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Encapsulation of DNA and non-viral protein changes the structure of murine polyomavirus virus-like particles

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Abstract

Asymmetrical-flow field flow fractionation with multiple-angle light scattering (AFFFF-MALS) was, for the first time, used to characterize the size of murine polyomavirus virus-like particles (MPV VLPs) packaged with either insect cell genomic DNA or non-viral protein. Encapsidation of both genomic DNA and non-viral protein were found to cause a contraction in VLP radii of gyration by approximately 1 nm. Non-viral protein packaged into VLPs consisted of a series of glutathione-S-transferase, His and S tags attached to the N-terminal end of the MPV structural protein VP2 (M r = 67108). Transmission electron microscopy analysis of MPV VLPs packaging non-viral protein suggested that VLPs grew in diameter by approximately 5 nm, highlighting the differences between this invasive technique and the relatively non-invasive AFFFF-MALS technique. Encapsulation of non-viral protein into MPV VLPs was found to prevent co-encapsidation of genomic DNA. Further investigation into why this occurred led to the discovery that encapsulation of non-viral protein alters the nuclear localization of MPV VLPs during in vivo assembly. VLPs were relocated away from the ring zone and the nuclear membrane towards the centre of the nucleus amongst the virogenic stroma. The change in nuclear localization away from the site where VLP assembly usually occurs is a likely reason why encapsidation of genomic DNA did not take place.

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Abbreviations

AcMNPV:

Autographa californica multiple nucleopolyhedrovirus

AFFFF:

Asymmetrical-flow field flow fractionation

ELISA:

Enzyme-linked immunosorbent assay

GST:

Glutathione-S-transferase

His:

Polyhistidine tag

MALS:

Multiple-angle light scattering

MOI:

Multiplicity of infection

MPV:

Murine polyomavirus

p.i.:

Post-infection

RI:

Refractive index

S:

S Tag (EMD Biosciences, Inc., Madison, WI, USA)

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

TAG:

Sequence of glutathione-S-transferase, His and S tags from pET-41a(+) vector (EMD Biosciences, Inc.)

TEM:

Transmission electron microscopy

VLP:

Virus-like particle

References

  1. Abbing A, Blaschke UK, Grein S, Kretschmar M, Stark CM, Thies MJ, Walter J, Weigand M, Woith DC, Hess J, Reiser CO (2004) Efficient intracellular delivery of a protein and a low molecular weight substance via recombinant polyomavirus-like particles. J Biol Chem 279:27410–27421

    Article  PubMed  CAS  Google Scholar 

  2. An K, Gillock ET, Sweat JA, Reeves WM, Consigli RA (1999) Use of the baculovirus system to assemble polyomavirus capsid-like particles with different polyomavirus structural proteins: analysis of the recombinant assembled capsid-like particles. J Gen Virol 80:1009–1016

    PubMed  CAS  Google Scholar 

  3. Baker TS, Caspar DLD, Murakami WT (1983) Polyoma-virus hexamer tubes consist of paired pentamers. Nature 303:446–448

    Article  PubMed  CAS  Google Scholar 

  4. Barouch DH, Harrison SC (1994) Interactions among the major and minor coat proteins of polyomavirus. J Virol 68:3982–3989

    PubMed  CAS  Google Scholar 

  5. Bellomo G, Vairetti M, Stivala L, Mirabelli F, Richelmi P, Orrenius S (1992) Demonstration of nuclear compartmentalization of glutathione in hepatocytes. PNAS 89:4412–4416

    Article  PubMed  CAS  Google Scholar 

  6. Bellomo G, Palladini G, Vairetti M (1997) Intranuclear distribution, function and fate of glutathione and glutathione-S-conjugate in living rat hepatocytes studied by fluorescence microscopy. Microsc Res Tech 36:243–252

