Mutagenic spectrum resulting from DNA damage by oxygen radicals.
Oxygen free radicals are highly reactive species that damage DNA and cause mutations. We determined the mutagenic spectrum of oxygen free radicals produced by the aerobic incubation of single-stranded M13mp2 DNA with Fe2+. The Fe2(+)-treated DNA was transfected into component Escherichia coli, and mutants within the nonessential lac Z alpha gene for beta-galactosidase were identified by decreased alpha-complementation. The frequency of mutants obtained with 10 microM Fe2+ was 20- to 80-fold greater than that obtained with untreated DNA. Mutagenesis was greater after the host cells were exposed to UV irradiation to induce the SOS "error-prone" response. The ability of catalase, mannitol, and superoxide dismutase to diminish mutagenesis indicates the involvement of oxygen free radicals. The sequence data on 94 of the mutants establish that mutagenesis results primarily from an increase in single-base substitutions. Ninety-four percent of the mutants with detectable changes in nucleotide sequence were single-base substitutions, the most frequent being G----C transversions, followed by C----T transitions and G----T transversions. The clustering of mutations at distinct gene positions suggests that Fe2+/oxygen damage to DNA is nonrandom. This mutational spectrum provides evidence that a multiplicity of DNA lesions produced by oxygen free radicals in vitro are promutagenic and could be a source of spontaneous mutations.
Joseph Gottstein Memorial Cancer Research Laboratory, Department of Pathology, University of Washington, Seattle 98195.
T J
TJ
McBride
B D
BD
Preston
L A
LA
Loeb
D001483:Base Sequence; D003090:Coliphages / Q000235:genetics; D004249:DNA Damage; D004279:DNA, Viral / Q000187:drug effects* / Q000235:genetics; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004706:Endodeoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D005609:Free Radicals; D005816:Genetic Complementation Test; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D007501:Iron / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010100:Oxygen; D013014:SOS Response, Genetics; D014162:Transfection
D004279:DNA, Viral; D029968:Escherichia coli Proteins; D005609:Free Radicals; D007501:Iron; D004706:Endodeoxyribonucleases; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D010100:Oxygen