Advertisement

Molecular characterization of an aquaporin-2 mutation causing a severe form of nephrogenic diabetes insipidus

  • Emel Saglar OzerEmail author
  • Hanne B. MoellerEmail author
  • Tugce Karaduman
  • Robert A. Fenton
  • Hatice Mergen
Original Article

Abstract

The water channel aquaporin 2 (AQP2) is responsible for water reabsorption by kidney collecting duct cells. A substitution of amino acid leucine 137 to proline in AQP2 (AQP2-L137P) causes Nephrogenic Diabetes Insipidus (NDI). This study aimed to determine the cell biological consequences of this mutation on AQP2 function. Studies were performed in HEK293 and MDCK type I cells, transfected with wildtype (WT) AQP2 or an AQP2-L137P mutant. AQP2-L137P was predominantly detected as a high-mannose form of AQP2, whereas AQP2-WT was observed in both non-glycosylated and complex glycosylated forms. In contrast to AQP2-WT, the AQP2-L137P mutant did not accumulate on the apical plasma membrane following stimulation with forskolin. Ubiquitylation of AQP2-L137P was different from AQP2-WT, with predominance of non-distinct protein bands at various molecular weights. The AQP2-L137P mutant displayed reduced half-life compared to AQP2-WT. Treatment of cells with chloroquine increased abundance of AQP2-WT, but not AQP2-L137P. In contrast, treatment with MG132 increased abundance of AQP2-L137P but not AQP2-WT. Xenopus oocytes injected with AQP2-WT had increased osmotic water permeability when compared to AQP2-L137P, which correlated with lack of the mutant form in the plasma membrane. From the localization of the mutation and nature of the substitution it is likely that AQP2-L137P causes protein misfolding, which may be responsible for the observed functional defects. The data suggest that the L137P mutation results in altered AQP2 protein maturation, increased AQP2 degradation via the proteasomal pathway and limited plasma membrane expression. These combined mechanisms are likely responsible for the phenotype observed in this class of NDI patients.

Keywords

Aquaporin 2 Water channel Nephrogenic diabetes insipidus Mutation Xenopus oocytes 

Notes

Acknowledgments

Christian Westberg, Helle Høyer and Tina Drejer are thanked for technical assistance. Christian Brix Folsted Andersen and Kristian Stødkilde-Jørgensen are thanked for discussion and insights into Pymol Software. Nuhan Puralı, Berk Saglam and Bora Ergin are thanked for help with oocyte experiments.

Funding

This research was funded by The Scientific and Technological Research Council of Turkey (Project number: 115S499). Emel Saglar Ozer was supported by an EMBO Short Term Fellowship at the Department of Biomedicine, Aarhus University, Aarhus, Denmark (ASFT No: 583-2014). Further funding is provided by the Danish Medical Research Council, The Novo Nordisk Foundation and the Lundbeck Foundation.

