Abstract
8-Hydroxy-2'-deoxyguanosine 5'-triphosphate (8-OH-dGTP), oxidatively damaged DNA precursor, is produced by reactive oxygen species formed by ionizing radiation, and is incorporated by DNA polymerase(s) opposite incorrect bases to form mispairs. It has been shown that 8-OH-dGTP is highly mutagenic in Escherichia coli and various in vitro replication systems. Here we studied the mutagenicity of 8-OH-dGTP in living mammalian cells by direct introduction using cationic lipids, Lipofectamine reagent. 8-OH-dGTP and plasmid DNA, including supF gene as a mutational target, were co-introduced into COS-7 cells. After 48 hr incubation, replicated plasmid DNA was recovered and were electroporated into indicator E. coli, and then mutant frequencies were determined. 8-OH-dGTP efficiently induced A:T to C:G transversion mutations, although total mutant frequency was not increased compared with dGTP control. This result is consistent with the previous observations that DNA polymerases misincorporate 8-OH-dGTP opposite A in vitro and that 8-OH-dGTP induces A:T to C:G mutations in E. coli. Thus, we showed the first direct evidence that 8-OH-dGTP was actually mutagenic in living mammalian cells.
