The Journal of Microbiology

, Volume 46, Issue 4, pp 429–435 | Cite as

Promoter analysis of Bombyx mori nucleopolyhedrovirus Ubiquitin gene

  • Xu’ai Lin
  • Yin Chen
  • Yongzhu Yi
  • Jie Yan
  • Zhifang Zhang


The aim of this study was to analyze the characteristics of Bombyx mori nucleopolyhedrovirus (BmNPV) ubiquitin gene promoter and the effects of conserved motifs, such as TAAG, TATA, and CAAT, along with baculovirus enhancer homologous region 3 (hr3), on promoter activity. Ubiquitin gene of BmNPV was expressed during the late phase of virus infection. In the presence of viral factors, significant reduction of promoter activity was observed by deletion of −382 to −124 bp upstream of ATG. The fragment between −187 and −383 bp upstream of ATG, including distal TAAG, CAAT motif, and TATA box, could also drive expression of the reporter gene. The mutation of cis-elements TATA boxes and TAAG motifs significantly decreased the promoter’s activity, while CAAT mutations enhanced promoter activity by 2-or 3-fold, as compared with the native promoter. In the presence of BmNPV, hr3, both located downstream of the reporter gene of the same vector, and separate vector, could significantly enhance transcription activity of ubiquitin promoter as compared to the control. We concluded that BmNPV ubiquitin gene might be regulated by dual sets of promoter elements, where TAAG and TATA box may positively regulate the expression of ubiquitin, while CAAT motif functions as a negative regulator. Viral factor(s) play an important role in the co-activation of hr3 and promoter.


