scholarly journals Inherited MUTYH mutations cause elevated somatic mutation rates and distinctive mutational signatures in normal human cells

2021 ◽  
Author(s):  
Philip S Robinson ◽  
Laura E. Thomas ◽  
Federico Abascal ◽  
Hyunchul Jung ◽  
Luke Harvey ◽  
...  
Keyword(s):  
Excision Repair ◽  
Functional Decline ◽  
Fold Increase ◽  
Colorectal Adenomas ◽  
Mutation Rates ◽  
Base Substitution ◽  
Intestinal Epithelial ◽  
Mutational Signatures ◽  
Base Excision ◽  
Cellular Dna

Cellular DNA damage caused by reactive oxygen species is repaired by the base excision repair (BER) pathway which includes the DNA glycosylase MUTYH. Inherited biallelic MUTYH mutations cause predisposition to colorectal adenomas and carcinoma. However, the mechanistic progression from germline MUTYH mutations to MUTYH-Associated Polyposis (MAP) is incompletely understood. Here, we sequenced normal tissue DNAs from 10 individuals with MAP. Somatic base substitution mutation rates in intestinal epithelial cells were elevated 2 to 5-fold in all individuals, except for one showing a 33-fold increase, and were also increased in other tissues. The increased mutation burdens were of multiple mutational signatures characterised by C>A changes. Different mutation rates and signatures between individuals were likely due to different MUTYH mutations or additional inherited mutations in other BER pathway genes. The elevated base substitution rate in normal cells likely accounts for the predisposition to neoplasia in MAP. Despite ubiquitously elevated mutation rates, individuals with MAP do not display overt evidence of premature ageing. Thus, accumulation of somatic mutations may not be sufficient to cause the global organismal functional decline of ageing.

scholarly journals Evaluating the utility of tumour mutational signatures for identifying hereditary colorectal cancer and polyposis syndrome carriers

Gut ◽  
2021 ◽  
pp. gutjnl-2019-320462
Author(s):  
Peter Georgeson ◽  
Bernard J Pope ◽  
Christophe Rosty ◽  
Mark Clendenning ◽  
Khalid Mahmood ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Excision Repair ◽  
Area Under The Curve ◽  
Hereditary Colorectal Cancer ◽  
Base Substitution ◽  
Mutational Signatures ◽  
Repair Gene ◽  
Dna Mismatch ◽  
Pathogenic Variants ◽  
Base Excision

ObjectiveGermline pathogenic variants (PVs) in the DNA mismatch repair (MMR) genes and in the base excision repair gene MUTYH underlie hereditary colorectal cancer (CRC) and polyposis syndromes. We evaluated the robustness and discriminatory potential of tumour mutational signatures in CRCs for identifying germline PV carriers.DesignWhole-exome sequencing of formalin-fixed paraffin-embedded (FFPE) CRC tissue was performed on 33 MMR germline PV carriers, 12 biallelic MUTYH germline PV carriers, 25 sporadic MLH1 methylated MMR-deficient CRCs (MMRd controls) and 160 sporadic MMR-proficient CRCs (MMRp controls) and included 498 TCGA CRC tumours. COSMIC V3 single base substitution (SBS) and indel (ID) mutational signatures were assessed for their ability to differentiate CRCs that developed in carriers from non-carriers.ResultsThe combination of mutational signatures SBS18 and SBS36 contributing >30% of a CRC’s signature profile was able to discriminate biallelic MUTYH carriers from all other non-carrier control CRCs with 100% accuracy (area under the curve (AUC) 1.0). SBS18 and SBS36 were associated with specific MUTYH variants p.Gly396Asp (p=0.025) and p.Tyr179Cys (p=5×10-5), respectively. The combination of ID2 and ID7 could discriminate the 33 MMR PV carrier CRCs from the MMRp control CRCs (AUC 0.99); however, SBS and ID signatures, alone or in combination, could not provide complete discrimination (AUC 0.79) between CRCs from MMR PV carriers and sporadic MMRd controls.ConclusionAssessment of SBS and ID signatures can discriminate CRCs from biallelic MUTYH carriers and MMR PV carriers from non-carriers with high accuracy, demonstrating utility as a potential diagnostic and variant classification tool.

scholarly journals Evaluating the utility of tumour mutational signatures for identifying hereditary colorectal cancer and polyposis syndrome carriers

