Nuclear import/export of hRPF1/Nedd4 regulates the ubiquitin-dependent degradation of its nuclear substrates

The ubiquitin-protein ligase (E3), hRPF1/Nedd4, is a component of the ubiqu itin-proteasome pathway responsible for substrate recognition and specifici ty, Although previously characterized as a regulator of the stability of cy toplasmic proteins, hRPF1/Nedd4 has also been suggested to have a role in t he nucleus. However, in light of the cytoplasmic localization of hRPF1/Nedd 4, it is unclear whether bona fide nuclear substrates of hRPF1/Nedd4 exist, and if so, what mechanism may allow a cytoplasmic ubiquitin ligase to mani fest nuclear activity. Our search for nuclear substrates led to the identif ication of the human proline-rich transcript, brain-expressed (hPRTB) prote in, the ubiquitination and degradation of which is regulated by hRPF1/Nedd4 . interestingly; hPRTB colocalizes with the splicing factor SC35 in nuclear speckles. Finally, we demonstrate that hRPF1/Nedd4 is indeed capable of en tering the nucleus; however, the presence of a functional Rev-like nuclear export sequence in hRPF1/Nedd4 ensures a predominant cytoplasmic localizati on, Cumulatively, these findings highlight a nuclear role for the ubiquitin ligase hRPF1/Nedd4 and underscore cytoplasmic/nuclear localization as an i mportant regulatory component of hRPF1/Nedd4-substrate recognition.