References

  1. 1.
    Bourque CW (2008) Central mechanisms of osmosensation and systemic osmoregulation. Nat Rev Neurosci 9(7):519–531CrossRefGoogle Scholar
  2. 2.
    Jung HJ, Kwon TH (2016) Molecular mechanisms regulating aquaporin-2 in kidney collecting duct. Am J Physiol Renal Physiol 311(6):F1318–F1328CrossRefGoogle Scholar
  3. 3.
    Hoffert JD et al (2006) Quantitative phosphoproteomics of vasopressin-sensitive renal cells: regulation of aquaporin-2 phosphorylation at two sites. Proc Natl Acad Sci USA 103(18):7159–7164CrossRefGoogle Scholar
  4. 4.
    Fushimi K, Sasaki S, Marumo F (1997) Phosphorylation of serine 256 is required for cAMP-dependent regulatory exocytosis of the aquaporin-2 water channel. J Biol Chem 272(23):14800–14804CrossRefGoogle Scholar
  5. 5.
    Hoffert JD et al (2008) Vasopressin-stimulated increase in phosphorylation at Ser269 potentiates plasma membrane retention of aquaporin-2. J Biol Chem 283(36):24617–24627CrossRefGoogle Scholar
  6. 6.
    Kamsteeg EJ et al (2006) Short-chain ubiquitination mediates the regulated endocytosis of the aquaporin-2 water channel. Proc Natl Acad Sci USA 103(48):18344–18349CrossRefGoogle Scholar
  7. 7.
    Wu Q et al (2018) CHIP regulates aquaporin-2 quality control and body water homeostasis. J Am Soc Nephrol 29(3):936–948Google Scholar
  8. 8.
    Bockenhauer D, Bichet DG (2015) Pathophysiology, diagnosis and management of nephrogenic diabetes insipidus. Nat Rev Nephrol 11(10):576CrossRefGoogle Scholar
  9. 9.
    Milano S et al (2017) Hereditary nephrogenic diabetes insipidus: pathophysiology and possible treatment. An update. Int J Mol Sci 18(11):2385CrossRefGoogle Scholar
  10. 10.
    Moeller HB, Rittig S, Fenton RA (2013) Nephrogenic diabetes insipidus: essential insights into the molecular background and potential therapies for treatment. Endocr Rev 34(2):278–301CrossRefGoogle Scholar
  11. 11.
    Bichet DG et al (2012) Aquaporin-2: new mutations responsible for autosomal-recessive nephrogenic diabetes insipidus-update and epidemiology. Clin Kidney J 5(3):195–202CrossRefGoogle Scholar
  12. 12.
    Marr N et al (2002) Cell-biologic and functional analyses of five new aquaporin-2 missense mutations that cause recessive nephrogenic diabetes insipidus. J Am Soc Nephrol 13(9):2267–2277CrossRefGoogle Scholar
  13. 13.
    Mulders SM et al (1998) An aquaporin-2 water channel mutant which causes autosomal dominant nephrogenic diabetes insipidus is retained in the Golgi complex. J Clin Invest 102(1):57–66CrossRefGoogle Scholar
  14. 14.
    Marr N et al (2002) Heteroligomerization of an aquaporin-2 mutant with wild-type aquaporin-2 and their misrouting to late endosomes/lysosomes explains dominant nephrogenic diabetes insipidus. Hum Mol Genet 11(7):779–789CrossRefGoogle Scholar
  15. 15.
    Calvanese L et al (2018) Structural basis for mutations of human aquaporins associated to genetic diseases. Int J Mol Sci 19(6):1577CrossRefGoogle Scholar
  16. 16.
    Duzenli D et al (2012) Mutations in the AVPR2, AVP-NPII, and AQP2 genes in Turkish patients with diabetes insipidus. Endocrine 42(3):664–669CrossRefGoogle Scholar
  17. 17.
    Rosenbaek LL et al (2014) Phosphorylation decreases ubiquitylation of the thiazide-sensitive cotransporter NCC and subsequent clathrin-mediated endocytosis. J Biol Chem 289(19):13347–13361CrossRefGoogle Scholar
  18. 18.
    Moeller HB et al (2014) Phosphorylation and ubiquitylation are opposing processes that regulate endocytosis of the water channel aquaporin-2. J Cell Sci 127(Pt 14):3174–3183CrossRefGoogle Scholar
  19. 19.
    Moeller HB et al (2010) Phosphorylation of aquaporin-2 regulates its endocytosis and protein–protein interactions. Proc Natl Acad Sci USA 107(1):424–429CrossRefGoogle Scholar
  20. 20.
