Biochemical and Biophysical Research Communications, Vol.521, No.4, 1030-1035, 2020
Hypertonicity-responsive ubiquitin ligase RNF183 promotes Na, K-ATPase lysosomal degradation through ubiquitination of its beta 1 subunit
We previously reported that RNF183, a member of the RING finger (RNF) protein family, is specifically expressed in the renal collecting duct and that RNF183 mRNA is induced by the activity of nuclear factor of activated T cells 5 (NFAT5), which regulates the transcription of essential proteins for adaptation to hypertonic conditions. The renal medulla is the only tissue that is continuously hypertonic; therefore, RNF183 possibly plays an important role in adaptation to continuous hypertonic conditions. However, the mechanism of how cells adapt to long-term hypertonicity via RNF183 remains unclear. In this study, the Na, K-ATPase alpha 1 subunit was identified as a candidate substrate of RNF183 by the BirA proximitybiotinylation technique. The Na, K-ATPase alpha 1 subunit acts as an ion transporter along with the Na, K-ATPase beta 1 subunit at the plasma membrane. We confirmed that RNF183 interacted with both alpha 1 and beta 1 subunits; however, we found that RNF183 ubiquitinated only the beta 1 subunit, not the alpha 1 subunit. Furthermore, RNF183 translocated both alpha 1 and beta 1 subunits from the plasma membrane to lysosomes. In addition, the expression levels of alpha 1 and beta 1 subunits in HEK293 cells stably expressing RNF183 were significantly decreased compared with mock control cells, and were restored by siRNA-mediated knockdown of RNF183. Moreover, in RNF183-expressing cells, chloroquine treatment increased the protein levels of the alpha 1 and beta 1 subunits. Therefore, our results suggest that Na, K-ATPase alpha 1 and beta 1 subunits are degraded in lysosomes by RNF183-mediated ubiquitination of beta 1 subunit. (C) 2019 Elsevier Inc. All rights reserved.