    Article  PubMed  CAS  Google Scholar 

  7. Belnap DM, Olson NH, Cladel NM, Newcomb WW, Brown JC, Kreider JW, Christensen ND, Baker TS (1996) Conserved features in papillomavirus and polyomavirus capsids. J Mol Biol 259:249–263

    Article  PubMed  CAS  Google Scholar 

  8. Bennett CF, Spector DL, Yeoman LC (1986) Nonhistone protein BA is a glutathione-S-transferase localized to interchromatinic regions of the cell-nucleus. J Cell Biol 102:600–609

    Article  PubMed  CAS  Google Scholar 

  9. Boura E, Liebl D, Spisek R, Fric J, Marek M, Stokrova J, Holan V, Forstova J (2005) Polyomavirus EGFP-pseudocapsids: analysis of model particles for introduction of proteins and peptides into mammalian cells. FEBS Lett 579:6549–6558

    Article  PubMed  CAS  Google Scholar 

  10. Bousse L, Mouradian S, Minalla A, Yee H, Williams K, Dubrow R (2001) Protein sizing on a microchip. Anal Chem 73:1207–1212

    Article  PubMed  CAS  Google Scholar 

  11. Cavaldesi M, Caruso M, Sthandier O, Amati P, Garcia MI (2004) Conformational changes of murine polyomavirus capsid proteins induced by sialic acid binding. J Biol Chem 279:41573–41579

    Article  PubMed  CAS  Google Scholar 

  12. Charpilienne A, Nejmeddine M, Berois M, Parez N, Neumann E, Hewat E, Trugnan G, Cohen J (2001) Individual rotavirus-like particles containing 120 molecules of fluorescent protein are visible in living cells. J Biol Chem 276:29361–29367

    Article  PubMed  CAS  Google Scholar 

  13. Chen XS, Stehle T, Harrison SC (1998) Interaction of polyomavirus internal protein VP2 with the major capsid protein VP1 and implications for participation of VP2 in viral entry. EMBO J 17:3233–3240

    Article  PubMed  CAS  Google Scholar 

  14. Chuan YP, Fan YY, Lua LHL, Middelberg APJ (2008) Quantitative analysis of virus-like particle size and distribution by field-flow fractionation. Biotechnol Bioeng 99:1425–1433

    Article  PubMed  CAS  Google Scholar 

  15. Citkowicz A, Petry H, Harkins RN, Ast O, Cashion L, Goldmann C, Bringmann P, Plummer K, Larsen BR (2008) Characterization of virus-like particle assembly for DNA delivery using asymmetrical flow field-flow fractionation and light scattering. Anal Biochem 376:163–172

    Article  PubMed  CAS  Google Scholar 

  16. Debye P (1947) Molecular-weight determination by light scattering. J Phys Colloid Chem 51:18–32

    Article  CAS  Google Scholar 

  17. Delos SE, Montross L, Moreland RB, Garcea RL (1993) Expression of the polyomavirus VP2 and VP3 proteins in insect cells: coexpression with the major capsid protein VP1 alters VP2/VP3 subcellular localization. Virology 194:393–398

    Article  PubMed  CAS  Google Scholar 

  18. Everett RD (2001) DNA viruses and viral proteins that interact with PML nuclear bodies. Oncogene 20:7266–7273

    Article  PubMed  CAS  Google Scholar 

  19. Fligge C, Schafer F, Selinka HC, Sapp C, Sapp M (2001) DNA-induced structural changes in the papillomavirus capsid. J Virol 75:7727–7731

    Article  PubMed  CAS  Google Scholar 

  20. Forstova J, Krauzewicz N, Sandig V, Elliott J, Palkova Z, Strauss M, Griffin BE (1995) Polyoma virus pseudocapsids as efficient carriers of heterologous DNA into mammalian cells. Hum Gene Ther 6:297–306

    Article  PubMed  CAS  Google Scholar 

  21. Garcea RL, Gissmann L (2004) Virus-like particles as vaccines and vessels for the delivery of small molecules. Curr Opin Biotechnol 15:513–517

