Regulation of activity and localization of the WNK1 protein kinase by hyperosmotic stress.
Mutations within the WNK1 (with-no-K[Lys] kinase-1) gene cause Gordon's hypertension syndrome. Little is known about how WNK1 is regulated. We demonstrate that WNK1 is rapidly activated and phosphorylated at multiple residues after exposure of cells to hyperosmotic conditions and that activation is mediated by the phosphorylation of its T-loop Ser382 residue, possibly triggered by a transautophosphorylation reaction. Activation of WNK1 coincides with the phosphorylation and activation of two WNK1 substrates, namely, the protein kinases STE20/SPS1-related proline alanine-rich kinase (SPAK) and oxidative stress response kinase-1 (OSR1). Small interfering RNA depletion of WNK1 impairs SPAK/OSR1 activity and phosphorylation of residues targeted by WNK1. Hyperosmotic stress induces rapid redistribution of WNK1 from the cytosol to vesicular structures that may comprise trans-Golgi network (TGN)/recycling endosomes, as they display rapid movement, colocalize with clathrin, adaptor protein complex 1 (AP-1), and TGN46, but not the AP-2 plasma membrane-coated pit marker nor the endosomal markers EEA1, Hrs, and LAMP1. Mutational analysis suggests that the WNK1 C-terminal noncatalytic domain mediates vesicle localization. Our observations shed light on the mechanism by which WNK1 is regulated by hyperosmotic stress.
Zagórska
Anna
A
Medical Research Council Protein Phosphorylation Unit, School of Life Sciences, Medical Sciences Institute/Wellcome Trust Biocentre Complex, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Pozo-Guisado
Eulalia
E
Boudeau
Jérôme
J
Vitari
Alberto C
AC
Rafiqi
Fatema H
FH
Thastrup
Jacob
J
Deak
Maria
M
Campbell
David G
DG
Morrice
Nick A
NA
Prescott
Alan R
AR
Alessi
Dario R
DR
D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002966:Clathrin / Q000378:metabolism; D022162:Cytoplasmic Vesicles / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D015778:Minor Histocompatibility Antigens; D008969:Molecular Sequence Data; D009997:Osmotic Pressure; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013012:Sorbitol / Q000494:pharmacology*; D000076204:WNK Lysine-Deficient Protein Kinase 1
D002966:Clathrin; D047908:Intracellular Signaling Peptides and Proteins; D015778:Minor Histocompatibility Antigens; D011993:Recombinant Fusion Proteins; D010768:Phosphoserine; D013012:Sorbitol; C118135:OXSR1 protein, human; D017346:Protein Serine-Threonine Kinases; C486409:STK39 protein, human; D000076204:WNK Lysine-Deficient Protein Kinase 1; C498743:WNK1 protein, human