Carcinogenesis Advance Access originally published online on February 14, 2008
Carcinogenesis 2008 29(4):866-874; doi:10.1093/carcin/bgn030
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Mgmt deficiency alters the in vivo mutational spectrum of tissues exposed to the tobacco carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)
Department of Biochemistry and Molecular Biology, The McCaig Institute for Bone and Joint Health, University of Calgary, 3330 Hospital Drive N.W., Calgary, Alberta, Canada T2N 4N1
1 Division of Environmental Health Sciences and the Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA
* To whom correspondence should be addressed. Tel: +403 220 8666; Fax: +403 210 8100;Email: jirik{at}ucalgary.ca
It has been proposed that O6-methylguanine DNA methyltransferase (MGMT) gene silencing in premalignant lesions and cancers of the lung might result in the acquisition of a ‘mutator’ phenotype. Previously, however, we found that Mgmt–/– mouse DNA failed to show an increase in spontaneous mutations. We thus hypothesized that only during exposure to specific environmental carcinogens would the consequences of MGMT deficiency become evident. Metabolism of the tobacco-derived nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) generates alkylating species that can react with the O6 position of deoxyguanine, thereby yielding substrates for MGMT-mediated repair. To investigate how MGMT might regulate the mutational effects of NNK, Mgmt–/– mice were crossed with a lacI-based transgenic reporter line (Big BlueTM) thus enabling an assessment of the in vivo mutagenic effects of this agent. We observed the induction of a complex spectrum of NNK-dependent lacI mutations in both control and Mgmt–/– tissues, but only a trend in the mutant frequency increases that could be attributed to MGMT deficiency. The mutational spectra of NNK-treated Mgmt–/– lungs revealed an increase in the absolute number of G:C to A:T changes accompanied by a shift in these from CpG to GpG sites, consistent with an SN1 alkylation mechanism. In keeping with the high levels of MGMT expressed in the liver, more pronounced mutagenic effects and greater differences in O6 position of deoxyguanosine adduct levels following NNK were observed in Mgmt–/– versus wild-type mice. Extrapolating to humans, MGMT-deficient cells would likely exhibit an increased mutational burden, but only following exposures to specific environmental mutagens such as NNK.
Abbreviations: dG, deoxyguanosine; dT, deoxythymidine; MCA, Monte Carlo estimation of the P-value of the hypergeometric test; Mf, mutation frequency; MGMT, O6-methylguanine DNA methyltransferase; NNK, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone; O6-mG, O6-methylguanine; O6-pobG, O6-[4-oxo-4-(3-pyridyl)butyl]guanine; WT, wild-type
Received March 21, 2007; revised December 28, 2007; accepted January 24, 2008.