1973 andJamieson et al., 2021). The second model, however, stressed the inability of the polymerase to begin new DNA synthesis itself, rather it is capable of elongating already existing oligonucleotide (Lie et al., 2018). However, recent data from the artificially created linear chromosome of the SV40 virus, have shown that in vitro, the leading strand was completely synthesised to the 5' end  ., 2021). The Apollo nucleases is believed to play a leading role, as a mutation or interference in the gene coding the Apollo nucleases Roy et al., 2021) and have been shown to lead to loss of the 3' overhang, induce loss of cell viability, senescence and apoptosis (Shi Y., Hellinga and Beese 2017 and Edera et al., 2021). The over hangs constitutes the telomeric loops (t-loops), which protects the telomeric DNA architecture from been recognize as double strand breaks (DSBs) which will initiate check point function and DNA repairs Roy et al., 2021). The architecture and profile of the end of linear chromosomes such as eukaryotic chromosomes, the overhangs, mimics the DNA double strand breaks (DSB) (Roy et al., 2021); and could be recognize as DSB by the DNA repair mechanisms (Sandell andZakian, 1993 andRoy et al., 2021). This recognition could activate checkpoint function and DNA repair pathways, which could degrade the end of the chromosomes, triggering cell arrest and genomic instability (Sandell andZakian, 1993 andDe Lange, 2010). They could also be processed by nucleases for repair either by homologous recombination (HR) or non-homologous end joining (NHEJ) (Sandell and Zakian,1993) thereby, initiating cell cycle arrest, with cell undergoing senescence (Roy et al., 2021) and apoptosis (Sandell andZakian, 1993 andRoy et al., 2021). This risk to the physical ends of the chromosomes occasioned by the difficulty of the cell to distinguish it's chromosomes natural ends from the DNA DSBs in the genome is known as the 'end protection problem' (Sandell and Zakian 1993

Transcription-associated instability
In both E. coli and Saccromyces pombe, transcription sites tend to have higher recombination and mutation rates. The coding or non-transcribed strand accumulates more mutations than the template strand (Gandini et al., 2019). This is due to the fact that the coding strand is single-stranded during transcription, which is chemically more unstable than doublestranded DNA. During elongation of transcription, supercoiling can occur behind an elongating RNA polymerase, leading to single-stranded breaks (Timashev and De Lange 2020). When the coding strand is singlestranded, it can also hybridize with itself, creating DNA secondary structures that can compromise replication . In E. coli, when attempting to transcribe GAA triplets such as those found in Friedrich's ataxia, the resulting RNA and template strand can form mismatched loops between different repeats (Gandini et al., 2019), leading the complementary segment in the coding-strand available to form its own loops which impede replication. Furthermore, replication of DNA and transcription of DNA are not temporally independent (Gandini et al., 2021); they can occur at the same time and lead to collisions between the replication fork and RNA polymerase complex. In S. cerevisiae, Rrm3 helicase is found at highly transcribed genes in the yeast genome, which is recruited to stabilize a stalling replication fork as described above. This suggests that transcription is an obstacle to replication (Holmes et al., 2020), which can lead to increased stress in the chromatin spanning the short distance between the unwound replication fork and transcription start site (Garcia et