Mutagenic properties of the 8-amino-2'-deoxyguanosine DNA adduct in mammalian cells

The DNA adduct 8-amino-2'-deoxyguanosine (8-amino-dG) is found in liver DNA of rats treated with the hepatocarcinogen 2-nitropropane. Site-specifically modified oligodeoxynucleotides were used to explore the mutagenic potential of 8-amino-dG in simian kidney (COS-7) cells. Oligodeoxynucleotides (5'-TCCTCCTX₁G₂CCTCTC and 5'-TCCTCCTG₁X₂CCTCTC, X = dG or 8-amino-dG) with the lesion positioned at codon 60 or 61 of the non-coding strand of the human c-Ha-ras1 gene were inserted into single-stranded phagemid vectors and transfected into COS-7 cells. The progeny plasmid obtained was used to transform Escherichia coli DH1OB. Transformants were analyzed by oligodeoxynucleotide hybridization and DNA sequencing to establish the mutation frequency and spectrum produced by the modified base. The correct base, dCMP, was incorporated preferentially opposite 8-amino-dG at X₁ and X₂. When 8-amino-dG was at X₁, targeted G(NH2)→T transversions were detected, along with smaller numbers of G(NH2)→A transitions and G(NH2)→C transversions. When the adduct was at X₂, only G(NH2)→T transversions were observed. The frequencies of targeted mutation at X₁ and X₂ were 2.7 and 1.7%, respectively. Mutation frequency and mutagenic spectrum were sequence context dependent. In addition, non-targeted G→T transversions, accompanied by some G→A transitions, were detected 5' to 8-amino-dG when the lesion was at X₂. We conclude that 8-amino-dG is a mutagenic lesion, generating G→T and G→C transversions and G→A transitions in mammalian cells.