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Peroxynitrite-induced DNA damage in the supF gene: correlation with the mutational spectrum
N Y Tretyakova1, S Burney, B Pamir
1Division of Bioengineering and Environmental Health, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Mutation Research
|February 7, 2001
Summary
Peroxynitrite (ONOO(-)) causes DNA damage and mutations linked to cancer risk. This study identifies specific DNA lesions, like 8-nitroguanine, induced by ONOO(-) in inflamed tissues.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Tissue inflammation and chronic infection elevate nitric oxide and superoxide, forming peroxynitrite (ONOO(-)).
- ONOO(-) is a potent oxidizing and nitrating agent implicated in cell death and cancer risk in inflamed tissues.
- ONOO(-) induces DNA damage, including single-strand breaks and base modifications, and is mutagenic, primarily causing G to T transversions.
Purpose of the Study:
- To identify biologically significant DNA modifications induced by peroxynitrite (ONOO(-)).
- To elucidate the chemical mechanisms underlying ONOO(-)-induced DNA damage and mutagenicity.
- To investigate the role of specific DNA lesions in the mutagenic effects of ONOO(-).
Main Methods:
- Treatment of pUC19 plasmid and supF gene oligonucleotides with ONOO(-).
- Analysis using liquid chromatography-negative ion electrospray ionization mass spectrometry (LC-ESI(-) MS) and tandem mass spectrometry.
- Polyacrylamide gel electrophoresis (PAGE) analysis of [32P]-end-labeled DNA fragments.
Main Results:
- ONOO(-) treatment caused single-strand breaks, abasic sites, and nucleobase modifications in DNA.
- 8-Nitroguanine (8-nitro-G) was identified as a dose-dependent product, with spontaneous release from DNA.
- Sequence-specific lesions at guanine bases, distinct from 8-nitro-G, were observed and repaired by Fpg glycosylase, suggesting alternative mutagenic lesions.
Conclusions:
- ONOO(-) induces diverse DNA modifications, including sugar damage and base lesions.
- While 8-nitroguanine is formed, other ONOO(-)-induced DNA lesions likely contribute significantly to its mutagenicity.
- Further research is needed to fully characterize the DNA modifications responsible for ONOO(-)'s adverse biological effects.
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