Major tegument protein VP8 of bovine herpesvirus 1 is phosphorylated by viral US3 and cellular CK2 protein kinases

• Published in: Journal of general virology Vol. 90; no. 12; pp. 2829 - 2839
• Main Authors: Labiuk, Shaunivan L, Babiuk, Lorne A, Van Drunen Littel-van den Hurk, Sylvia
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.12.2009; Society for General Microbiology
• Subjects: Animals; autophosphorylation; Benzimidazoles - pharmacology; Biological and medical sciences; Bovine alphaherpesvirus 1; Bovine herpesvirus 1; Capsid Proteins - metabolism; Casein Kinase II - antagonists & inhibitors; Casein Kinase II - metabolism; Cattle; Cell Line; cell lines; Chlorocebus aethiops; COS Cells; cytopathogenicity; enzyme activity; Fundamental and applied biological sciences. Psychology; Herpesvirus 1, Bovine - genetics; Herpesvirus 1, Bovine - growth & development; Herpesvirus 1, Bovine - metabolism; host-pathogen relationships; Immunoprecipitation; infection; Kidney - cytology; Mass Spectrometry; Microbiology; Miscellaneous; Phosphorylation; protein kinases; protein phosphorylation; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; tegument protein; viral proteins; Viral Proteins - genetics; Viral Proteins - metabolism; Virology; VP8 protein; Virus; Phosphorylation; Microbiology; Enzyme; Herpesviridae; Transferases; Non-specific serine/threonine protein kinase; Alphaherpesvirinae; Bovine herpesvirus 1; Veterinary; In vitro; Protein
• DOI: 10.1099/vir.0.013532-0
• ISSN: 0022-1317; 1465-2099

The UL47 gene product, VP8, is one of the major tegument proteins of bovine herpesvirus 1 (BoHV-1) and is subject to phosphorylation. Analysis of protein bands co-immunoprecipitated with VP8 from BoHV-1-infected cells by mass spectroscopy suggested that VP8 interacts with two protein kinases: cellular CK2 and viral US3. CK2 is a highly conserved cellular protein, expressed ubiquitously and known to phosphorylate numerous proteins. The US3 gene product is one of the viral kinases produced by BoHV-1 during infection. Interactions of CK2 and US3 with VP8 were confirmed outside the context of infection when FLAG–VP8 was expressed alone or co-expressed with US3–haemagglutinin tag in Cos-7 cells. Furthermore, VP8 and US3 were found to co-localize in the nucleus during viral infection. To explore the significance of these interactions, an in vitro kinase assay was performed, which demonstrated that VP8 is heavily phosphorylated by CK2. In the presence of the highly specific CK2 kinase inhibitor 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT), phosphorylation of VP8 was significantly reduced. Phosphorylation of VP8 was also inhibited by the presence of kenpaullone, a less specific CK2 inhibitor, but not by protein kinase CK1 or protein kinase C inhibitors. When VP8 and US3 were both included in the kinase assay in the presence of DMAT, phosphorylation of VP8 was again observed. Autophosphorylation of US3 was also detected and was not inhibited by DMAT. Based on these results, it is proposed that VP8 interacts with cellular CK2 and viral US3 in BoHV-1-infected cells, and is in turn subject to kinase activities associated with both of these proteins.