| First Author | Oberfeld B | Year | 2011 |
| Journal | Am J Physiol Renal Physiol | Volume | 301 |
| Issue | 1 | Pages | F189-96 |
| PubMed ID | 21478478 | Mgi Jnum | J:389211 |
| Mgi Id | MGI:8386173 | Doi | 10.1152/ajprenal.00487.2010 |
| Citation | Oberfeld B, et al. (2011) Ubiquitin-specific protease 2-45 (Usp2-45) binds to epithelial Na+ channel (ENaC)-ubiquitylating enzyme Nedd4-2. Am J Physiol Renal Physiol 301(1):F189-96 |
| abstractText | Regulation of the epithelial Na(+) channel (ENaC) by ubiquitylation is controlled by the activity of two counteracting enzymes, the E3 ubiquitin-protein ligase Nedd4-2 (mouse ortholog of human Nedd4L) and the ubiquitin-specific protease Usp2-45. Previously, Usp2-45 was shown to decrease ubiquitylation and to increase surface function of ENaC in Xenopus laevis oocytes, whereas the splice variant Usp2-69, which has a different N-terminal domain, was inactive toward ENaC. It is shown here that the catalytic core of Usp2 lacking the N-terminal domain has a reduced ability relative to Usp2-45 to enhance ENaC activity in Xenopus oocytes. In contrast, its catalytic activity toward the artificial substrate ubiquitin-AMC is fully maintained. The interaction of Usp2-45 with ENaC exogenously expressed in HEK293 cells was tested by coimmunoprecipitation. The data indicate that different combinations of ENaC subunits, as well as the alpha-ENaC cytoplasmic N-terminal but not C-terminal domain, coprecipitate with Usp2-45. This interaction is decreased but not abolished when the cytoplasmic ubiquitylation sites of ENaC are mutated. Importantly, coimmunoprecipitation in HEK293 cells and GST pull-down of purified recombinant proteins show that both the catalytic domain and the N-terminal tail of Usp2-45 physically interact with the HECT domain of Nedd4-2. Taken together, the data support the conclusion that Usp2-45 action on ENaC is promoted by various interactions, including through binding to Nedd4-2 that is suggested to position Usp2-45 favorably for ENaC deubiquitylation. |
