1887

Abstract

The gene product of bovine herpesvirus-1 (BoHV-1) is a protein kinase that is expressed early during infection and capable of autophosphorylation. By examining differentially labelled US3 moieties by co-immunoprecipitation, we demonstrated that the protein kinase interacts with itself , which supports autophosphorylation by US3. Based on its homology to other serine/threonine protein kinases, we defined two highly conserved lysines in US3, at position 195 within the ATP-binding pocket and at position 282 within the catalytic loop; altering either residue resulted in kinase-dead mutants, demonstrating that these two residues are critical for the catalytic activity of BoHV-1 US3. During immunoprecipitation experiments, US3 interacted weakly with VP22, another tegument protein of BoHV-1. Furthermore, VP22 co-localized with US3 inside the nucleus in BoHV-1-infected cells. kinase assays demonstrated that VP22 is phosphorylated not only by US3, but also by the cellular casein kinase 2 (CK2) protein. The selective CK2 protein kinase inhibitor, 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT) and the less specific CK2 inhibitor Kenpaullone reduced VP22 phosphorylation, while CK1, protein kinase C or protein kinase A inhibitors did not affect phosphorylation. When US3 was included with VP22 in the kinase assay in the presence of DMAT, a low level of VP22 phosphorylation was observed. These data demonstrate that BoHV-1 VP22 interacts with both CK2 and US3, and that CK2 is the major kinase phosphorylating VP22, with US3 playing a minor role.

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2010-05-01
2024-03-28
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References

  1. Asai, R., Ohno, T., Kato, A. & Kawaguchi, Y.(2007). Identification of proteins directly phosphorylated by UL13 protein kinase from herpes simplex virus 1. Microbes Infect 9, 1434–1438.[CrossRef] [Google Scholar]
  2. Birchall, A. M., Bishop, J., Bradshaw, D., Cline, A., Coffey, J., Elliott, L. H., Gibson, V. M., Greenham, A., Hallam, T. J. & other authors(1994). Ro 32–0432, a selective and orally active inhibitor of protein kinase C prevents T-cell activation. J Pharmacol Exp Ther 268, 922–929. [Google Scholar]
  3. Blouin, A. & Blaho, J. A.(2001). Assessment of the subcellular localization of the herpes simplex virus structural protein VP22 in the absence of other viral gene products. Virus Res 81, 57–68.[CrossRef] [Google Scholar]
  4. Brignati, M. J., Loomis, J. S., Wills, J. W. & Courtney, R. J.(2003). Membrane association of VP22, a herpes simplex virus type 1 tegument protein. J Virol 77, 4888–4898.[CrossRef] [Google Scholar]
  5. Coulter, L. J., Moss, H. W., Lang, J. & McGeoch, D. J.(1993). A mutant of herpes simplex virus type 1 in which the UL13 protein kinase gene is disrupted. J Gen Virol 74, 387–395.[CrossRef] [Google Scholar]
  6. Daikoku, T., Yamashita, Y., Tsurumi, T., Maeno, K. & Nishiyama, Y.(1993). Purification and biochemical characterization of the protein kinase encoded by the US3 gene of herpes simplex virus type 2. Virology 197, 685–694.[CrossRef] [Google Scholar]
  7. Elliott, G. & O'Hare, P.(1997). Intercellular trafficking and protein delivery by a herpesvirus structural protein. Cell 88, 223–233.[CrossRef] [Google Scholar]
  8. Elliott, G. & O'Hare, P.(1998). Herpes simplex virus type 1 tegument protein VP22 induces the stabilization and hyperacetylation of microtubules. J Virol 72, 6448–6455. [Google Scholar]
