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.

Cucurbiturils in nucleic acids research

“…Abasic DNA sites (apurinic/apyrimidinic (AP) sites) are mutagenic and toxic DNA lesions resulting from the loss of nucleobases. 53 AP sites are formed in course of enzymatic repair of damaged DNA or by the influence of external factors. Considering the relevance of AP sites to carcinogenesis, these lesions represent an important analytical and therapeutic target.…”

“…Considering the relevance of AP sites to carcinogenesis, these lesions represent an important analytical and therapeutic target. 53,54 In 2018, Y. Shao and co-workers described a supramolecular multicolor DNA sensor for the selective recognition of abasic DNA sites based on the ICA concept with CB [7]. 55 Two natural alkaloids, namely coptisine (21) and palmatine (22) have been used as fluorescent indicators for AP DNA sites (Scheme 3).…”

EGFP Reporters for Direct and Sensitive Detection of Mutagenic Bypass of DNA Lesions

“…This corresponds to the GenBank entries AAB08064 (protein) and U57609 (DNA). The pZAJ_5c vector encoding the functional EGFP protein [33] and the derived pEGFP_Q205* vector carrying the c.613C>T point mutation coding for the non-fluorescent truncated EGFP 1-204 protein, were described previously [30]. The procedure for site-specific incorporation of synthetic oligonucleotides into a single-stranded gap generated in the TS with the Nb.Bpu10I nicking endonuclease (Thermo Fisher Scientific Inc., St. Leon-Rot, Germany) was described previously [30].…”

“…We therefore hoped that an actionable reporter approach would be equally useful for characterization of mutations introduced by DNA TLS. We recently described a new enhanced green fluorescent protein (EGFP) c.613C>T point mutation, which leads to the synthesis of a non-fluorescent EGFP Q205* truncated protein, and showed that any subsequent nucleotide substitution at the affected position results in reversion to a fluorescent phenotype, thus offering a sensitive reporter for detection of TM [30]. Importantly, the mutated base pair in the EGFP Q205* gene is flanked by tandem Bpu10I sites retained from the original EGFP coding sequence.…”

Nucleosomal embedding reshapes the dynamics of abasic sites

“…They are generated by hydrolysis of the N-glycosylic bond between a nucleobase and a deoxyribose 3 , spontaneously or as intermediates in the excision repair of damaged bases. Their repair by dedicated enzymes turns out to be most efficient by base excision repair (BER) but also nucleotide excision repair (NER) 4 . Yet accumulation of AP sites, notably triggered by exogenous oxidative stress, represents a threat for genome integrity.…”