A Single Amino Acid Substitution Makes WNK4 Susceptible to SB 203580 and SB 202190

Regulation of the SLC12 family of membrane transporters including NCCT involves a scaffold of interacting proteins including the STE 20 kinase SPAK and the WNK kinases, WNK 1 and WNK 4, which are mutated in the hypertensive syndrome of pseudohypoaldosteronism type 2 (PHAII). WNK4 regulates NCCT by affecting forward trafficking to the surface membrane. Studies in Xenopus using kinase dead WNK4 site mutants have produced inconsistent results with regard to the necessity of kinase function for NCCT regulation. Dynamic inhibition of WNK4 by small molecules may bring clarity to this issue however WNK4 is naturally resistant to commercial MAP kinase inhibitors owing to steric constraints prohibiting entry of small molecules to the active site. Using an approach similar to that used in p38 and ERK, we show that a single substitution in WNK4 (T261G) dramatically enhances its susceptibility to the inhibitors SB 202190 and SB 203580.