2019 ◽  
Author(s):  
Peter Georgeson ◽  
Bernard J. Pope ◽  
Christophe Rosty ◽  
Mark Clendenning ◽  
Khalid Mahmood ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Excision Repair ◽  
Base Substitution ◽  
List Type ◽  
Mutational Signatures ◽  
Dna Mismatch ◽  
Pathogenic Variants ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Base Excision

ABSTRACTObjectiveGermline pathogenic variants (PVs) in the DNA mismatch repair (MMR) genes and in the base excision repair gene MUTYH underlie hereditary colorectal cancer (CRC) and polyposis syndromes. We evaluated the robustness and discriminatory potential of tumour mutational signatures in CRCs for identifying germline PV carriers.DesignWhole exome sequencing of formalin-fixed paraffin embedded (FFPE) CRC tissue was performed on 33 MMR germline PV carriers, 12 biallelic MUTYH germline PV carriers, 25 sporadic MLH1 methylated MMR-deficient CRCs (MMRd controls) and 160 sporadic MMR-proficient CRCs (MMRp controls) and included 498 TCGA CRC tumours. COSMIC V3 single base substitution (SBS) and indel (ID) mutational signatures were assessed for their ability to differentiate CRCs that developed in carriers from non-carriers.ResultsThe combination of mutational signatures SBS18 and SBS36 contributing >30% of a CRC’s signature profile was able to discriminate biallelic MUTYH carriers from all other non-carrier control CRCs with 100% accuracy (area under the curve (AUC) 1.0). SBS18 and SBS36 were associated with specific MUTYH variants p.Gly396Asp (p=0.025) and p.Tyr179Cys (p=5×10−5), respectively. The combination of ID2 and ID7 could discriminate the 33 MMR PV carrier CRCs from the MMRp control CRCs (AUC 0.99), however, SBS and ID signatures, alone or in combination, could not provide complete discrimination (AUC 0.79) between CRCs from MMR PV carriers and sporadic MMRd controls.ConclusionAssessment of SBS and ID signatures can discriminate CRCs from biallelic MUTYH carriers and MMR PV carriers from non-carriers with high accuracy, demonstrating utility as a potential diagnostic and variant classification tool.SIGNIFICANCE OF THE STUDYWhat is already known about this subject?Identifying carriers of pathogenic variants (PVs) in moderate/high-risk colorectal cancer (CRC) and polyposis susceptibility genes has clinical relevance for diagnosis, targeted screening and prevention strategies, prognosis, and treatment options. However, challenges still remain in the identification of carriers and the classification of rare variants in these genes.Previous studies have identified tumour mutational signatures that result from defective DNA repair including DNA mismatch repair (MMR) deficiency and base excision repair defects, DNA repair mechanisms that underlie the common hereditary CRC and polyposis syndromes but their diagnostic utility in CRC is unknown.What are the new findings?Single base substitution (SBS)-related mutational signatures derived from whole exome sequencing of formalin-fixed paraffin embedded (FFPE)-derived CRC tissue DNA can effectively discriminate CRCs that developed in biallelic MUTYH PV carriers from CRC-affected non-carriers.CRCs that develop in MMR PV carriers (Lynch syndrome) can be effectively differentiated from sporadic MMR-proficient CRC by a combination of indel (ID) signatures, but the SBS and ID tumour mutational signatures are less effective at discriminating Lynch syndrome-related CRC from sporadic MMR-deficient CRC resulting from MLH1 gene promoter hypermethylation.The SBS and ID mutational signatures associated with biallelic MUTYH PV carriers and MMR PV carriers are robust to changes in experimental settings.We demonstrate the optimal experimental settings for calculating mutational signatures and define thresholds that optimise sensitivity and specificity for classifying CRC aetiology as hereditary or non-hereditary.How might it impact on clinical practice in the foreseeable future?Deriving SBS- and ID-related mutational signatures from CRCs can identify carriers of PVs in hereditary CRC and polyposis susceptibility genes.The application of mutational signatures has the potential to improve the diagnosis of hereditary CRC and aid in variant classification, leading to improved clinical management and CRC prevention.

scholarly journals Heritable pattern of oxidized DNA base repair coincides with pre-targeting of repair complexes to open chromatin

2020 ◽  
Author(s):  
Albino Bacolla ◽  
Shiladitya Sengupta ◽  
Zu Ye ◽  
Chunying Yang ◽  
Joy Mitra ◽  
...  
Keyword(s):  
Genome Stability ◽  
High Throughput Sequencing ◽  
Excision Repair ◽  
Essential Gene ◽  
Atmospheric Oxygen ◽  
Super Resolution ◽  
Population Variation ◽  
Mutation Rates ◽  
Open Chromatin ◽  
Base Excision

