{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,22]],"date-time":"2026-07-22T23:32:42Z","timestamp":1784763162172,"version":"3.55.0"},"reference-count":35,"publisher":"Wiley","issue":"13","license":[{"start":{"date-parts":[[2023,3,14]],"date-time":"2023-03-14T00:00:00Z","timestamp":1678752000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"content-domain":{"domain":["physoc.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["The Journal of Physiology"],"published-print":{"date-parts":[[2024,7]]},"abstract":"<jats:sec><jats:title>Abstract<\/jats:title><jats:p>Nephrogenic syndrome of inappropriate antidiuresis (NSIAD) is a rare X\u2010linked disease caused by gain\u2010of\u2010function mutations of arginine vasopressin receptor 2 (V2R). Patients with NSIAD are characterized by the inability to excrete a free water load and by inappropriately increased urinary osmolality despite very low levels of plasma vasopressin, resulting in euvolaemic hyponatraemia. To dissect the signalling downstream V2R constitutively active variants, Flp\u2010In T\u2010REx Madin\u2010Darby canine kidney (FTM) cells, stably transfected with V2R mutants (R137L, R137C and F229V) and AQP2\u2010wt or non\u2010phosphorylatable AQP2\u2010S269A\/AQP2\u2010S256A, were used as cellular models. All three activating V2R mutations presented constitutive plasma membrane expression of AQP2\u2010wt and significantly higher basal water permeability. In addition, V2R\u2010R137L\/C showed significantly higher activity of Rho\u2010associated kinase (ROCK), a serine\/threonine kinase previously suggested to be involved in S269\u2010AQP2 phosphorylation downstream of these V2R mutants. Interestingly, FTM cells expressing V2R\u2010R137L\/C mutants and AQP2\u2010S269A showed a significant reduction in AQP2\u00a0membrane abundance and a significant reduction in ROCK activity, indicating the crucial importance of S269\u2010AQP2 phosphorylation in the gain\u2010of\u2010function phenotype. Conversely, V2R\u2010R137L\/C mutants retained the gain\u2010of\u2010function phenotype when AQP2\u2010S256A was co\u2010expressed. In contrast, cells expressing the F229V mutant and the non\u2010phosphorylatable AQP2\u2010S256A had a significant reduction in AQP2\u00a0membrane abundance along with a significant reduction in basal osmotic water permeability, indicating a crucial role of Ser256 for this mutant. These data indicate that the constitutive AQP2 trafficking associated with the gain\u2010of\u2010function V2R\u2010R137L\/C mutants causing NSIAD is protein kinase\u00a0A independent and requires an intact Ser269 in AQP2 under the control of ROCK phosphorylation.\n<jats:boxed-text content-type=\"graphic\" position=\"anchor\"><jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" mimetype=\"image\/png\" position=\"anchor\" specific-use=\"enlarged-web-image\" xlink:href=\"graphic\/tjp15463-gra-0001-m.png\"><jats:alt-text>image<\/jats:alt-text><\/jats:graphic><\/jats:boxed-text><\/jats:p><\/jats:sec><jats:sec><jats:title>Key points<\/jats:title><jats:p><jats:list list-type=\"bullet\">\n<jats:list-item><jats:p>Nephrogenic syndrome of inappropriate antidiuresis is caused by two constitutively active variant phenotypes of <jats:italic>AVPR2<\/jats:italic>, one sensitive to vaptans (V2R\u2010F229V) and the other vaptan resistant (V2R\u2010R137C\/L). In renal cells, all three activating arginine vasopressin receptor 2 (V2R) variants display constitutive AQP2 plasma membrane expression and high basal water permeability.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>In cells expressing V2R\u2010R137L\/C mutants, disruption of the AQP2\u2010S269 phosphorylation site caused the loss of the gain\u2010of\u2010function phenotype, which, in contrast, was retained in V2R\u2010F229V\u2010expressing cells.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>Cells expressing the V2R\u2010F229V mutant were instead sensitive to disruption of the AQP2\u2010S256 phosphorylation site.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>The serine\/threonine kinase Rho\u2010associated kinase (ROCK) was found to be involved in AQP2\u2010S269 phosphorylation downstream of the V2R\u2010R137L\/C mutants.<\/jats:p><\/jats:list-item>\n<jats:list-item><jats:p>These findings might have clinical relevance for patients with nephrogenic syndrome of inappropriate antidiuresis.<\/jats:p><\/jats:list-item>\n<\/jats:list><\/jats:p><\/jats:sec>","DOI":"10.1113\/jp284039","type":"journal-article","created":{"date-parts":[[2023,2,24]],"date-time":"2023-02-24T05:57:22Z","timestamp":1677218242000},"page":"3169-3189","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Novel signalling pathways in nephrogenic syndrome of inappropriate antidiuresis: functional implication of site\u2010specific AQP2 phosphorylation"],"prefix":"10.1113","volume":"602","author":[{"given":"Maria","family":"Venneri","sequence":"first","affiliation":[{"name":"Department of Biosciences, Biotechnology and Environment University of Bari Aldo Moro  Bari Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vanessa","family":"Vezzi","sequence":"additional","affiliation":[{"name":"National Center for Drug Research and Evaluation Istituto Superiore di Sanit\u00e0  Rome 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