The Functional characterization of Msh6 missense variants in Saccharomyces cerevisiae.
Board Location: #34
Discipline: Biological Sciences
Subcategory: Genetics
Session: 4
Maya H. Garner - Xavier University of Louisiana
Co-Author(s): Mohammed Mona, Xavier University of Louisiana, New Orleans, LA; Naser Raslan, Xavier University of Louisiana, New Orleans, LA; Joanna E. Haye-Bertolozzi, Xavier University of Louisiana, New Orleans, LA
During DNA replication mismatches between bases that are not fixed by the DNA polymerase are recognized by the DNA mismatch repair (MMR) proteins. Failure to fix mismatches can result in increased mutation frequency and result in disease. Mutations in DNA MMR genes can result in cancer syndromes such as Lynch syndrome (LS). LS accounts for 3% to 5% of colorectal cancers and other cancers such as ovarian, and endometrial. In eukaryotes, mismatches are recognized by MutSa (Msh6 and Msh2). Msh6 missense variants currently classified as Variants of Unknown Significance (VUS) require further testing to understand the role they may play in cancer development. Due to the high degree of conservation between Saccharomyces cerevisiae (yeast) and humans, yeast can be used to aid in the understanding of the pathogenicity of Msh6 missense variants. The InSiGHT database was used to select Msh6 missense VUSs that were identified in LS patients. Using Site-directed mutagenesis, mutations were created at the equivalent position in the yeast MSH6 gene. Msh6 missense variants were tested for MMR functionality using the CAN1 forward mutation and fluctuation assays. Variants were compared to wild-type MSH6 and a strain that lacked the MSH6 gene. From this analysis, we have identified variants that are classified as wild-type like, as well as mutants that are defective in DNA MMR. Though we have successfully identified some defective missense variants of Msh6, further analysis will include testing the effect of these mutations on Msh6 protein levels in the cell and nuclear localization. These experiments will aid in the understanding of the role these variants play in cancer development, which could further inform diagnoses of patients with LS.
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Funder Acknowledgement(s): Support for this research was partially provided by the LSAMP and the NIH SuRE-FIRST grant to J. Haye.
Faculty Advisor: Joanna E. Haye-Bertolozzi, jhaye@xula.edu
Role: I have performed both qualitative and quantitative analyses of a subset of the Msh6 missense variants.