Molecular cloning of Nedd4 from Xenopus laevis

The tryptophan-bounded WW domains of Nedd4 bind to the proline-tyrosine (PY ) motifs contained in the C-terminal cytoplasmic region of the beta and gam ma subunits of the rat amiloride-sensitive sodium channel (ENaC). In patien ts with Liddle's syndrome, the PY motif is mutated and the channel remains constitutively activated leading to sodium retention and hypertension. Alth ough the function of Nedd4 is unknown, it contains a highly conserved ubiqu itin protein ligase domain that may attach ubiquitin to ENaC, targeting it for degradation or it may modulate ENaC activity through another undetermin ed pathway. Xenopus laevis-derived cells, such as oocytes and the A6 kidney cell line, are important models currently used for the study of ENaC regul ation. We describe the X. laevis homologue of Nedd4 (xNedd4). A partial clo ne, similar to 2.6 Kb, was isolated from an aldosterone-treated A6 cell cDN A library. Further 5' sequence, similar to 1.2 Kb, was obtained using a mod ified 5' rapid amplification of cDNA (RACE) protocol and cDNA from untreate d A6 cells as the substrate. The identity and similarity of xNedd4 with hum an Nedd4 are approximately 63 and 71%, respectively. xNedd4 contains the C2 , ubiquitin protein ligase, and 4 WW domains previously described for Nedd4 from other species.