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In Vitro Analysis of PDZ-dependent CFTR Macromolecular Signaling Complexes
10:05

In Vitro Analysis of PDZ-dependent CFTR Macromolecular Signaling Complexes

Published on: August 13, 2012

Rescue of DeltaF508-CFTR by the SGK1/Nedd4-2 signaling pathway

Hung Caohuy1, Catherine Jozwik, Harvey B Pollard

  • 1Department of Anatomy, Physiology, and Genetics, Uniformed Services University School of Medicine, Bethesda, Maryland 20814, USA. hcaohuy@usuhs.mil

The Journal of Biological Chemistry
|July 21, 2009
PubMed

Insights

Dexamethasone treatment rescues the DeltaF508-cystic fibrosis transmembrane conductance regulator (CFTR) mutation by enhancing its trafficking and stability. This occurs via the SGK1/Nedd4-2 pathway, which also regulates epithelial sodium channel (ENaC) trafficking in CF cells.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Physiology

Background:

  • Cystic Fibrosis (CF) is primarily caused by the DeltaF508 mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) protein.
  • DeltaF508-CFTR fails to traffic correctly to the plasma membrane, leading to epithelial sodium channel (ENaC) over-activity.
  • The glucocorticoid receptor, SGK1, and Nedd4-2 pathway are known regulators of ENaC trafficking.

Purpose of the Study:

  • To investigate the effect of dexamethasone on DeltaF508-CFTR trafficking and function.
  • To elucidate the role of the SGK1/Nedd4-2 signaling pathway in CFTR and ENaC regulation.

Main Methods:

  • Dexamethasone treatment of cells expressing DeltaF508-CFTR.
  • Pharmacological inhibition of glucocorticoid receptor and phosphatidylinositol 3-kinase.
  • Co-immunoprecipitation to assess protein interactions.
  • Small interfering RNA (siRNA) to knock down SGK1 and Nedd4-2 expression.

Main Results:

  • Dexamethasone rescued functional expression and enhanced the half-life of DeltaF508-CFTR.
  • Dexamethasone's effect was blocked by glucocorticoid receptor or PI3K inhibitors.
  • Dexamethasone treatment reduced Nedd4-2 binding to CFTR and decreased CFTR ubiquitination.
  • Nedd4-2 knockdown corrected DeltaF508-CFTR trafficking, while SGK1 knockdown blocked dexamethasone-induced rescue.

Conclusions:

  • The SGK1/Nedd4-2 signaling pathway is a key regulator of both CFTR and ENaC trafficking in cystic fibrosis.
  • Dexamethasone-mediated rescue of DeltaF508-CFTR involves modulation of this pathway.

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