1887

Abstract

Recently, class II fusion proteins have been identified on the surface of alpha- and flaviviruses. These proteins have two functions besides membrane fusion: they generate an isometric lattice on the viral surface and they form ion-permeable pores at low pH. An attempt was made to identify inhibitors for the ion pores generated by the fusion proteins of the alphaviruses and . These pores can be detected and analysed in three situations: (i) in the target membrane during virus entry, by performing patch-clamp measurements of membrane currents; (ii) in the virus particle, by studying the entry of propidium iodide; and (iii) in the plasma membrane of infected cells, by Fura-2 fluorescence imaging of Ca entry into infected cells. It is shown here that, at a concentration of 0·1 mM, rare earth ions block the ion permeability of alphavirus ion pores in all three situations. Even at a concentration of 0·5 mM, these ions do not block formation of the viral fusion pore, as they do not inhibit entry or multiplication of alphaviruses. The data indicate that ions flow through the ion pores into the virus particle in the endosome and from the endosome into the cytoplasm after fusion of the viral envelope with the endosomal membrane. These ion flows, however, are not necessary for productive infection. The possibility that the ability of class II fusion proteins to form ion-permeable pores reflects their origin from protein toxins that form ion-permeable pores, and that entry via class II fusion proteins may resemble the entry of non-enveloped viruses, is discussed.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.81096-0
2005-12-01
2019-11-16
Loading full text...

Full text loading...

/deliver/fulltext/jgv/86/12/3311.html?itemId=/content/journal/jgv/10.1099/vir.0.81096-0&mimeType=html&fmt=ahah

