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Nucleic Acids Res. 2015 Jan;43(2):1012-8. doi: 10.1093/nar/gku1391. Epub 2015 Jan 8.

Transcriptional inhibition and mutagenesis induced by N-nitroso compound-derived carboxymethylated thymidine adducts in DNA.

Author information

1
Department of Chemistry, University of California, Riverside, CA 92521-0403, USA.
2
Department of Chemistry, University of California, Riverside, CA 92521-0403, USA Yinsheng.Wang@ucr.edu.

Abstract

N-nitroso compounds represent a common type of environmental and endogenous DNA-damaging agents. After metabolic activation, many N-nitroso compounds are converted into a diazoacetate intermediate that can react with nucleobases to give carboxymethylated DNA adducts such as N3-carboxymethylthymidine (N3-CMdT) and O(4)-carboxymethylthymidine (O(4)-CMdT). In this study, we constructed non-replicative plasmids carrying a single N3-CMdT or O(4)-CMdT, site-specifically positioned in the transcribed strand, to investigate how these lesions compromise the flow of genetic information during transcription. Our results revealed that both N3-CMdT and O(4)-CMdT substantially inhibited DNA transcription mediated by T7 RNA polymerase or human RNA polymerase II in vitro and in human cells. In addition, we found that N3-CMdT and O(4)-CMdT were miscoding lesions and predominantly directed the misinsertion of uridine and guanosine, respectively. Our results also suggested that these carboxymethylated thymidine lesions may constitute efficient substrates for transcription-coupled nucleotide excision repair in human cells. These findings provided important new insights into the biological consequences of the carboxymethylated DNA lesions in living cells.

PMID:
25572317
PMCID:
PMC4333421
DOI:
10.1093/nar/gku1391
[Indexed for MEDLINE]
Free PMC Article

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