title: Escherichia coli DNA polymerase III is responsible for the high level of spontaneous mutations in mutT strains.
abstract: Reactive oxygen species induce oxidative damage in DNA precursors, i.e. dNTPs, leading to point mutations upon incorporation. Escherichia coli mutT strains, deficient in the activity hydrolysing 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphate (8-oxo-dGTP), display more than a 100-fold higher spontaneous mutation frequency over the wild-type strain. 8-oxo-dGTP induces A to C transversions when misincorporated opposite template A. Here, we report that DNA pol III incorporates 8-oxo-dGTP ≈ 20 times more efficiently opposite template A compared with template C. Single, double or triple deletions of pol I, pol II, pol IV or pol V had modest effects on the mutT mutator phenotype. Only the deletion of all four polymerases led to a 70% reduction of the mutator phenotype. While pol III may account for nearly all 8-oxo-dGTP incorporation opposite template A, it only extends ≈ 30% of them, the remaining 70% being extended by the combined action of pol I, pol II, pol IV or pol V. The unique property of pol III, a C-family DNA polymerase present only in eubacteria, to preferentially incorporate 8-oxo-dGTP opposite template A during replication might explain the high spontaneous mutation frequency in E. coli mutT compared with the mammalian counterparts lacking the 8-oxo-dGTP hydrolysing activities.
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, Tokyo 158-8501, Japan.
authors.forename: Masami
authors.initials: M
authors.lastname: Yamada
authors.forename: Masatomi
authors.initials: M
authors.lastname: Shimizu
authors.forename: Atsushi
authors.initials: A
authors.lastname: Katafuchi
authors.forename: Petr
authors.initials: P
authors.lastname: Grúz
authors.forename: Shingo
authors.initials: S
authors.lastname: Fujii
authors.forename: Yukio
authors.initials: Y
authors.lastname: Usui
authors.forename: Robert P
authors.initials: RP
authors.lastname: Fuchs
authors.forename: Takehiko
authors.initials: T
authors.lastname: Nohmi
mesh_terms: D004258:DNA Polymerase III / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D029968:Escherichia coli Proteins; D009154:Mutation; D059645:Mutation Rate; D011755:Pyrophosphatases / Q000172:deficiency*
chemical_list: D004269:DNA, Bacterial; D003848:Deoxyguanine Nucleotides; D029968:Escherichia coli Proteins; C078206:8-oxodeoxyguanosine triphosphate; D004258:DNA Polymerase III; D011755:Pyrophosphatases; C052211:mutT protein, E coli