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Src Family Protein Tyrosine Kinase Regulates the Basolateral K Channel in the Distal Convoluted Tubule (DCT) by Phosphorylation of KCNJ10 Protein
Authors:Chengbiao Zhang  Lijun Wang  Sherin Thomas  Kemeng Wang  Dao-Hong Lin  Jesse Rinehart  Wen-Hui Wang
Affiliation:From the Jiangsu Province Key Laboratory of Anesthesiology, Xuzhou Medical College, Xuzhou, Jiangsu 221002, China.;the §Department of Pharmacology, New York Medical College, Valhalla, New York 10595.;the Department of Cellular and Molecular Physiology and ;Systems Biology Institute, Yale University, New Haven, Connecticut 06520
Abstract:The loss of function of the basolateral K channels in the distal nephron causes electrolyte imbalance. The aim of this study is to examine the role of Src family protein tyrosine kinase (SFK) in regulating K channels in the basolateral membrane of the mouse initial distal convoluted tubule (DCT1). Single-channel recordings confirmed that the 40-picosiemen (pS) K channel was the only type of K channel in the basolateral membrane of DCT1. The suppression of SFK reversibly inhibited the basolateral 40-pS K channel activity in cell-attached patches and decreased the Ba2+-sensitive whole-cell K currents in DCT1. Inhibition of SFK also shifted the K reversal potential from −65 to −43 mV, suggesting a role of SFK in determining the membrane potential in DCT1. Western blot analysis showed that KCNJ10 (Kir4.1), a key component of the basolateral 40-pS K channel in DCT1, was a tyrosine-phosphorylated protein. LC/MS analysis further confirmed that SFK phosphorylated KCNJ10 at Tyr8 and Tyr9. The single-channel recording detected the activity of a 19-pS K channel in KCNJ10-transfected HEK293T cells and a 40-pS K channel in the cells transfected with KCNJ10+KCNJ16 (Kir.5.1) that form a heterotetramer in the basolateral membrane of the DCT. Mutation of Tyr9 did not alter the channel conductance of the homotetramer and heterotetramer. However, it decreased the whole-cell K currents, the probability of finding K channels, and surface expression of KCNJ10 in comparison to WT KCNJ10. We conclude that SFK stimulates the basolateral K channel activity in DCT1, at least partially, by phosphorylating Tyr9 on KCNJ10. We speculate that the modulation of tyrosine phosphorylation of KCNJ10 should play a role in regulating membrane transport function in DCT1.
Keywords:Ion Channels   Kidney   Membrane Transport   Phosphorylation   Potassium Channels
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