  9. Elliott, G. & O'Hare, P.(1999). Live-cell analysis of a green fluorescent protein-tagged herpes simplex virus infection. J Virol 73, 4110–4119. [Google Scholar]
  10. Elliott, G. & O'Hare, P.(2000). Cytoplasm-to-nucleus translocation of a herpesvirus tegument protein during cell division. J Virol 74, 2131–2141.[CrossRef] [Google Scholar]
  11. Elliott, G., Mouzakitis, G. & O'Hare, P.(1995). VP16 interacts via its activation domain with VP22, a tegument protein of herpes simplex virus, and is relocated to a novel macromolecular assembly in coexpressing cells. J Virol 69, 7932–7941. [Google Scholar]
  12. Elliott, G., O'Reilly, D. & O'Hare, P.(1996). Phosphorylation of the herpes simplex virus type 1 tegument protein VP22. Virology 226, 140–145.[CrossRef] [Google Scholar]
  13. Elliott, G., O'Reilly, D. & O'Hare, P.(1999). Identification of phosphorylation sites within the herpes simplex virus tegument protein VP22. J Virol 73, 6203–6206. [Google Scholar]
  14. Frame, M. C., Purves, F. C., McGeoch, D. J., Marsden, H. S. & Leader, D. P.(1987). Identification of the herpes simplex virus protein kinase as the product of viral gene US3. J Gen Virol 68, 2699–2704.[CrossRef] [Google Scholar]
  15. Geiss, B. J., Tavis, J. E., Metzger, L. M., Leib, D. A. & Morrison, L. A.(2001). Temporal regulation of herpes simplex virus type 2 VP22 expression and phosphorylation. J Virol 75, 10721–10729.[CrossRef] [Google Scholar]
  16. Goshima, F., Daikoku, T., Yamada, H., Oshima, S., Tsurumi, T. & Nishiyama, Y.(1998). Subcellular localization of the US3 protein kinase of herpes simplex virus type 2. Arch Virol 143, 613–622.[CrossRef] [Google Scholar]
  17. Harms, J. S., Ren, X., Oliveira, S. C. & Splitter, G. A.(2000). Distinctions between bovine herpesvirus 1 and herpes simplex virus type 1 VP22 tegument protein subcellular associations. J Virol 74, 3301–3312.[CrossRef] [Google Scholar]
  18. Hung, C. F., Cheng, W. F., Chai, C. Y., Hsu, K. F., He, L., Ling, M. & Wu, T. C.(2001). Improving vaccine potency through intercellular spreading and enhanced MHC class I presentation of antigen. J Immunol 166, 5733–5740.[CrossRef] [Google Scholar]
  19. Ioannou, X. P., Gomis, S. M., Hecker, R., Babiuk, L. A. & van Drunen Littel-van den Hurk, S.(2003). Safety and efficacy of CpG-containing oligodeoxynucleotides as immunological adjuvants in rabbits. Vaccine 21, 4368–4372.[CrossRef] [Google Scholar]
  20. Jones, C. & Chowdhury, S.(2007). A review of the biology of bovine herpesvirus type 1 (BHV-1), its role as a cofactor in the bovine respiratory disease complex and development of improved vaccines. Anim Health Res Rev 8, 187–205.[CrossRef] [Google Scholar]
  21. Kato, A., Yamamoto, M., Ohno, T., Kodaira, H., Nishiyama, Y. & Kawaguchi, Y.(2005). Identification of proteins phosphorylated directly by the Us3 protein kinase encoded by herpes simplex virus 1. J Virol 79, 9325–9331.[CrossRef] [Google Scholar]
  22. Kato, A., Tanaka, M., Yamamoto, M., Asai, R., Sata, T., Nishiyama, Y. & Kawaguchi, Y.(2008). Identification of a physiological phosphorylation site of the herpes simplex virus 1-encoded protein kinase Us3 which regulates its optimal catalytic activity in vitro and influences its function in infected cells. J Virol 82, 6172–6189.[CrossRef] [Google Scholar]
