Miscoding Properties of Model Estrogen−DNA Adducts in Reactions Catalyzed by Mammalian and Escherichia coli DNA Polymerases,

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Abstract
The miscoding properties of the model estrogen-derived DNA adducts, N2-[3-methoxyestra-1,3,5(10)-trien-6-yl]-2‘-deoxyguanosine (dG-N2-3MeE) and N6-[3-methoxyestra-1,3,5(10)-trien-6-yl]-2‘-deoxyadenosine (dA-N6-3MeE), have been explored, using an in vitro experimental system to quantify base substitutions and deletions. Site-specifically modified oligodeoxynucleotides containing a single dG-N2-3MeE or dA-N6-3MeE were prepared postsynthetically and used as templates in primer extension reactions catalyzed by Escherichia coli and mammalian DNA polymerases. When the 3‘ → 5‘ exonuclease free (exo-) Klenow fragment of DNA polymerase I was used, dG-N2-3MeE promoted mostly one- and two-base deletions, along with small amounts of incorporation of dAMP, dGMP, and dCMP opposite the lesion. dA-N6-3MeE promoted the incorporation of dTMP opposite the lesion as well as two-base deletions, accompanied by the incorporation of dAMP. Using pol α, primer extension reactions were blocked at dG-N2-3MeE; however, dA-N6-3MeE promoted preferential incorporation of dTMP opposite the lesion with small amounts of incorporation of dCMP and deletions. Primer extension reactions catalyzed by pol δ were blocked at these lesions. When pol β was used, dG-N2-3MeE produced small amounts of incorporation of dAMP and deletions. dA-N6-3MeE promoted preferential incorporation of dTMP, along with incorporation of dCMP and two-base deletions. The miscoding specificities and frequencies varied depending on the DNA polymerase used. These results indicate that estrogen−DNA adducts have miscoding potential.