    Article  PubMed  CAS  Google Scholar 

  22. Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins MR, Appel RD, A. B (2005) Protein Identification and Analysis Tools on the ExPASy Server. In: Walker JM (ed) The Proteomics Protocols Handbook. Humana Press, pp 571–607

  23. Gillock ET, Rottinghaus S, Chang D, Cai X, Smiley SA, An K, Consigli RA (1997) Polyomavirus major capsid protein VP1 is capable of packaging cellular DNA when expressed in the baculovirus system. J Virol 71:2857–2865

    PubMed  CAS  Google Scholar 

  24. Gillock ET, An K, Consigli RA (1998) Truncation of the nuclear localization signal of polyomavirus VP1 results in a loss of DNA packaging when expressed in the baculovirus system. Virus Res 58:149–160

    Article  PubMed  CAS  Google Scholar 

  25. Gleiter S, Lilie H (2003) Cell-type specific targeting and gene expression using a variant of polyoma VP1 virus-like particles. Biol Chem 384:247–255

    Article  PubMed  CAS  Google Scholar 

  26. Greenstone HL, Nieland JD, de Visser KE, De Bruijn MLH, Kirnbauer R, Roden RBS, Lowy DR, Kast WM, Schiller JT (1998) Chimeric papillomavirus virus-like particles elicit antitumor immunity against the E7 oncoprotein in an HPV16 tumor model. Proc Natl Acad Sci USA 95:1800–1805

    Article  PubMed  CAS  Google Scholar 

  27. Gunther C, Schmidt U, Rudolph R, Bohm G (2001) Protein and peptide delivery via engineered polyomavirus-like particles. FASEB J. doi:10.1096/fj.1000-0645fje

  28. Hagensee ME, Olson NH, Baker TS, Galloway DA (1994) 3-Dimensional structure of vaccinia virus-produced human papillomavirus type-1 capsids. J Virol 68:4503–4505

    PubMed  CAS  Google Scholar 

  29. Huglin MB (1972) Specific refractive index increments. In: Huglin MB (ed) Light scattering from polymer solutions. Academic Press, London, pp 165–331

    Google Scholar 

  30. Inoue T, Kawano M-a, Takahashi R-u, Tsukamoto H, Enomoto T, Imai T, Kataoka K, Handa H (2007) Engineering of SV40-based nano-capsules for delivery of heterologous proteins as fusions with the minor capsid proteins VP2/3. Journal of Biotechnology: doi:10.1016/j.jbiotec.2007.1012.1006

  31. Ishizu KI, Watanabe H, Han SI, Kanesashi SN, Hoque M, Yajima H, Kataoka K, Handa H (2001) Roles of disulfide linkage and calcium ion-mediated interactions in assembly and disassembly of virus-like particles composed of simian virus 40 VP1 capsid protein. J Virol 75:61–72

    Article  PubMed  CAS  Google Scholar 

  32. Kawana K, Yoshikawa H, Taketani Y, Yoshiike K, Kanda T (1998) In vitro construction of pseudovirions of human papillomavirus type 16: incorporation of plasmid DNA into reassembled L1/L2 capsids. J Virol 72:10298–10300

    PubMed  CAS  Google Scholar 

  33. Krauzewicz N, Stokrova J, Jenkins C, Elliott M, Higgins CF, Griffin BE (2000) Virus-like gene transfer into cells mediated by polyoma virus pseudocapsids. Gene Ther 7:2122–2131

    Article  PubMed  CAS  Google Scholar 

  34. Lua LHL, Reid S (2000) Virus morphogenesis of Helicoverpa armigera nucleopolyhedrovirus in Helicoverpa zea serum-free suspension culture. J Gen Virol 81:2531–2543

