Functional characterization of pathogenic human MSH2 missense mutations in Saccharomyces cerevisiae
Alison E Gammie 1*, Naz Erdeniz 2, Julia Beaver 3, Barbara Devlin 1, Afshan Nanji 4 and Mark D Rose 1
1 Princeton University
2 Oregon Health & Sciences University
3 University of Pennsylvania School of Medicine
4 Johns Hopkins School of Medicine
* To whom correspondence should be addressed. E-mail: agammie{at}princeton.edu.
Submitted on January 17, 2007
Revised on February 12, 2007
Accepted on 13 August 2007
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Abstract |
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Hereditary Non-Polyposis Colorectal Cancer (HNPCC) is associated with defects in DNA mismatch repair. Mutations in either hMSH2 or hMLH1 underlie the majority of HNPCC cases. Approximately 25% of annotated hMSH2 disease alleles are missense mutations, resulting in a single change out of 934 amino acids. We engineered 54 missense mutations in the cognate positions in yeast MSH2 and tested for function. Of the human alleles, 58% conferred strong defects, 5% displayed intermediate defects and 37% showed no defects in mismatch repair assays. 50% of the defective alleles resulted in decreased steady-state levels of the variant Msh2 protein, and 49% of the Msh2 variants lost crucial protein-protein interactions. Finally, 9 positions are predicted to influence the mismatch recognition complex ATPase activity. In summary, the missense mutations leading to loss of mismatch repair defined important structure-function relationships and the molecular analysis revealed the nature of the deficiency for Msh2 variants expressed in the tumors. Of medical relevance are 15 human alleles annotated as pathogenic in public databases that conferred no obvious defects in mismatch repair assays. This analysis underscores the importance of functional characterization of missense alleles to ensure that they are the causative factor for disease.
Key Words:
HNPCC, colorectal cancer, mismatch repair, missense, yeast
