Developing a new platform to characterize and treat disease-associated polycystin variants

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Autosomal dominant polycystic kidney disease (ADPKD) results from mutations in the genes encoding polycystin 1 (PC1) and polycystin 2 (PC2). The disease places an extraordinary burden on patient quality of life and results in estimated annual healthcare costs of $5.7 billion in the US due to the necessity for renal replacement therapy. Despite knowing the identity of the causative genes, we lack a treatment that directly targets the polycystins. A poor understanding of the molecular defects underlying PC1 and PC2 mutations has thus far prevented the development of tailored therapeutics to treat ADPDK. Disease-causing mutations in PC1 and PC2 lead to the accumulation of large fluid-filled renal cysts, which is followed by an eventual loss of kidney function. PC1 and PC2 are believed to assemble, traffic to, and permit ion passage and signaling from the epithelial cell membrane and the primary apical cilium. Therefore, the absence of functional polycystins at these locations is primarily responsible for ADPKD. To reach the cell surface and function, PC1 and PC2 must be synthesized and fold in the endoplasmic reticulum (ER). Next, the proteins must traffic to the cell surface and function. Although ADPKD-causing mutations can affect any of these steps, this proposal will develop, optimize, and then deploy a tool to classify which disease-causing mutations impede the appearance of functional PC1 at the cell surface. Based on previous analysis of select mutants in PC2, our preliminary data, and emerging work from other labs, this approach will provide new reagents and assays to the scientific and PKD community to study the molecular defects of PC1 mutants. Our novel research tool employs yeast, which will provide a rapid and quantitative read-out for PC1’s impact on channel function. Yeast are an ideal model system due to their rapid generation, genetic malleability, use in high-throughput formats, and the many examples in which studies on disease- causing mutations were translated into human cells and in vivo. To provide a proof-of-principle for the continued development of this tool, we will rapidly and quantitatively classify how ~70 identified mutations in PC1 disrupt PC1:PC2 channel activity. Background data provided in this application indicate that the yeast system faithfully reports on how mutations alter PC2 activity. Preliminary data in the application also show that PC1 can be co- expressed with PC2 in yeast and form a functional channel. This project will help the PKD community test the defects associated with new and emerging PC1 mutants to help springboard the development of personalized therapies for ADPKD.