Rotavirus protein expression is important for virus assembly and pathogenesis

  • P. Tian
  • J. M. Ball
  • C. Q-Y Zeng
  • M. K. Estes
Conference paper
Part of the Archives of Virology book series (ARCHIVES SUPPL, volume 12)


Rotaviruses have a unique morphogenesis in which particles obtain a transient membrane-envelope as newly made subviral particles bud into the endoplasmic reticulum (ER). This process is mediated by a viral nonstructural glycoprotein, NSP4. We have found that NSP4 has pleiotropic properties that became evident following expression of this protein in eukaryotic cells. NSP4 expressed in insect cells bound double-layered rotavirus particles in a manner similar to receptor-ligand interactions and this interaction is thought to trigger the particle budding process. Expression of NSP4 in insect cells also increases intracellular calcium ([Ca2+]i) levels and this effect may explain the toxicity of this protein in eukaryotic cells. Increases in [Ca2+]i levels in insect cells also are observed following exogenous addition to cells of purified NSP4 or of a synthetic peptide of NSP4. Experiments to determine the mechanism by which NSP4 causes an increase in [Ca2+]i showed that Ca2+ is released from a subset of the thapsigargin-sensitive store [endoplasmic reticulum (ER)]. However, exogenously added and endogenously expressed NSP4 use different mechanisms to alter the Ca2+ permeability of the ER membrane. We hypothesize that NSP4-mediated changes in ER membrane permeability trigger viral budding into the lumen of the ER, and eventually induce cell death and release of virus particles from infected cells. We also propose that release of NSP4 following cell lysis and the concomitant stimulation of a Ca2+ signal transduction pathway in neighboring cells contributes to altered ion transport in intestinal epithelium resulting in diarrheal disease.


Insect Cell Baculovirus Recombinant Outer Capsid Protein Subviral Particle Mature Viral Particle 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


