Resistance to Nucleotide Excision Repair of Bulky Guanine Adducts Opposite Abasic Sites in DNA Duplexes and Relationships between Structure and Function.

Zhi Liu,Konstantin Kropachev,Shantu Amin,Jia Lei,Shuang Ding,Suse Broyde,Nicholas E. Geacintov,Maria Spies

doi:10.1371/journal.pone.0137124

Zhi Liu, Konstantin Kropachev + Show 6 more

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https://doi.org/10.1371/journal.pone.0137124

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Journal: PloS one	Publication Date: Sep 4, 2015
Citations: 21	License type: CC BY 4.0

Affiliation: New York University, Pennsylvania State University

Abstract

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-N 2-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.

Highlights

Eukaryotic nucleotide excision repair (NER) is an important mammalian defense system against DNA lesions derived from environmental genotoxic agents that include polycyclic aromatic hydrocarbons (PAH)
In order to elucidate how features of the local DNA binding site and adduct conformation affect NER activity, we explored the impact of positioning the trans-B[a]P-dG adduct opposite a tertrahydrofuran doi:10.1371/journal.pone.0137124.g001 (THF) abasic site (AB); the AB site lacks only the cytosine base in the complementary strand opposite the modified guanine, but not its 2’-deoxyribose residue and phosphodiester group (Fig 1)
The assignment of the majority of the chemical shifts was based on total correlation spectroscopy (TOCSY) (75ms mixing time) and nuclear Overhauser effect spectroscopy (NOESY) (300 ms mixing time with solutions of different concentrations) data sets, and confirmed by water NOESY (250 ms mixing time) and correlation spectroscopy (COSY) methods

Summary

Introduction

Eukaryotic nucleotide excision repair (NER) is an important mammalian defense system against DNA lesions derived from environmental genotoxic agents that include polycyclic aromatic hydrocarbons (PAH). The NER process involves ~ 30 proteins and entails the excision of a 24–30-mer oligonucleotide that contains the lesion, followed by repair synthesis of the resulting gap [1]. The XPC-RAD23B—DNA complex stimulates the recruitment of subsequent NER factors [5, 8], which leads to the characteristic dual incisions of the damaged strand that yield the single-stranded, 24–32 nucleotide-long DNA sequence that contains the lesion. The yields of dual incision products depends on factors that include: (1) the base sequence context in which the DNA lesions are embedded [9,10,11], (2) the presence or absence of the Watson-Crick partner nucleotides opposite the modified nucleotide [12], and (3) the identity of a mismatched base opposite the modified nucleotide [13]

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