1887

Abstract

Alphaviruses are small enveloped viruses that have been used extensively as model enveloped viruses. During infection, virus particles are taken up into endosomes, where a low pH activates the viral fusion protein, E1. Fusion of the viral and the endosomal membranes releases the viral core into the cytoplasm where cores are disassembled by interaction with 60S ribosomal subunits. Recently, we have shown that this disassembly is strongly stimulated by low pH. We have proposed that after entry of the core into the cytoplasm, the viral membrane proteins that have been transferred to the endosomal membrane form an ion-permeable pore in the endosome. The resulting flow of protons from the endosome into the cytoplasm through this pore could generate a low-pH environment for core disassembly . Here we report two types of analysis aimed at the identification of such pores. First, the release of [H]choline from the interior of liposomes was analysed in the presence of virus particles and viral proteins. Secondly, cells were infected with Sindbis or Semliki Forest alphaviruses at the plasma membrane and the possible generation of ion-permeable pores during this process was analysed by whole-cell voltage clamp analysis of the membrane current. The results obtained indicated that the proposed pores are in fact generated and allowed us to identify the formation of individual pores. Available evidence indicates that the alphavirus E1 protein probably forms these pores. Proteins homologous to the alphavirus E1 protein are present in flaviviruses and hepatitis C virus.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.18696-0
2003-01-01
2020-04-04
Loading full text...

Full text loading...

/deliver/fulltext/jgv/84/1/vir840173.html?itemId=/content/journal/jgv/10.1099/vir.0.18696-0&mimeType=html&fmt=ahah

