| First Author | Lagnaz D | Year | 2014 |
| Journal | Am J Physiol Renal Physiol | Volume | 307 |
| Issue | 3 | Pages | F275-86 |
| PubMed ID | 24920754 | Mgi Jnum | J:212100 |
| Mgi Id | MGI:5578075 | Doi | 10.1152/ajprenal.00574.2013 |
| Citation | Lagnaz D, et al. (2014) WNK3 abrogates the NEDD4-2-mediated inhibition of the renal Na+-Cl- cotransporter. Am J Physiol Renal Physiol 307(3):F275-86 |
| abstractText | The serine/threonine kinase WNK3 and the ubiquitin-protein ligase NEDD4-2 are key regulators of the thiazide-sensitive Na(+)-Cl(-) cotransporter (NCC), WNK3 as an activator and NEDD2-4 as an inhibitor. Nedd4-2 was identified as an interacting partner of WNK3 through a glutathione-S-transferase pull-down assay using the N-terminal domain of WNK3, combined with LC-MS/MS analysis. This was validated by coimmunoprecipitation of WNK3 and NEDD4-2 expressed in HEK293 cells. Our data also revealed that the interaction between Nedd4-2 and WNK3 does not involve the PY-like motif found in WNK3. The level of WNK3 ubiquitylation did not change when NEDD4-2 was expressed in HEK293 cells. Moreover, in contrast to SGK1, WNK3 did not phosphorylate NEDD4-2 on S222 or S328. Coimmunoprecipitation assays showed that WNK3 does not regulate the interaction between NCC and NEDD4-2. Interestingly, in Xenopus laevis oocytes, WNK3 was able to recover the SGK1-resistant NEDD4-2 S222A/S328A-mediated inhibition of NCC and further activate NCC. Furthermore, elimination of the SPAK binding site in the kinase domain of WNK3 (WNK3-F242A, which lacks the capacity to bind the serine/threonine kinase SPAK) prevented the WNK3 NCC-activating effect, but not the Nedd4-2-inhibitory effect. Together, these results suggest that a novel role for WNK3 on NCC expression at the plasma membrane, an effect apparently independent of the SPAK kinase and the aldosterone-SGK1 pathway. |
