Detection of Epstein–Barr virus BGLF4 protein kinase in virus replication compartments and virus particles Free

Abstract

BGLF4 is the only serine/threonine protein kinase identified in Epstein–Barr virus (EBV); it is known to phosphorylate viral DNA polymerase processivity factor, EA-D (BMRF1), EBNA-LP, EBNA-2, cellular EF-1 and nucleoside analogue ganciclovir. However, the expression and biological functions of BGLF4 have not yet been clearly demonstrated in EBV-infected cells. To reveal authentic functions of BGLF4 protein within viral-replicating cells, a panel of specific monoclonal antibodies was generated and characterized. The major immunogenic regions of BGLF4 were mapped to aa 27–70 and 327–429. Using these antibodies, the expression kinetics and localization of BGLF4 were analysed in reactivated EBV-positive lymphoid and epithelial cells. BGLF4 was expressed as a phosphoprotein at the early lytic stage and was detected predominantly in the nucleus of EBV-positive cells, but small amounts of BGLF4 were observed in cytosolic and heavy membrane fractions at the late phase of virus replication. Additionally, it was demonstrated that BGLF4 co-localizes with viral DNA polymerase processivity factor, EA-D (BMRF1), in the virus replication compartment and that it is a virion component. Finally, possible functional domains at the N terminus of BGLF4 were analysed and it was found that aa 1–26 of BGLF4 are dispensable for EA-D phosphorylation, whereas deletion of aa 27–70 reduced kinase activity.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.81313-0
2005-12-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/jgv/86/12/3215.html?itemId=/content/journal/jgv/10.1099/vir.0.81313-0&mimeType=html&fmt=ahah

