Src-Family Protein Tyrosine Kinase Phosphorylates WNK4 and Modulates Its Inhibitory Effect on KCNJ1 (ROMK)

Authors

Dao-Hong Lin, Department of Pharmacology, New York Medical College, Valhalla, NY 10595;
Peng Yue, Department of Pharmacology, New York Medical College, Valhalla, NY 10595;
Orlando Yarborough, Department of Genetics and Howard Hughes Medical Institute, and.
Ute I. Scholl, Department of Genetics and Howard Hughes Medical Institute, and.
Gerhard Giebisch, Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT 06520; and wenhui_wang@nymc.edu gerhard.giebisch@yale.edu.
Richard P. Lifton, Department of Genetics and Howard Hughes Medical Institute, and.
Jesse Rinehart, Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT 06520; and Systems Biology Institute, Yale University, West Haven, CT 06516.
Wen-Hui Wang, Department of Pharmacology, New York Medical College, Valhalla, NY 10595; wenhui_wang@nymc.edu gerhard.giebisch@yale.edu.

Author Type(s)

Faculty

DOI

10.1073/pnas.1503437112

Journal Title

Proceedings of the National Academy of Sciences of the United States of America

First Page

4495

Last Page

500

Document Type

Article

Publication Date

4-7-2015

Abstract

With-no-lysine kinase 4 (WNK4) inhibits the activity of the potassium channel KCNJ1 (ROMK) in the distal nephron, thereby contributing to the maintenance of potassium homeostasis. This effect is inhibited via phosphorylation at Ser1196 by serum/glucocorticoid-induced kinase 1 (SGK1), and this inhibition is attenuated by the Src-family protein tyrosine kinase (SFK). Using Western blot and mass spectrometry, we now identify three sites in WNK4 that are phosphorylated by c-Src: Tyr(1092), Tyr(1094), and Tyr(1143), and show that both c-Src and protein tyrosine phosphatase type 1D (PTP-1D) coimmunoprecipitate with WNK4. Mutation of Tyr(1092) or Tyr(1143) to phenylalanine decreased the association of c-Src or PTP-1D with WNK4, respectively. Moreover, the Tyr1092Phe mutation markedly reduced ROMK inhibition by WNK4; this inhibition was completely absent in the double mutant WNK4(Y1092/1094F). Similarly, c-Src prevented SGK1-induced phosphorylation of WNK4 at Ser(1196), an effect that was abrogated in the double mutant. WNK4(Y1143F) inhibited ROMK activity as potently as wild-type (WT) WNK4, but unlike WT, the inhibitory effect of WNK4(Y1143F) could not be reversed by SGK1. The failure to reverse WNK4(Y1143F)-induced inhibition of ROMK by SGK1 was possibly due to enhancing endogenous SFK effect on WNK4 by decreasing the WNK4-PTP-1D association because inhibition of SFK enabled SGK1 to reverse WNK4(Y1143F)-induced inhibition of ROMK. We conclude that WNK4 is a substrate of SFKs and that the association of c-Src and PTP-1D with WNK4 at Tyr(1092) and Tyr(1143) plays an important role in modulating the inhibitory effect of WNK4 on ROMK.

Recommended Citation

Lin, D., Yue, P., Yarborough, O., Scholl, U. I., Giebisch, G., Lifton, R. P., Rinehart, J., & Wang, W. (2015). Src-Family Protein Tyrosine Kinase Phosphorylates WNK4 and Modulates Its Inhibitory Effect on KCNJ1 (ROMK). Proceedings of the National Academy of Sciences of the United States of America, 112 (14), 4495-500. https://doi.org/10.1073/pnas.1503437112