Role of DNA repair in carcinogen-induced ras mutation

Abstract

In this contribution we discuss the gene- and cell type-specific repair of miscoding DNA alkylation products as a risk parameter in both mutation induction and malignant transformation by N-nitroso carcinogens. Upon exposure to N-nitroso compounds such as N-methyl- N-nitrosourea (MeNU) or N-ethyl- N-nitrosourea (EtNU), about a dozen different alkylation products are formed in cellular DNA. Among these are O 6-methylguanine ( O 6-MeGua) and O 6-ethylguanine ( O 6-EtGua), respectively, which differ only by one CH 2 group in their alkyl residue and, when unrepaired, cause G:C→A:T transition mutations by anomalous base pairing during DNA replication. We have analyzed the global and gene-specific repair of O 6-MeGua and O 6-EtGua in target cell DNA, ras gene mutation frequencies, and tumor incidence, in the model of mammary carcinogenesis induced in 50-day-old female Sprague–Dawley rats by a single application of MeNU or EtNU. Both carcinogens induce histologically indistinguishable mammary adenocarcinomas at high yield. In the target mammary epithelia, O 6-MeGua is repaired at similar slow rates in both transcriptionally active genes (Ha- ras, β-actin), silent genes ( lgE heavy chain), and in bulk DNA, by the one-step repair protein O 6-alkylguanine-DNA alkyltransferase (MGMT; low level of expression in the target cells). The slow repair of O 6-MeGua translates into a high frequency of mutations at the central position of Ha- ras codon 12 (G G A) in MeNU-induced tumors. O 6-EtGua, however, is removed ∼20 times faster than O 6-MeGua selectively from transcribed genes via an MGMT independent, as yet uncharacterized excision mechanism. Accordingly, no Ha- ras codon 12 mutations are found in the EtNU-induced mammary tumors. Neither MeNU- nor EtNU-induced tumors exhibit mutations at codons 13 and 61 of Ha- ras or at codons 12, 13 and 61 of Ki- ras. While a moderate surplus MGMT activity of the target cells — contributed by a bacterial MGMT transgene ( ada) — significantly counteracts mammary tumorigenesis in MeNU-exposed rats, this is not the case in the EtNU-treated animals. Differential repair of structurally distinct DNA lesions in transcribed or (temporarily) silent genes thus determines the probability of mutation and, together with cell type-specific and interindividual differences in DNA repair capacity, influences carcinogenic risk.