Human Nedd4 interacts with the human epithelial Na+ channel:: WW3 but not WW1 binds to Na+-channel subunits

The epithelial Na+ channel (ENaC) regulates Na+ absorption in epithelial tissues including the lung, colon and sweat gland, and in the distal nephrons of the kidney. When Na+-channel function is disrupted, salt and water homoeostasis is affected. The cytoplasmic regions of the Na+-channel subunits provide binding sites for other proteins to interact with and potentially regulate Na+-channel activity. Previously we showed that a proline-rich region of the alpha subunit of the Na+ channel bound to a protein of 116 kDa from human lung cells. Here we report the identification of this protein as human Nedd4, a ubiquitin-protein ligase that binds to the Na+-channel subunits via its WW domains. Further, we show that WW domains 2, 3 and 4 of human Nedd4 bind to the alpha, beta and gamma Na+-channel subunits but not to a mutated beta subunit. In addition, when co-expressed in Xenopus oocytes, human Nedd4 down-regulates Na+-channel activity.