Abstract Human genome stability requires efficient repair of oxidized bases, which is initiated via damage recognition and excision by NEIL1 and other base excision repair (BER) pathway DNA glycosylases (DGs). However, the biological mechanisms underlying detection of damaged bases among the million-fold excess of undamaged bases remain enigmatic. Indeed, mutation rates vary greatly within individual genomes, and lesion recognition by purified DGs in the chromatin context is inefficient. Employing super-resolution microscopy and co-immunoprecipitation assays, we find that acetylated NEIL1 (AcNEIL1), but not its non-acetylated form, is predominantly localized in the nucleus in association with epigenetic marks of uncondensed chromatin. Furthermore, chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) revealed non-random AcNEIL1 binding near transcription start sites of weakly transcribed genes and along highly transcribed chromatin domains. Bioinformatic analyses revealed a striking correspondence between AcNEIL1 occupancy along the genome and mutation rates, with AcNEIL1-occupied sites exhibiting fewer mutations compared to AcNEIL1-free domains, both in cancer genomes and in population variation. Intriguingly, from the evolutionarily conserved unstructured domain that targets NEIL1 to open chromatin, its damage surveillance of highly oxidation-susceptible sites to preserve essential gene function and to limit instability and cancer likely originated ∼500 million years ago during the buildup of free atmospheric oxygen.

scholarly journals DNA Damage and Base Excision Repair in Mitochondria and Their Role in Aging

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Ricardo Gredilla
Keyword(s):  
Dna Damage ◽  
Mitochondrial Dna ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Functional Decline ◽  
Dna Integrity ◽  
Mitochondrial Dna Mutations ◽  
Mitochondrial Dna Damage ◽  
Dna Mutations ◽  
Base Excision

During the last decades, our knowledge about the processes involved in the aging process has exponentially increased. However, further investigation will be still required to globally understand the complexity of aging. Aging is a multifactorial phenomenon characterized by increased susceptibility to cellular loss and functional decline, where mitochondrial DNA mutations and mitochondrial DNA damage response are thought to play important roles. Due to the proximity of mitochondrial DNA to the main sites of mitochondrial-free radical generation, oxidative stress is a major source of mitochondrial DNA mutations. Mitochondrial DNA repair mechanisms, in particular the base excision repair pathway, constitute an important mechanism for maintenance of mitochondrial DNA integrity. The results reviewed here support that mitochondrial DNA damage plays an important role in aging.

scholarly journals Genetic Analysis of Transcription-Associated Mutation in Saccharomyces cerevisiae

Genetics ◽  
2000 ◽  
Vol 154 (1) ◽  
pp. 109-120 ◽  
Author(s):  
Natalie J Morey ◽  
Christopher N Greene ◽  
Sue Jinks-Robertson
Keyword(s):  
Dna Damage ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Mutation Rates ◽  
Base Pairs ◽  
Genetic Studies ◽  
Dna Damage And Repair ◽  
Nucleotide Excision ◽  
Translesion Bypass ◽  
Base Excision

Abstract High levels of transcription are associated with elevated mutation rates in yeast, a phenomenon referred to as transcription-associated mutation (TAM). The transcription-associated increase in mutation rates was previously shown to be partially dependent on the Rev3p translesion bypass pathway, thus implicating DNA damage in TAM. In this study, we use reversion of a pGAL-driven lys2ΔBgl allele to further examine the genetic requirements of TAM. We find that TAM is increased by disruption of the nucleotide excision repair or recombination pathways. In contrast, elimination of base excision repair components has only modest effects on TAM. In addition to the genetic studies, the lys2ΔBgl reversion spectra of repair-proficient low and high transcription strains were obtained. In the low transcription spectrum, most of the frameshift events correspond to deletions of AT base pairs whereas in the high transcription strain, deletions of GC base pairs predominate. These results are discussed in terms of transcription and its role in DNA damage and repair.

scholarly journals Structures of LIG1 engaging with mutagenic mismatches inserted by polβ in base excision repair

2022 ◽  
Author(s):  
Qun Tang ◽  
Robert McKenna ◽  
Melike Caglayan
Keyword(s):  
Base Excision Repair ◽  
Excision Repair ◽  
Initial Step ◽  
Base Substitution ◽  
Dna Structures ◽  
Dna Ligation ◽  
Nucleotide Insertion ◽  
Base Excision ◽  
Similar Conformation ◽  
Ligation Step

