Mechanism of Translesion Synthesis Past an Equine Estrogen-DNA Adduct by Y-Family DNA Polymerases

4-Hydroxyequilenin (4-OHEN)-dC is a major, potentially mutagenic DNA adduct induced by equine estrogens used for hormone replacement therapy. To study the miscoding property of 4-OHEN-dC and the involvement of Y-family human DNA polymerases (pols) η, κ and ι in that process, we incorporated 4-OHEN-dC into oligodeoxynucleotides and used them as templates in primer extension reactions catalyzed by pol η, κ and ι. Pol η inserted dAMP opposite 4-OHEN-dC, accompanied by lesser amounts of dCMP and dTMP incorporation and base deletion. Pol κ promoted base deletions as well as direct incorporation of dAMP and dCMP. Pol ι worked in conjunction with pol κ, but not with pol η, at a replication fork stalled by the adduct, resulting in increased dTMP incorporation. Our results provide a direct evidence that Y-family DNA pols can switch with one another during synthesis past the lesion. No direct incorporation of dGMP, the correct base, was observed with Y-family enzymes. The miscoding potency of 4-OHEN-dC may be associated with the development of reproductive cancers observed in women receiving hormone replacement therapy.