Endogenous DNA Damage as a Source of Genomic Instability in Cancer

Abstract

Genome instability, defined as higher than normal rates of mutation, is a double-edged sword. As a source of genetic diversity and natural selection, mutations are beneficial for evolution. On the other hand, genomic instability can have catastrophic consequences for age-related diseases such as cancer. Mutations arise either from inactivation of DNA repair pathways or in a repair-competent background due to genotoxic stress from celluar processes such as transcription and replication that overwhelm high-fidelity DNA repair. Here, we review recent studies that shed light on endogenous sources of mutation and epigenomic features that promote genomic instability during cancer evolution.

Keywords: DNA damage; DNA repair; DNA replication; Genome instability; cancer; mutagenesis; transcription.

Figure 1.

Estimated Frequencies of DNA Lesions and Mutations Associated with Dysfunctional DNA Repair

Figure 2.. DNA Repair Pathways and Mutations Associated with Dysfunctional Repair

(A) Spontaneous deamination of 5-methylcytosine (C*) generates an aging signature, characterized by CpG>TpG transition mutations. 5-methylcytosine deamination converts C>T, which is recognized by thymine DNA glycosylase (TDG). Apurinic/apyrimidinic endodeoxyribonuclease (APE) excises the abasic nucleotide, which is replaced by polymerase beta (POLβ). (B) Reactive oxygen species (ROS) oxidizes guanine to form 8-oxoG. 8-oxoG can pair with adenine and may generate G>T mutations if 8-oxoG:C pairing is not detected by OGG1 before replication or if the 8-oxoG:A pairing is not recognized by MUTYH after the first round of replication. (C) Exposure to UV light generates cyclobutane pyrimidine dimers, which are recognized by nucleotide excision repair (NER) factors and excised by XPG and XPF flap endonucleases. Polymerase delta (POLδ) synthesizes DNA to restore C:G pairing. Failure to complete NER results in C>T transition mutations at CC dinucleotides. (D) DNA polymerase errors induce mismatch repair (MMR) through MSH2/MSH6. Exonuclease I (EXO1) removes a short patch of DNA that is resynthesized by POLδ. Failure to complete MMR results in base substitutions. (E) DNA DSBs are repaired using non-homologous end joining (NHEJ) or homologous recombination (HR). Normal NHEJ may result in small insertions or deletions, as does an inherently error-prone form of end-joining (alternative end joining), which relies on short homologous sequences (microhomologies) for base pairing. Normal HR is an error-free repair mechanism, but abortive HR may yield chromosomal rearrangements, such as tandem duplications or large deletions, which are common in HR-deficient cancers. BRCA1 and CtIP promote DNA end resection during HR, whereas BRCA2 promotes strand invasion to facilitate normal HR.

Figure 3.. Causes and Consequences of Enzyme-Induced Cytosine Deamination

(A) ssDNA exposed during replication fork progression is a substrate for cytosine deamination. Active replication forks expose ssDNA on the lagging strand. Stalled replication forks expose ssDNA on both the leading and lagging strand, which activates ATR to stop replication and stabilize the fork through controlled fork reversal. Endonuclease cleavage causes replication fork collapse into one-ended DNA DSBs, which must be resected before repair by HR. (B) ssDNA exposed during transcription is a substrate for cytosine deamination. RNA polymerase (RNAP) exposes ssDNA while synthesizing nascent RNA (red) at the transcription bubble and behind the RNAP as a result of negative supercoiling (left). Transcription stress induces the formation of R-loops (middle), which exposes long stretches of ssDNA on the non-template strand. R-loops may be resolved in DSBs through cleavage by TC-NER components XPF and XPG (right). (C) Mechanism of enzyme-induced somatic hypermutation (SHM). Cytosine deamination generates a U:G mismatch, which is recognized by uracil glycosylase UNG to generate an abasic site opposite the G. Apurinic/apyrimidinic endodeoxyribonuclease (APE) excises the abasic nucleotide which is repaired by POLb. If the abasic site is replicated over prior to repair, this can give rise to transition or transversion mutations (N). If the U:G mismatch is replicated prior to repair, C>T substitution occurs. Mutations at neighboring A:T base pairs can occur if the U:G pair is recognized by the MMR pathway and repaired by error prone polymerase eta (Polη). (D) Mechanisms of enzyme-induced rearrangements and class-switch recombination (CSR). If cytosine deamination occurs nearby on opposite strands, repair through BER or MMR will form DNA DSBs, leading to chromosomal rearrangements or CSR.

Figure 4.. Replication Timing and Protein-DNA Interactions Influence Regional Mutation Rates

(A) Early replicating regions of the genome are associated with high levels of H3K4me3, transcription, increased chromatin accessibility, and low levels of H3K9me3. (B) In cancer genomes, accessible early replicating regions have fewer base-pair substitutions. MMR is more likely to recognize mismatches in early replication regions. NER is also more efficient in transcribed, accessible chromatin. (C) DNA accessibility determines efficiency of NER (adapted from Sabarinathan et al., 2016). Repair of UV-induced lesions is efficient within DNase I hyper-sensitive sites (DHSs), but is impaired by nucleosome occupancy and transcription factor binding. Conversely, UV-induced mutations are more frequent where DNA-binding proteins interfere with NER machinery. (D) Whereas DSBs are more efficiently repaired in accessible chromatin, chromosome rearrangements in cancer genomes accumulate at early replicating regions.

Figure 5.. Canonical and Non-canonical Topoisomerase Activity and Gene Activation

(A) Upon gene activation, canonical topoisomerase activity promotes gene expression. The enzymes TOP1 and TOP2 cleave one or both strands to alleviate positive supercoiling ahead of and negative supercoiling behind RNA polymerase (Pol II). ssDNA behind Pol II can be a substrate for cytosine deamination. (B) Non-canonical, abortive TOP2 reactions result in protein-linked DNA breaks that are resolved the NHEJ repair pathway. Abortive TOP2 activity also promotes transcription by an unknown mechanism. Persistent DNA damage may also result in chromosomal alterations.

Figure 6.. Replication and Transcription Stress Contribute to the Mutational Landscape of Cancer

An emerging model for how mitogenic signaling promotes genome instability at early replicating fragile sites (ERFSs). Oncogenes, hormones, and inflammation can promote unscheduled activation of replication and transcription programs. This can result in replication stress and transcription stress through mechanisms that are largely unknown. The generation of ssDNA and DSBs at ERFSs can lead to genomic instability.