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  1. 1.
    Dubois-Dalcq M, Holmes KV, Rentier B (1984) Assembly of enveloped RNA viruses. Springer, New YorkCrossRefGoogle Scholar
  2. 2.
    Au K-S, Chan W-K, Burns JW, Estes MK (1989) Receptor activity of rotavirus nonstructural glycoprotein NS28. J Virol 63: 4553–4562PubMedGoogle Scholar
  3. 3.
    Au K-S, Mattion NM, Estes MK (1993) A subviral particle binding domain on the rotavirus nonstructural glycoprotein NS28. Virology 194: 665–673PubMedCrossRefGoogle Scholar
  4. 4.
    Au K-S, Chan W-K, Estes MK (1988) Rotavirus morphogenesis involves an endoplasmic reticulum transmembrane glycoprotein. In: Compans R, Helenius A, Oldston M (eds) Cell Biology of Virus Entry, Replication and Pathogenesis. Alan R Lill, New YorkGoogle Scholar
  5. 5.
    Meyer JC, Bergmann CC, Bellamy AR (1989) Interaction of rotavirus cores with the nonstructural glycoprotein NS28. Virology 171: 98–107PubMedCrossRefGoogle Scholar
  6. 6.
    Petrie BL, Estes MK, Graham DY (1983) Effects of tunicamycin on rotavirus morphogenesis and infectivity. J Virol 46: 270–274PubMedGoogle Scholar
  7. 7.
    Ericson BL, Petrie BL, Graham DY, Mason BB, Estes MK (1983) Rotaviruses code for two types of glycoprotein precursors. J Cell Biochem 22: 151–160PubMedCrossRefGoogle Scholar
  8. 8.
    Chan W-K, Au K-S, Estes MK (1988) Topography of the simian rotavirus nonstructural glycoprotein (NS28) in the endoplasmic reticulum membrane. Virology 164: 435–442PubMedCrossRefGoogle Scholar
  9. 9.
    Holmes IH (1993) Rotaviruses. In: Joklik WK (ed) The Reoviridae. Plenum Publishing, New York, pp 359–423Google Scholar
  10. 10.
    Petrie BL (1983) Biologic activity of rotavirus particles lacking glycosylated proteins. In: Compans RW, Bishop DHL (eds) Double-Stranded RNA Viruses. Elsevier, New York, pp 146–156Google Scholar
  11. 11.
    Tian P, Hu Y, Schilling WP, Lindsay DA, Eiden J, Estes MK (1994) The nonstructural glycoprotein of rotavirus affects intracellular calcium levels. J Virol 68: 251–257PubMedGoogle Scholar
  12. 11a.
    Taylor JA, O’Brien JA, Lord VJ, Meyer JC, Bellamy AR (1993) The RER-localized intracellular receptor: A truncated purified soluble form is multivalent and binds virus particles. Virology 194: 807–814PubMedCrossRefGoogle Scholar
  13. 12.
    Cohen J, Laprote J, Charpilienne A, Scherrer R (1979) Activation of rotavirus RNA polymerase by calcium chelation. Arch Virol 60: 177–186PubMedCrossRefGoogle Scholar
  14. 13.
    Estes MK, Graham DY, Mason BB (1981) Proteolytic enhancement of rotavirus infectivity: molecular mechanisms. J Virol 39: 879–888PubMedGoogle Scholar
  15. 14.
    Shahrabadi MS, Lee PW (1986) Bovine rotavirus maturation is a calcium-dependent process. Virology 152: 298–307PubMedCrossRefGoogle Scholar
  16. 15.
    Ludert JE, Michelangeli F, Gil F, Liprandi F, Esparza J (1987) Penetration and uncoating of rotaviruses in cultured cells. Intervirology 27: 95–101PubMedCrossRefGoogle Scholar
  17. 16.
    Dormitzer PR, Greenberg HB (1992) Calcium chelation induces a conformational change in recombinant herpes simplex virus-1-expressed rotavirus VP7. Virology 189: 828–832PubMedCrossRefGoogle Scholar
  18. 17.
    Michelangeli F, Ruiz M-C, Del Castillo JR, Ernesto Ludert J, Liprandi F (1991) Effect of rotavirus infection on intracellular calcium homeostasis in cultured cells. Virology 181: 520–527PubMedCrossRefGoogle Scholar
  19. 18.
    Tian P, Estes MK, Hu Y, Ball JM, Zeng CQ, and Schilling WP (1995) The rotavirus nonstructural glycoprotein NSP4 mobilizes Ca2+ from the endoplasmic reticulum. J Virol 69: 5763–5772PubMedGoogle Scholar
  20. 19.
    Crawford SE, Labbe M, Cohen J, Burroughs MH, Zhou Y, Estes MK (1994) Characterization of virus-like particles produced by the expression of rotavirus capsid proteins in insect cells. J Virol 68(9): 5945–5952PubMedGoogle Scholar
  21. 20.
    Belyaev AS, Roy P (1993) Development of baculovirus triple and quadruple expression vectors: Co-expression of three or four bluetongue virus proteins and the synthesis of bluetongue virus-like particles in insect cells. Nucleic Acids Research 21: 1219–1223PubMedCrossRefGoogle Scholar
  22. 21.
    Hu Y, Rajan L, Schilling WP (1994) Ca2+ signaling in Sf9 insect cells and the functional expression of a rat brain M5 muscarinic receptor. Am J Physiol 266 (June): C1736–C1743PubMedGoogle Scholar
  23. 22.
    Schilling WP, Chen X, Sinkins WG, Rajan L (1994) Expression of a human bradykinin B2 receptor in Sf9 insect cells using baculovirus expression vector. FASEB J 8: A352 (Abstract)Google Scholar
  24. 23.
    Chen X, Schilling WP (1994) Desensitization of recombinant human thrombin receptor: evidence for active tethered ligand. FASEB J 8:A89 (Abstract)Google Scholar
  25. 24.
    Ball JM, Tian P, Zeng CQ, Morris A, Estes MK (1996) Age-dependent diarrhea is induced by a viral nonstructural glycoprotein. Science 272: 101–104PubMedCrossRefGoogle Scholar
  26. 25.
    Mattion NM, Cohen J, Estes MK (1994) The Rotavirus Proteins. In: Kapikian A (ed) Viral Infections of the Gastrointestinal Tract, 2nd edn. Marcel Dekker, New YorkGoogle Scholar
  27. 26.
    Tian P (1994) Pleiotropic properties of rotavirus nonstructural glycoprotein NSP4 in viral morphogenesis and pathogenesis. Baylor College of Medicine, PhD Thesis, p 119Google Scholar

Copyright information

© Springer-Verlag Wien 1996

Authors and Affiliations

  • P. Tian
    • 1
    • 2
  • J. M. Ball
    • 1
  • C. Q-Y Zeng
    • 1
  • M. K. Estes
    • 1
  1. 1.Division of Molecular VirologyBaylor College of MedicineHoustonUSA
  2. 2.Howard Hughes Medical Institute and Department of Cell BiologyBaylor College of MedicineHoustonUSA

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