    PubMed  CAS  Google Scholar 

  35. Magnuson B, Rainey EK, Benjamin T, Baryshev M, Mkrtchian S, Tsai B (2005) ERp29 triggers a conformational change in polyomavirus to stimulate membrane binding. Mol Cell 20:289–300

    Article  PubMed  CAS  Google Scholar 

  36. Mannova P, Liebl D, Krauzewicz N, Fejtova A, Stokrova J, Palkova Z, Griffin BE, Forstova J (2002) Analysis of mouse polyomavirus mutants with lesions in the minor capsid proteins. J Gen Virol 83:2309–2319

    PubMed  CAS  Google Scholar 

  37. Mcmillen J, Consigli RA (1974) Characterization of polyoma DNA-protein complexes.1. Electrophoretic identification of proteins in a nucleoprotein complex isolated from polyoma-Infected cells. J Virol 14:1326–1336

    PubMed  CAS  Google Scholar 

  38. Montross L, Watkins S, Moreland RB, Mamon H, Caspar DL, Garcea RL (1991) Nuclear assembly of polyomavirus capsids in insect cells expressing the major capsid protein VP1. J Virol 65:4991–4998

    PubMed  CAS  Google Scholar 

  39. Moreland RB, Montross L, Garcea RL (1991) Characterization of the DNA-binding properties of the polyomavirus capsid protein-VP1. J Virol 65:1168–1176

    PubMed  CAS  Google Scholar 

  40. Nagamine T, Kawasaki Y, Matsumoto S (2006) Induction of a subnuclear structure by the simultaneous expression of baculovirus proteins, IE1, LEF3, and P143 in the presence of hr. Virology 352:400–407

    Article  PubMed  CAS  Google Scholar 

  41. Noad R, Roy P (2003) Virus-like particles as immunogens. Trends Microbiol 11:438–444

    Article  PubMed  CAS  Google Scholar 

  42. Ou WC, Wang M, Fung CY, Tsai RT, Chao PC, Hseu TH, Chang D (1999) The major capsid protein, VP1, of human JC virus expressed in Escherichia coli is able to self-assemble into a capsid-like particle and deliver exogenous DNA into human kidney cells. J Gen Virol 80:39–46

    PubMed  CAS  Google Scholar 

  43. Pawlita M, Muller M, Oppenlander M, Zentgraf H, Herrmann M (1996) DNA encapsidation by viruslike particles assembled in insect cells from the major capsid protein VP1 of B-lymphotropic papovavirus. J Virol 70:7517–7526

    PubMed  CAS  Google Scholar 

  44. Rayment I, Baker TS, Caspar DL, Murakami WT (1982) Polyoma virus capsid structure at 22.5 A resolution. Nature 295:110–115

    Article  PubMed  CAS  Google Scholar 

  45. Sambrook J, Russell D (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbour Laboratory Press, New York

    Google Scholar 

  46. Schafer F, Florin L, Sapp M (2002) DNA binding of L1 is required for human papillomavirus morphogenesis in vivo. Virology 295:172–181

    Article  PubMed  CAS  Google Scholar 

  47. Schmidt U, Rudolph R, Bohm G (2000) Mechanism of assembly of recombinant murine polyomavirus-like particles. J Virol 74:1658–1662

    Article  PubMed  CAS  Google Scholar 

  48. Seebeck FP, Woycechowsky KJ, Zhuang W, Rabe JP, Hilvert D (2006) A simple tagging system for protein encapsulation. J Am Chem Soc 128:4516–4517

    Article  PubMed  CAS  Google Scholar 

  49. Shishido-Hara Y, Ichinose S, Higuchi K, Hara Y, Yasui K (2004) Major and minor capsid proteins of human polyomavirus JC cooperatively accumulate to nuclear domain 10 for assembly into virions. J Virol 78:9890–9903

