1887

Abstract

Repeated baculovirus infections in cultured insect cells lead to the generation of defective interfering viruses (DIs), which accumulate at the expense of the intact helper virus and compromise heterologous protein expression. In particular, multicapsid nucleopolyhedovirus (AcMNPV) DIs are enriched in an origin of viral DNA replication () not associated with the homologous regions (s). This non- is located within the coding sequence of the non-essential gene. We investigated the effect of a deletion of the AcMNPV non- on the heterologous protein expression levels following serial passage in Sf21 insect cells. Using homologous ET recombination in , deletions within the gene were made in a bacterial artificial chromosome (BAC) containing the entire AcMNPV genome (bacmid). All bacmids were equipped with an expression cassette containing the green fluorescent protein gene and a gene encoding the classical swine fever virus E2 glycoprotein (CSFV-E2). For the parental (intact) bacmid only, a strong accumulation of DIs with reiterated non-s was observed. This was not observed for the mutants, indicating that removal of the non- enhanced the genetic stability of the viral genome upon passaging. However, for all passaged viruses it was found that the entire BAC vector including the expression cassette was spontaneously deleted from the viral genome, leading to a rapid decrease in GFP and CSFV-E2 production. The rationale for the (intrinsic) genetic instability of the BAC vector in insect cells and the implications with respect to large-scale production of proteins with bacmid-derived baculoviruses are discussed.

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2003-10-01
2019-10-18
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References

  1. Adler, H., Messerle, M. & Koszinowski, U. H. ( 2001; ). Virus reconstituted from infectious bacterial artificial chromosome (BAC)-cloned murine gammaherpesvirus 68 acquires wild-type properties in vivo only after excision of BAC vector sequences. J Virol 75, 5692–5696.[CrossRef]
    [Google Scholar]
  2. Altschul, S. F., Madden, T. L., Schaffer, A. A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D. J. ( 1997; ). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25, 3389–3402.[CrossRef]
    [Google Scholar]
  3. Ayres, M. D., Howard, S. C., Kuzio, J., Lopez Ferber, M. & Possee, R. D. ( 1994; ). The complete DNA sequence of Autographa californica nuclear polyhedrosis virus. Virology 202, 586–605.[CrossRef]
    [Google Scholar]
  4. Bett, A. J., Prevec, L. & Graham, F. L. ( 1993; ). Packaging capacity and stability of human adenovirus type 5 vectors. J Virol 67, 5911–5921.
    [Google Scholar]
  5. Bigot, Y., Stasiak, K., Rouleux-Bonnin, F. & Federici, B. A. ( 2000; ). Characterization of repetitive DNA regions and methylated DNA in ascovirus genomes. J Gen Virol 81, 3073–3082.
    [Google Scholar]
  6. Bloss, T. A. & Sugden, B. ( 1994; ). Optimal lengths for DNAs encapsidated by Epstein–Barr virus. J Virol 68, 8217–8222.
    [Google Scholar]
  7. Brune, W., Messerle, M. & Koszinowski, U. H. ( 2000; ). Forward with BACs – new tools for herpesvirus genomics. Trends Genet 16, 254–259.[CrossRef]
    [Google Scholar]
  8. Clem, R. J., Robson, M. & Miller, L. K. ( 1994; ). Influence of infection route on the infectivity of baculovirus mutants lacking the apoptosis-inhibiting gene p35 and the adjacent gene p94. J Virol 68, 6759–6762.
    [Google Scholar]
  9. Dai, X. J., Hajós, J. P., Joosten, N. N., van Oers, M. M., IJkel, W. F. J., Zuidema, D., Pang, Y. & Vlak, J. M. ( 2000; ). Isolation of a Spodoptera exigua baculovirus recombinant with a 10·6 kbp genome deletion that retains biological activity. J Gen Virol 81, 2545–2554.
