SPAK and OSR1 Dependent Down-Regulation of Murine Renal Outer Medullary K+ Channel ROMK1

Abstract

Background/Aims: The kinases SPAK (SPS1-related proline/alanine-rich kinase) and OSR1 (oxidative stress-responsive kinase 1) participate in the regulation of the NaCl cotransporter NCC and the Na⁺,K⁺,2Cl^- cotransporter NKCC2. The kinases are regulated by WNK (with-no-K[Lys]) kinases. Mutations of genes encoding WNK kinases underly Gordon's syndrome, a monogenic disease leading to hypertension and hyperkalemia. WNK kinases further regulate the renal outer medullary K⁺ channel ROMK1. The present study explored, whether SPAK and/or OSR1 have similarly the potential to modify the activity of ROMK1. Methods: ROMK1 was expressed in Xenopus oocytes with or without additional expression of wild-type SPAK, constitutively active ^T233ESPAK, catalytically inactive ^D212ASPAK, wild-type OSR1, constitutively active ^T185EOSR1 and catalytically inactive ^D164AOSR1. Channel activity was determined utilizing dual electrode voltage clamp and ROMK1 protein abundance in the cell membrane utilizing chemiluminescence of ROMK1 containing an extracellular hemagglutinin epitope (ROMK1-HA). Results: ROMK1 activity and ROMK1-HA protein abundance were significantly down-regulated by wild-type SPAK and ^T233ESPAK, but not by ^D212ASPAK. Similarly, ROMK1 activity and ROMK1-HA protein abundance were significantly down-regulated by wild-type OSR1 and ^T185EOSR1, but not by ^D164AOSR1. Conclusion: ROMK1 protein abundance and activity are down-regulated by SPAK and OSR1.