AMP-activated kinase inhibits the epithelial Na+ channel through functional regulation of the ubiquitin ligase Nedd4-2

J Biol Chem. 2006 Sep 8;281(36):26159-69. doi: 10.1074/jbc.M606045200. Epub 2006 Jul 14.

Abstract

We recently found that the metabolic sensor AMP-activated kinase (AMPK) inhibits the epithelial Na+ channel (ENaC) through decreased plasma membrane ENaC expression, an effect requiring the presence of a binding motif in the cytoplasmic tail of the beta-ENaC subunit for the ubiquitin ligase Nedd4-2. To further examine the role of Nedd4-2 in the regulation of ENaC by AMPK, we studied the effects of AMPK activation on ENaC currents in Xenopus oocytes co-expressing ENaC and wild-type (WT) or mutant forms of Nedd4-2. ENaC inhibition by AMPK was preserved in oocytes expressing WT Nedd4-2 but blocked in oocytes expressing either a dominant-negative (DN) or constitutively active (CA) Nedd4-2 mutant, suggesting that AMPK-dependent modulation of Nedd4-2 function is involved. Similar experiments utilizing WT or mutant forms of the serum- and glucocorticoid-regulated kinase (SGK1), modulators of protein kinase A (PKA), or extracellular-regulated kinase (ERK) did not affect ENaC inhibition by AMPK, suggesting that these pathways known to modulate the Nedd4-2-ENaC interaction are not responsible. AMPK-dependent phosphorylation of Nedd4-2 expressed in HEK-293 cells occurred both in vitro and in vivo, suggesting a potential mechanism for modulation of Nedd4-2 and thus cellular ENaC activity. Moreover, cellular AMPK activation significantly enhanced the interaction of the beta-ENaC subunit with Nedd4-2, as measured by co-immunoprecipitation assays in HEK-293 cells. In summary, these results suggest a novel mechanism for ENaC regulation in which AMPK promotes ENaC-Nedd4-2 interaction, thereby inhibiting ENaC by increasing Nedd4-2-dependent ENaC retrieval from the plasma membrane. AMPK-dependent ENaC inhibition may limit cellular Na+ loading under conditions of metabolic stress when AMPK becomes activated.

Publication types

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

MeSH terms

  • Animals
  • Cell Line
  • Cell Membrane / metabolism
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Endosomal Sorting Complexes Required for Transport
  • Enzyme Activation
  • Epithelial Sodium Channel Blockers*
  • Epithelial Sodium Channels / metabolism
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Humans
  • Immediate-Early Proteins / metabolism
  • Nedd4 Ubiquitin Protein Ligases
  • Oocytes / physiology
  • Patch-Clamp Techniques
  • Phosphorylation
  • Protein Subunits / genetics
  • Protein Subunits / metabolism
  • Protein-Serine-Threonine Kinases / metabolism
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*
  • Xenopus Proteins / genetics
  • Xenopus Proteins / metabolism*
  • Xenopus laevis

Substances

  • Endosomal Sorting Complexes Required for Transport
  • Epithelial Sodium Channel Blockers
  • Epithelial Sodium Channels
  • Immediate-Early Proteins
  • Protein Subunits
  • Xenopus Proteins
  • Nedd4 Ubiquitin Protein Ligases
  • Nedd4 protein, Xenopus
  • Nedd4 protein, human
  • Nedd4L protein, human
  • nedd4l protein, Xenopus
  • Ubiquitin-Protein Ligases
  • Protein-Serine-Threonine Kinases
  • serum-glucocorticoid regulated kinase
  • Cyclic AMP-Dependent Protein Kinases
  • Extracellular Signal-Regulated MAP Kinases