References

  1. Abell, B. A. & Brown, D. T. ( 1993; ). Sindbis virus membrane fusion is mediated by reduction of glycoprotein disulfide bridges at the cell surface. J Virol 67, 5496–5501.
    [Google Scholar]
  2. Aidley, D. J. & Stanfield, P. R. ( 1996; ). Ion Channels: Molecules in Action. Cambridge: Cambridge University Press.
  3. Alouf, J. E. & Freer, J. H. (editors) ( 1999; ). The Comprehensive Sourcebook of Bacterial Protein Toxins, 2nd edn. London: Academic Press.
  4. Benz, R., Schmid, A., Wagner, W. & Goebel, W. ( 1989; ). Pore formation by the Escherichia coli hemolysin: evidence for an association-dissociation equilibrium of the pore-forming aggregates. Infect Immun 57, 887–895.
    [Google Scholar]
  5. Burns, D. L., Barbieri, J. T., Iglewski, B. H. & Rappuoli, R. (editors) ( 2003; ). Bacterial Protein Toxins. Washington, DC: American Society for Microbiology.
  6. Carrasco, L. ( 1995; ). Modification of membrane permeability by animal viruses. Adv Virus Res 45, 61–112.
    [Google Scholar]
  7. Collier, R. J. ( 2001; ). Understanding the mode of action of diphtheria toxin: a perspective on progress during the 20th century. Toxicon 39, 1793–1803.[CrossRef]
    [Google Scholar]
  8. Corver, J., Bron, R., Snippe, H., Kraaijeveld, C. & Wilschut, J. ( 1997; ). Membrane fusion activity of Semliki Forest virus in a liposomal model system: specific inhibition by Zn2+ ions. Virology 238, 14–21.[CrossRef]
    [Google Scholar]
  9. Dick, M., Barth, B. U. & Kempf, C. ( 1996; ). The E1 protein is mandatory for pore formation by Semliki Forest virus spikes. Virology 220, 204–207.[CrossRef]
    [Google Scholar]
  10. Garoff, H., Wilschut, J., Liljeström, P. & 7 other authors ( 1994; ). Assembly and entry mechanisms of Semliki Forest virus. Arch Virol Suppl 9, 329–338.
    [Google Scholar]
  11. Gibbons, D. L., Reilly, B., Ahn, A., Vaney, M.-C., Vigouroux, A., Rey, F. A. & Kielian, M. ( 2004; ). Purification and crystallization reveal two types of interactions of the fusion protein homotrimer of Semliki Forest virus. J Virol 78, 3514–3523.[CrossRef]
    [Google Scholar]
  12. Heinz, F. X. & Allison, S. L. ( 2003; ). Flavivirus structure and membrane fusion. Adv Virus Res 59, 63–97.
    [Google Scholar]
  13. Helenius, A., Marsh, M. & White, J. ( 1982; ). Inhibition of Semliki Forest virus penetration by lysosomotropic weak bases. J Gen Virol 58, 47–61.[CrossRef]
    [Google Scholar]
  14. Hernandez, R., Luo, T. & Brown, D. T. ( 2001; ). Exposure to low pH is not required for penetration of mosquito cells by Sindbis virus. J Virol 75, 2010–2013.[CrossRef]
    [Google Scholar]
  15. Hille, B. ( 2001; ). Ion Channels of Excitable Membranes, 3rd edn. Sunderland, MA: Sinauer Associates.
  16. Käsermann, F. & Kempf, C. ( 1996; ). Low pH-induced pore formation by spike proteins of enveloped viruses. J Gen Virol 77, 3025–3032.[CrossRef]
    [Google Scholar]
  17. Kielian, M. ( 1995; ). Membrane fusion and the alphavirus life cycle. Adv Virus Res 45, 113–151.
    [Google Scholar]
  18. Kielian, M. C. & Helenius, A. ( 1984; ). Role of cholesterol in the fusion of Semliki Forest virus with membranes. J Virol 52, 281–283.
    [Google Scholar]
  19. Kielian, M., Chatterjee, P. K., Gibbons, D. L. & Lu, Y. E. ( 2000; ). Specific roles for lipids in virus fusion and exit. Examples from the alphaviruses. Subcell Biochem 34, 409–455.
    [Google Scholar]
  20. Koschinski, A., Wengler, G., Wengler, G. & Repp, H. ( 2003; ). The membrane proteins of flaviviruses form ion-permeable pores in the target membrane after fusion: identification of the pores and analysis of their possible role in virus infection. J Gen Virol 84, 1711–1721.[CrossRef]
    [Google Scholar]
  21. Lanzrein, M., Weingart, R. & Kempf, C. ( 1993; ). pH-dependent pore formation in Semliki Forest virus-infected Aedes albopictus cells. Virology 193, 296–302.[CrossRef]
    [Google Scholar]
  22. 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. Cell 105, 137–148.[CrossRef]
    [Google Scholar]
  23. Lindenbach, B. D. & Rice, C. M. ( 2001; ). Flaviviridae: the viruses and their replication. In Fields Virology, 4th edn, pp. 991–1041. Edited by D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott Williams & Wilkins.
  24. Madan, V., Sanz, M. A. & Carrasco, L. ( 2005; ). Requirement of the vesicular system for membrane permeabilization by Sindbis virus. Virology 332, 307–315.[CrossRef]
