West Nile virus capsid protein interaction with importin and HDM2 protein is regulated by protein kinase C-mediated phosphorylation.
West Nile virus (WNV) capsid (C) protein was shown to enter the nucleus via importin-mediated pathway and induce apoptosis although the precise regulatory mechanisms for such events have remained elusive. In this study, it was shown that WNV C protein was phosphorylated by protein kinase C (PKC). PKC-mediated phosphorylation influenced nuclear trafficking of C protein by modulating the efficiency of C protein-importin-alpha binding. Combination of bio-informatics, site-directed mutagenesis, co-immunoprecipitation, immuno-fluorescence and mammalian two-hybrid analyses showed that phosphorylation at amino acid residues residing near (Ser83) or within (Ser99 and Thr100) the bipartite nuclear localization motif of WNV C protein was essential for efficient interaction between C protein and importin-alpha. In addition, phosphorylation of WNV C protein by PKC was shown to enhance its binding to HDM2 and could subsequently induce p53-dependent apoptosis. Collectively, this study highlighted that phosphorylation is an important post-translational modification required to execute the functions of C protein.
Bhuvanakantham
Raghavan
R
Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Cheong
Yuen Kuen
YK
Ng
Mah-Lee
ML
D000818:Animals; D017209:Apoptosis; D036022:Capsid Proteins / Q000378:metabolism*; D002460:Cell Line; D006224:Cricetinae; D047468:Immunoprecipitation; D028884:Karyopherins / Q000378:metabolism*; D018613:Microscopy, Confocal; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014902:West Nile virus / Q000472:pathogenicity*
D036022:Capsid Proteins; D028884:Karyopherins; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D011493:Protein Kinase C