Journal Article

Vasopressin increases S261 phosphorylation in AQP2-P262L, a mutant in recessive nephrogenic diabetes insipidus

Christiane Trimpert, Dennis T.M. van den Berg, Robert A. Fenton, Enno Klussmann and Peter M.T. Deen

in Nephrology Dialysis Transplantation

Published on behalf of European Renal Association - European Dialysis and Transplant Assoc

Volume 27, issue 12, pages 4389-4397
Published in print December 2012 | ISSN: 0931-0509
Published online July 2012 | e-ISSN: 1460-2385 | DOI: http://dx.doi.org/10.1093/ndt/gfs292
Vasopressin increases S261 phosphorylation in AQP2-P262L, a mutant in recessive nephrogenic diabetes insipidus

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Background

Mutations in the aquaporin-2 (AQP2) gene cause nephrogenic diabetes insipidus (NDI), a renal disorder characterized by polyuria due to a lacking antidiuretic response to vasopressin. While most AQP2 mutants in recessive NDI are misfolded and retained in the endoplasmic reticulum, AQP2-P262L in NDI was impaired in its vasopressin-dependent translocation from vesicles to the plasma membrane.

Methods

Vasopressin-induced translocation of AQP2 coincides with AQP2 phosphorylation at S256, S264 and T269 and dephosphorylation at S261. Since P262 lies adjacent to S261, we tested whether a changed phosphorylation could underlie AQP-P262L missorting in NDI.

Results

In polarized cells, AQP2-P262L expressed as a double 29/30 kDa band, whereas wt-AQP2 expressed only as a 29 kDa band. Phosphatase treatment revealed that the 30 kDa AQP2-P262L band was due to changed phosphorylation. The use of newly developed phospho-specific antibodies showed that forskolin not only increased pS256 and pT269, but, in contrast to wt-AQP2, also pS261 in AQP2-P262L. The expression of AQP2-P262L proteins in which S261 phosphorylation was prevented (S261A), however, was still missorted to vesicles/basolateral membrane, despite the absence of the 30 kDa band.

Conclusions

Together, our data reveal that vasopressin induces instead of reduces the phosphorylation of S261 in AQP2-P262L, but it remains to be established whether the changed phosphorylation causes its missorting in NDI.

Keywords: AQP2 water channel; nephrogenic diabetes insipidus; phosphorylation; water transport

Journal Article.  5151 words.  Illustrated.

Subjects: Nephrology

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