The function(s) of the genes responsible for the vast majority of cases of autosomal dominant polycystic kidney disease (ADPKD) are unknown. Based on sequence analysis, the gene product of the polycystic kidney disease gene 1 (PKD1) has been proposed to encode a protein with a role in cell-cell and/or cell - extracellular matrix interactions while PKD2 is thought to function as a cation channel. We have shown that PKD2 physically associated with PKD1 and only in the presence of PKD1, PKD2 was able to form a Ca++ permeable cation channel. In addition to PKD 1, PKD2 was also able to associate with the transient receptor potential channel 1 (TRPC1). We now show that TRPC1 has a widespread distribution in epithelial structures, primarily the ductal cells of the kidney and liver. TRPC 1 was shown to enhance Ca++ entry in response to store depletion (or capacitative Ca++ entry, CCE). CCE is the major route by which non-excitable cells regulate their intracellular Ca++ concentration. Among the many cellular functions regulated by CCE, regulation of cAMP accumulation is a well-characterized and specific physiological target of CCE. Notably, the involvement of cAMP in cyst formation in kidney and liver epithelial cells has been well established. We propose that PKD1, PKD2 and TRPC1 assemble to a functional complex to enhance CCE. Naturally occurring mutations in PKD2 may result in the disruption of this complex and thereby in alterations in CCE. We will test our model by showing the existence of an endogenous complex and identifying protein-protein interactions responsible for complex assembly. Next, we will measure CCE in cells transfected with PKD1, PKD2 and TRPC1. We will evaluate an effect of PKD2 on CCE by introducing dominant negative constructs of PKD2 in cell lines that endogenously express PKD1, PKD2 and TRPC1 and testing whether wild type or mutant PKD2 can regulate cAMP accumulation in kidney epithelial cells. Our ultimate goal is to develop a biologically significant system that would allow us to probe the mechanisms by which pathogenic mutations in PKD2 alter its normal function, and to design therapeutic interventions in diseases such as ADPKD.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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General Medicine B Study Section (GMB)
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Rasooly, Rebekah S
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University of Oklahoma Health Sciences Center
Anatomy/Cell Biology
Schools of Medicine
Oklahoma City
United States
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