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Cdk5 phosphorylation of EFhd2 at S74 affects its calcium binding activity
Author(s) -
VázquezRosa Edwin,
RodríguezCruz Eva N.,
Serrano Sybelle,
RodríguezLaureano Lucelenie,
Vega Irving E.
Publication year - 2014
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.2499
Subject(s) - cyclin dependent kinase 5 , phosphorylation , kinase , microbiology and biotechnology , biology , protein phosphorylation , activator (genetics) , phosphoproteomics , chemistry , protein kinase a , biochemistry , cyclin dependent kinase 2 , receptor
EFhd2 is a calcium binding protein, which is highly expressed in the central nervous system and associated with pathological forms of tau proteins in tauopathies. Previous phosphoproteomics studies and bioinformatics analysis suggest that EFhd2 may be phosphorylated. Here, we determine whether Cdk5, a hyperactivated kinase in tauopathies, phosphorylates EFhd2 and influence its known molecular activities. The results indicated that EFhd2 is phosphorylated by brain extract of the transgenic mouse CK‐p25, which overexpresses the Cdk5 constitutive activator p25. Consistently, in vitro kinase assays demonstrated that Cdk5, but not GSK3β, directly phosphorylates EFhd2. Biomass, tandem mass spectrometry, and mutagenesis analyses indicated that Cdk5 monophosphorylates EFhd2 at S74, but not the adjacent S76. Furthermore, Cdk5‐mediated phosphorylation of EFhd2 affected its calcium binding activity. Finally, a phospho‐specific antibody was generated against EFhd2 phosphorylated at S74 and was used to detect this phosphorylation event in postmortem brain tissue from Alzheimer's disease and normal‐aging control cases. Results demonstrated that EFhd2 is phosphorylated in vivo at S74. These results imply that EFhd2's physiological and/or pathological function could be regulated by its phosphorylation state.