DNA ligase I (LIG1) catalyzes final ligation step following DNA polymerase (pol) β gap filling and an incorrect nucleotide insertion by polβ creates a nick repair intermediate with mismatched end at the downstream steps of base excision repair (BER) pathway. Yet, how LIG1 discriminates against the mutagenic 3'-mismatches at atomic resolution remains undefined. Here, we determined X-ray structures of LIG1/nick DNA complexes with G:T and A:C mismatches and uncovered the ligase strategies that favor or deter ligation of base substitution errors. Our structures revealed that LIG1 active site can accommodate G:T mismatch in a similar conformation with A:T base pairing, while it stays in the LIG1-adenylate intermediate during initial step of ligation reaction in the presence of A:C mismatch at 3'-strand. Moreover, we showed mutagenic ligation and aberrant nick sealing of the nick DNA substrates with 3'-preinserted dG:T and dA:C mismatches, respectively. Finally, we demonstrated that AP-Endonuclease 1 (APE1), as a compensatory proofreading enzyme, interacts and coordinates with LIG1 during mismatch removal and DNA ligation. Our overall findings and ligase/nick DNA structures provide the features of accurate versus mutagenic outcomes at the final BER steps where a multi-protein complex including polβ, LIG1, and APE1 can maintain accurate repair.

scholarly journals The mutational signatures of formalin fixation on the human genome

2021 ◽  
Author(s):  
Qingli Guo ◽  
Eszter Lakatos ◽  
Ibrahim Al Bakir ◽  
Kit Curtius ◽  
Trevor A. Graham ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
Excision Repair ◽  
Clinical Pathology ◽  
Dna Lesions ◽  
Computational Method ◽  
Formalin Fixation ◽  
Sequencing Data ◽  
Mutational Signatures ◽  
Cytosine Deamination ◽  
Base Excision

AbstractBackgroundFormalin fixation and paraffin embedding (FFPE) of patient material remains standard practice in clinical pathology labs around the world. Clinical archives of patient material near-exclusively consist of FFPE blocks. The ability to perform high quality genome sequencing on FFPE-derived DNA would accelerate a broad spectrum of medical research. However, formalin is a recognised mutagen and sequencing of DNA derived from FFPE material is known to be riddled with artefactual mutations.ResultsHere we derive genome-wide mutational signatures caused by formalin fixation, and provide a computational method to correct mutational profiles for these formalin-induced artefacts. We show that the FFPE-signature is dominated by C>T transitions caused by cytosine deamination, and has very high similarity to COSMIC signature SBS30 (base excision repair deficiency due to inactivation mutations in NTHL1). Further, we demonstrate that chemical repair of formalin-induced DNA lesions, a process that is routinely performed as part of sequencing library preparation, leads to a signature highly similar to COSMIC signature SBS1 (spontaneous deamination of methylated cytosine). Next, we design FFPEsig, a computational method to remove the formalin-induced artefacts from mutational counts. We prove the efficacy of this method by generating synthetic FFPE samples using 2,780 cancer genomes from the Pan-Cancer Analysis of Whole Genome (PCAWG) project, and via analysis of FFPE-derived genome sequencing data from colorectal cancers.ConclusionsFormalin fixation leaves a predictable mutational footprint across the genome. The application of our FFPEsig software corrects the mutational profile for the influence of formalin, enabling robust mutational signature analysis in FFPE-derived patient material.

scholarly journals Formation and Repair of Tobacco Carcinogen-Derived Bulky DNA Adducts

2010 ◽  
Vol 2010 ◽  
pp. 1-29 ◽  
Author(s):  
Bo Hang
Keyword(s):  
Dna Adducts ◽  
Excision Repair ◽  
Complex Structure ◽  
Chemical Carcinogenesis ◽  
Tobacco Carcinogen ◽  
Polycyclic Aromatic ◽  
Carcinogenic Effects ◽  
Bulky Dna Adducts ◽  
Base Excision ◽  
Cellular Dna