    Article  PubMed  CAS  Google Scholar 

  50. Shortt DW, Roessner D, Wyatt PJ (1996) Absolute measurement of diameter distributions of particles using a multiangle light scattering photometer coupled with flow field-flow fractionation. Am Lab 17:21

    Google Scholar 

  51. Tsukamoto H, M-a Kawano, Inoue T, Enomoto T, R-u Takahashi, Yokoyama N, Yamamoto N, Imai T, Kataoka K, Yamaguchi Y, Handa H (2007) Evidence that SV40 VP1-DNA interactions contribute to the assembly of 40-nm spherical viral particles. Genes Cells 12:1267–1279

    Article  PubMed  CAS  Google Scholar 

  52. Voronkova T, Kazaks A, Ose V, Ozel M, Scherneck S, Pumpens P, Ulrich R (2007) Hamster polyomavirus-derived virus-like particles are able to transfer in vitro encapsidated plasmid DNA to mammalian cells. Virus Genes 34:303–314

    Article  PubMed  CAS  Google Scholar 

  53. Williams GV, Faulkner P (1997) Cytological changes and viral morphogenesis during baculovirus infection. In: Miller LK (ed) The Baculoviruses. Plenum, New York, pp 61–108

    Google Scholar 

  54. Windram O, Weber B, Jaffer M, Rybicki E, Shepherd D, Varsani A (2008) An investigation into the use of human papillomavirus type 16 virus-like particles as a delivery vector system for foreign proteins: N- and C-terminal fusion of GFP to the L1 and L2 capsid proteins. Archives of Virology: DOI 10.1007/s00705-00007-00025-00702

  55. Wyatt PJ, Villalpando DN (1997) High-precision measurement of submicrometer particle size distributions. Langmuir 13:3913–3914

    Article  CAS  Google Scholar 

  56. Yamada T, Iwasaki Y, Tada H, Iwabuki H, Chuah MKL, VandenDriessche T, Fukuda H, Kondo A, Ueda M, Seno M, Tanizawa K, Kuroda S (2003) Nanoparticles for the delivery of genes and drugs to human hepatocytes. Nat Biotechnol 21:885–890

    Article  PubMed  CAS  Google Scholar 

  57. Zimm BH (1948) The scattering of light and the radial distribution function of high polymer solutions. J Chem Phys 16:1093–1099

    Article  CAS  Google Scholar 

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Acknowledgments

L. H. L. L. and A. P. J. M. acknowledge the support of the Australian Research Council (Grants FF0348465 and DP0773111) and the Australian National Health and Medical Research Council (Grant 409976). D. I. L. is a recipient of University of Queensland Graduate School and Center for Biomolecular Engineering scholarships. A. P. J. M. acknowledges support from the Australian Research Council through the award of a Federation Fellowship. Y. P. C. is a recipient of Australian Postgraduate Award and Australian Institute of Bioengineering and Nanotechnology scholarships.

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Authors and Affiliations

  1. Centre for Biomolecular Engineering, Australian Institute of Bioengineering and Nanotechnology, The University of Queensland, Saint Lucia, QLD, 4072, Australia

    D. I. Lipin, Y. P. Chuan, L. H. L. Lua & A. P. J. Middelberg

  2. SRC Protein Expression Facility, Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Saint Lucia, QLD, 4072, Australia

    L. H. L. Lua

  3. Centre for Biomolecular Engineering, School of Engineering, The University of Queensland, Saint Lucia, QLD, 4072, Australia

    D. I. Lipin, Y. P. Chuan & A. P. J. Middelberg

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  1. D. I. Lipin
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Correspondence to A. P. J. Middelberg.

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Lipin, D.I., Chuan, Y.P., Lua, L.H.L. et al. Encapsulation of DNA and non-viral protein changes the structure of murine polyomavirus virus-like particles. Arch Virol 153, 2027–2039 (2008). https://doi.org/10.1007/s00705-008-0220-9

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  • DOI: https://doi.org/10.1007/s00705-008-0220-9

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