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Abstract Na-K-2Cl cotransporter (NKCC) and K-Cl cotransporter (KCC) play key roles in cell volume regulation and epithelial Cl − transport. Reductions in either cell volume or cytosolic Cl − concentration (Cl − i ) stimulate a corrective uptake of KCl and water via NKCC, whereas cell swelling triggers KCl loss via KCC. The dependence of these transporters on volume and Cl − i was evaluated in model duck red blood cells. Replacement of Cl − i with methanesulfonate elevated the volume set point at which NKCC activates and KCC inactivates. The set point was insensitive to cytosolic ionic strength. Reducing Cl − i at a constant driving force for inward NKCC and outward KCC caused the cells to adopt the new set point volume. Phosphopeptide maps of NKCC indicated that activation by cell shrinkage or low Cl − i is associated with phosphorylation of a similar constellation of Ser/Thr sites. Like shrinkage, reduction of Cl − i accelerated NKCC phosphorylation after abrupt inhibition of the deactivating phosphatase with calyculin A in vivo, whereas Cl − had no specific effect on dephosphorylation in vitro. Our results indicate that NKCC and KCC are reciprocally regulated by a negative feedback system dually modulated by cell volume and Cl − . The major effect of Cl − on NKCC is exerted through the volume-sensitive kinase that phosphorylates the transport protein. sodium-potassium-chloride cotransport intracellular chloride cell volume regulation ionic strength cell water content sodium-potassium-chloride cotransporter phosphorylation Footnotes This work was supported by the National Science Foundation (NSF-MCB9904605). Address for reprint requests and other correspondence: C. Lytle, Div. of Biomedical Sciences, 2226 Webber Hall, Univ. of California, Riverside, Riverside, CA 92521 (E-mail: christian.lytle@ucr.edu ). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “ advertisement ” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 10.1152/ajpcell.00130.2002 Copyright © 2002 the American Physiological Society
AJP - Cell Physiology – The American Physiological Society
Published: Nov 1, 2002
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