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

Dao-Hong Lin1, Peng Yue, Jesse Rinehart

  • 1Department of Pharmacology, New York Medical College, Valhalla, New York 10595, USA.

American Journal of Physiology. Renal Physiology
|April 20, 2012
PubMed

Insights

Src-family tyrosine kinase (SFK) regulates With-no-Lysine kinase 4 (WNK4) and serum-glucocorticoid-induced kinase 1 (SGK1) interaction, impacting ROMK channel activity in the kidney. This SFK-mediated regulation is crucial for renal potassium secretion.

Area of Science:

  • Nephrology
  • Molecular Biology
  • Cell Signaling

Background:

  • With-no-Lysine kinase 4 (WNK4) inhibits renal potassium ROMK (Kir1.1) channels.
  • Serum-glucocorticoid-induced kinase 1 (SGK1) abolishes WNK4's inhibitory effect, while c-Src restores it.
  • The interplay between WNK4, SGK1, and Src-family tyrosine kinases (SFKs) in regulating ROMK channels requires elucidation.

Purpose of the Study:

  • To investigate the mechanism by which SFK modulates SGK1's effect on WNK4.
  • To determine the role of the SFK-WNK4-SGK1 interaction in regulating kidney ROMK channels.
  • To explore the role of phosphatase activity in SFK-mediated regulation of WNK4 and ROMK channels.

Main Methods:

  • Immunoprecipitation to identify protein binding sites between WNK4 and protein phosphatase 1 (PP1).
  • Site-directed mutagenesis to create WNK4 variants lacking PP1 binding sites (WNK4(-PP1#1), WNK4(-PP1#2)).
  • Expression studies in cells to assess the effects of c-Src and SGK1 on WNK4 phosphorylation and ROMK channel activity, and in rat kidney tubules.

Main Results:

  • PP1 binds to WNK4 at two distinct sites (aa 695-699 and aa 1211-1215).
  • c-Src restored WNK4's inhibitory effect on ROMK channels in the presence of SGK1, but only when the PP1(#1) binding site was intact.
  • c-Src inhibited SGK1-induced WNK4 phosphorylation at Ser(1196) via PP1 binding at aa 695-9, suggesting SFK activates PP1 to decrease WNK4 phosphorylation.
  • Inhibition of SFK or phosphatases increased ROMK channel activity in rat kidney tubules, particularly during volume depletion.

Conclusions:

  • SFK, likely through activating PP1 binding to WNK4 at aa 695-9, decreases WNK4 phosphorylation by SGK1, thereby restoring WNK4's inhibitory effect on ROMK channels.
  • This SFK-WNK4-SGK1-PP1 pathway plays a role in suppressing ROMK channel activity during physiological states like volume depletion.
  • Regulation of phosphatase activity by SFK is critical for aldosterone's effect on ROMK channels and renal potassium secretion.

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