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Enhanced Repair of O⁶-Methylguanine DNA Adducts in the Liver of Transgenic Mice Expressing the ada Gene¹

Department of Medicine and the Ireland Cancer Center, Case Western Reserve University and University Hospitals of Cleveland, Cleveland, Ohio 44106 [L. L. D., C. A., K. N., S. L. G.], and the Edison Biotechnology Center, Ohio University, Athens, Ohio 45701 [J. Y., T. W.]

The capacity to repair O⁶-methylguanine-DNA adducts was measured in the liver of transgenic mice expressing a chimeric gene consisting of the inducible P-enolpyruvate carboxykinase (GTP) promoter linked to the bacterial O⁶-alkylguanine-DNA alkyltransferase (ada) gene. Under induced conditions, total hepatic alkyltransferase reached 32.8 ± 4.2 (SE) fmol/µg DNA compared to 7.8 ± 1.1 fmol/µg DNA in nontransgenic mice. Administration of methylnitrosourea or nitrosodimethylamine to both groups of mice produced O⁶-methylguanine-DNA adducts which resulted in repair-mediated depletion of total hepatic alkyltransferase in a dose-dependent fashion. In nontransgenic mice, depletion of hepatic alkyltransferase occurred at lower doses of carcinogen, and recovery of alkyltransferase activity occurred later than in ada⁺ transgenic mice. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of residual alkyltransferase activity after methylating agent exposure indicated that the bacterial as well as endogenous mammalian alkyltransferases were functioning as DNA repair proteins in hepatocytes in vivo. Analysis of O⁶-methylguanine- and N⁷-methylguanine-DNA adducts in the liver of transgenic and nontransgenic mice after treatment with one dose of 50 mg/kg methylnitrosourea i.p. revealed that trangenic mice repaired in situ O⁶-methylguanine-DNA adducts approximately 3 times faster than nontransgenic mice, commensurate with the increase in alkyltransferase activity. Thus, ada⁺ transgenic mice treated with methylnitrosourea have lower levels of persistent mutagenic O⁶-methylguanine adducts than ada^- nontransgenic mice. Hepatic expression of bacterial alkyltransferase appears to protect mice from the DNA-damaging effects of N-nitroso compounds in vivo.

¹ Supported by Grants CA45609 (S. L. G.), ESCA00134 (S. L. G.), CA08644 (L. L. D.), and P30CA-43703 from the NIH and RD-314 (S. L. G.), CN-34 (S. L. G.), and CG1962 (L. L. D.) from the American Cancer Society.

² Recipient of an American Society of Clinical Oncology Young Investigator Award.

³ To whom requests for reprints should be addressed, at Departments of Medicine, Ireland Cancer Center, University Hospitals of Cleveland, 2074 Abingdon Rd., Cleveland, OH 44106. Recipient of a Mallinckrodt Foundation Scholar Award.

Received 12/ 3/90. Accepted 4/23/91.

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				L. Dumenco, E Allay, K Norton, and S. Gerson The prevention of thymic lymphomas in transgenic mice by human O6-alkylguanine-DNA alkyltransferase Science, January 8, 1993; 259(5092): 219 - 222. [Abstract] [PDF]