Protein phosphatase 1 modulates the inhibitory effect of With-no-Lysine kinase 4 on ROMK channels

Am J Physiol Renal Physiol. 2012 Jul 1;303(1):F110-9. doi: 10.1152/ajprenal.00676.2011. Epub 2012 Apr 18.

Abstract

With-no-Lysine kinase 4 (WNK4) inhibited ROMK (Kir1.1) channels and the inhibitory effect of WNK4 was abolished by serum-glucocorticoid-induced kinase 1 (SGK1) but restored by c-Src. The aim of the present study is to explore the mechanism by which Src-family tyrosine kinase (SFK) modulates the effect of SGK1 on WNK4 and to test the role of SFK-WNK4-SGK1 interaction in regulating ROMK channels in the kidney. Immunoprecipitation demonstrated that protein phosphatase 1 (PP1) binds to WNK4 at amino acid (aa) residues 695-699 (PP1(#1)) and at aa 1211-1215 (PP1(#2)). WNK4(-PP1#1) and WNK4(-PP1#2), in which the PP1(#1) or PP1(#2) binding site was deleted or mutated, inhibited ROMK channels as potently as WNK4. However, c-Src restored the inhibitory effect of WNK4 but not WNK4(-PP1#1) on ROMK channels in the presence of SGK1. Moreover, expression of c-Src inhibited SGK1-induced phosphorylation of WNK4 but not WNK4(-PP1#1) at serine residue 1196 (Ser(1196)). In contrast, coexpression of c-Src restored the inhibitory effect of WNK4(-PP1#2) on ROMK in the presence of SGK1 and diminished SGK1-induced WNK4 phosphorylation at Ser(1196) in cells transfected with WNK4(-PP1#2). This suggests the possibility that c-Src regulates the interaction between WNK4 and SGK1 through activating PP1 binding to aa 695-9 thereby decreasing WNK4 phosphorylation and restoring the inhibitory effect of WNK4. This mechanism plays a role in suppressing ROMK channel activity during the volume depletion because inhibition of SFK or serine/threonine phosphatases increases ROMK channel activity in the cortical collecting duct of rats on a low-Na diet. We conclude that regulation of phosphatase activity by SFK plays a role in determining the effect of aldosterone on ROMK channels and on renal K secretion.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • HEK293 Cells
  • Humans
  • Immediate-Early Proteins / metabolism
  • Kidney / metabolism*
  • Kidney Tubules, Collecting / metabolism
  • Male
  • Phosphorylation
  • Potassium Channels, Inwardly Rectifying / metabolism*
  • Protein Phosphatase 1 / metabolism*
  • Protein-Serine-Threonine Kinases / metabolism*
  • Rats
  • Rats, Sprague-Dawley
  • Transfection
  • src-Family Kinases / metabolism

Substances

  • Immediate-Early Proteins
  • KCNJ1 protein, human
  • Potassium Channels, Inwardly Rectifying
  • ROMK2 protein, human
  • src-Family Kinases
  • Protein-Serine-Threonine Kinases
  • WNK4 protein, human
  • serum-glucocorticoid regulated kinase
  • Protein Phosphatase 1