Mutational Strand Asymmetries in Cancer Genomes Reveal Mechanisms of DNA Damage and Repair

Haradhvala, Nicholas J.; Polak, Paz; Stojanov, Petar; Covington, Kyle R.; Shinbrot, Eve; Hess, Julian M.; Rheinbay, Esther; Kim, Jaegil; Maruvka, Yosef E.; Braunstein, Lior Z.; Kamburov, Atanas; Hanawalt, Philip C.; Wheeler, David A.; Koren, Amnon; Lawrence, Michael S.; Getz, Gad

doi:10.1016/j.cell.2015.12.050

Cited by 386 publications

(555 citation statements)

References 67 publications

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“…The pattern of repair, however, suggests that mutations originated from the nontemplate strand. Higher mutation levels on the nontranscribed strand would be consistent with reports of transcription-based strand-asymmetries in mutagenesis (21,39).…”

Section: Slow Repair Is Associated With Higher Somatic Mutagenesis Insupporting

confidence: 78%

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Genome-wide kinetics of DNA excision repair in relation to chromatin state and mutagenesis

Adar

Lieb

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

154

226

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We recently developed a high-resolution genome-wide assay for mapping DNA excision repair named eXcision Repair-sequencing (XR-seq) and have now used XR-seq to determine which regions of the genome are subject to repair very soon after UV exposure and which regions are repaired later. Over a time course, we measured repair of the UV-induced damage of cyclobutane pyrimidine dimers (CPDs) (at 1, 4, 8, 16, 24, and 48 h) and (6-4)pyrimidinepyrimidone photoproducts [(6-4)PPs] (at 5 and 20 min and 1, 2, and 4 h) in normal human skin fibroblasts. Each type of damage has distinct repair kinetics. The (6-4)PPs are detected as early as 5 min after UV treatment, with the bulk of repair completed by 4 h. Repair of CPDs, which we previously showed is intimately coupled to transcription, is slower and in certain regions persists even 2 d after UV irradiation. We compared our results to the Encyclopedia of DNA Elements data regarding histone modifications, chromatin state, and transcription. For both damage types, and for both transcription-coupled and general excision repair, the earliest repair occurred preferentially in active and open chromatin states. Conversely, repair in regions classified as "heterochromatic" and "repressed" was relatively low at early time points, with repair persisting into the late time points. Damage that remains during DNA replication increases the risk for mutagenesis. Indeed, laterepaired regions are associated with a higher level of cancer-linked mutations. In summary, we show that XR-seq is a powerful approach for studying relationships among chromatin state, DNA repair, genome stability, mutagenesis, and carcinogenesis.DNA repair | DNA damage | chromatin | transcription | mutation D NA damage blocks transcription and replication and compromises the integrity of the genome. Bulky adducts in DNA, the focus of this work, are caused by a variety of genotoxic agents including UV radiation in sunlight. In human cells, this damage is removed exclusively by the nucleotide excision repair mechanism (1-3). In nucleotide excision repair, a single-stranded oligomer that contains the bulky adduct is excised by dual incisions bracketing the lesion. The resulting gap is filled in by DNA polymerases, and the newly synthesized repair patch is then sealed by DNA ligase. In general excision repair, damage recognition is achieved by the repair factors xeroderma pigmentosum (XP) complementation group C (XPC) together with replication protein A (RPA) and XPA (4-6). In DNA that is being actively transcribed, damage recognition can also be achieved by a stalled RNA polymerase II, which, with the aid of the Cockayne Syndrome B (CSB) protein, accelerates the recruitment of the excision repair factors. This recognition process leads to transcription-coupled repair (7,8). The subsequent dual-incision reaction is carried out by the core excision repair factors RPA, XPA, transcription factor II H (TFIIH), XPG, and XPF-ERCC1, releasing an excised oligomer that is 24-32 nucleotides (nt) in length (6, 9).The mechanism of DNA...

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Section: Slow Repair Is Associated With Higher Somatic Mutagenesis Insupporting

confidence: 78%

“…DNA replication may also affect repair efficiencies and, in addition, has been implicated in mutagenesis strand asymmetries (39). Future work in synchronized cells will be able to address the effect of replication timing on DNA repair efficiencies.…”

Section: Slow Repair Is Associated With Higher Somatic Mutagenesis Inmentioning

confidence: 99%

Genome-wide kinetics of DNA excision repair in relation to chromatin state and mutagenesis

Adar

Lieb

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

154

226

View full text Add to dashboard Cite

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“…The data presented here suggest that the LGST at the replication forks of rapidly dividing cancer cells would be accessible to these enzymes, and hence the cytosines mutated in cancer genomes should be found preferentially in the LGST compared with the LDST. Recent results from whole-genome sequencing of mutations in yeast expressing APOBEC3A or APOBEC3B (43) and in 590 human tumors (44) found a strand bias in mutations consistent with this prediction.…”

mentioning

confidence: 60%

Strand-biased cytosine deamination at the replication fork causes cytosine to thymine mutations in Escherichia coli

