All information required for the life of a cell or organism is encoded in its DNA. As DNA is heritable, it also encodes information that shapes how populations of related organisms change over time. As such, the capacity for high-fidelity DNA replication is a requirement for the genomic stability of organisms and populations. DNA is constantly exposed to environmental and endogenous causes of lesions that make high-fidelity replication of DNA difficult. DNA repair is therefore necessary for the viability of life, and failure to repair damage accurately or rapidly can have catastrophic consequences. This strong evolutionary pressure has produced a variety of DNA repair pathways that can have mutagenic effects upon the genomes of cells and organisms. These changes can be harmful to the survival or health of the lifeform, but also consist of the basis of evolution that allows populations to adapt to changes in their environment with overall positive effects on long-term survival. Error-prone DNA repair can be thought of as a double-edged sword that requires tight regulation. This essay will focus on the positive and negative consequences of mutagenic DNA repair on cellular, organismal, and population scales.
In humans, cells are estimated to experience on average 50,000 DNA lesions per cell per day. The accumulation of DNA damage, and mutations resulting from inaccurate repair, can lead to cellular apoptosis, senescence, or uncontrolled proliferation. One of the most common forms of DNA lesion is the formation of cyclobutane pyrimidine dimers (CPDs). CPDs are the result of UV-light catalysed covalent linkage between adjacent cysteine or thymine residues. They are repaired in cells by the nucleotide excision repair (NER) pathway; often CPDs are detected via transcription-coupled repair, an NER sub-pathway in which the stalling of a transcribing RNA polymerase II on the CPD results in the non-allosteric recruitment of NER factors. Eukaryotic cells also detect UV lesions by testing for the presence of unpaired bases by contorting DNA, mediated by the XPC-hHR23B-centrin 2 complex. Once this complex binds to the lesion, TFIIH, XPA and RPA proteins are recruited. The TFIIH complex unwinds the DNA to generate a single-strand bubble around the CPD, which is excised by DNA nucleases XPF/XPG. The resulting gap is filled in by a high-fidelity DNA polymerase and sealed by DNA ligase.
While NER is a highly accurate repair pathway, CPDs that aren’t detected or repaired quickly enough can have mutagenic consequences during DNA replication. High-fidelity replicative polymerases stall when encountering CPDs because the active site of the DNA polymerase is only large enough to accommodate a single nucleotide at a time, and is incapable of ‘reading’ the pyrimidine dimer. Stalling risks replication fork collapse and cytotoxic double-strand breaks (DSBs) in the DNA. To prevent this, cells can use low-fidelity trans-lesion DNA synthesis (TLS) to allow the replication fork to overcome the CPD. The molecular switch that determines the repair pathway used is PCNA. The high-fidelity polymerase dissociates from the DNA and Rad6/Rad18 proteins are recruited. Rad6/18 monoubiquitylate PCNA on Lys164. TLS factors such as DNA Pol η have high affinity for monoubiquitylated PCNA, and so are recruited to the lesion. The bypass polymerase Pol η has a large active site capable of accommodating the CPD, at the cost of low-fidelity base-pairing. DNA Pol η also lacks the capacity to proof-read DNA, resulting in increased frequency of mutation. Often these mutations are benign, but can result in the loss of critical protein/gene function depending on the location and quantity of CPDs, ultimately inducing cell death.
TLS is also necessary for correct functioning of B-cells in the immune system, allowing generation of high-affinity antibodies via somatic hypermutation (SHM). AID-catalysed base deamination and subsequent repair by TLS polymerases enables rapid mutation in the immunoglobulin variable region DNA of proliferating B-cells. This highlights the positive consequences of inaccurate DNA repair in enabling cells to fulfil their functions within the eukaryotic immune system.
