Abstract: PO1510
Mechanisms of Tethered-Ligand Mediated Polycystin 1 GPCR Signaling
Session Information
- Cystic Kidney Diseases: Mechanisms, Genetics, and Treatment
October 22, 2020 | Location: On-Demand
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1001 Genetic Diseases of the Kidneys: Cystic
Authors
- Pawnikar, Shristi, University of Kansas, Lawrence, Kansas, United States
- Magenheimer, Brenda S., University of Kansas Medical Center, Kansas City, Kansas, United States
- Maser, Robin L., University of Kansas Medical Center, Kansas City, Kansas, United States
- Miao, Yinglong, University of Kansas, Lawrence, Kansas, United States
Background
Polycystin-1 (PC1) is the most commonly mutated protein in autosomal dominant polycystic kidney disease (ADPKD) thought to function as an atypical GPCR. In vivo studies have demonstrated PC1-regulated G protein signaling is a critical function for preventing renal cystogenesis. Like the Adhesion class of GPCRs, PC1 undergoes auto-catalyzed cleavage at a GPS motif which generates an extracellular N-terminal fragment and a membrane-embedded C-terminal fragment (CTF) composed of 11 transmembrane domains preceded by an N-terminal extracellular stalk of 25 residues. We previously reported that CTF-mediated signaling to an NFAT promoter-luciferase reporter is dependent on the presence of the stalk, is reduced by ADPKD-associated missense mutations within the stalk, and can be rescued by synthetic, stalk-derived peptides, supporting a tethered ligand mechanism of PC1-G protein signaling. In this study, we have utilized a combination of computational molecular dynamics (MD) simulations and mutation-function analyses to investigate the mechanism of PC1 signaling.
Methods
A computer model of the human PC1 CTF was generated using the cryo-EM structure of the PC1-PC2 complex (Su et al, 2019) and the I-TASSER protein structure prediction tool. All-atom enhanced simulations (> 900 ns) using a robust Gaussian accelerated molecular dynamics (GaMD) technique were performed followed by calculation of residue correlation matrices and free energy profiles for residue-residue interactions.
Results
All-atom simulations with the wildtype, ADPKD mutant and stalk-less versions of the PC1 CTF model were consistent with the previous functional signaling data. Key residue interactions between different domains of the CTF predicted from the GaMD simulations suggest an allosteric mechanism for PC1-G protein activation for which mutagenesis and functional signaling and expression assays are underway.
Conclusion
Complementary experiments and simulations have provided important insights into a mechanism of PC1 GPCR signaling at an atomic level and support an important role of the stalk region as a tethered ligand. This in-depth knowledge is expected to facilitate future drug design efforts targeting this function of PC1 for more effective treatments of ADPKD.
Funding
- NIDDK Support