baculovirus ubiquitin promoter transient expression mutation enhancer hr3 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Blissard, G.W. and G.F. Rohrmann. 1989. Location, sequence, transcriptional mapping, and temporal expression of the gp64 envelope glycoprotein gene of the Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus. Virology 170, 537–555.PubMedCrossRefGoogle Scholar
  2. Blissard, G.W. and G.F. Rohrmann. 1990. Baculovirus diversity and molecular biology. Annu. Rev. Entomol. 35, 127–155.PubMedCrossRefGoogle Scholar
  3. Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.PubMedCrossRefGoogle Scholar
  4. Brown, T. and K. Mackey. 1997. Analysis of RNA by Northern and slot blot hybridization, p. 491. In F.M. Ausubel, R. Brent, R.E. Kingston, D.D. Moore, J.G. Seidman, J.A. Smith, and K. Struhl (eds.), Current protocols in molecular biology, vol. 1. John Wiley, New York, N.Y., USA.Google Scholar
  5. Garcia-Maruniak, A., J. Maruniak, P. Zanotto, A. Doumbouya, J. Liu, T. Merritt, and J. Lanoie. 2004. Sequence analysis of the genome of the Neodiprion sertifer nucleopolyhedrovirus. J. Virol. 78, 7036–7051.PubMedCrossRefGoogle Scholar
  6. Guarino, L.A. 1990. Identification of a viral gene encoding a ubiquitin-like protein. Proc. Natl. Acad. Sci. USA 87, 409–413.PubMedCrossRefGoogle Scholar
  7. Guarino, L.A. and M.D. Summers. 1986. Interspersed homologous DNA of Autographa californica nuclear polyhedrosis virus enhances delayed-early gene expression. J. Virol. 60, 215–223.PubMedGoogle Scholar
  8. Haas, A.L., D.J. Katzung, P.M. Reback, and L.A. Guarino. 1996. Functional characterization of the ubiquitin variant encoded by the baculovirus Autographa californica. Biochemistry 35, 5385–5394.PubMedCrossRefGoogle Scholar
  9. Kumar, M., S. Shanker, V.P. Rawat, and R.H. Das. 2001. Characterization of the antiapoptotic (p35) gene homologue of Spodoptera litura nucleopolyhedrosis virus (SlNPV). Mol. Biol. Rep. 28, 167–173.PubMedCrossRefGoogle Scholar
  10. Lauzon, H., C. Lucarotti, P. Krell, Q. Feng, A. Retnakaran, and B. Arif. 2004. Sequence and organization of the Neodiprion lecontei nucleopolyhedrovirus genome. J. Virol. 78, 7023–7035.PubMedCrossRefGoogle Scholar
  11. Lei, X.D., Y.D. Mi, X.F. Wu, Z.P. Li, and Z.Y. Yuan. 1993. Expression of luciferase gene in insect cells. Acta Biochim. Biophys. Sin. 25, 65–69.Google Scholar
  12. Lin, X.A., Y. Chen, W.H. Xu, Y.Z. Yi, and Z.F. Zhang. 2007. Characterization of Autographa californica nucleopolyhedrovirus ubiquitin gene promoter. Zeitschrift für Naturforschung C. Article in press.Google Scholar
  13. Lu, A. and E.B. Carstens. 1993. Immediate-early baculovirus genes transactivate the p143 gene promoter of Autographa californica nuclear polyhedrosis virus. Virology 195, 710–718.PubMedCrossRefGoogle Scholar
  14. Lu, A., P.J. Krell, J.M. Vlak, and G.F. Rohrmann. 1997. Regulation of baculovirus late and very late expression, p. 193–211. In L.K. Miller (ed.), The Baculoviruses. Plenum Press, New York, N.Y., USA.Google Scholar
  15. Mans, R.M. and D. Knebel-Morsdorf. 1999. Mitochondrial DNA acts as potential promoter of the baculovirus RNA polymerase. Biol. Chem. 380, 579–583.PubMedCrossRefGoogle Scholar
  16. Morris, T.D. and L.K. Miller. 1994. Mutational analysis of a baculovirus major late promoter. Gene 140, 147–153.PubMedCrossRefGoogle Scholar
  17. Okano, K., A. Vanarsdall, V. Mikhailov, and G. Rohrmann. 2006. Conserved molecular systems of the baculoviridae. Virology 344, 77–87.PubMedCrossRefGoogle Scholar
  18. Ooi, B.G., C. Rankin, and L.K. Miller. 1989. Downstream sequences augment transcription from the essential initiation site of a baculovirus polyhedrin gene. J. Mol. Biol. 210, 721–736.PubMedCrossRefGoogle Scholar
  19. Reilly, L.M. and L.A. Guarino. 1996. The viral ubiquitin gene of Autographa californica nuclear polyhedrosis virus is not essential for viral replication. Virology 218, 243–247.PubMedCrossRefGoogle Scholar
  20. Summers, M.D. and G.E. Smith. 1987. A manual of methods for baculovirus vectors and insect cell culture procedures. Texas Agric. Exp. Stat. Bull, No.1555.Google Scholar
  21. Thiem, S.M. and L.K. Miller. 1990. Differential gene expression mediated by late, very late and hybrid baculovirus promoters. Gene 91, 87–94.PubMedCrossRefGoogle Scholar
  22. Todd, J.W., A.L. Passarelli, and L.K. Miller. 1995. Eighteen baculovirus genes, including lef-11, p35, 39K, and p47, support late gene expression. J. Virol. 69, 968–974.PubMedGoogle Scholar
  23. Xiao, Q.L., Z.F. Zhang, Y.Z. Yi, J.L. He, and X.F. Wu. 2001. Functional analysis of helicase gene promoter and homologus region 3 enhancer in Bombyx mori nuclear polyhedrosis virus. Acta Biochim. Biophys. Sin. 33, 525–530.PubMedGoogle Scholar
  24. Yao, J., S.S. Chen, K.G. Yang, X. Di, A.Q. Xiong, and Y.H. Zhang. 2002. The transcriptional regulation study on human delta globin gene with CAAT box C→T point mutation in its promoter. Zhongguo Yi Xue Ke Xue Yuan Xue Bao. 24, 140–143.PubMedGoogle Scholar
  25. Zhang, Z.F., Y.Z. Zhang, H.S. Lü, and X.F. Wu. 1995. Structure and function of DNA replication origin hr3 of Bombyx mori nuclear polyhedrosis virus. Sci. China Ser. B 25, 949–955.Google Scholar

Copyright information

© The Microbiological Society of Korea and Springer-Verlag Berlin Heidelber GmbH 2008

Authors and Affiliations

  • Xu’ai Lin
    • 1
    • 2
  • Yin Chen
    • 2
    • 3
  • Yongzhu Yi
    • 2
  • Jie Yan
    • 1
  • Zhifang Zhang
    • 2
  1. 1.Department of Medical Microbiology and Parasitology, School of MedicineZhejiang UniversityHangzhouP. R. China
  2. 2.Biotechnology Research Institute, National Key Facility for Crop Gene Resources and Genetic ImprovementChinese Academy of Agricultural SciencesBeijingP. R. China
  3. 3.Zhejiang Provincial Center for Disease Control and PreventionHangzhouP. R. China

Personalised recommendations