  23. Kim, T. W., Hung, C. F., Kim, J. W., Juang, J., Chen, P. J., He, L., Boyd, D. A. & Wu, T. C.(2004). Vaccination with a DNA vaccine encoding herpes simplex virus type 1 VP22 linked to antigen generates long-term antigen-specific CD8-positive memory T cells and protective immunity. Hum Gene Ther 15, 167–177.[CrossRef] [Google Scholar]
  24. Labiuk, S. L., Babiuk, L. A. & van Drunen Littel-van den Hurk, S.(2009). The major tegument protein VP8 of bovine herpesvirus-1 is phosphorylated by viral US3 and cellular CK2 protein kinases. J Gen Virol 90, 2829–2839.[CrossRef] [Google Scholar]
  25. Leach, N., Bjerke, S. L., Christensen, D. K., Bouchard, J. M., Mou, F., Park, R., Baines, J., Haraguchi, T. & Roller, R. J.(2007). Emerin is hyperphosphorylated and redistributed in herpes simplex virus type 1-infected cells in a manner dependent on both UL34 and US3. J Virol 81, 10792–10803.[CrossRef] [Google Scholar]
  26. Leader, D. P., Deana, A. D., Marchiori, F., Purves, F. C. & Pinna, L. A.(1991). Further definition of the substrate specificity of the alpha-herpesvirus protein kinase and comparison with protein kinases A and C. Biochim Biophys Acta 1091, 426–431.[CrossRef] [Google Scholar]
  27. Lemken, M. L., Wolf, C., Wybranietz, W. A., Schmidt, U., Smirnow, I., Buhring, H. J., Mack, A. F., Lauer, U. M. & Bitzer, M.(2007). Evidence for intercellular trafficking of VP22 in living cells. Mol Ther 15, 310–319.[CrossRef] [Google Scholar]
  28. Leopardi, R., Van Sant, C. & Roizman, B.(1997). The herpes simplex virus 1 protein kinase US3 is required for protection from apoptosis induced by the virus. Proc Natl Acad Sci U S A 94, 7891–7896.[CrossRef] [Google Scholar]
  29. Liang, X., Chow, B. & Babiuk, L. A.(1997). Study of immunogenicity and virulence of bovine herpesvirus 1 mutants deficient in the UL49 homolog, UL49.5 homolog and dUTPase genes in cattle. Vaccine 15, 1057–1064.[CrossRef] [Google Scholar]
  30. Lobanov, V. A., Babiuk, L. A. & van Drunen Littel-van den Hurk, S.(2009). Intracellular trafficking of VP22 in bovine herpesvirus-1 infected cells. Virology 396, 189–202. [Google Scholar]
  31. Marchler-Bauer, A. & Bryant, S. H.(2004). CD-Search: protein domain annotations on the fly. Nucleic Acids Res 32, W327–W331 [Google Scholar]
  32. Matsuzaki, A., Yamauchi, Y., Kato, A., Goshima, F., Kawaguchi, Y., Yoshikawa, T. & Nishiyama, Y.(2005). US3 protein kinase of herpes simplex virus type 2 is required for the stability of the UL46-encoded tegument protein and its association with virus particles. J Gen Virol 86, 1979–1985.[CrossRef] [Google Scholar]
  33. Meggio, F. & Pinna, L. A.(2003). One-thousand-and-one substrates of protein kinase CK2? FASEB J 17, 349–368.[CrossRef] [Google Scholar]
  34. Misra, V., Babiuk, L. A. & Darcel, C. L.(1983). Analysis of bovine herpes virus-type 1 isolates by restriction endonuclease fingerprinting. Arch Virol 76, 341–354.[CrossRef] [Google Scholar]
  35. Morris, J. B., Hofemeister, H. & O'Hare, P.(2007). Herpes simplex virus infection induces phosphorylation and delocalization of emerin, a key inner nuclear membrane protein. J Virol 81, 4429–4437.[CrossRef] [Google Scholar]
  36. Morrison, E. E., Wang, Y. F. & Meredith, D. M.(1998). Phosphorylation of structural components promotes dissociation of the herpes simplex virus type 1 tegument. J Virol 72, 7108–7114. [Google Scholar]