References

  1. Ahn A., Gibbons D. L., Kielian M.. 2002; The fusion peptide of Semliki Forest virus associates with sterol-rich membrane domains. J Virol76:3267–3275
    [Google Scholar]
  2. Carrasco L.. 1981; Modification of membrane permeability induced by animal viruses early in infection. Virology113:623–629
    [Google Scholar]
  3. Carrasco L.. 1995; Modification of membrane permeability by animal viruses. Adv Virus Res45:61–112
    [Google Scholar]
  4. Dick M., Barth B. U., Kempf C.. 1996; The E1 protein is mandatory for pore formation by Semliki Forest virus spikes. Virology220:204–207
    [Google Scholar]
  5. Forsell K., Xing L., Kozlovska T., Cheng R. H., Garoff H.. 2000; Membrane proteins organize a symmetrical virus. EMBO J19:5081–5091
    [Google Scholar]
  6. Froshauer S., Kartenbeck J., Helenius A.. 1988; Alphavirus RNA replicase is located on the cytoplasmic surface of endosomes and lysosomes. J Cell Biol107:2075–2086
    [Google Scholar]
  7. Garoff H., Wilschut J., Liljeström P.. 7 other authors 1994; Assembly and entry mechansims of Semliki Forest virus. Arch Virol9:329–338
    [Google Scholar]
  8. Hamill O. P., Marty A., Neher E., Sakmann B., Sigworth F. J.. 1981; Improved patch-clamp techniques for high resolution current recording from cells and cell-free membrane patches. Pflügers Arch391:85–100
    [Google Scholar]
  9. Harrison S. C.. 1986; Alphavirus structure. In The Togaviridae and Flaviviridae pp 21–34 Edited by Schlesinger S., Schlesinger M. J.. New York: Plenum;
    [Google Scholar]
  10. Helenius A.. 1995; Alphavirus and flavivirus glycoproteins: structures and functions. Cell81:651–653
    [Google Scholar]
  11. Helenius A., Bondsdorff C. H.. 1976; Semliki Forest virus membrane proteins. Preparation and characterization of spike complexes soluble in detergent-free medium. Biochim Biophys Acta436:895–899
    [Google Scholar]
  12. Helenius A., Kartenbeck J., Simons K., Fries E.. 1980; On the entry of Semliki Forest virus into BHK-21 cells. J Cell Biol84:404–420
    [Google Scholar]
  13. Hescheler J., Schultz G.. 1993; Nonselective cation channels: physiological and pharmacological modulations of channel activity. In Nonselective Cation Channels pp 27–43 Edited by Hescheler J., Schultz G.. Basel: Birkhäuser;
    [Google Scholar]
  14. Käsermann F., Kempf C.. 1996; Low pH-induced pore formation by spike proteins of enveloped viruses. J Gen Virol77:3025–3032
    [Google Scholar]
  15. Kempf C., Michel M. R., Kohler U., Koblet H.. 1987; Can viral envelope proteins act as or induce proton channels?. Biosci Rep7:761–769
    [Google Scholar]
  16. Kielian M.. 1995; Membrane fusion and the alphavirus life cycle. Adv Virus Res45:113–151
    [Google Scholar]
  17. Lanzrein M., Weingart R., Kempf C.. 1993; pH-dependent pore formation in Semliki Forest virus-infected Aedes albopictus cells. Virology193:296–302
    [Google Scholar]
  18. Lescar J., Roussel A., Wien M. W., Navaza J., Fuller S. D., Wengler G., Wengler G., Rey F. A.. 2001; The fusion glycoprotein shell of Semliki Forest virus: an icosahedral assembly primed for fusogenic activation at endosomal pH. Cell105:137–148
    [Google Scholar]
  19. Marsh M., Helenius A.. 1989; Virus entry into animal cells. Adv Virus Res36:107–151
    [Google Scholar]
  20. Morein B., Helenius A., Simons K., Pettersson R., Kääriäinen L., Schirrmacher V.. 1978; Effective subunit vaccines against an enveloped animal virus. Nature276:715–718
    [Google Scholar]
  21. Nieva J. L., Bron R., Corver J., Wilschut J.. 1994; Membrane fusion of Semliki Forest virus requires sphingolipids in the target membrane. EMBO J13:2797–2804
    [Google Scholar]
  22. Nyfeler S., Senn K., Kempf C.. 2001; Expression of Semliki Forest virus E1 protein in Escherichia coli . J Biol Chem276:15453–15457
    [Google Scholar]
  23. Omar A., Koblet H.. 1988; Semliki Forest virus particles containing only the E1 envelope glycoprotein are infectious and can induce cell–cell fusion. Virology166:17–23
    [Google Scholar]
  24. Pérez L., Carrasco L.. 1994; Involvement of the vacuolar H+-ATPase in animal virus entry. J Gen Virol75:2595–2606
    [Google Scholar]
  25. Pletnev S. V., Zhang W., Mukhopadhyay S., Fisher B. R., Hernandez R., Brown D. T., Baker T. S., Rossmann M. G., Kuhn R. J.. 2001; Locations of carbohydrate sites on alphavirus glycoproteins show that E1 forms an icosahedral scaffold. Cell105:127–136
    [Google Scholar]
  26. Rey F. A., Heinz F. X., Mandl C., Kunz C., Harrison S. C.. 1995; The envelope glycoprotein from tick-borne encephalitis virus at 2 Å resolution. Nature375:291–298
    [Google Scholar]
  27. Schlegel A., Omar A., Jentsch P., Morell A., Kempf C.. 1991; Semliki Forest virus envelope proteins function as proton channels. Biosci Rep11:243–255
    [Google Scholar]
  28. Schlesinger M. J., Schlesinger S.. 1986; Formation and assembly of alphavirus glycoproteins. In The Togaviridae and Flaviviridae pp 121–148 Edited by Schlesinger S., Schlesinger M. J.. New York: Plenum;
    [Google Scholar]
  29. Schlesinger S., Schlesinger M. J.. 2001; Togaviridae : the viruses and their replication. In Fields Virology , 4th edn. pp 895–916 Edited by Knipe D. M., Howley P. M.. Philadelphia: Lippincott Williams & Wilkins;
    [Google Scholar]
  30. Singh I. R., Helenius A.. 1992; Role of ribosomes in Semliki Forest virus nucleocapsid uncoating. J Virol66:7049–7058
    [Google Scholar]
  31. Singh I. R., Suomalainen M., Varadarajan S., Garoff H., Helenius A.. 1997; Multiple mechanisms for the inhibition of entry and uncoating of superinfecting Semliki Forest virus. Virology231:59–71
    [Google Scholar]
  32. Smit J. M., Bittmann R., Wilschut J.. 1999; Low pH-dependent fusion of Sindbis virus with receptor-free cholesterol- and sphingolipid-containing liposomes. J Virol73:8476–8484
    [Google Scholar]
  33. Smit J. M., Li G., Schoen P., Corver J., Bittman R., Lin K.-C., Wilschut J.. 2002; Fusion of alphaviruses with liposomes is a non-leaky process. FEBS Lett521:62–66
    [Google Scholar]
  34. Strauss J. H., Strauss E. G.. 1994; The alphviruses: gene expression, replication and evolution. Microbiol Rev58:491–562
    [Google Scholar]
  35. Van Regenmortel M. H. V., Fauquet C. M., Bishop D. H. L.. 8 other editors 2000; Virus Taxonomy. Seventh Report of the International Committee on Taxonomy of Viruses San Diego: Academic Press;
    [Google Scholar]
  36. Wengler G., Wengler G.. 1984; Identification of a transfer of viral core protein to cellular ribosomes during the early stages of alphavirus infection. Virology134:435–442
    [Google Scholar]
  37. Wengler G., Wengler G.. 2002; In vitro analyses of factors involved in the disassembly of Sindbis virus cores by 60S ribosomal subunits identify a possible role of low pH in this process. J Gen Virol83:2417–2426
    [Google Scholar]
  38. Wengler G., Wengler G., Rey F. A.. 1999; The isolation of the ectodomain of the alphavirus E1 protein as a soluble hemagglutinin and its crystallization. Virology257:472–482
    [Google Scholar]
  39. White J., Helenius A.. 1980; PH-dependent fusion between the Semliki Forest virus membrane and liposomes. Proc Natl Acad Sci U S A77:3273–3277
    [Google Scholar]
  40. White J., Kartenbeck J., Helenius A.. 1980; Fusion of Semliki Forest virus with the plasma membrane can be induced by low pH. J Cell Biol87:264–272
    [Google Scholar]
  41. Yagnik A. T., Lahm A., Meola A., Roccasecca R. M., Ercole B. B., Nicosia A., Tramontano A.. 2000; A model for the hepatitis C virus envelope glycoprotein E2. Proteins Struc Funct Genet40:355–366
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.18696-0
Loading
/content/journal/jgv/10.1099/vir.0.18696-0
Loading

Data & Media loading...

Most cited this month

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