MISCODING PROPERTIES OF MODEL ESTROGEN-DNA ADDUCTS IN REACTIONS CATALYZED BY MAMMALIAN AND ESCHERICHIA-COLI DNA-POLYMERASES

The miscoding properties of the model estrogen-derived DNA adducts, th oxyestra-1,3,5(10)-trien-6-yl]-2'-deoxyguanosine (dG-N-2-3MeE) and N-6 -[3-methoxyestra-1,3,5(10)-trien-6-yl]-2'- deoxyadenosine (dA-N-6-3MeE ), have been explored, using an in vitro experimental system to quanti fy base substitutions and deletions. Site-specifically modified oligod eoxynucleotides containing a single dC-N-2-3MeE or dA-N-6-3MeE were pr epared postsynthetically and used as templates in primer extension rea ctions catalyzed by Escherichia coli and mammalian DNA polymerases. Wh en the 3' --> 5' exonuclease free (exo(-)) Klenow fragment of DNA poly merase I was used, dG-N-2-3MeE promoted mostly one- and two-base delet ions, along with small amounts of incorporation of dAMP, dGMP, and dCM P opposite the lesion. dA-N-6-3MeE promoted the incorporation of dTMP opposite the lesion as well as two-base deletions, accompanied by the incorporation of dAMP. Using pol alpha, primer extension reactions wer e blocked at dG-N-2-3MeE; however, dA-N-6-3MeE promoted preferential i ncorporation of dTMP opposite the lesion with small amounts of incorpo ration of dCMP and deletions. Primer extension reactions catalyzed by pol delta were blocked at these lesions. When pol beta was used, dG-N- 2-3MeE produced small amounts of incorporation of dAMP and deletions. dA-N-6-3MeE promoted preferential incorporation of dTMP, along with in corporation of dCMP and two-base deletions. The miscoding specificitie s and frequencies varied depending on the DNA polymerase used. These r esults indicate that estrogen-DNA adducts have miscoding potential.