References

  1. Bortz E., Whitelegge J. P., Jia Q., Zhou Z. H., Stewart J. P., Wu T. T., Sun R. 2003; Identification of proteins associated with murine gammaherpesvirus 68 virions. J Virol 77:13425–13432 [CrossRef]
    [Google Scholar]
  2. Chang Y., Tung C. H., Huang Y. T., Lu J., Chen J. Y., Tsai C. H. 1999; Requirement for cell-to-cell contact in Epstein–Barr virus infection of nasopharyngeal carcinoma cells and keratinocytes. J Virol 73:8857–8866
    [Google Scholar]
  3. Chang Y., Cheng S. D., Tsai C. H. 2002; Chromosomal integration of Epstein–Barr virus genomes in nasopharyngeal carcinoma cells. Head Neck 24:143–150 [CrossRef]
    [Google Scholar]
  4. Chen C., Okayama H. 1987; High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol 7:2745–2752
    [Google Scholar]
  5. Chen M. R., Tsai C. H., Wu F. F., Kan S. H., Yang C. S., Chen J. Y. 1999; The major immunogenic epitopes of Epstein–Barr virus (EBV) nuclear antigen 1 are encoded by sequence domains which vary among nasopharyngeal carcinoma biopsies and EBV-associated cell lines. J Gen Virol 80:447–455
    [Google Scholar]
  6. Chen M. R., Chang S. J., Huang H., Chen J. Y. 2000a; A protein kinase activity associated with Epstein–Barr virus BGLF4 phosphorylates the viral early antigen EA-D in vitro. J Virol 74:3093–3104 [CrossRef]
    [Google Scholar]
  7. Chen M. R., Huang H., Fen C. Y., Chen J. Y. 2000b; A novel EBNA-1 tag system for high level expression and efficient detection of fusion proteins in vitro and in vivo. J Virol Methods 85:35–41 [CrossRef]
    [Google Scholar]
  8. Daikoku T., Kudoh A., Fujita M., Sugaya Y., Isomura H., Shirata N., Tsurumi T. 2005; Architecture of replication compartments formed during Epstein–Barr virus lytic replication. J Virol 79:3409–3418 [CrossRef]
    [Google Scholar]
  9. 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]
  10. Feng W. H., Hong G., Delecluse H. J., Kenney S. C. 2004; Lytic induction therapy for Epstein–Barr virus-positive B-cell lymphomas. J Virol 78:1893–1902 [CrossRef]
    [Google Scholar]
  11. Gershburg E., Pagano J. S. 2002; Phosphorylation of the Epstein–Barr virus (EBV) DNA polymerase processivity factor EA-D by the EBV-encoded protein kinase and effects of the l-riboside benzimidazole 1263W94. J Virol 76:998–1003 [CrossRef]
    [Google Scholar]
  12. Gershburg E., Marschall M., Hong K., Pagano J. S. 2004; Expression and localization of the Epstein–Barr virus-encoded protein kinase. J Virol 78:12140–12146 [CrossRef]
    [Google Scholar]
  13. Hamza M. S., Reyes R. A., Izumiya Y., Wisdom R., Kung H. J., Luciw P. A. 2004; ORF36 protein kinase of Kaposi's sarcoma herpesvirus activates the c-Jun N-terminal kinase signaling pathway. J Biol Chem 279:38325–38330 [CrossRef]
    [Google Scholar]
  14. Hanks S. K., Hunter T. 1995; Protein kinases 6. The eukaryotic protein kinase superfamily: kinase (catalytic) domain structure and classification. FASEB J 9:576–596
    [Google Scholar]
  15. Hinuma Y., Konn M., Yamaguchi J., Wudarski D. J., Blakeslee J. R. Jr, Grace J. T. Jr 1967; Immunofluorescence and herpes-type virus particles in the P3HR-1 Burkitt lymphoma cell line. J Virol 1:1045–1051
    [Google Scholar]
  16. Holley-Guthrie E. A., Seaman W. T., Bhende P., Merchant J. L., Kenney S. C. 2005; The Epstein–Barr virus protein BMRF1 activates gastrin transcription. J Virol 79:745–755 [CrossRef]
    [Google Scholar]
  17. Hopp T. P., Woods K. R. 1981; Prediction of protein antigenic determinants from amino acid sequences. Proc Natl Acad Sci U S A 78:3824–3828 [CrossRef]
    [Google Scholar]
  18. Israel B. F., Kenney S. C. 2003; Virally targeted therapies for EBV-associated malignancies. Oncogene 22:5122–5130 [CrossRef]
    [Google Scholar]
  19. Johannsen E., Luftig M., Chase M. R., Weicksel S., Cahir-McFarland E., Illanes D., Sarracino D., Kieff E. 2004; Proteins of purified Epstein–Barr virus. Proc Natl Acad Sci U S A 101:16286–16291 [CrossRef]
    [Google Scholar]
  20. Kato K., Yokoyama A., Tohya Y., Akashi H., Nishiyama Y., Kawaguchi Y. 2003; Identification of protein kinases responsible for phosphorylation of Epstein–Barr virus nuclear antigen leader protein at serine-35, which regulates its coactivator function. J Gen Virol 84:3381–3392 [CrossRef]
    [Google Scholar]
  21. Kawaguchi Y., Kato K., Tanaka M., Kanamori M., Nishiyama Y., Yamanashi Y. 2003; Conserved protein kinases encoded by herpesviruses and cellular protein kinase cdc2 target the same phosphorylation site in eukaryotic elongation factor 1delta. J Virol 77:2359–2368 [CrossRef]
    [Google Scholar]
  22. Kieff E., Rickinson A. B. 2001; Epstein–Barr virus and its replication. In Fields Virology , 4th edn. pp  2511–2573 Edited by Knipe D. M., Howley P. M. Philadelphia: Lippincott Williams & Wilkins;
    [Google Scholar]
  23. Kiehl A., Dorsky D. I. 1991; Cooperation of EBV DNA polymerase and EA-D(BMRF1) in vitro and colocalization in nuclei of infected cells. Virology 184:330–340 [CrossRef]
    [Google Scholar]
  24. Kolaskar A. S., Tongaonkar P. C. 1990; A semi-empirical method for prediction of antigenic determinants on protein antigens. FEBS Lett 276:172–174 [CrossRef]
    [Google Scholar]
