title: Src family protein tyrosine kinase regulates the basolateral K channel in the distal convoluted tubule (DCT) by phosphorylation of KCNJ10 protein.
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 Ba(2+)-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 Tyr(8) and Tyr(9). 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 Tyr(9) 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 Tyr(9) on KCNJ10. We speculate that the modulation of tyrosine phosphorylation of KCNJ10 should play a role in regulating membrane transport function in DCT1.
authors.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.
authors.forename: Chengbiao
authors.initials: C
authors.lastname: Zhang
authors.affiliation: the Department of Pharmacology, New York Medical College, Valhalla, New York 10595.
authors.forename: Lijun
authors.initials: L
authors.lastname: Wang
authors.affiliation: the Department of Pharmacology, New York Medical College, Valhalla, New York 10595.
authors.forename: Sherin
authors.initials: S
authors.lastname: Thomas
authors.affiliation: the Department of Pharmacology, New York Medical College, Valhalla, New York 10595.
authors.forename: Kemeng
authors.initials: K
authors.lastname: Wang
authors.affiliation: the Department of Pharmacology, New York Medical College, Valhalla, New York 10595.
authors.forename: Dao-Hong
authors.initials: DH
authors.lastname: Lin
authors.affiliation: the Department of Cellular and Molecular Physiology and; Systems Biology Institute, Yale University, New Haven, Connecticut 06520.
authors.forename: Jesse
authors.initials: J
authors.lastname: Rinehart
authors.affiliation: the Department of Pharmacology, New York Medical College, Valhalla, New York 10595,. Electronic address: wenhui_wang@nymc.edu.
authors.forename: Wen-Hui
authors.initials: WH
authors.lastname: Wang
mesh_terms: D000818:Animals; D002460:Cell Line; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D017136:Ion Transport / Q000502:physiology; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology*; D051379:Mice; D020125:Mutation, Missense; D010766:Phosphorylation / Q000502:physiology; D011188:Potassium / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*; D000098854:Kcnj10 Channel; D000097062:Kir5.1 Channel
chemical_list: D011188:Potassium; D024661:Potassium Channels, Inwardly Rectifying; D019061:src-Family Kinases; D000098854:Kcnj10 Channel; C576242:KCNJ16 protein, human; D000097062:Kir5.1 Channel
keywords: Ion Channels; Kidney; Membrane Transport; Phosphorylation; Potassium Channels