    [Google Scholar]
  10. Davis, S. J. & Vierstra, R. D. ( 1998; ). Soluble, highly fluorescent variants of green fluorescent protein (GFP) for use in higher plants. Plant Mol Biol 36, 521–528.[CrossRef]
    [Google Scholar]
  11. Friesen, P. D. ( 1997; ). Regulation of baculovirus early gene expression. In The Baculoviruses, pp. 141–170. Edited by L. K. Miller. New York: Plenum Press.
  12. Friesen, P. D. & Miller, L. K. ( 1987; ). Divergent transcription of early 35- and 94-kilodalton protein genes encoded by the HindIII K genome fragment of the baculovirus Autographa californica nuclear polyhedrosis virus. J Virol 61, 2264–2272.
    [Google Scholar]
  13. Friesen, P. D. & Nissen, M. S. ( 1990; ). Gene organization and transcription of TED, a lepidopteran retrotransposon integrated within the baculovirus genome. Mol Cell Biol 10, 3067–3077.
    [Google Scholar]
  14. Hayakawa, T., Ko, R., Okano, K., Seong, S. I., Goto, C. & Maeda, S. ( 1999; ). Sequence analysis of the Xestia c-nigrum granulovirus genome. Virology 262, 277–297.[CrossRef]
    [Google Scholar]
  15. Heldens, J. G., van Strien, E. A., Feldmann, A. M., Kulcsar, P., Muñoz, D., Leisy, D. J., Zuidema, D., Goldbach, R. W. & Vlak, J. M. ( 1996; ). Spodoptera exigua multicapsid nucleopolyhedrovirus deletion mutants generated in cell culture lack virulence in vivo. J Gen Virol 77, 3127–3134.[CrossRef]
    [Google Scholar]
  16. Heldens, J. G., Broer, R., Zuidema, D., Goldbach, R. W. & Vlak, J. M. ( 1997; ). Identification and functional analysis of a non-hr origin of DNA replication in the genome of Spodoptera exigua multicapsid nucleopolyhedrovirus. J Gen Virol 78, 1497–1506.
    [Google Scholar]
  17. Hu, Z. H., Arif, B. M., Sun, J. S., Chen, X. W., Zuidema, D., Goldbach, R. W. & Vlak, J. M. ( 1998; ). Genetic organization of the HindIII-I region of the single-nucleocapsid nucleopolyhedrovirus of Buzura suppressaria. Virus Res 55, 71–82.[CrossRef]
    [Google Scholar]
  18. Huang, J. & Levin, D. B. ( 1999; ). Identification and functional analysis of a putative non-hr origin of DNA replication from the Spodoptera littoralis type B multinucleocapsid nucleopolyhedrovirus. J Gen Virol 80, 2263–2274.
    [Google Scholar]
  19. IJkel, W. F., van Strien, E. A., Heldens, J. G., Broer, R., Zuidema, D., Goldbach, R. W. & Vlak, J. M. ( 1999; ). Sequence and organization of the Spodoptera exigua multicapsid nucleopolyhedrovirus genome. J Gen Virol 80, 3289–3304.
    [Google Scholar]
  20. Jehle, J. A. ( 2002; ). The expansion of a hypervariable, non-hr ori-like region in the genome of Cryptophlebia leucotreta granulovirus provides in vivo evidence for the utilization of baculovirus non-hr oris during replication. J Gen Virol 83, 2025–2034.
    [Google Scholar]
  21. Kamita, S. G., Majima, K. & Maeda, S. ( 1993; ). Identification and characterization of the p35 gene of Bombyx mori nuclear polyhedrosis virus that prevents virus-induced apoptosis. J Virol 67, 455–463.
