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Kidney Week

Abstract: FR-OR006

Regulation of PKD2 Channel Function by PKD1

Session Information

Category: Genetic Diseases of the Kidneys

  • 1001 Genetic Diseases of the Kidneys: Cystic

Authors

  • Padhy, Biswajit, University of Iowa Carver College of Medicine, Iowa City, Iowa, United States
  • An, Sung Wan, University of Iowa Carver College of Medicine, Iowa City, Iowa, United States
  • Xie, Jian, University of Iowa Carver College of Medicine, Iowa City, Iowa, United States
  • Huang, Chou-Long, University of Iowa Carver College of Medicine, Iowa City, Iowa, United States
Background

Mutations of PKD1 and 2 cause autosomal-dominant polycystic kidney disease (ADPKD). PKD1 peptide has a very large extracellular N-terminus (Nt), 11 transmembrane (TM) domains, and intracellular C-terminus (Ct). PKD2 is 6-TM domains channel with intracellular N and C-terminus. The function and relationship of PKD1 and PKD2 and in the pathogenesis of ADPKD remain elusive. Previous studies show PKD1 and PKD2 interact with its C-terminus. Recent cryo-EM structural studies reveal the last 6 TM of PKD1 can interact with PKD2 to form presumably non-functional channel complexes.

Methods

Wild-type (WT) PKD2 and phenylalanine-604 to proline (F604P) mutant PKD2 were expressed in Xenopus oocytes with or without WT and mutant PKD1. PKD2 channel function was studied by using two-electrode voltage-clamp. Extracellular Ca2+-inhibitable inward K+ currents were measured.

Results

We found that currents in oocytes expressing WT- PKD2 were not different from in control oocytes. WT-PKD1 itself did not produce currents, and had no effects on WT-PKD2 currents when coexpression with it. It is known that F604P mutation on the 5th TM domain of PKD2 leads to widening of the lower gate and constitutive activation of the channel. We then used F604P-PKD2 to study the regulation by PKD1. Oocytes expressing F604P-PKD2 showed ~3 fold higher currents than background currents in control oocytes, confirming gain-of-function of F604P. Coexpression with WT-PKD1 completely inhibited F604P-PKD2, to the level of control oocytes. To map regions of PKD1 that regulate PKD2, we made five PKD1 mutant constructs with deletion of Nt, deletion of Ct, deletion of the last 6 TM domains, deletion of both Nt and Ct or deletion of Nt as well as the first 5 TM domain. Comparing with WT-PKD1, deletion of Nt and/or the first 5 TM did not alter PKD1's ability to inhibit F604P-PKD2. Deletion of Ct or the last 6 TM resulted in partial reduction in the ability of PKD1 to inhibit F604P-PKD2.

Conclusion

PKD1 inhibits PKD2 channel activity. The C-terminus and last 6 TM domains of PKD1 are involved. The results support the implication of structural studies that PKD1 and PKD2 complexes are non-conducting. Further, they support the hypothesis that ligands or factors target PKD1 to activate channel activities of PKD1/PKD2 complexes.

Funding

  • Private Foundation Support