    Poulsen SB et al (2013) Long-term vasopressin-V2-receptor stimulation induces regulation of aquaporin 4 protein in renal inner medulla and cortex of Brattleboro rats. Nephrol Dial Transplant 28(8):2058–2065CrossRefGoogle Scholar
  21. 21.
    Moeller HB et al (2016) Regulation of the water channel Aquaporin-2 via 14-3-3theta and -zeta. J Biol Chem 291(5):2469–2484CrossRefGoogle Scholar
  22. 22.
    Zeuthen T et al (1997) Water transport by the Na +/glucose cotransporter under isotonic conditions. Biol Cell 89(5–6):307–312Google Scholar
  23. 23.
    Duquette PP, Bissonnette P, Lapointe JY (2001) Local osmotic gradients drive the water flux associated with Na(+)/glucose cotransport. Proc Natl Acad Sci USA 98(7):3796–3801CrossRefGoogle Scholar
  24. 24.
    Guyon C et al (2009) Characterization of D150E and G196D aquaporin-2 mutations responsible for nephrogenic diabetes insipidus: importance of a mild phenotype. Am J Physiol Renal Physiol 297(2):F489–F498CrossRefGoogle Scholar
  25. 25.
    El Tarazi A et al (2016) Functional recovery of AQP2 recessive mutations through hetero-oligomerization with wild-type counterpart. Sci Rep 6:33298CrossRefGoogle Scholar
  26. 26.
    Leduc-Nadeau A et al (2007) Elaboration of a novel technique for purification of plasma membranes from Xenopus laevis oocytes. Am J Physiol Cell Physiol 292(3):C1132–C1136CrossRefGoogle Scholar
  27. 27.
    Janson G et al (2017) PyMod 2.0: improvements in protein sequence-structure analysis and homology modeling within PyMOL. Bioinformatics 33(3):444–446Google Scholar
  28. 28.
    Frick A et al (2014) X-ray structure of human aquaporin 2 and its implications for nephrogenic diabetes insipidus and trafficking. Proc Natl Acad Sci USA 111(17):6305–6310CrossRefGoogle Scholar
  29. 29.
    Hirano K et al (2003) The proteasome is involved in the degradation of different aquaporin-2 mutants causing nephrogenic diabetes insipidus. Am J Pathol 163(1):111–120CrossRefGoogle Scholar
  30. 30.
    Deen PM et al (1995) Water channels encoded by mutant aquaporin-2 genes in nephrogenic diabetes insipidus are impaired in their cellular routing. J Clin Invest 95(5):2291–2296CrossRefGoogle Scholar
  31. 31.
    Baumgarten R et al (1998) Glycosylation is not essential for vasopressin-dependent routing of aquaporin-2 in transfected Madin–Darby canine kidney cells. J Am Soc Nephrol 9(9):1553–1559Google Scholar
  32. 32.
    Mulders SM et al (1997) New mutations in the AQP2 gene in nephrogenic diabetes insipidus resulting in functional but misrouted water channels. J Am Soc Nephrol 8(2):242–248Google Scholar
  33. 33.
    Preston GM, Brodsky JL (2017) The evolving role of ubiquitin modification in endoplasmic reticulum-associated degradation. Biochem J 474(4):445–469CrossRefGoogle Scholar
  34. 34.
    Feige MJ, Hendershot LM (2011) Disulfide bonds in ER protein folding and homeostasis. Curr Opin Cell Biol 23(2):167–175CrossRefGoogle Scholar
  35. 35.
    Moeller HB et al (2009) Role of multiple phosphorylation sites in the COOH-terminal tail of aquaporin-2 for water transport: evidence against channel gating. Am J Physiol Renal Physiol 296(3):F649–F657CrossRefGoogle Scholar
  36. 36.
    Tamarappoo BK, Verkman AS (1998) Defective aquaporin-2 trafficking in nephrogenic diabetes insipidus and correction by chemical chaperones. J Clin Invest 101(10):2257–2267CrossRefGoogle Scholar
  37. 37.
    Yang B, Zhao D, Verkman AS (2009) Hsp90 inhibitor partially corrects nephrogenic diabetes insipidus in a conditional knock-in mouse model of aquaporin-2 mutation. FASEB J 23(2):503–512CrossRefGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Department of Biology, Faculty of ScienceHacettepe UniversityAnkaraTurkey
  2. 2.Department of BiomedicineAarhus UniversityAarhusDenmark

Personalised recommendations