  37. Murray, A. J.(2008). Pharmacological PKA inhibition: all may not be what it seems. Sci Signal 1, re4 [Google Scholar]
  38. Pagano, M. A., Meggio, F., Ruzzene, M., Andrzejewska, M., Kazimierczuk, Z. & Pinna, L. A.(2004). 2-Dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole: a novel powerful and selective inhibitor of protein kinase CK2. Biochem Biophys Res Commun 321, 1040–1044.[CrossRef] [Google Scholar]
  39. Pinna, L. A.(2002). Protein kinase CK2: a challenge to canons. J Cell Sci 115, 3873–3878.[CrossRef] [Google Scholar]
  40. Pomeranz, L. E. & Blaho, J. A.(1999). Modified VP22 localizes to the cell nucleus during synchronized herpes simplex virus type 1 infection. J Virol 73, 6769–6781. [Google Scholar]
  41. Purves, F. C., Deana, A. D., Marchiori, F., Leader, D. P. & Pinna, L. A.(1986). The substrate specificity of the protein kinase induced in cells infected with herpesviruses: studies with synthetic substrates [corrected] indicate structural requirements distinct from other protein kinases. Biochim Biophys Acta 889, 208–215.[CrossRef] [Google Scholar]
  42. Purves, F. C., Longnecker, R. M., Leader, D. P. & Roizman, B.(1987). Herpes simplex virus 1 protein kinase is encoded by open reading frame US3 which is not essential for virus growth in cell culture. J Virol 61, 2896–2901. [Google Scholar]
  43. Rena, G., Bain, J., Elliott, M. & Cohen, P.(2004). D4476, a cell-permeant inhibitor of CK1, suppresses the site-specific phosphorylation and nuclear exclusion of FOXO1a. EMBO Rep 5, 60–65.[CrossRef] [Google Scholar]
  44. Retamal, C. A., Thiebaut, P. & Alves, E. W.(1999). Protein purification from polyacrylamide gels by sonication extraction. Anal Biochem 268, 15–20.[CrossRef] [Google Scholar]
  45. Reynolds, A. E., Wills, E. G., Roller, R. J., Ryckman, B. J. & Baines, J. D.(2002). Ultrastructural localization of the herpes simplex virus type 1 UL31, UL34, and US3 proteins suggests specific roles in primary envelopment and egress of nucleocapsids. J Virol 76, 8939–8952.[CrossRef] [Google Scholar]
  46. Takashima, Y., Tamura, H., Xuan, X. & Otsuka, H.(1999). Identification of the US3 gene product of BHV-1 as a protein kinase and characterization of BHV-1 mutants of the US3 gene. Virus Res 59, 23–34.[CrossRef] [Google Scholar]
  47. Turin, L., Russo, S. & Poli, G.(1999). BHV-1: new molecular approaches to control a common and widespread infection. Mol Med 5, 261–284. [Google Scholar]
  48. Zaharevitz, D. W., Gussio, R., Leost, M., Senderowicz, A. M., Lahusen, T., Kunick, C., Meijer, L. & Sausville, E. A.(1999). Discovery and initial characterization of the paullones, a novel class of small-molecule inhibitors of cyclin-dependent kinases. Cancer Res 59, 2566–2569. [Google Scholar]
  49. Zheng, C., Babiuk, L. A. & van Drunen Littel-van den Hurk, S.(2005). Bovine herpesvirus 1 VP22 enhances the efficacy of a DNA vaccine in cattle. J Virol 79, 1948–1953.[CrossRef] [Google Scholar]
  50. Zheng, C. F., Brownlie, R., Huang, D. Y., Babiuk, L. A. & van Drunen Littel-van den Hurk, S.(2006). Intercellular trafficking of the major tegument protein VP22 of bovine herpesvirus-1 and its application to improve a DNA vaccine. Arch Virol 151, 985–993.[CrossRef] [Google Scholar]
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vol. , part 5, pp. 1117 - 1126

List of mutagenesis primers used for the mutation of pUS3–HA [PDF](36 KB)



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