    [Google Scholar]
  22. King, L. A. & Possee, R. D. ( 1992; ). The baculovirus expression system. Dordrecht: Kluwer.
  23. Kitts, P. A. ( 1996; ). Construction of baculovirus recombinants. Cytotechnology 20, 111–123.[CrossRef]
    [Google Scholar]
  24. Kool, M., Voncken, J. W., van Lier, F. L., Tramper, J. & Vlak, J. M. ( 1991; ). Detection and analysis of Autographa californica nuclear polyhedrosis virus mutants with defective interfering properties. Virology 183, 739–746.[CrossRef]
    [Google Scholar]
  25. Kool, M., van den Berg, P. M., Tramper, J., Goldbach, R. W. & Vlak, J. M. ( 1993; ). Location of two putative origins of DNA replication of Autographa californica nuclear polyhedrosis virus. Virology 192, 94–101.[CrossRef]
    [Google Scholar]
  26. Kool, M., Goldbach, R. W. & Vlak, J. M. ( 1994; ). A putative non-hr origin of DNA replication in the HindIII-K fragment of Autographa californica multiple nucleocapsid nuclear polyhedrosis virus. J Gen Virol 75, 3345–3352.[CrossRef]
    [Google Scholar]
  27. Krell, P. J. ( 1996; ). Passage effect of virus infection in insect cells. Cytotechnology 20, 125–137.[CrossRef]
    [Google Scholar]
  28. Kumar, S. & Miller, L. K. ( 1987; ). Effects of serial passage of Autographa californica nuclear polyhedrosis virus in cell culture. Virus Res 7, 335–349.[CrossRef]
    [Google Scholar]
  29. Lee, H. Y. & Krell, P. J. ( 1992; ). Generation and analysis of defective genomes of Autographa californica nuclear polyhedrosis virus. J Virol 66, 4339–4347.
    [Google Scholar]
  30. Lee, H. & Krell, P. J. ( 1994; ). Reiterated DNA fragments in defective genomes of Autographa californica nuclear polyhedrosis virus are competent for AcMNPV-dependent DNA replication. Virology 202, 418–429.[CrossRef]
    [Google Scholar]
  31. Lu, A., Krell, P. J., Vlak, J. M. & Rohrmann, G. F. ( 1997; ). Baculovirus DNA replication. In The Baculoviruses, pp. 171–216. Edited by L. K. Miller. New York: Plenum Press.
  32. Luckow, V. A., Lee, S. C., Barry, G. F. & Olins, P. O. ( 1993; ). Efficient generation of infectious recombinant baculoviruses by site-specific transposon-mediated insertion of foreign genes into a baculovirus genome propagated in Escherichia coli. J Virol 67, 4566–4579.
    [Google Scholar]
  33. Luque, T., Finch, R., Crook, N., O'Reilly, D. R. & Winstanley, D. ( 2001; ). The complete sequence of the Cydia pomonella granulovirus genome. J Gen Virol 82, 2531–2547.
    [Google Scholar]
  34. Muyrers, J. P., Zhang, Y., Testa, G. & Stewart, A. F. ( 1999; ). Rapid modification of bacterial artificial chromosomes by ET-recombination. Nucleic Acids Res 27, 1555–1557.[CrossRef]
    [Google Scholar]
  35. Pearson, M. N., Bjornson, R. M., Ahrens, C. & Rohrmann, G. F. ( 1993; ). Identification and characterization of a putative origin of DNA replication in the genome of a baculovirus pathogenic for Orgyia pseudotsugata. Virology 197, 715–725.[CrossRef]
    [Google Scholar]
  36. Pijlman, G. P., van den Born, E., Martens, D. E. & Vlak, J. M. ( 2001; ). Autographa californica baculoviruses with large genomic deletions are rapidly generated in infected insect cells. Virology 283, 132–138.[CrossRef]
    [Google Scholar]
  37. Pijlman, G. P., Dortmans, J. C. F. M., Vermeesch, A. M. G., Yang, K., Martens, D. E., Goldbach, R. W. & Vlak, J. M. ( 2002; ). Pivotal role of the non-hr origin of DNA replication in the genesis of defective interfering baculoviruses. J Virol 76, 5605–5611.[CrossRef]
    [Google Scholar]
  38. Possee, R. D. ( 1997; ). Baculoviruses as expression vectors. Curr Opin Biotechnol 8, 569–572.[CrossRef]
    [Google Scholar]
  39. Roosien, J., Usmany, M., Klinge-Roode, E. C., Meijerink, P. H. S. & Vlak, J. M. ( 1986; ). Heterologous recombination between baculoviruses. In Fundamental and Applied Aspects of Invertebrate Pathology, pp. 389–392. Edited by R. A. Samson, J. M. Vlak & D. Peters. ICIP86 Foundation.
