Developing the Polycystic Kidney Disease Interactome

Candidate: Mackenzie Brauer

Supervisor: Dr. Gagan Gupta

Abstract:
Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a genetic disease that causes numerous renal cysts to form and enlarge with time. This results in renal failure in adults and a reduced life expectancy. The PKD-causing genes, polycystic kidney disease-1 and -2 (PKD1 and PKD2), encode for proteins polycystin-1 and -2 (PC1 and PC2)­­. PC1 is a large membrane bound receptor involved in the regulation of various signaling pathways and biological functions. It has a large extracellular N-terminal region, 11 transmembrane domains and a short intracellular C-terminal. PC2 is a non-selective cation channel of the transient receptor potential family. It has short intracellular N- and C-termini, 6 transmembrane domains and a pore-forming region for channeling cations. PC1 and PC2 form a complex through a coiled-coil domain on their C-termini at the primary cilia. This complex has been shown to be necessary in renal cells as mutations abolishing the interaction impair polycystin localization to cilia and contribute to cystogenesis. We sought to characterize the PC1 PC2 complex binding partners, with the hope of identifying novel protein interactors. We report the first use of proximity-dependent biotinylation for identification (BioID) to characterize the PC1 PC2 coiled-coil complex interactome in HEK 293 cells. We identify 303 high confidence PKD1-interacting partners along its C-terminus, 611 high confidence PKD2-interactin partners along its C-terminus and 446 high confidence PKD2-interacting partners along its N-terminus. Of these interactions, gene ontology reveals an enrichment of cilia-related, transporter activity and trafficking genes. We report interesting interactions, specifically with PC1, which include: the Biogenesis of Lysosome-Related Organelles Complex-1 (BLOC-1) subunits, the BLOC-One-Related Complex (BORC) subunits, Nesprin-2 (SYNE2), SEC63 Protein Translation Regulator and Oxysterol Binding Protein Like 8 (OSBPL8). We hypothesize that each interaction likely contributes to either PC1 membrane traffic or function. We also report several endoplasmic reticulum (ER)-related proteins, specifically with PC2, that likely contribute to its ER localization and function. Overall, this research hopes to contribute information to how PC1 and PC2 traffic to cilia, as well as their function as a complex. Future localization, knock out, and interaction techniques will be used to characterize each gene’s association with PKD1 and/or PKD2, ultimately contributing information towards understanding disease mechanism and find a cure.