DNA adducts play a central role in chemical carcinogenesis. The analysis of formation and repair of smoking-related DNA adducts remains particularly challenging as both smokers and nonsmokers exposed to smoke are repetitively under attack from complex mixtures of carcinogens such as polycyclic aromatic hydrocarbons andN-nitrosamines. The bulky DNA adducts, which usually have complex structure, are particularly important because of their biological relevance. Several known cellular DNA repair pathways have been known to operate in human cells on specific types of bulky DNA adducts, for example, nucleotide excision repair, base excision repair, and direct reversal involvingO6-alkylguanine DNA alkyltransferase or AlkB homologs. Understanding the mechanisms of adduct formation and repair processes is critical for the assessment of cancer risk resulting from exposure to cigarette smoke, and ultimately for developing strategies of cancer prevention. This paper highlights the recent progress made in the areas concerning formation and repair of bulky DNA adducts in the context of tobacco carcinogen-associated genotoxic and carcinogenic effects.

scholarly journals A Multi-Endpoint Approach to Base Excision Repair Incision Activity Augmented by PARylation and DNA Damage Levels in Mice: Impact of Sex and Age

2020 ◽  
Vol 21 (18) ◽  
pp. 6600
Author(s):  
Nicola Winkelbeiner ◽  
Viktoria K. Wandt ◽  
Franziska Ebert ◽  
Kristina Lossow ◽  
Ezgi E. Bankoglu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Decisive Role ◽  
Dna Lesions ◽  
Dna Integrity ◽  
Repair Mechanisms ◽  
Murine Liver ◽  
Base Excision ◽  
Cellular Dna

Investigation of processes that contribute to the maintenance of genomic stability is one crucial factor in the attempt to understand mechanisms that facilitate ageing. The DNA damage response (DDR) and DNA repair mechanisms are crucial to safeguard the integrity of DNA and to prevent accumulation of persistent DNA damage. Among them, base excision repair (BER) plays a decisive role. BER is the major repair pathway for small oxidative base modifications and apurinic/apyrimidinic (AP) sites. We established a highly sensitive non-radioactive assay to measure BER incision activity in murine liver samples. Incision activity can be assessed towards the three DNA lesions 8-oxo-2’-deoxyguanosine (8-oxodG), 5-hydroxy-2’-deoxyuracil (5-OHdU), and an AP site analogue. We applied the established assay to murine livers of adult and old mice of both sexes. Furthermore, poly(ADP-ribosyl)ation (PARylation) was assessed, which is an important determinant in DDR and BER. Additionally, DNA damage levels were measured to examine the overall damage levels. No impact of ageing on the investigated endpoints in liver tissue were found. However, animal sex seems to be a significant impact factor, as evident by sex-dependent alterations in all endpoints investigated. Moreover, our results revealed interrelationships between the investigated endpoints indicative for the synergetic mode of action of the cellular DNA integrity maintaining machinery.

scholarly journals HECTD1 promotes base excision repair in nucleosomes through chromatin remodelling

2019 ◽  
Vol 48 (3) ◽  
pp. 1301-1313 ◽  
Author(s):  
Laura Bennett ◽  
Eleanor C E T Madders ◽  
Jason L Parsons
Keyword(s):  
Dna Damage ◽  
Base Excision Repair ◽  
Genome Stability ◽  
Excision Repair ◽  
Chromatin Remodelling ◽  
Abasic Site ◽  
Cell Extracts ◽  
Base Excision ◽  
Cellular Dna

Abstract Base excision repair (BER) is the major cellular DNA repair pathway that recognises and excises damaged DNA bases to help maintain genome stability. Whilst the major enzymes and mechanisms co-ordinating BER are well known, the process of BER in chromatin where DNA is compacted with histones, remains unclear. Using reconstituted mononucleosomes containing a site-specific synthetic abasic site (tetrahydrofuran, THF), we demonstrate that the DNA damage is less efficiently incised by recombinant AP endonuclease 1 (APE1) when the DNA backbone is facing the histone core (THF-in) compared to that orientated away (THF-out). However, when utilizing HeLa whole cell extracts, the difference in incision of THF-in versus THF-out is less pronounced suggesting the presence of chromatin remodelling factors that stimulate THF accessibility to APE1. We subsequently purified an activity from HeLa cell extracts and identify this as the E3 ubiquitin ligase, HECTD1. We demonstrate that a recombinant truncated form of HECTD1 can stimulate incision of THF-in by APE1 in vitro by histone ubiquitylation, and that siRNA-mediated depletion of HECTD1 leads to deficiencies in DNA damage repair and decreased cell survival following x-ray irradiation, particularly in normal fibroblasts. Thus, we have now identified HECTD1 as an important factor in promoting BER in chromatin.

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