  40. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  41. Shizuya, H., Birren, B., Kim, U. J., Mancino, V., Slepak, T., Tachiiri, Y. & Simon, M. ( 1992; ). Cloning and stable maintenance of 300-kilobase-pair fragments of human DNA in Escherichia coli using an F-factor-based vector. Proc Natl Acad Sci U S A 89, 8794–8797.[CrossRef]
    [Google Scholar]
  42. Smith, G. A. & Enquist, L. W. ( 1999; ). Construction and transposon mutagenesis in Escherichia coli of a full-length infectious clone of pseudorabies virus, an alphaherpesvirus. J Virol 73, 6405–6414.
    [Google Scholar]
  43. Smith, G. A. & Enquist, L. W. ( 2000; ). A self-recombining bacterial artificial chromosome and its application for analysis of herpesvirus pathogenesis. Proc Natl Acad Sci U S A 97, 4873–4878.[CrossRef]
    [Google Scholar]
  44. Summers, M. D. & Smith, G. E. ( 1987; ). A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures. Texas Agricultural Experiment Station Bulletin No. 1555.
  45. Van Hulten, M. C. W., Witteveldt, J., Peters, S., Kloosterboer, N., Tarchini, R., Fiers, M., Sandbrink, H., Lankhorst, R. K. & Vlak, J. M. ( 2001; ). The white spot syndrome virus DNA genome sequence. Virology 286, 7–22.[CrossRef]
    [Google Scholar]
  46. Van Oers, M. M., Thomas, A. A. M., Moormann, R. J. M. & Vlak, J. M. ( 2001; ). Secretory pathway limits the enhanced expression of classical swine fever virus E2 glycoprotein in insect cells. J Biotechnol 86, 31–38.[CrossRef]
    [Google Scholar]
  47. Van Rijn, P. A., van Gennip, H. G. & Moormann, R. J. ( 1999; ). An experimental marker vaccine and accompanying serological diagnostic test both based on envelope glycoprotein E2 of classical swine fever virus (CSFV). Vaccine 17, 433–440.[CrossRef]
    [Google Scholar]
  48. Vaughn, J. L., Goodwin, R. H., Tompkins, G. J. & McCawley, P. ( 1977; ). The establishment of two cell lines from the insect Spodoptera frugiperda (Lepidoptera; Noctuidae). In Vitro 13, 213–217.[CrossRef]
    [Google Scholar]
  49. Vialard, J. E., Arif, B. M. & Richardson, C. D. ( 1995; ). Introduction to the molecular biology of baculoviruses. Methods Mol Biol 39, 1–24.
    [Google Scholar]
  50. Vlak, J. M. ( 1979; ). The proteins of nonoccluded Autographa californica nuclear polyhedrosis virus produced in an established cell line of Spodoptera frugiperda. J Invertebr 34, 110–118.[CrossRef]
    [Google Scholar]
  51. Wagner, M., Jonjic, S., Koszinowski, U. H. & Messerle, M. ( 1999; ). Systematic excision of vector sequences from the BAC-cloned herpesvirus genome during virus reconstitution. J Virol 73, 7056–7060.
    [Google Scholar]
  52. Wickham, T. J., Davis, T., Granados, R. R., Hammer, D. A., Shuler, M. L. & Wood, H. A. ( 1991; ). Baculovirus defective interfering particles are responsible for variations in recombinant protein production as a function of multiplicity of infection. Biotechnol Lett 13, 483–488.[CrossRef]
    [Google Scholar]
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