A novel role for Escherichia coli endonuclease VIII in prevention of spontaneous G -> T transversions

In the bacterium Escherichia coli, oxidized pyrimidines are removed by two DNA glycosylases, endonuclease III and endonuclease Vm (endo Vm), encoded b y the nth and nei genes, respectively. Double mutants lacking both of these activities exhibit a high spontaneous mutation frequency, and here we show that all of the mutations observed in the double mutants were G:C-->A:T tr ansitions; no thymine mutations were found. These findings are in agreement with the preponderance of C-->T transitions in the oxidative and spontaneo us mutational databases. The major oxidized purine lesion in DNA, 7,8-dihyd ro-8-oxoguanine (8-oxoG), is processed by two DNA glycosylases, formamidopy rimidine DNA glycosylase (Fpg), which removes 8-oxoG opposite C, and MutY D NA glycosylase, which removes misincorporated A opposite 8-oxoG, The high s pontaneous mutation frequency previously observed in fpg mutY double mutant s was significantly enhanced by the addition of the nei mutation, suggestin g an overlap in the substrate specificities between endo VIII and Fpg/MutY. When the mutational specificity was examined, all of the mutations observe d were G:C-->T:A transversions, indicating that in the absence of Fpg and M utY, endo VIII serves as a backup activity to remove 8-oxoG. This was confi rmed by showing that, indeed, endo Vm can recognize 8-oxoG in vitro.