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Correction of Hypermutability, N-Methyl-N'-nitro-N-nitrosoguanidine Resistance, and Defective DNA Mismatch Repair by Introducing Chromosome 2 into Human Tumor Cells with Mutations in MSH2 and MSH6¹

Laboratories of Molecular Genetics [A. U., T. A. K.], Molecular Carcinogenesis [M. K., J. I. R., J. C. B.], and Environmental Carcinogenesis and Mutagenesis [W. E. G., K. R. T.], National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709; Curricula in Genetics and Molecular Biology [J. I. R., J. C. B.] and Toxicology [W. E. G., K. R. T.], University of North Carolina, Chapel Hill, North Carolina 27599; Department of Medicine, University of California at San Diego, La Jolla, California 92093-0688 [C. R. B.]; and Charles A. Dana Division of Human Cancer Genetics, Dana-Farber Cancer Institute, Boston, Massachusetts 02115 [R. D. K.]

The human DNA mismatch repair genes hMSH2 and hMSH6 encode the proteins that, together, bind to mismatches to initiate repair of replication errors. Human tumor cells containing mutations in these genes have strongly elevated mutation rates in selectable genes and at microsatellite loci, although mutations in these genes cause somewhat different mutator phenotypes. These cells are also resistant to killing by certain drugs and are defective in mismatch repair. Because the elevated mutation rates in these cells may lead to mutations in additional genes that are causally related to the other defects, here we attempt to establish a cause-effect relationship between the hMSH2 and hMSH6 gene mutations and the observed phenotypes. The endometrial tumor cell line HEC59 contains mutations in both alleles of hMSH2. The colon tumor cell line HCT15 contains mutations in hMSH6 and also has a sequence change in a conserved region of the coding sequence for DNA polymerase , a replicative DNA polymerase. We introduced human chromosome 2 containing the wild-type hMSH2 and hMSH6 genes into HEC59 and HCT15 cells. Introduction of chromosome 2 to HEC59 cells restored microsatellite stability, sensitivity to N-methyl-N'-nitro-N-nitrosoguanidine treatment, and mismatch repair activity. Transfer of chromosome 2 to HCT15 cells also reduced the mutation rate at the HPRT locus and restored sensitivity to N-methyl-N'-nitro-N-nitrosoguanidine treatment and mismatch repair activity. The results demonstrate that the observed defects are causally related to mutations in genes on chromosome 2, probably hMSH2 or hMSH6, but are not related to sequence changes in other genes, including the gene encoding DNA polymerase .

¹ This work was supported in part by NIH Grant CA72851 (to C. R. B.).

² To whom requests for reprints should be addressed. Phone: (919) 541-2644; Fax: (919) 541-7613; E-mail: kunkel@niehs.nih.gov.

Received 2/27/97. Accepted 7/15/97.

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