A Cdc2-related protein kinase hPFTAIRE1 from human brain interacting with 14-3-3 proteins.
hPFTAIRE1 (PFTK1), a Cdc2-related protein kinase, is highly expressed in human brain. It exhibits cytoplasmic distribution in Hela cells, although it contains two nuclear localization signals (NLSs) in its N-terminus. To search for its substrates and regulatory components, we screened a two-hybrid library by using the full-length hPFTAIRE1 as a bait. Four 14-3-3 isoforms (beta, epsilon, eta, tau) were identified interacting with the hPFTAIRE1. We found a putative 14-3-3 binding consensus motif (RHSSPSS) in the hPFTAIRE1, which overlapped with its second NLS. Deletion of the RHSSPSS motif or substitution of Ser119 with Ala in the conserved binding motif abolished the specific interaction between the hPFTAIRE1 and the 14-3-3 proteins. The mutant S120A hPFTAIRE1 also showed a weak interaction to the 14-3-3 proteins. The results suggested that the Ser119 is crucial for the interaction between hPFTAIRE1 and the 14-3-3 proteins. All the hPFTAIRE1 mutants distributed in cytoplasm of Hela cells and human neuroblastoma cells (SH-SY5Y) when fused to the C-terminus of a green fluorescent protein (GFP), indicating that binding with the 14-3-3 proteins does not contribute to the subcellular localization of the hPFTAIRE1, although the binding may be involved in its signaling regulation.
Gao
Yankun
Y
State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yue-yang Road, Shanghai 200031, China.
Jiang
Mei
M
Yang
Tao
T
Ni
Jian
J
Chen
Jiangye
J
D048948:14-3-3 Proteins / Q000378:metabolism*; D000595:Amino Acid Sequence; D001921:Brain / Q000378:metabolism*; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D045744:Cell Line, Tumor; D018844:Cyclin-Dependent Kinases; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D019913:Nuclear Localization Signals / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques
D048948:14-3-3 Proteins; D019913:Nuclear Localization Signals; D020033:Protein Isoforms; D016203:CDC2 Protein Kinase; C515076:CDK14 protein, human; D018844:Cyclin-Dependent Kinases