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The Cul4‐DDB1‐WDR3/WDR6 Complex Binds SPAK and OSR1 Kinases in a Phosphorylation‐Dependent Manner
Author(s) -
Dhiani Binar A.,
Mehellou Youcef
Publication year - 2020
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201900454
Subject(s) - phosphorylation , kinase , ubiquitin ligase , cullin , protein serine threonine kinases , microbiology and biotechnology , biochemistry , chemistry , serine , protein kinase a , ubiquitin , biology , gene
SPAK and OSR1 are two protein kinases that play critical roles in regulating ion homeostasis. They are activated under osmotic stress through phosphorylation by their upstream WNK kinases at a conserved threonine site on their T‐loops. Additionally, WNK kinases phosphorylate SPAK and OSR1 at a highly conserved serine residue on their S‐motif, the function of which remains elusive. Using affinity pull down and mass spectrometry, we identified the E3 ubiquitin ligase complex Cullin 4‐DDB1‐WDR3/WDR6 as a binder to OSR1 kinase in a SPAK/OSR1 S‐motif phosphorylation‐dependent manner. This binding was found to be compromised by S‐motif phosphorylation following osmotic stress. Using proteasomal and neddylation inhibitors, we subsequently showed that OSR1 ubiquitylation was abolished under osmotic stress when its S‐motif is phosphorylated. These results provide the first example of an E3 ubiquitin ligase system that binds the OSR1 kinase and, thus, links the CRL4 complex to ion homeostasis.