Cleavage of N-terminus of polycystin-1 increases calcium permeability of polycystin-1/2 receptor channel complexes
Runping Wang, Danish Idrees, Mohammad Amir, Biswajit Padhy, Jian Xie, Chou-Long Huang
Runping Wang, Danish Idrees, Mohammad Amir, Biswajit Padhy, Jian Xie, Chou-Long Huang
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Research Article Cell biology Nephrology

Cleavage of N-terminus of polycystin-1 increases calcium permeability of polycystin-1/2 receptor channel complexes

Abstract

Mutations on genes encoding polycystin-1 (PC1) and PC2 cause autosomal-dominant polycystic kidney disease. How these 2 proteins work together to exert anticystogenesis remains elusive. PC1 resembles adhesion G-protein coupled receptors and undergoes autocleavage in the extracellular N-terminus to expose a hidden “stalk” region, which is hypothesized to act as a “tethered agonist.” Here, we show that WT PC1 and PC2 formed functional heteromeric channel complexes in Xenopus oocytes with different biophysical properties from PC2 homomeric channels. Deletion of PC1 N-terminus, which exposed the stalk, increased calcium permeability in PC1/PC2 heteromers that required the presence of stalk. Extracellular application of synthetic stalk peptide increased calcium permeation in stalkless PC1/PC2. Application of Wnt9B protein increased calcium permeability in PC1/PC2 but not in heteromers containing cleavage-resistant mutant PC1. Wnt9B interacted with N-terminal leucine-rich repeat (LRR) of PC1. Pretreatment with LRR blunted the increase in calcium permeability by Wnt9B. Thus, PC1 and PC2 form receptor-channel complexes that is activated by exposure of the stalk region following ligand binding to the PC1 N-terminus. The stalk peptide acts as a tethered agonist to activate PC1/PC2 by affecting ion selectivity of the complexes.

Authors

Runping Wang, Danish Idrees, Mohammad Amir, Biswajit Padhy, Jian Xie, Chou-Long Huang

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Figure 3

GOF mutation F604P-PC2 mutant is different from WT PC2.

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GOF mutation F604P-PC2 mutant is different from WT PC2. (A) Representati...
(A) Representative immunofluorescence (IF) images and mean ± SEM of normalized IF intensity of surface expression of PC2F604P (labeled F604P302HA, n = 11) or F604P302HA/sPC1 (n = 14) as indicated. IF staining using anti-HA antibody against HA tag on the TOP domain as in Figure 2C. Note that the experiments in A were conducted concurrently with those shown in Figure 2, C and D; thus, the intensity of the IF images were comparable. Scale bar: 150 μm. (B) Representative current steps, steady-state I-V curve (at 400 ms), and mean ± SEM for inward and outward currents at –100 and 100 mV in oocytes injected with mRNA for F604P-PC2 (PC2F604P) or PC2F604P/sPC1 as indicated. n = 5 for each group. Experiments were conducted concurrently with those shown in Figure 2B; refer to Figure 2B for the values of the vehicle and sPC1-injected groups. (C) Rectification ratio of PC2 and sPC1/PC2 currents recorded in 100 mM KCl bath. The ratio of outward K+ current (at +100 mV) to inward K+ current (at –100 mV) is shown. n = 13 each group. Inset: I-V curve. (D) Rectification ratio for PC2F604P and sPC1/PC2F604P as illustrated in C. n = 13 each group, inset: I-V curve. (E) Effect of sPC1 carrying R4100E and R4107E mutation (sPC1RR) on inward and outward currents of heteromeric channels formed with PC2 and PC2F604P. n = 4–5 each group. Two-tailed unpaired Student’s t test for A–E. In all panels, experimental number (n) is number of oocytes as shown by scatter plots. All experiments were repeated 2 or more times with similar results.