Interaction of serum- and glucocorticoid regulated kinase 1 (SGK1) with the WW-domains of Nedd4-2 is required for epithelial sodium channel regulation

• Published in: PloS one 2010-08, Vol.5 (8), p.e12163
• Main Authors: Wiemuth, Dominik, Lott, J Shaun, Ly, Kevin, Ke, Ying, Teesdale-Spittle, Paul, Snyder, Peter M, McDonald, Fiona J
• Format: Article
• Language: English
• Subjects: 14-3-3 protein; Aldosterone; Amino Acid Motifs; Animals; Binding; Biochemistry/Cell Signaling and Trafficking Structures; Biochemistry/Experimental Biophysical Methods; Cercopithecus aethiops; COS Cells; Cystic fibrosis; Dose-Response Relationship, Drug; Endosomal Sorting Complexes Required for Transport - chemistry; Endosomal Sorting Complexes Required for Transport - metabolism; Endosomal Sorting Complexes Required for Transport - pharmacology; Enzyme Activation; Epithelial cells; Epithelial Sodium Channel Blockers; Epithelial Sodium Channels - metabolism; Glucocorticoids; Immediate-Early Proteins - chemistry; Immediate-Early Proteins - metabolism; Kinases; Ligases; Models, Molecular; Nedd4 Ubiquitin Protein Ligases; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Phosphorylation; Physiology; Physiology/Renal, Fluid, and Electrolyte Physiology; Protein Binding; Protein Structure, Tertiary; Protein-Serine-Threonine Kinases - chemistry; Protein-Serine-Threonine Kinases - metabolism; Proteins; Rodents; Sodium; Steroids (Organic compounds); Studies; Tryptophan; Ubiquitin; Ubiquitin-Protein Ligases - chemistry; Ubiquitin-Protein Ligases - metabolism; Ubiquitin-Protein Ligases - pharmacology
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0012163

The epithelial sodium channel (ENaC) is an integral component of the pathway for Na(+) absorption in epithelial cells. The ubiquitin ligases Nedd4 and Nedd4-2 bind to ENaC and decrease its activity. Conversely, Serum- and Glucocorticoid regulated Kinase-1 (SGK1), a downstream mediator of aldosterone, increases ENaC activity. This effect is at least partly mediated by direct interaction between SGK and Nedd4-2. SGK binds both Nedd4 and Nedd4-2, but it is only able to phosphorylate Nedd4-2. Phosphorylation of Nedd4-2 reduces its ability to bind to ENaC, due to the interaction of phosphorylated Nedd4-2 with 14-3-3 proteins, and hence increases ENaC activity. WW-domains in Nedd4-like proteins bind PY-motifs (PPXY) present in ENaC subunits, and SGK also has a PY-motif. Here we show that single or tandem WW-domains of Nedd4 and Nedd4-2 mediate binding to SGK and that different WW-domains of Nedd4 and Nedd4-2 are involved. Our data also show that WW-domains 2 and 3 of Nedd4-2 mediate the interaction with SGK in a cooperative manner, that activated SGK has increased affinity for the WW-domains of Nedd4-2 in vitro, and a greater stimulatory effect on ENaC Na(+) transport compared to wildtype SGK. Further, SGK lacking a PY motif failed to stimulate ENaC activity in the presence of Nedd4-2. Binding of Nedd4-2 WW-domains to SGK is necessary for SGK-induced ENaC activity.