Dataset Information

Nucleotide excision repair of abasic DNA lesions.

ABSTRACT: Apurinic/apyrimidinic (AP) sites are a class of highly mutagenic and toxic DNA lesions arising in the genome from a number of exogenous and endogenous sources. Repair of AP lesions takes place predominantly by the base excision pathway (BER). However, among chemically heterogeneous AP lesions formed in DNA, some are resistant to the endonuclease APE1 and thus refractory to BER. Here, we employed two types of reporter constructs accommodating synthetic APE1-resistant AP lesions to investigate the auxiliary repair mechanisms in human cells. By combined analyses of recovery of the transcription rate and suppression of transcriptional mutagenesis at specifically positioned AP lesions, we demonstrate that nucleotide excision repair pathway (NER) efficiently removes BER-resistant AP lesions and significantly enhances the repair of APE1-sensitive ones. Our results further indicate that core NER components XPA and XPF are equally required and that both global genome (GG-NER) and transcription coupled (TC-NER) subpathways contribute to the repair.

SUBMITTER: Kitsera N

PROVIDER: S-EPMC6895268 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Nucleotide excision repair of abasic DNA lesions.

Kitsera Nataliya N Rodriguez-Alvarez Marta M Emmert Steffen S Carell Thomas T Khobta Andriy A

Nucleic acids research 20190901 16

Apurinic/apyrimidinic (AP) sites are a class of highly mutagenic and toxic DNA lesions arising in the genome from a number of exogenous and endogenous sources. Repair of AP lesions takes place predominantly by the base excision pathway (BER). However, among chemically heterogeneous AP lesions formed in DNA, some are resistant to the endonuclease APE1 and thus refractory to BER. Here, we employed two types of reporter constructs accommodating synthetic APE1-resistant AP lesions to investigate the ...[more]

PMID: 31226203

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Project description:The nucleotide excision repair of certain bulky DNA lesions is abrogated in some specific non-canonical DNA base sequence contexts, while the removal of the same lesions by the nucleotide excision repair mechanism is efficient in duplexes in which all base pairs are complementary. Here we show that the nucleotide excision repair activity in human cell extracts is moderate-to-high in the case of two stereoisomeric DNA lesions derived from the pro-carcinogen benzo[a]pyrene (cis- and trans-B[a]P-N2-dG adducts) in a normal DNA duplex. By contrast, the nucleotide excision repair activity is completely abrogated when the canonical cytosine base opposite the B[a]P-dG adducts is replaced by an abasic site in duplex DNA. However, base excision repair of the abasic site persists. In order to understand the structural origins of these striking phenomena, we used NMR and molecular spectroscopy techniques to evaluate the conformational features of 11mer DNA duplexes containing these B[a]P-dG lesions opposite abasic sites. Our results show that in these duplexes containing the clustered lesions, both B[a]P-dG adducts adopt base-displaced intercalated conformations, with the B[a]P aromatic rings intercalated into the DNA helix. To explain the persistence of base excision repair in the face of the opposed bulky B[a]P ring system, molecular modeling results suggest how the APE1 base excision repair endonuclease, that excises abasic lesions, can bind productively even with the trans-B[a]P-dG positioned opposite the abasic site. We hypothesize that the nucleotide excision repair resistance is fostered by local B[a]P residue-DNA base stacking interactions at the abasic sites, that are facilitated by the absence of the cytosine partner base in the complementary strand. More broadly, this study sets the stage for elucidating the interplay between base excision and nucleotide excision repair in processing different types of clustered DNA lesions that are substrates of nucleotide excision repair or base excision repair mechanisms.

Project description:The packaging of DNA in nucleosomes presents a barrier for biological transactions including replication, transcription and repair. However, despite years of research, how the DNA is freed from the histone proteins and thereby allows the molecular machines to access the DNA remains poorly understood. We are interested in global genomic nucleotide excision repair (GG-NER). It is established that the histones are obstacles to this process, and DNA lesions are repaired less efficiently in nucleosomes than in free DNA. In the present study, we utilized molecular dynamics simulations to elucidate the nature of the distortions and dynamics imposed in the nucleosome by a set of three structually different lesions that vary in GG-NER efficiencies in free DNA, and in nucleosomes [Shafirovich, Geacintov, et. al, 2019]. Two of these are bulky lesions derived from metabolic activation of the environmental carcinogen benzo[a]pyrene, the 10R (+)-cis-anti-B[a]P-N2-dG and the stereoisomeric 10S (+)-trans-anti-B[a]P-N2-dG, which respectively adopt base-displaced/intercalated and minor groove-aligned conformations in DNA. The third is a non-bulky lesion, the 5'R-8-cyclo-2'-deoxyguanosine cross-link, produced by reactive oxygen and nitrogen species; cyclopurine lesions are highly mutagenic. These adducts are placed near the dyad axis, and rotationally with the lesion-containing strand facing towards or away from the histones. While each lesion has distinct conformational characteristics that are retained in the nucleosome, a spectrum of structural and dynamic disturbances, from slight to substantial, are displayed that depend on the lesion's structure and position in the nucleosome. We hypothesize that these intrinsic structural and dynamic distinctions provide different signals to initiate the cascade of chromatin-opening processes, including acetylation and other post translational modifications, remodeling by ATP-dependent complexes and spontaneous unwrapping that regulate the rate of access to the lesion; this may translate ultimately into varying GG-NER efficiencies, including repair resistance when signals for access are too weak.

Dataset Information

Nucleotide excision repair of abasic DNA lesions.

Publications

Nucleotide excision repair of abasic DNA lesions.

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