{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,19]],"date-time":"2026-10-19T17:45:17Z","timestamp":1792431917116,"version":"4.5.0"},"reference-count":47,"publisher":"The Company of Biologists","issue":"5","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2011,3,1]]},"abstract":"<jats:p>Ion cotransporters, such as the Na+\/Cl\u2212 cotransporter (NCC), control renal salt re-absorption and are regulated by the WNK-signalling pathway, which is over-stimulated in patients suffering from Gordon's hypertension syndrome. Here, we study the regulation of the NKCC2 (SLC12A1) ion cotransporter that contributes towards ~25% of renal salt re-absorption and is inhibited by loop-diuretic hypertensive drugs. We demonstrate that hypotonic low-chloride conditions that activate the WNK1-SPAK and OSR1 pathway promote phosphorylation of NKCC2 isoforms (A, B and F) at five residues (Ser91, Thr95, Thr100, Thr105 and Ser130). We establish that the SPAK and OSR1 kinases activated by WNK interact with an RFQV motif on NKCC2 and directly phosphorylate Thr95, Thr100, Thr105 and, possibly, Ser91. Our data indicate that a SPAK-OSR1-independent kinase, perhaps AMP-activated protein kinase (AMPK), phosphorylates Ser130 and that phosphorylation of Thr105 and Ser130 plays the most important roles in stimulating NKCC2 activity. In contrast with NCC, whose membrane translocation is triggered by SPAK-OSR1 phosphorylation, NKCC2 appears to be constitutively at the membrane. Our findings provide new insights into how NKCC2 is regulated and suggest that inhibitors of SPAK and\/or OSR1 for the treatment of hypertension would be therapeutically distinct from thiazide or loop diuretics, as they would suppress the activity of both NCC and NKCC2.<\/jats:p>","DOI":"10.1242\/jcs.077230","type":"journal-article","created":{"date-parts":[[2011,2,15]],"date-time":"2011-02-15T01:06:18Z","timestamp":1297731978000},"page":"789-800","source":"Crossref","is-referenced-by-count":170,"title":["Regulation of the NKCC2 ion cotransporter by SPAK-OSR1-dependent and -independent pathways"],"prefix":"10.1242","volume":"124","author":[{"given":"Ciaran","family":"Richardson","sequence":"first","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kei","family":"Sakamoto","sequence":"additional","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paola","family":"de los Heros","sequence":"additional","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria","family":"Deak","sequence":"additional","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David G.","family":"Campbell","sequence":"additional","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alan R.","family":"Prescott","sequence":"additional","affiliation":[{"name":"Division of Cell Signalling and Immunology, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dario R.","family":"Alessi","sequence":"additional","affiliation":[{"name":"MRC Protein Phosphorylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"237","reference":[{"key":"2021042521413052800_R1","doi-asserted-by":"crossref","first-page":"10883","DOI":"10.1073\/pnas.0604607103","article-title":"WNK1 and OSR1 regulate the Na+, K+, 2Cl- cotransporter in HeLa cells","volume":"103","author":"Anselmo","year":"2006","journal-title":"Proc. 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Med."},{"key":"2021042521413052800_R32","doi-asserted-by":"crossref","first-page":"3293","DOI":"10.1242\/jcs.029223","article-title":"The regulation of salt transport and blood pressure by the WNK-SPAK\/OSR1 signalling pathway","volume":"121","author":"Richardson","year":"2008","journal-title":"J. Cell Sci."},{"key":"2021042521413052800_R33","doi-asserted-by":"crossref","first-page":"675","DOI":"10.1242\/jcs.025312","article-title":"Activation of the thiazide-sensitive Na+-Cl- cotransporter by the WNK-regulated kinases SPAK and OSR1","volume":"121","author":"Richardson","year":"2008","journal-title":"J. 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Commun."},{"key":"2021042521413052800_R39","doi-asserted-by":"crossref","first-page":"238","DOI":"10.1093\/ndt\/16.2.238","article-title":"Vasopressin-stimulated chloride transport in transimmortalized mouse cell lines derived from the distal convoluted tubule and cortical and inner medullary collecting ducts","volume":"16","author":"Van Huyen","year":"2001","journal-title":"Nephrol. Dial. Transplant."},{"key":"2021042521413052800_R40","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1042\/BJ20051180","article-title":"The WNK1 and WNK4 protein kinases that are mutated in Gordon's hypertension syndrome phosphorylate and activate SPAK and OSR1 protein kinases","volume":"391","author":"Vitari","year":"2005","journal-title":"Biochem. 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USA"},{"key":"2021042521413052800_R44","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1016\/j.cmet.2007.03.009","article-title":"Molecular pathogenesis of pseudohypoaldosteronism type II: generation and analysis of a Wnk4(D561A\/+) knockin mouse model","volume":"5","author":"Yang","year":"2007","journal-title":"Cell Metab."},{"key":"2021042521413052800_R45","doi-asserted-by":"crossref","first-page":"37922","DOI":"10.1074\/jbc.M006218200","article-title":"Low chloride stimulation of prostaglandin E2 release and cyclooxygenase-2 expression in a mouse macula densa cell line","volume":"275","author":"Yang","year":"2000","journal-title":"J. Biol. Chem."},{"key":"2021042521413052800_R46","doi-asserted-by":"crossref","first-page":"89","DOI":"10.1083\/jcb.200605093","article-title":"Regulation of activity and localization of the WNK1 protein kinase by hyperosmotic stress","volume":"176","author":"Zagorska","year":"2007","journal-title":"J. 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