Søren Kragh Moestrup

Regulation of the Water Channel Aquaporin-2 via 14-3-3θ and -ζ

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review


  • Hanne B Moeller
  • Joachim Slengerik-Hansen
  • ,
  • Takwa Aroankins
  • ,
  • Mette Assentoft, the Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark.
  • ,
  • Nanna MacAulay, the Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark.
  • ,
  • Soeren K Moestrup
  • Vivek Bhalla, the Division of Nephrology, Department of Medicine, Stanford University, Palo Alto, California 94305.
  • ,
  • Robert A Fenton

The 14-3-3 family of proteins are multifunctional proteins that interact with many of their cellular targets in a phosphorylation-dependent manner. Here, we determined that 14-3-3 proteins interact with phosphorylated forms of the water channel aquaporin-2 (AQP2) and modulate its function. With the exception of σ, all 14-3-3 isoforms were abundantly expressed in mouse kidney and mouse kidney collecting duct cells (mpkCCD14). Long-term treatment of mpkCCD14 cells with the type 2 vasopressin receptor agonist dDAVP increased mRNA and protein levels of AQP2 alongside 14-3-3β and -ζ, whereas levels of 14-3-3η and -θ were decreased. Co-immunoprecipitation (co-IP) studies in mpkCCD14 cells uncovered an AQP2/14-3-3 interaction that was modulated by acute dDAVP treatment. Additional co-IP studies in HEK293 cells determined that AQP2 interacts selectively with 14-3-3ζ and -θ. Use of phosphatase inhibitors in mpkCCD14 cells, co-IP with phosphorylation deficient forms of AQP2 expressed in HEK293 cells, or surface plasmon resonance studies determined that the AQP2/14-3-3 interaction was modulated by phosphorylation of AQP2 at various sites in its carboxyl terminus, with Ser-256 phosphorylation critical for the interactions. shRNA-mediated knockdown of 14-3-3ζ in mpkCCD14 cells resulted in increased AQP2 ubiquitylation, decreased AQP2 protein half-life, and reduced AQP2 levels. In contrast, knockdown of 14-3-3θ resulted in increased AQP2 half-life and increased AQP2 levels. In conclusion, this study demonstrates phosphorylation-dependent interactions of AQP2 with 14-3-3θ and -ζ. These interactions play divergent roles in modulating AQP2 trafficking, phosphorylation, ubiquitylation, and degradation.

Original languageEnglish
JournalJournal of Biological Chemistry
Pages (from-to)2469-84
Number of pages16
Publication statusPublished - 29 Jan 2016

    Research areas

  • 14-3-3 Proteins, Animals, Aquaporin 2, Biotinylation, Deamino Arginine Vasopressin, Gene Expression Regulation, Glutathione Transferase, HEK293 Cells, Humans, Kidney, Kidney Tubules, Mice, Phosphorylation, Protein Processing, Post-Translational, Protein Structure, Tertiary, Protein Transport, RNA, Messenger, RNA, Small Interfering, Surface Plasmon Resonance, Ubiquitin, Vasopressins, Journal Article, Research Support, Non-U.S. Gov't

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