Mutagenic specificity of model estrogen-DNA adducts in mammalian cells.
Site-specifically modified oligodeoxynucleotides were used to explore the mutagenic properties of the model estrogen-DNA adducts N2-[3-methoxyestra-1,3,5(10)-trien-6(alpha, beta)-yl]-2'-deoxyguanosine (dG-N2-3MeE) and N6-[3-methoxyestra-1,3, 5(10)-trien-6(alpha,beta)-yl]-2'-deoxyadenosine (dA-N6-3MeE) in simian kidney (COS-7) cells. Oligodeoxynucleotides (5'TCCTCCTCXCCTCTC; X = dG, dA, dG-N2-3MeE, or dA-N6-3MeE) containing an unmodified or model estrogen lesion were inserted into single-stranded (ss) phagemid vectors. These ss vectors were transfected into COS-7 cells. The progeny plasmid obtained were used to transform Escherichia coli DH10B. The transformants were analyzed by oligodeoxynucleotide hybridization and sequencing to determine the mutation frequency and spectrum. Preferential incorporation of dCMP, the correct base, was observed opposite the dG-N2-3MeE lesion. Targeted mutations showing G --> T transversions were detected, along with a small number of G --> C transversions. When a dA-N6-3MeE-modified oligodeoxynucleotide was used, preferential incorporation of dTMP, the correct base, was also observed. Targeted mutations representing A --> T transversions were detected, accompanied by a small amount of A --> G transitions. The frequency of mutation observed opposite dA-N6-3MeE (17.5%) was 2.3 times higher than that observed opposite dG-N2-3MeE (7.5%). These results indicate that estrogen DNA adducts have mutagenic potential in mammalian cells.
Terashima
I
I
Department of Pharmacological Sciences, State University of New York at Stony Brook, Stony Brook, New York 11794-8651, USA.
Suzuki
N
N
Itoh
S
S
Yoshizawa
I
I
Shibutani
S
S
D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D018736:DNA Adducts / Q000235:genetics* / Q000378:metabolism*; D004967:Estrogens / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D014162:Transfection
D018736:DNA Adducts; D004967:Estrogens