  25. Krajewski S., Tanaka S., Takayama S., Schibler M. J., Fenton W., Reed J. C. 1993; Investigation of the subcellular distribution of the bcl-2 oncoprotein: residence in the nuclear envelope, endoplasmic reticulum, and outer mitochondrial membranes. Cancer Res 53:4701–4714
    [Google Scholar]
  26. Krosky P. M., Baek M. C., Coen D. M. 2003; The human cytomegalovirus UL97 protein kinase, an antiviral drug target, is required at the stage of nuclear egress. J Virol 77:905–914 [CrossRef]
    [Google Scholar]
  27. Liao G., Wu F. Y., Hayward S. D. 2001; Interaction with the Epstein–Barr virus helicase targets Zta to DNA replication compartments. J Virol 75:8792–8802 [CrossRef]
    [Google Scholar]
  28. Lin C. T., Chan W. Y., Chen W., Huang H. M., Wu H. C., Hsu M. M., Chuang S. M., Wang C. C. 1993; Characterization of seven newly established nasopharyngeal carcinoma cell lines. Lab Invest 68:716–727
    [Google Scholar]
  29. Marschall M., Marzi A., Aus dem Siepen P., Jochmann R., Kalmer M., Auerochs S., Lischka P., Leis M., Stamminger T. 2005; Cellular p32 recruits cytomegalovirus kinase pUL97 to redistribute the nuclear lamina. JBiol Chem (Jun 23; Epub ahead of print) doi: 10.1074/jbc.M502672200
    [Google Scholar]
  30. Michel D., Mertens T. 2004; The UL97 protein kinase of human cytomegalovirus and homologues in other herpesviruses: impact on virus and host. Biochim Biophys Acta 1697169–180 [CrossRef]
    [Google Scholar]
  31. 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]
  32. Pulvertaft R. J. 1964; Phytohaemagglutinin in relation to Burkitt's tumour (African lymphoma). Lancet 14:552–554
    [Google Scholar]
  33. Purves F. C., Roizman B. 1992; The UL13 gene of herpes simplex virus 1 encodes the functions for posttranslational processing associated with phosphorylation of the regulatory protein α 22. Proc Natl Acad Sci U S A 89:7310–7314 [CrossRef]
    [Google Scholar]
  34. Purves F. C., Ogle W. O., Roizman B. 1993; Processing of the herpes simplex virus regulatory protein α 22 mediated by the UL13 protein kinase determines the accumulation of a subset of α and γ mRNAs and proteins in infected cells. Proc Natl Acad Sci U S A 90:6701–6705 [CrossRef]
    [Google Scholar]
  35. Takada K., Horinouchi K., Ono Y., Aya T., Osato T., Takahashi M., Hayasaka S. 1991; An Epstein–Barr virus-producer line Akata: establishment of the cell line and analysis of viral DNA. Virus Genes 5:147–156 [CrossRef]
    [Google Scholar]
  36. Tsai C. H., Glaser R. 1991; A comparison of Epstein–Barr virus specific proteins expressed by three Epstein–Barr virus isolates using specific monoclonal antibodies. Intervirology 32:376–382
    [Google Scholar]
  37. Tsai C. H., Williams M. V., Glaser R. 1991; Characterization of two monoclonal antibodies to Epstein–Barr virus diffuse early antigen which react to two different epitopes and have different biological function. J Virol Methods 33:47–52
    [Google Scholar]
  38. Tsai C. H., Liu M. T., Chen M. R., Lu J., Yang H. L., Chen J. Y., Yang C. S. 1997; Characterization of monoclonal antibodies to the Zta and DNase proteins of Epstein–Barr virus. J Biomed Sci 4:69–77 [CrossRef]
    [Google Scholar]
  39. Tsurumi T. 1993; Purification and characterization of the DNA-binding activity of the Epstein–Barr virus DNA polymerase accessory protein BMRF1 gene products, as expressed in insect cells by using the baculovirus system. J Virol 67:1681–1687
    [Google Scholar]
  40. van Zeijl M., Fairhurst J., Baum E. Z., Sun L., Jones T. R. 1997; The human cytomegalovirus UL97 protein is phosphorylated and a component of virions. Virology 231:72–80 [CrossRef]
    [Google Scholar]
  41. Walther R. F., Atlas E., Carrigan A. 7 other authors 2005; A serine/threonine-rich motif is one of three nuclear localization signals that determine unidirectional transport of the mineralocorticoid receptor to the nucleus. J Biol Chem 280:17549–17561 [CrossRef]
    [Google Scholar]
  42. Westphal E. M., Mauser A., Swenson J., Davis M. G., Talarico C. L., Kenney S. C. 1999; Induction of lytic Epstein–Barr virus (EBV) infection in EBV-associated malignancies using adenovirus vectors in vitro and in vivo. Cancer Res 59:1485–1491
    [Google Scholar]
  43. Wolf D. G., Honigman A., Lazarovits J., Tavor E., Panet A. 1998; Characterization of the human cytomegalovirus UL97 gene product as a virion-associated protein kinase. Arch Virol 143:1223–1232 [CrossRef]
    [Google Scholar]
  44. Wolf D. G., Courcelle C. T., Prichard M. N., Mocarski E. S. 2001; Distinct and separate roles for herpesvirus-conserved UL97 kinase in cytomegalovirus DNA synthesis and encapsidation. Proc Natl Acad Sci U S A 98:1895–1900 [CrossRef]
    [Google Scholar]
  45. Yue W., Gershburg E., Pagano J. S. 2005; Hyperphosphorylation of EBNA2 by Epstein–Barr virus protein kinase suppresses transactivation of the LMP1 promoter. J Virol 79:5880–5885 [CrossRef]
    [Google Scholar]
  46. Zhang Q., Holley-Guthrie E., Ge J. Q., Dorsky D., Kenney S. 1997; The Epstein–Barr virus (EBV) DNA polymerase accessory protein, BMRF1, activates the essential downstream component of the EBV oriLyt. Virology 230:22–34 [CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.81313-0
Loading
/content/journal/jgv/10.1099/vir.0.81313-0
Loading

Data & Media loading...

Most cited Most Cited RSS feed