Carcinogenesis, Vol. 20, No. 2, 205-214,
February 1999
© 1999 Oxford University Press
Mismatch repair and differential sensitivity of mouse and human cells to methylating agents
1 Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Hertfordshire EN6 3LD, UK and
2 Istituto Superiore di Sanitá, Laboratory of Comparative Toxicology and Ecotoxicology, Viale Regina Elena 299, 00161 Rome, Italy
The long-patch mismatch repair pathway contributes to the cytotoxic effect of methylating agents and loss of this pathway confers tolerance to DNA methylation damage. Two methylation-tolerant mouse cell lines were identified and were shown to be defective in the MSH2 protein by in vitro mismatch repair assay. A normal copy of the human MSH2 gene, introduced by transfer of human chromosome 2, reversed the methylation tolerance. These mismatch repair defective mouse cells together with a fibroblast cell line derived from an MSH2–/– mouse, were all as resistant to N-methyl-N-nitrosourea as repair-defective human cells. Although long-patch mismatch repair-defective human cells were 50- to 100-fold more resistant to methylating agents than repair-proficient cells, loss of the same pathway from mouse cells conferred only a 3-fold increase. This discrepancy was accounted for by the intrinsic N-methyl-N-nitrosourea resistance of normal or transformed mouse cells compared with human cells. The >20-fold differential resistance between mouse and human cells could not be explained by the levels of either DNA methylation damage or the repair enzyme O6-methylguanine–DNA methyltransferase. The resistance of mouse cells to N-methyl-N-nitrosourea was selective and no cross-resistance to unrelated DNA damaging agents was observed. Pathways of apoptosis were apparently intact and functional after exposure to either N-methyl-N-nitrosourea or ultraviolet light. Extracts of mouse cells were found to perform 2-fold less long-patch mismatch repair. The reduced level of mismatch repair may contribute to their lack of sensitivity to DNA methylation damage.
Abbreviations: bzGua, O6-benzylguanine; DMSO, dimethyl sulphoxide; ES, embryonic stem; FISH, fluorescence in situ hybridization; HNPCC, hereditary non-polyposis colorectal cancer; MGMT, O6-methylguanine–DNA methyltransferase; MNNG, N-methyl-N'-nitro-N-nitrosoguanidine; MNU, N-methyl-N-nitrosourea; O6-meGua, O6-methylguanine; PBS, phosphatebuffered saline; SDS, sodium dodecyl sulphate; UV, ultraviolet; 3-meAde, 3-methyladenine.
3 To whom correspondence should be addressed Email: bignami{at}net.iss.it
4 These authors contributed equally to this work
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