Cleavage and trafficking of polycystin-1

Polycystin-1 (PC1) and polycystin-2 (PC2) are membrane proteins implicated in Autosomal Dominant Polycystic Kidney Disease, a genetic disease that causes the progressive formation of fluid-filled cysts in the kidney. Disease-causing mutations alter the function, localization or expression of PC1 or PC2, and can decrease the ability of either protein to regulate signaling pathways involved in cellular growth and differentiation. Given that disease correlates with PC1 mis-localization, I set out to understand the factors that regulate PC1 trafficking to the plasma membrane. Correct PC1 physiologic function requires autocatalytic cleavage at its G protein-coupled receptor proteolytic site (GPS). My results suggest that this cleavage is stimulated by co-expression of PC1 with PC2. Importantly, this increased cleavage correlates with the appearance of PC1 at the plasma membrane, and specifically at the apical and ciliary membranes in polarized cells.;PC2 is a member of the trp family of cation channels and is an important binding partner of PC1. Targeted mutations in the PC2 sequence show that PC2's effect on PC1 localization is apparently independent of its channel activity. Although PC1 and PC2 have been shown to interact with one another through their C-terminal tail domains, I use truncated PC1 and PC2 constructs to show that the surface localization of PC1 is independent of PC2 interactions mediated by the either protein's C-terminal tail. I also describe results using a novel protocol that shows PC1 molecules delivered to the plasma membrane over a specific time frame. This technique reveals that blocking purinergic signaling decreases the delivery of PC1 to the plasma membrane. In further analysis of PC1's trafficking I show that the delivery of PC1 to the apical membrane delivery is selectively and negatively affected by the transport inhibitor Brefeldin A, but not affected by blocking protein synthesis. In sum, my work shows that PC2 expression is required for the movement of PC1 to the plasma membrane, including the primary cilium. This localization correlates with increased PC1 GPS cleavage but is independent of PC2's channel activity or binding of PC1. Delivery of PC1 to the plasma membrane is negatively affected by inhibiting purinergic signaling or anterograde trafficking to the Golgi, but delivery to the ciliary membrane is apparently insensitive to blocking Golgi transport.