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Mutational-reporter transgenes rescued from mice lacking either Mgmt, or both Mgmt and Msh6 suggest that O6-alkylguanine-induced miscoding does not contribute to the spontaneous mutational spectrum

Oncogene volume 23, pages 5931–5940 (2004)Cite this article

Abstract

O6-methylguanine methyltransferase, Mgmt, constitutes the first line of defense against O6-alkylguanine, which can result in G : C to A : T transitions upon DNA replication. Mgmt has been found in organisms as diverse as archaebacteria and mammals. This evolutionary conservation suggests that all organisms may be exposed to either endogenous or environmental alkylating agents. We thus hypothesized that tissues of Mgmt−/− mice would exhibit elevated mutant frequencies. Employing the Big Blue™ transgenic system, we evaluated lacI mutants rescued from liver and small intestinal DNA of young Mgmt−/− mice. Interestingly, while there was a small difference between Mgmt−/− mice and controls with respect to lacI mutant frequency, no differences attributable to Mgmt deficiency were apparent in the mutational spectra. Although mutations stemming from O6-guanine alkylations would be predicted to be cumulative, we found no evidence of an Mgmt-dependent alteration in mutation spectrum in DNA samples from 12 month-old mice. To optimize our ability to detect mutations resulting from O6-alkylguanine-induced G : T mismatches, mice with combined deficiencies of Mgmt and the DNA mismatch repair molecule, Msh6, were analysed. In spite of this strategy, we observed no significant differences between Mgmt−/− Msh6−/− and Msh6−/− mouse lacI mutations, except for a trend towards a greater percentage (of total transitions) of G : C to A : T changes in Mgmt−/−Msh6−/− livers. Therefore, despite the striking evolutionary conservation of Mgmt, deficiency of this gene did not significantly impact the spontaneous lacI mutational spectrum in vivo.

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Abbreviations

IDLs:

insertion/deletion loops

MCA:

Monte Carlo analysis

Mf:

mutant frequency

Mgmt:

O6-methylguanine-DNA methyltransferase

MMR:

mismatch repair

MSH:

MutS homolog

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Acknowledgements

We are grateful to W Edelmann for the Msh6-deficient mice, and to S Lines and S Chan for generating the various genetic crosses and caring for the animal colonies. This work was supported by the National Cancer Institute of Canada with funds from the Canadian Cancer Society, and by an Alberta Heritage Foundation for Medical Research (AHFMR) establishment grant (both to FRJ). LES and HAL were supported by AHFMR post-doctoral training awards, and FRJ was the recipient of AHFMR Medical Scientist and Canada Research Chair awards. LDS is the Ellison American Cancer Society Research Professor. Generation of the Mgmt-null mouse was supported by National Institutes of Health Grants CA75576 and CA55042 (to LDS).

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of Calgary, Calgary, T2N 4N1, Alberta, Canada

    Linda E Sandercock, Melvin CH Kwok, H Artee Luchman, Sean C Mark, Jennette L Giesbrecht & Frank R Jirik

  2. Biological Engineering Division and Center for Environmental Health Sciences, Massachusetts Institute of Technology, Cambridge, 02139, MA, USA

    Leona D Samson

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  1. Linda E Sandercock
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Correspondence to Frank R Jirik.

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Sandercock, L., Kwok, M., Artee Luchman, H. et al. Mutational-reporter transgenes rescued from mice lacking either Mgmt, or both Mgmt and Msh6 suggest that O6-alkylguanine-induced miscoding does not contribute to the spontaneous mutational spectrum. Oncogene 23, 5931–5940 (2004). https://doi.org/10.1038/sj.onc.1207791

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