1887

Abstract

The virion glycoproteins Gn and Gc of Bunyamwera orthobunyavirus (family ) are encoded by the M RNA genome segment and have roles in both viral attachment and membrane fusion. To investigate further the structure and function of the Gc protein in viral replication, we generated 12 mutants that contain truncations from the N terminus. The effects of these deletions were analysed with regard to Golgi targeting, low pH-dependent membrane fusion, infectious virus-like particle (VLP) formation and virus infectivity. Our results show that the N-terminal half (453 residues) of the Gc ectodomain (909 residues in total) is dispensable for Golgi trafficking and cell fusion. However, deletions in this region resulted in a significant reduction in VLP formation. Four mutant viruses that contained N-terminal deletions in their Gc proteins were rescued, and found to be attenuated to different degrees in BHK-21 cells. Taken together, our data indicate that the N-terminal half of the Gc ectodomain is dispensable for replication in cell culture, whereas the C-terminal half is required to mediate cell fusion. A model for the domain structure of the Gc ectodomain is proposed.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.013540-0
2009-10-01
2024-04-27
Loading full text...

Full text loading...

/deliver/fulltext/jgv/90/10/2483.html?itemId=/content/journal/jgv/10.1099/vir.0.013540-0&mimeType=html&fmt=ahah

References

  1. Brandenburg B., Zhuang X. 2007; Virus trafficking – learning from single-virus tracking. Nat Rev Microbiol 5:197–208 [CrossRef]
    [Google Scholar]
  2. Bridgen A., Elliott R. M. 1996; Rescue of a segmented negative-strand RNA virus entirely from cloned complementary DNAs. Proc Natl Acad Sci U S A 93:15400–15404 [CrossRef]
    [Google Scholar]
  3. Buchholz U. J., Finke S., Conzelmann K. K. 1999; Generation of bovine respiratory syncytial virus (BRSV) from cDNA: BRSV NS2 is not essential for virus replication in tissue culture, and the human RSV leader region acts as a functional BRSV genome promoter. J Virol 73:251–259
    [Google Scholar]
  4. Cortez I., Aires A., Pereira A. M., Goldbach R., Peters D., Kormelink R. 2002; Genetic organisation of iris yellow spot virus M RNA: indications for functional homology between the G(C) glycoproteins of tospoviruses and animal-infecting bunyaviruses. Arch Virol 147:2313–2325 [CrossRef]
    [Google Scholar]
  5. Elliott R. M. 1990; Molecular biology of the Bunyaviridae . J Gen Virol 71:501–522 [CrossRef]
    [Google Scholar]
  6. Elliott R. M. editor 1996 The Bunyaviridae New York: Plenum;
    [Google Scholar]
  7. Elliott R. M., Lees J. F., Watret G. E., Clark W., Pringle C. R. 1984; Genetic diversity of bunyaviruses and mechanisms of genetic variation. In Mechanisms of Viral Pathogenesis: from Gene to Pathogen pp 61–76Edited by Kohn A., Fuchs P. Boston: Martinus Nijhoff;
    [Google Scholar]
  8. Elliott R. M., Bouloy M., Calisher C. H., Goldbach R., Moyer J. T., Nichol S. T., Pettersson R., Plyusnin A., Schmaljohn C. S. 2000; Bunyaviridae . In Virus Taxonomy: Seventh Report of the International Committee on Taxonomy of Viruses . pp 599–621Edited by van Regenmortel M. H. V., Fauquet C. M., Bishop D. H. L., Carstens E. B., Estes M. K., Lemon S. M., Maniloff J., Mayo M. A., McGeoch D. J., Pringle C. R., Wickner R. B. San Diego, CA: Academic Press;
  9. Filone C. M., Heise M., Doms R. W., Bertolotti-Ciarlet A. 2006; Development and characterization of a Rift Valley fever virus cell-cell fusion assay using alphavirus replicon vectors. Virology 356:155–164 [CrossRef]
    [Google Scholar]
  10. Fritz R., Stiasny K., Heinz F. X. 2008; Identification of specific histidines as pH sensors in flavivirus membrane fusion. J Cell Biol 183:353–361 [CrossRef]
    [Google Scholar]
  11. Garry C. E., Garry R. F. 2004; Proteomics computational analyses suggest that the carboxyl terminal glycoproteins of bunyaviruses are class II viral fusion protein (beta-penetrenes). Theor Biol Med Model 1:10 [CrossRef]
    [Google Scholar]
  12. Hacker J. K., Hardy J. L. 1997; Adsorptive endocytosis of California encephalitis virus into mosquito and mammalian cells: a role for G1. Virology 235:40–47 [CrossRef]
