Human nedd-4 as a family member of ubiquitin ligases

Ubiquitin-proteasome proteolytic pathway is involved in a number of cellular physiology. In this system, a ubiquitin ligase, or E3 enzyme, is a major component which recognizes a specific substrate protein for targeted degradation. E6-associated protein (E6-AP) has been recently found to be a member of the E3 enzyme family that is characterized by a conserved carboxy-terminal sequence, termed Aectf-domain. Using human EST data bases, we identified five uncharacterized genes of this family. By FISH and radiation hybrid panel these genes were mapped to subchromosomal locations as follows; E6-AP (15ql2-13), NEDD-4 (15q22; D15S209), D28476 (2q35-36; D2S396), D13635 (7q36; FB16B4),and D25215 (4q21-22; WI-6336). NEDD-4 gene was mapped close to the translocation breakpoint of acute promyelocytic leukemia. Interestingly, NEDD-4 was highly conserved among species from yeast to human, suggesting its significant function on basic biomodulation. Northern blot analysis showed that NEDD-4 mRNA was much higher expressed in lung carcinoma A549 and chronic myelogenous leukemia K562 than in other cancer cell lines tested. The rest of the members, except D25215, were ubiquitously expressed in all cells. In immunoblot analysis, human NEDD-4 protein was down-regulated during neural differentiation in BDNFtreated neuroblastoma cells. Collectively, our data suggest that NEDD-4 protein may function as a ubiquitin ligase closely related to development, differentiation and tumorigenesis.