    [Google Scholar]
  25. Major, M. E., Rehermann, B. & Feinstone, S. M. ( 2001; ). Hepatitis C viruses. In Fields Virology, 4th edn, pp. 1127–1161. Edited by D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott Williams & Wilkins.
  26. Modis, Y., Ogata, S., Clements, D. & Harrison, S. C. ( 2003; ). A ligand-binding pocket in the dengue virus envelope glycoprotein. Proc Natl Acad Sci U S A 100, 6986–6991.[CrossRef]
    [Google Scholar]
  27. Nieva, J. L., Bron, R., Corver, J. & Wilschut, J. ( 1994; ). Membrane fusion of Semliki Forest virus requires sphingolipids in the target membrane. EMBO J 13, 2797–2804.
    [Google Scholar]
  28. Nyfeler, S., Senn, K. & Kempf, C. ( 2001; ). Expression of Semliki Forest virus E1 protein in Escherichia coli: low pH-induced pore formation. J Biol Chem 276, 15453–15457.[CrossRef]
    [Google Scholar]
  29. Omar, A. & Koblet, H. ( 1988; ). Semliki Forest virus particles containing only the E1 envelope glycoprotein are infectious and can induce cell-cell fusion. Virology 166, 17–23.[CrossRef]
    [Google Scholar]
  30. Panchal, R. G., Smart, M. L., Bowser, D. N., Williams, D. A. & Petrou, S. ( 2002; ). Pore-forming proteins and their application in biotechnology. Curr Pharm Biotechnol 3, 99–115.[CrossRef]
    [Google Scholar]
  31. Paredes, A. M., Ferreira, D., Horton, M. & 8 other authors ( 2004; ). Conformational changes in Sindbis virions resulting from exposure to low pH and interactions with cells suggest that cell penetration may occur at the cell surface in the absence of membrane fusion. Virology 324, 373–386.[CrossRef]
    [Google Scholar]
  32. 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. Cell 105, 127–136.[CrossRef]
    [Google Scholar]
  33. 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. Nature 375, 291–298.[CrossRef]
    [Google Scholar]
  34. Sandvig, K. & van Deurs, B. ( 2002; ). Membrane traffic exploited by protein toxins. Annu Rev Cell Dev Biol 18, 1–24.[CrossRef]
    [Google Scholar]
  35. Schlegel, A., Omar, A., Jentsch, P., Morell, A. & Kempf, C. ( 1991; ). Semliki Forest virus envelope proteins function as proton channels. Biosci Rep 11, 243–255.[CrossRef]
    [Google Scholar]
  36. Schlesinger, S. & Schlesinger, M. J. ( 2001; ). Togaviridae: the viruses and their replication. In Fields Virology, 4th edn, pp. 895–916. Edited by D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott Williams & Wilkins.
  37. Silver, R. B. ( 1998; ). Ratio imaging: practical considerations for measuring intracellular calcium and pH in living tissue. Methods Cell Biol 56, 237–251.
    [Google Scholar]
  38. 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 Lett 521, 62–66.[CrossRef]
    [Google Scholar]
  39. Spyr, C. A., Käsermann, F. & Kempf, C. ( 1995; ). Identification of the pore forming element of Semliki Forest virus spikes. FEBS Lett 375, 134–136.[CrossRef]
    [Google Scholar]
  40. Strauss, J. H. & Strauss, E. G. ( 2001; ). Virus evolution: how does an enveloped virus make a regular structure? Cell 105, 5–8.[CrossRef]
    [Google Scholar]
  41. 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. J Gen Virol 83, 2417–2426.
    [Google Scholar]
  42. 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. Virology 257, 472–482.[CrossRef]
    [Google Scholar]
  43. Wengler, G., Koschinski, A., Wengler, G. & Dreyer, F. ( 2003; ). Entry of alphaviruses at the plasma membrane converts the viral surface proteins into an ion-permeable pore that can be detected by electrophysiological analyses of whole-cell membrane currents. J Gen Virol 84, 173–181.[CrossRef]
    [Google Scholar]
  44. Wengler, G., Koschinski, A., Wengler, G. & Repp, H. ( 2004; ). During entry of alphaviruses, the E1 glycoprotein molecules probably form two separate populations that generate either a fusion pore or ion-permeable pores. J Gen Virol 85, 1695–1701.[CrossRef]
    [Google Scholar]
  45. White, J. & Helenius, A. ( 1980; ). pH-dependent fusion between the Semliki Forest virus membrane and liposomes. Proc Natl Acad Sci U S A 77, 3273–3277.[CrossRef]
    [Google Scholar]
  46. White, J., Kartenbeck, J. & Helenius, A. ( 1980; ). Fusion of Semliki Forest virus with the plasma membrane can be induced by low pH. J Cell Biol 87, 264–272.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.81096-0
Loading
/content/journal/jgv/10.1099/vir.0.81096-0
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