    [Google Scholar]
  13. Hacker J. K., Volkman L. E., Hardy J. L. 1995; Requirement for the G1 protein of California encephalitis virus in infection in vitro and in vivo . Virology 206:945–953 [CrossRef]
    [Google Scholar]
  14. Harrison S. C. 2005; Mechanism of membrane fusion by viral envelope proteins, advances in virus research. In Virus Structure and AssemblyAdvances in Virus Research vol. 64 pp 231–261Edited by Roy P. San Diego, CA: Elsevier Academic Press;
    [Google Scholar]
  15. Harrison S. C. 2008; Viral membrane fusion. Nat Struct Mol Biol 15:690–698 [CrossRef]
    [Google Scholar]
  16. Jacoby D. R., Cooke C., Prabakaran I., Boland J., Nathanson N., Gonzalez-Scarano F. 1993; Expression of the La Crosse M segment proteins in a recombinant vaccinia expression system mediates pH-dependent cellular fusion. Virology 193:993–996 [CrossRef]
    [Google Scholar]
  17. Kelley L. A., Sternberg M. J. 2009; Protein structure prediction on the web: a case study using the Phyre server. Nat Protoc 4:363–371 [CrossRef]
    [Google Scholar]
  18. Kielian M., Rey F. A. 2006; Virus membrane-fusion proteins: more than one way to make a hairpin. Nat Rev Microbiol 4:67–76 [CrossRef]
    [Google Scholar]
  19. Kingsford L., Ishizawa L. D., Hill D. W. 1983; Biological activities of monoclonal antibodies reactive with antigenic sites mapped on the G1 glycoprotein of La Crosse virus. Virology 129:443–455 [CrossRef]
    [Google Scholar]
  20. Lappin D. F., Nakitare G. W., Palfreyman J. W., Elliott R. M. 1994; Localization of Bunyamwera bunyavirus G1 glycoprotein to the Golgi requires association with G2 but not with NSm. J Gen Virol 75:3441–3451 [CrossRef]
    [Google Scholar]
  21. Lees J. F., Pringle C. R., Elliott R. M. 1986; Nucleotide sequence of the Bunyamwera virus M RNA segment: conservation of structural features in the bunyavirus glycoprotein gene product. Virology 148:1–14 [CrossRef]
    [Google Scholar]
  22. Lowen A. C., Noonan C., McLees A., Elliott R. M. 2004; Efficient bunyavirus rescue from cloned cDNA. Virology 330:493–500 [CrossRef]
    [Google Scholar]
  23. Ludwig G. V., Christensen B. M., Yuill T. M., Schultz K. T. 1989; Enzyme processing of La Crosse virus glycoprotein G1: a bunyavirus-vector infection model. Virology 171:108–113 [CrossRef]
    [Google Scholar]
  24. Ludwig G. V., Israel B. A., Christensen B. M., Yuill T. M., Schultz K. T. 1991; Role of La Crosse virus glycoproteins in attachment of virus to host cells. Virology 181:564–571 [CrossRef]
    [Google Scholar]
  25. Marsh M., Helenius A. 2006; Virus entry: open sesame. Cell 124:729–740 [CrossRef]
    [Google Scholar]
  26. Moller S., Croning M. D. R., Apweiler R. 2001; Evaluation of methods for the prediction of membrane spanning regions. Bioinformatics 17:646–653 [CrossRef]
    [Google Scholar]
  27. Murphy J., Pringle C. R. 1987; Bunyavirus mutants: reassortment group assignment and G1 protein variants. In The Biology of Negative Strand Viruses pp 357–362Edited by Mahy B., Kolakofsky D. Amsterdam: Elsevier;
    [Google Scholar]
  28. Nakamura N., Rabouille C., Watson R., Nilsson T., Hui N., Slusarewicz P., Kreis T. E., Warren G. 1995; Characterization of a cis-Golgi matrix protein, GM130. J Cell Biol 131:1715–1726 [CrossRef]
    [Google Scholar]
  29. Nichol S. T., Beaty B., Elliott R. M., Goldbach R., Plyusnin A., Schmaljohn A. L., Tesh R. B. 2005; Bunyaviridae . In Virus Taxonomy: Eighth Report of the International Committee on Taxonomy of Viruses pp 695–716Edited by Fauquet C. M., Mayo M. A., Maniloff J., Desselberger U., Ball L. A. San Diego, CA: Elsevier Academic Press;
    [Google Scholar]
  30. Ogino M., Yoshimatsu K., Ebihara H., Araki K., Lee B.-H., Okumura M., Arikawa J. 2004; Cell fusion activities of Hantaan virus envelope glycoproteins. J Virol 78:10776–10782 [CrossRef]
    [Google Scholar]
  31. Pekosz A., Griot C., Nathanson N., Gonzalez-Scarano F. 1995; Tropism of bunyaviruses: evidence for a G1 glycoprotein-mediated entry pathway common to the California serogroup. Virology 214:339–348 [CrossRef]
    [Google Scholar]
