title: Lack of arginine vasopressin-induced phosphorylation of aquaporin-2 mutant AQP2-R254L explains dominant nephrogenic diabetes insipidus.
abstract: Water homeostasis in humans is regulated by vasopressin, which induces the translocation of homotetrameric aquaporin-2 (AQP2) water channels from intracellular vesicles to the apical membrane of renal principal cells. For this process, phosphorylation of AQP2 at S256 by cAMP-dependent protein kinase A is thought to be essential. Mutations in the AQP2 gene cause recessive and dominant nephrogenic diabetes insipidus (NDI), a disease in which the kidney is unable to concentrate urine in response to vasopressin. Here, a family in which dominant NDI was caused by an exchange of arginine 254 by leucine in the intracellular C terminus of AQP2 (AQP2-R254L), which destroys the protein kinase A consensus site, was identified. Expressed in oocytes, AQP2-R254L appeared to be a functional water channel but was impaired in its transport to the cell surface to the same degree as AQP2-S256A, which mimics nonphosphorylated AQP2. In polarized renal cells, AQP2-R254L was retained intracellularly and was distributed similarly as AQP2-S256A or wild-type AQP2 in unstimulated cells. Upon co-expression in MDCK cells, AQP2-R254L interacted with and retained wild-type AQP2 in intracellular vesicles. Furthermore, AQP2-R254L had a low basal phosphorylation level, which was not increased with forskolin, and mimicking constitutive phosphorylation in AQP2-R254L with the S256D mutation shifted its expression to the basolateral and apical membrane. These data indicate that dominant NDI in this family is due to a R254L mutation, resulting in the loss of arginine vasopressin-mediated phosphorylation of AQP2 at S256, and illustrates the in vivo importance of phosphorylation of AQP2 at S256 for the first time.
authors.affiliation: Department of Cell Physiology, Radboud University Nijmegen Medical Center, P.O. Box 9101, 6500 HB Nijmegen, The Netherlands.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: de Mattia
authors.forename: Paul J M
authors.initials: PJ
authors.lastname: Savelkoul
authors.forename: Erik-Jan
authors.initials: EJ
authors.lastname: Kamsteeg
authors.forename: Irene B M
authors.initials: IB
authors.lastname: Konings
authors.forename: Peter
authors.initials: P
authors.lastname: van der Sluijs
authors.forename: Rudolf
authors.initials: R
authors.lastname: Mallmann
authors.forename: Alexander
authors.initials: A
authors.lastname: Oksche
authors.forename: Peter M T
authors.initials: PM
authors.lastname: Deen
mesh_terms: D051399:Aquaporin 2 / Q000235:genetics* / Q000378:metabolism*; D001127:Arginine Vasopressin / Q000502:physiology*; D002478:Cells, Cultured; D002648:Child; D018500:Diabetes Insipidus, Nephrogenic / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008297:Male; D009154:Mutation; D009865:Oocytes / Q000378:metabolism; D010375:Pedigree; D010766:Phosphorylation; D021381:Protein Transport
chemical_list: C496023:AQP2 protein, human; D051399:Aquaporin 2; D001127:Arginine Vasopressin