title: Mutator phenotype in Msh2-deficient murine embryonic fibroblasts.
abstract: Embryonic fibroblast cell lines were established from mice deficient, heterozygous, or proficient for Msh2, one of the three known DNA mismatch repair genes involved in hereditary nonpolyposis colon cancer (HNPCC). Cell lines were established by transfection of primary mouse embryo fibroblasts with E7 and Ras oncogenes or mutant p53. Spontaneously immortalized cells derived from the primary cultures were also studied. To determine whether these cells developed a mutator phenotype similar to that found in colon cancer cells deficient in mismatch repair, we measured mutation rates, microsatellite instability, and sensitivities to a range of DNA-damaging agents. The mutator phenotype detected in the E7 and Ras or mutant p53-immortalized Msh2-/- mouse cells was similar to that found in human mismatch repair-deficient colorectal carcinoma cell lines. Mutation rates to ouabain resistance were increased 8-12-fold relative to lines from Msh2+/+ mice, and microsatellite instability was detectable in 12-18% of subclones derived from the Msh2-/- line but was undetectable in subclones developed from the Msh2+/+ line. Furthermore, E7 and Ras or spontaneously immortalized Msh2-/- cells were significantly more resistant to the cytotoxic effects of 6-thioguanine relative to Msh2+/+ cells. In contrast, these lines showed various responses to UV light and cis-platinum, suggesting that mismatch repair deficiency was not the sole determinant for sensitivity to these DNA-damaging agents. Particular attention was paid to the properties of cells heterozygous for the Msh2 mutant gene, which would mimic the situation of an HNPCC carrier. However, our studies failed to reveal any properties of these cells that might provide a growth advantage or predispose them for the acquisition of further mutations. This observation is consistent with the model that inactivation of the wild-type Msh2 allele is a critical step for tumorigenesis in HNPCC patients.
authors.affiliation: Ontario Cancer Institute/Amgen Institute, Department of Medical Biophysics, University of Toronto, Canada.
authors.forename: A H
authors.initials: AH
authors.lastname: Reitmair
authors.forename: R
authors.initials: R
authors.lastname: Risley
authors.forename: R G
authors.initials: RG
authors.lastname: Bristow
authors.forename: T
authors.initials: T
authors.lastname: Wilson
authors.forename: A
authors.initials: A
authors.lastname: Ganesh
authors.forename: A
authors.initials: A
authors.lastname: Jang
authors.forename: J
authors.initials: J
authors.lastname: Peacock
authors.forename: S
authors.initials: S
authors.lastname: Benchimol
authors.forename: R P
authors.initials: RP
authors.lastname: Hill
authors.forename: T W
authors.initials: TW
authors.lastname: Mak
authors.forename: R
authors.initials: R
authors.lastname: Fishel
authors.forename: M
authors.initials: M
authors.lastname: Meuth
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents; D000970:Antineoplastic Agents / Q000494:pharmacology; D002460:Cell Line; D002934:Cinnamates; D002945:Cisplatin / Q000494:pharmacology; D003123:Colorectal Neoplasms, Hereditary Nonpolyposis / Q000235:genetics*; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D004351:Drug Resistance / Q000235:genetics; D005347:Fibroblasts / Q000187:drug effects / Q000473:pathology / Q000528:radiation effects; D016158:Genes, p53 / Q000235:genetics; D011905:Genes, ras / Q000235:genetics; D006801:Humans; D006921:Hygromycin B / Q000031:analogs & derivatives; D051379:Mice; D008810:Mice, Inbred C57BL; D018895:Microsatellite Repeats / Q000235:genetics; D051718:MutS Homolog 2 Protein; D016296:Mutagenesis / Q000235:genetics*; D010641:Phenotype; D011518:Proto-Oncogene Proteins / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology; D014162:Transfection; D014410:Tumor Stem Cell Assay
chemical_list: D000900:Anti-Bacterial Agents; D000970:Antineoplastic Agents; D002934:Cinnamates; D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D011838:Radiation-Sensitizing Agents; D006921:Hygromycin B; C026273:hygromycin A; C497172:MSH2 protein, human; C497173:Msh2 protein, mouse; D051718:MutS Homolog 2 Protein; D002945:Cisplatin