  32. Plassmeyer M. L., Soldan S. S., Stachelek K. M., Martin-Garcia J., Gonzalez-Scarano F. 2005; California serogroup Gc (G1) glycoprotein is the principal determinant of pH-dependent cell fusion and entry. Virology 338:121–132 [CrossRef]
    [Google Scholar]
  33. Plassmeyer M. L., Soldan S. S., Stachelek K. M., Roth S. M., Martin-Garcia J., Gonzalez-Scarano F. 2007; Mutagenesis of the La Crosse virus glycoprotein supports a role for Gc (1066–1087) as the fusion peptide. Virology 358:273–282 [CrossRef]
    [Google Scholar]
  34. Pollitt E., Zhao J., Muscat P., Elliott R. M. 2006; Characterization of Maguari orthobunyavirus mutants suggests the nonstructural protein NSm is not essential for growth in tissue culture. Virology 348:224–232 [CrossRef]
    [Google Scholar]
  35. Schmaljohn C., Hooper J. W. 2001; Bunyaviridae : the viruses and their replication. In Fields Virology , 4th edn. pp 1581–1602Edited by Knipe D. M., Howley P. M. Philadephia, PA: Lippincott Williams & Wilkins;
    [Google Scholar]
  36. Shi X., Elliott R. M. 2002; Golgi localization of Hantaan virus glycoproteins requires coexpression of G1 and G2. Virology 300:31–38 [CrossRef]
    [Google Scholar]
  37. Shi X., Elliott R. M. 2004; Analysis of N-linked glycosylation of Hantaan virus glycoproteins and the role of oligosaccharide side chains in protein folding and intracellular trafficking. J Virol 78:5414–5422 [CrossRef]
    [Google Scholar]
  38. Shi X., Lappin D. F., Elliott R. M. 2004; Mapping the Golgi targeting and retention signal of Bunyamwera virus glycoproteins. J Virol 78:10793–10802 [CrossRef]
    [Google Scholar]
  39. Shi X., Brauburger K., Elliott R. M. 2005; Role of N-linked glycans on bunyamwera virus glycoproteins in intracellular trafficking, protein folding, and virus infectivity. J Virol 79:13725–13734 [CrossRef]
    [Google Scholar]
  40. Shi X., Kohl A., Léonard V., Li P., McLees A., Elliott R. M. 2006; Requirement of the N-terminal region of the orthobunyavirus non-structural protein NSm for virus assembly and morphogenesis. J Virol 80:8089–8099 [CrossRef]
    [Google Scholar]
  41. Shi X., Kohl A., Li P., Elliott R. M. 2007; Role of the cytoplasmic tail domains of Bunyamwera orthobunyavirus glycoproteins Gn and Gc in virus assembly and morphogenesis. J Virol 81:10151–10160 [CrossRef]
    [Google Scholar]
  42. Smith A. E., Helenius A. 2004; How viruses enter animal cells. Science 304:237–242 [CrossRef]
    [Google Scholar]
  43. Sundin D. R., Beaty B. J., Nathanson N., Gonzalez-Scarano F. 1987; A G1 glycoprotein epitope of La Crosse virus: a determinant of infection of Aedes triseriatus . Science 235:591–593 [CrossRef]
    [Google Scholar]
  44. Tischler N. D., Gonzalez A., Perez-Acle T., Rosemblatt M., Valenzuela P. D. T. 2005; Hantavirus Gc glycoprotein: evidence for a class II fusion protein. J Gen Virol 86:2937–2947 [CrossRef]
    [Google Scholar]
  45. Watret G. E., Pringle C. R., Elliott R. M. 1985; Synthesis of bunyavirus-specific proteins in a continuous cell line (XTC-2) derived from Xenopus laevis . J Gen Virol 66:473–482 [CrossRef]
    [Google Scholar]
  46. Weber F., Dunn E. F., Bridgen A., Elliott R. M. 2001; The Bunyamwera virus nonstructural protein NSs inhibits viral RNA synthesis in a minireplicon system. Virology 281:67–74 [CrossRef]
    [Google Scholar]
  47. Weber F., Bridgen A., Fazakerley J. K., Streitenfeld H., Kessler N., Randall R. E., Elliott R. M. 2002; Bunyamwera bunyavirus nonstructural protein NSs counteracts the induction of alpha/beta interferon. J Virol 76:7949–7955 [CrossRef]
    [Google Scholar]
  48. Weissenhorn W., Hinz A., Gaudin Y. 2007; Virus membrane fusion. FEBS Lett 581:2150–2155 [CrossRef]
    [Google Scholar]
  49. White J., Matlin K., Helenius A. 1981; Cell fusion by Semliki Forest, influenza, and vesicular stomatitis viruses. J Cell Biol 89:674–679 [CrossRef]
    [Google Scholar]
  50. Whitfield A. E., Ullman D. E., German T. L. 2005; Tomato spotted wilt virus glycoprotein GC is cleaved at acidic pH. Virus Res 110:183–186 [CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.013540-0
Loading
/content/journal/jgv/10.1099/vir.0.013540-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error