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Mutator phenotypes of yeast strains heterozygous for mutations in the MSH2 gene
K Drotschmann1, A B Clark, H T Tran
1Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, P.O. Box 12233, Research Triangle Park, NC 27709, USA.
Abstract:
Heterozygosity for germ-line mutations in the DNA mismatch repair gene MSH2 predisposes humans to cancer. Here we use a highly sensitive reporter to describe a spontaneous mutator phenotype in diploid yeast cells containing a deletion of only one MSH2 allele. We also identify five MSH2 missense mutations that have dominant mutator effects in heterozygous cells when expressed at normal levels from the natural MSH2 promoter. For example, a 230-fold mutator effect is observed in an MSH2/msh2 diploid strain in which Gly693, which is invariant in MutS homologs and involved in ATP hydrolysis, is changed to alanine. DNA binding data suggest that mismatch repair is suppressed by binding of a mutant Msh2-Msh6 heterodimer to a mismatch with subsequent inability to dissociate from the mismatch in the presence of ATP. A dominant mutator effect also is observed in yeast when Gly693 is changed to serine. An early onset colorectal tumor is heterozygous for the analogous Gly --> Ser mutation in hMSH2, and a second hMSH2 mutation was not found, suggesting that this missense mutation may predispose to cancer via a dominant mutator effect. The mutator effects of the deletion mutant and the Gly --> Ala missense mutant in yeast MSH2 are enhanced by heterozygosity for a missense mutation in DNA polymerase delta that reduces its proofreading activity but is not a mutator in the heterozygous state. The synergistic effects of heterozygosity for mutations in two different genes that act in series to correct replication errors may be relevant to cancer predisposition.
Insights
Germline mutations in DNA mismatch repair gene MSH2 increase cancer risk. This study reveals dominant mutator effects of MSH2 missense mutations in yeast, suggesting a novel mechanism for cancer predisposition.
Area of Science:
- Genetics
- Molecular Biology
- Cancer Research
Background:
- Germline mutations in the DNA mismatch repair gene MSH2 are linked to hereditary cancer syndromes.
- Understanding the mechanisms by which MSH2 mutations contribute to cancer is crucial for risk assessment and prevention.
Purpose of the Study:
- To investigate the mutator effects of MSH2 heterozygous mutations in a diploid yeast model.
- To identify specific MSH2 missense mutations with dominant mutator phenotypes.
- To explore the synergistic effects of MSH2 mutations with DNA polymerase delta proofreading defects.
Main Methods:
- Utilized a sensitive reporter system to detect spontaneous mutations in diploid yeast.
- Generated and analyzed yeast strains with MSH2 deletions and missense mutations.
- Investigated DNA binding and dissociation kinetics of mutant Msh2-Msh6 heterodimers.
- Assessed the impact of combined MSH2 and DNA polymerase delta mutations.
Main Results:
- A single MSH2 allele deletion induced a spontaneous mutator phenotype in yeast.
- Five MSH2 missense mutations exhibited dominant mutator effects in heterozygous cells.
- A specific mutation (Gly693Ala) caused a 230-fold mutator effect, linked to impaired ATP-dependent dissociation of the Msh2-Msh6 complex.
- Synergistic mutator effects were observed when MSH2 mutations were combined with a DNA polymerase delta proofreading-deficient mutation.
Conclusions:
- Heterozygous MSH2 mutations can confer a dominant mutator phenotype, contributing to cancer predisposition.
- Impaired ATP hydrolysis or dissociation of the MSH2 repair complex may underlie the dominant mutator effect.
- Synergistic interactions between DNA mismatch repair and replication proofreading pathways are relevant to cancer development.