Base Excision Repair and its Role in Maintaining Genome Stability

Whilst avoidance of chemical modifications of DNA bases is essential to maintain genome stability, during evolution eukaryotic cells have evolved a chemically reversible modification of the cytosine base. These dynamic methylation and demethylation reactions on carbon-5 of cytosine regulate several cellular and developmental processes such as embryonic stem cell pluripotency, cell identity, differentiation or tumourgenesis. Whereas these physiological processes are well characterized, very little is known about the toxicity of these cytosine analogues when they incorporate during replication. Here, we report a role of the base excision repair factor XRCC1 in protecting replication fork upon incorporation of 5-hydroxymethyl-2′-deoxycytosine (5hmC) and its deamination product 5-hydroxymethyl-2′-deoxyuridine (5hmU) during DNA synthesis. In the absence of XRCC1, 5hmC exposure leads to increased genomic instability, replication fork impairment and cell lethality. Moreover, the 5hmC deamination product 5hmU recapitulated the genomic instability phenotypes observed by 5hmC exposure, suggesting that 5hmU accounts for the observed by 5hmC exposure. Remarkably, 5hmC-dependent genomic instability and replication fork impairment seen in Xrcc1−/− cells were exacerbated by the trapping of Parp1 on chromatin, indicating that XRCC1 maintains replication fork stability during processing of 5hmC and 5hmU by the base excision repair pathway. Our findings uncover natural epigenetic DNA bases 5hmC and 5hmU as genotoxic nucleosides that threaten replication dynamics and genome integrity in the absence of XRCC1.

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Nucleophosmin modulates stability, activity, and nucleolar accumulation of base excision repair proteins

Molecular Biology of the Cell ◽

10.1091/mbc.e13-12-0717 ◽

2014 ◽

Vol 25 (10) ◽

pp. 1641-1652 ◽

Cited By ~ 40

Author(s):

Mattia Poletto ◽

Lisa Lirussi ◽

David M. Wilson ◽

Gianluca Tell

Keyword(s):

Dna Damage Response ◽

Base Excision Repair ◽

Genome Stability ◽

Excision Repair ◽

Multifunctional Protein ◽

Protein Levels ◽

Exact Function ◽

Damage Response ◽

Base Excision ◽

Representative Protein

Nucleophosmin (NPM1) is a multifunctional protein that controls cell growth and genome stability via a mechanism that involves nucleolar–cytoplasmic shuttling. It is clear that NPM1 also contributes to the DNA damage response, yet its exact function is poorly understood. We recently linked NPM1 expression to the functional activation of the major abasic endonuclease in mammalian base excision repair (BER), apurinic/apyrimidinic endonuclease 1 (APE1). Here we unveil a novel role for NPM1 as a modulator of the whole BER pathway by 1) controlling BER protein levels, 2) regulating total BER capacity, and 3) modulating the nucleolar localization of several BER enzymes. We find that cell treatment with the genotoxin cisplatin leads to concurrent relocalization of NPM1 and BER components from nucleoli to the nucleoplasm, and cellular experiments targeting APE1 suggest a role for the redistribution of nucleolar BER factors in determining cisplatin toxicity. Finally, based on the use of APE1 as a representative protein of the BER pathway, our data suggest a function for BER proteins in the regulation of ribogenesis.

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Co-ordination of base excision repair and genome stability

DNA Repair ◽

10.1016/j.dnarep.2013.02.001 ◽

2013 ◽

Vol 12 (5) ◽

pp. 326-333 ◽

Cited By ~ 49

Author(s):

Jason L. Parsons ◽

Grigory L. Dianov

Keyword(s):

Base Excision Repair ◽

Genome Stability ◽

Excision Repair ◽

Base Excision

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HECTD1 promotes base excision repair in nucleosomes through chromatin remodelling

Nucleic Acids Research ◽

10.1093/nar/gkz1129 ◽

2019 ◽

Vol 48 (3) ◽

pp. 1301-1313 ◽

Cited By ~ 2

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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Apurinic/Apyrimidinic Endonuclease 2 (APE2): An ancillary enzyme for contextual base excision repair mechanisms to preserve genome stability

Biochimie ◽

10.1016/j.biochi.2021.07.006 ◽

2021 ◽

Author(s):

Sima Chaudhari ◽

Akshay P. Ware ◽

Pradyumna Jayaram ◽

Sankar Prasad Gorthi ◽

Sherif F. El-Khamisy ◽

...

Keyword(s):

Base Excision Repair ◽

Genome Stability ◽

Excision Repair ◽

Repair Mechanisms ◽

Base Excision

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The E295K DNA Polymerase Beta Gastric Cancer-Associated Variant Interferes with Base Excision Repair and Induces Cellular Transformation

Molecular and Cellular Biology ◽

10.1128/mcb.01883-06 ◽

2007 ◽

Vol 27 (15) ◽

pp. 5587-5596 ◽

Cited By ~ 66

Author(s):

Tieming Lang ◽

Shibani Dalal ◽

Anna Chikova ◽

Daniel DiMaio ◽

Joann B. Sweasy

Keyword(s):

Base Excision Repair ◽

Genome Stability ◽

Excision Repair ◽

Cellular Transformation ◽

Sister Chromatid Exchanges ◽

Dominant Negative ◽

Single Amino Acid ◽

Polymerase Beta ◽

Homology Directed Repair ◽

Base Excision

ABSTRACT Approximately 30% of human tumors examined for mutations in polymerase beta (pol β) appear to express pol β variant proteins (D. Starcevic, S. Dalal, and J. B. Sweasy, Cell Cycle 3:998-1001, 2004). Many of these variants result from a single amino acid substitution. We have previously shown that the K289M and I260M colon and prostate cancer variants, respectively, induce cellular transformation most likely due to sequence-specific mutator activity (S. Dalal et al., Biochemistry 44:15664-15673, 2005; T. Lang et al., Proc. Natl. Acad. Sci. USA 101:6074-6079, 2004; J. B. Sweasy et al., Proc. Natl. Acad. Sci. USA 102:14350-14355, 2005). In the work described here, we show that the E295K gastric carcinoma pol β variant acts in a dominant-negative manner by interfering with base excision repair. This leads to an increase in sister chromatid exchanges. Expression of the E295K variant also induces cellular transformation. Our data suggest that unfilled gaps are channeled into a homology-directed repair pathway that could lead to genomic instability. The results indicate that base excision repair is critical for maintaining genome stability and could therefore be a tumor suppressor mechanism.

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