title: Identification of protein kinases responsible for phosphorylation of Epstein-Barr virus nuclear antigen leader protein at serine-35, which regulates its coactivator function.
abstract: Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is a phosphoprotein suggested to play important roles in EBV-induced immortalization. Earlier studies have shown that the major site of phosphorylation of EBNA-LP by cellular kinase(s) is a serine residue at position 35 (Ser-35) and that the phosphorylation of Ser-35 is critical for regulation of the coactivator function of EBNA-LP (Yokoyama et al., J Virol 75, 5119-5128, 2001). In the present study, we have attempted to identify protein kinase(s) responsible for the phosphorylation of EBNA-LP at Ser-35. A purified chimeric protein consisting of glutathione S-transferase (GST) fused to a domain of EBNA-LP containing Ser-35 was found to be specifically phosphorylated by purified cdc2 in vitro, while GST fused to a mutated domain of EBNA-LP in which Ser-35 was replaced with alanine was not. In addition, overexpression of cdc2 in mammalian cells caused a significant increase in the phosphorylation of EBNA-LP, while this increased phosphorylation was eliminated if Ser-35 of EBNA-LP was replaced with alanine. These results indicate that the cellular protein kinase cdc2 mediates the phosphorylation of EBNA-LP at Ser-35. Recently, we reported that cdc2 and conserved protein kinases encoded by herpesviruses phosphorylate the same amino acid residue of target proteins (Kawaguchi et al., J Virol 77, 2359-2368, 2003). Consistent with this, the EBV-encoded conserved protein kinase BGLF4 specifically mediated the phosphorylation of EBNA-LP at Ser-35. These results indicate that the coactivator function of EBNA-LP can be regulated by the activity of these cellular and viral protein kinases.
authors.affiliation: Department of Veterinary Microbiology, Graduate School of Agricultural and Life Science, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan.|Department of Cell Regulation, Medical Research Institute, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Kato
authors.affiliation: Department of Cell Regulation, Medical Research Institute, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan.
authors.forename: Akihiko
authors.initials: A
authors.lastname: Yokoyama
authors.affiliation: Department of Veterinary Microbiology, Graduate School of Agricultural and Life Science, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan.
authors.forename: Yukinobu
authors.initials: Y
authors.lastname: Tohya
authors.affiliation: Department of Veterinary Microbiology, Graduate School of Agricultural and Life Science, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan.
authors.forename: Hiroomi
authors.initials: H
authors.lastname: Akashi
authors.affiliation: Department of Virology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan.
authors.forename: Yukihiro
authors.initials: Y
authors.lastname: Nishiyama
authors.affiliation: PRESTO, Japan Science and Technology Corporation, Tachikawa, Tokyo 190-0012, Japan.|Department of Virology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan.|Department of Cell Regulation, Medical Research Institute, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Kawaguchi
mesh_terms: D000409:Alanine / Q000378:metabolism; D019943:Amino Acid Substitution; D000818:Animals; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D002460:Cell Line; D002472:Cell Transformation, Viral; D004789:Enzyme Activation; D004854:Herpesvirus 4, Human / Q000737:chemistry / Q000502:physiology; D009154:Mutation; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine / Q000737:chemistry / Q000378:metabolism; D018411:Spodoptera; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C402640:EBNA-LP protein, Human herpesvirus 4; D011994:Recombinant Proteins; D014764:Viral Proteins; D012694:Serine; D011494:Protein Kinases; C406063:BGLF4 protein, Epstein-Barr virus; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; D000409:Alanine