| The Role of Mutagenic Metal Ions in Mediating In Vitro Mispairing by Alkylpyrimidines Environ Health Perspect Key words: alkylating agents, alkylpyrimidines, DNA adducts, divalent metal activator, misincorporation, DNA synthesis, mutation, nitroso compounds, DNA polymerase
This paper was presented at the Second International Meeting on Molecular Mechanisms of Metal Toxicity and Carcinogenicity held 10-17 January 1993 in Madonna di Campiglio, Italy.
We thank Dr. Peter C. Grevatt and Ms. Jean M. Donahue for their participation in the DNA replication studies of ethyldeoxythymidine adducts. The research described here was supported by research grant DMB-8607556 (OSB) from the National Science Foundation, grant ES 05694 (JJS) from the National Institute of Environmental Health Sciences, grant CIAR 90-12 (JJS) from the Center for Indoor Air Research, center grants CA 16087 and ES 00260 from NIH, and grant SIG 9A from the American Cancer Society.
Address correspondence to Dr. Opinder S. Bhanot, Department of Environmental Medicine, New York University Medical Center, 550 First Avenue, New York, NY 10016-6451. Telephone (914) 351-2204. Fax (914) 351-3489.
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A variety of alkylating mutagens and carcinogens produce pyrimidine adducts in DNA that block DNA synthesis in vitro. Since DNA synthesis past the lesion is a necessary step to produce mutations, we investigated the role of the mutagenic metal ion Mn++ in facilitating DNA synthesis past alkylpyrimidines. In the presence of the natural metal activator Mg++, N3-ethyldeoxythymidine (N3-Et-dT) and O2-ethyldeoxythymidine (O2-Et-dT) , present at a single site in DNA, blocked in vitro DNA synthesis 3' to the lesion and after incorporating dA opposite each lesion. The presence of Mn++ permitted postlesion synthesis with dT misincorporated opposite N3-Et-dT and O2-Et-dT, implicating these lesions in A-T->T-A transversion mutagenesis. The DNA synthesis block by O4-ethyldeoxythymidine (O4-Et-dT) in the presence of Mg++ was partial and was also removed by Mn++. Consistent with in vivo studies, dG was incorporated opposite O4-Et-dT during postlesion synthesis, leading to A-T->G-C transition mutagenesis. We also have discovered a new class of DNA adducts, N3-hydroxyalkyldeoxyuridine (3-HA-dU) lesions, which are produced by mutagenic and carcinogenic aliphatic epoxides. 3-HA-dU is formed after initial alkylation at the N3 position of dC followed by a rapid hydrolytic deamination. As observed with the analogous mutagenic N3-Et-dT, the ethylene oxide-induced 3-hydroxyethyldeoxyuridine (3-HE-dU) blocked in vitro DNA synthesis, which could be bypassed in the presence of Mn++. The nucleotide incorporated opposite 3-HE-dU during postlesion synthesis is being identified. These studies suggest a role for Mn++ in mediating mutagenic and carcinogenic effects of environmentally important ethylating agents and aliphatic epoxides. -- Environ Health Perspect 102(Suppl 3) :81-90 (1994) . Key words: alkylating agents, alkylpyrimidines, DNA adducts, divalent metal activator, misincorporation, DNA synthesis, mutation, nitroso compounds, DNA polymerase
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