Bhagwat

Hao

Townes

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The rate of cytosine deamination is much higher in single-stranded DNA (ssDNA) than in double-stranded DNA, and copying the resulting uracils causes C to T mutations. To study this phenomenon, the catalytic domain of APOBEC3G (A3G-CTD), an ssDNA-specific cytosine deaminase, was expressed in an Escherichia coli strain defective in uracil repair (ung mutant), and the mutations that accumulated over thousands of generations were determined by whole-genome sequencing. C:G to T:A transitions dominated, with significantly more cytosines mutated to thymine in the lagging-strand template (LGST) than in the leading-strand template (LDST). This strand bias was present in both repair-defective and repair-proficient cells and was strongest and highly significant in cells expressing A3G-CTD. These results show that the LGST is accessible to cellular cytosine deaminating agents, explains the well-known GC skew in microbial genomes, and suggests the APOBEC3 family of mutators may target the LGST in the human genome.uracil-DNA glycosylase | APOBEC3A | APOBEC3B | kataegis | cancer genome mutations P airing of complementary DNA strands protects the DNA bases against modification by a number of hydrolytic, oxidizing, and alkylating chemicals (1-4). For example, water reacts with cytosine, creating uracil, and the rate of this reaction in single-stranded DNA (ssDNA) is more than 100-fold the rate in double-stranded DNA [dsDNA (5-7)]. Uracil-DNA glycosylase (Ung) excises uracils created by cytosine deamination in both ssDNA and dsDNA, resulting in abasic (AP) sites. In dsDNA, the AP sites are replaced with cytosines as a result of copying of the guanine in the complementary strand during repair by the base-excision repair (BER) pathway (8). In contrast, cytosine deaminations occurring in ssDNA are problematic because the complementary strand is not available to the BER pathway. Uracils that escape repair create C:G to T:A mutations, and incomplete repair of uracils can result in persistent AP sites and strand breaks that can destabilize the genome. Hence, identifying ssDNA regions that are susceptible to damage will increase our understanding of causes of mutations and genome instability.The AID/APOBEC (activation-induced deaminase/apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like) family of DNA-cytosine deaminases are specific for ssDNA (9, 10). They are found only in vertebrates and are good probes of ssDNA in cells because of their relatively small size (about 190-amino acid catalytic domain). They are active in heterologous hosts such as Escherichia coli (11, 12) and yeast (13-15) and cause mutations in the same sequence context as in their known targets, such as Ig genes and the DNA copy of HIV-1 genome (11,16,17). In particular, the catalytic domain of human APOBEC3G (A3G-CTD) was expressed in an engineered yeast strain lacking the UNG gene and was shown to target ssDNA generated through aberrant resection of telomeric ends (13).To similarly probe ssDNA in E. coli, we expressed A3G-CTD on a plasmid (pA3G-CTD) in...

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“…This has been demonstrated by ectopic expression of human APOBEC3 enzymes in model organisms as well as analysis of cancer genomes. [30][31][32][33] Although these studies suggest ssDNA at replication forks is vulnerable to deamination by APOBECs, this has not been established experimentally in mammalian cells.…”

Section: Introductionmentioning

confidence: 92%

“…23 Additionally, expression of A3A and A3B in yeast can produce clusters of break-associated mutations indicative of deamination of resected DSBs. 22 The ssDNA at replication forks has been suggested as an additional substrate that is susceptible to APOBEC3 deamination based on recent data from genome sequencing analyses 32,33 and model organism systems. 30,31 These studies point to the potential for APOBEC3 enzymes to deaminate various cellular substrates, however the capacity for A3A to damage ssDNA during replication has not been previously demonstrated in mammalian cells.…”

Section: Discussionmentioning

confidence: 99%

APOBEC3A damages the cellular genome during DNA replication

et al. 2016

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The human APOBEC3 family of DNA-cytosine deaminases comprises 7 members (A3A-A3H) that act on single-stranded DNA (ssDNA). The APOBEC3 proteins function within the innate immune system by mutating DNA of viral genomes and retroelements to restrict infection and retrotransposition. Recent evidence suggests that APOBEC3 enzymes can also cause damage to the cellular genome. Mutational patterns consistent with APOBEC3 activity have been identified by bioinformatic analysis of tumor genome sequences. These mutational signatures include clusters of base substitutions that are proposed to occur due to APOBEC3 deamination. It has been suggested that transiently exposed ssDNA segments provide substrate for APOBEC3 deamination leading to mutation signatures within the genome. However, the mechanisms that produce single-stranded substrates for APOBEC3 deamination in mammalian cells have not been demonstrated. We investigated ssDNA at replication forks as a substrate for APOBEC3 deamination. We found that APOBEC3A (A3A) expression leads to DNA damage in replicating cells but this is reduced in quiescent cells. Upon A3A expression, cycling cells activate the DNA replication checkpoint and undergo cell cycle arrest. Additionally, we find that replication stress leaves cells vulnerable to A3A-induced DNA damage. We propose a model to explain A3A-induced damage to the cellular genome in which cytosine deamination at replication forks and other ssDNA substrates results in mutations and DNA breaks. This model highlights the risk of mutagenesis by A3A expression in replicating progenitor cells, and supports the emerging hypothesis that APOBEC3 enzymes contribute to genome instability in human tumors.

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Mutational Strand Asymmetries in Cancer Genomes Reveal Mechanisms of DNA Damage and Repair

Cited by 386 publications

References 67 publications

Genome-wide kinetics of DNA excision repair in relation to chromatin state and mutagenesis

Genome-wide kinetics of DNA excision repair in relation to chromatin state and mutagenesis

Strand-biased cytosine deamination at the replication fork causes cytosine to thymine mutations in Escherichia coli

APOBEC3A damages the cellular genome during DNA replication

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