Failure to repair CPDs by TLS can lead to replication fork collapse and DSB formation. DSBs contribute to the accumulation of genetic mutations in somatic cells, leading to aging and pathologies in organisms. They can also lead to major chromosomal structural abnormalities in daughter cells if they are left unrepaired, a driver of carcinogenesis. DSB repair pathways are typified by the homologous recombination (HR) and non-homologous end-joining (NHEJ) pathways. NHEJ is an inherently mutagenic repair pathway that can repair DNA at any point in the cell cycle. The Ku70/80 heterodimer threads onto either end of DNA at the site of the DSB and forms the DNA-PK bridging complex. This complex holds the ends of the DNA close together and recruits break-processing and ligation factors. The exonuclease and template-independent polymerase activity of the break-processing factors iteratively trim and expand the DNA ends to generate regions of microhomology. Once generated, these regions are composed of random sequences of DNA and are joined by DNA ligase. Depending on the location of the DSB, NHEJ repair can result in loss-of-function mutations in critical genes, including those coding DNA repair proteins. This can result in reduced repair efficiency, and the accumulation of more DNA lesions and mutations. This is the major driver of aging due to the onset of chronic cell senescence in tissues throughout an organism, and is a particular problem when progenitor stem cells enter senescence, limiting tissue regeneration.
Endogenous DNA lesions can also result in mutations during cell replication if they aren’t accurately repaired. Reactive oxygen species (ROS) produced by aerobic respiration or other metabolic pathways can result in various types of DNA lesions. The most damaging ROS is the hydroxyl radical, produced from the reaction of H2O2 with Fe2+. Hydroxyl radicals can react with guanine bases to produce 8-oxo-guanine (8-oxo-G). 8-oxo-G is capable of incorrectly pairing with adenine via Watson-Crick base-pairing. These lesions are usually repaired via the base excision repair (BER) pathway, but failure to do so before DNA replication can result in the mutation of GC to AT base pairs in daughter cells. As the DNA strands are separated during DNA replication, the incorrect pairing of 8-oxo-G with adenine will result in the production of a DNA molecule containing an AT base pair at the site of the original GC. This means that daughter cells produced by mitosis can contain heritable mutations that will be passed on exponentially to their offspring. The accumulation of mutations can have pathological consequences for the organism; for example, they can promote carcinogenesis if they result in the activation of proto-oncogenes, or drive the aberrant expression of oncogenic RNAs such as oncomir-1 miRNA. This shows that mutations can cause pathologies within an organism.
Mutations arising from inaccurate DNA repair can be passed onto the next generation in a population. This can have negative consequences for the long-term survival of the population, as in the case of hereditary disease. The Founder effect can result in a high frequency of hereditary disease amongst organisms of small populations due to inbreeding between closely related individuals. This has been noted in the Afrikaner population in South Africa, which exhibits a high frequency of a mutant Htt allele, responsible for Huntington’s disease. The mutant allele is formed by trinucleotide repeat expansion, a DNA mutation that can occur as a result of DNA strand slippage during HR, NHEJ, and BER repair pathways. Htt encodes a CAG trinucleotide repeat that varies between 7-35 repeats. When DNA polymerase encounters a direct repeat during repair it pauses and transiently dissociates from the DNA, allowing the nascent DNA strand to unpair from the template and re-anneal at a different repeat upstream of the site of separation. The polymerase then re-assembles on the template strand, but in doing so backtracks and re-inserts the CAG trinucleotide that was already added. This expands the trinucleotide repeat region on the nascent strand compared to the template strand, meaning the strands no longer anneal correctly. This is corrected by the NER pathway by either deletion of the inserted repeat on the nascent strand, or insertion of an extra repeat on the template strand. The increase in number of CAG trinucleotide repeats above 35 results in the Huntington’s disease phenotype. Conversely, heritable mutations also allow the proliferation of beneficial phenotypic changes throughout a species by natural selection. This can occur through intergenerational changes, but also by horizontal gene transfer in bacterial populations. Antibiotic resistance in bacteria is acquired through mutations arising from the incorrect repair of DNA lesions. These mutations confer a survival advantage to the bacterium by making it resistant to an antibiotic, meaning that the offspring of the bacterium are more likely to survive and reproduce than antibiotic-susceptible bacteria in the population. Over time, the allelic frequency of the antibiotic resistance mutation will increase in the population as a result of the evolutionary pressure. Furthermore, bacterial conjugation allows the transfer of DNA in the form of plasmids between bacteria. This allows the antibiotic resistance allele to be transferred to genetically different bacteria in the same population. This highlights how heritable mutations can have positive and negative consequences on the survival of populations.
Newscleus 