Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Identification of nucleolin as a cellular receptor for human respiratory syncytial virus

Abstract

Human respiratory syncytial virus (RSV) causes a large burden of disease worldwide1. There is no effective vaccine or therapy, and the use of passive immunoprophylaxis with RSV-specific antibodies is limited to high-risk patients2,3,4,5. The cellular receptor (or receptors) required for viral entry and replication has yet to be described; its identification will improve understanding of the pathogenesis of infection and provide a target for the development of novel antiviral interventions. Here we show that RSV interacts with host-cell nucleolin via the viral fusion envelope glycoprotein and binds specifically to nucleolin at the apical cell surface in vitro. We observed decreased RSV infection in vitro in neutralization experiments using nucleolin-specific antibodies before viral inoculation, in competition experiments in which virus was incubated with soluble nucleolin before inoculation of cells, and upon RNA interference (RNAi) to silence cellular nucleolin expression. Transfection of nonpermissive Spodoptera frugiperda Sf9 insect cells with human nucleolin conferred susceptibility to RSV infection. RNAi-mediated knockdown of lung nucleolin was associated with a significant reduction in RSV infection in mice (P = 0.0004), confirming that nucleolin is a functional RSV receptor in vivo.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: RSV VOPBA and coimmunoprecipitation of RSV fusion protein with nucleolin.
Figure 2: RSV and nucleolin colocalized on cell surfaces, and inhibition interventions decreased infection.
Figure 3: Sf9 cells are made permissive to RSV infection by heterologous expression of human nucleolin.
Figure 4: Silencing nucleolin reduces RSV infection in mice.

References

  1. Collins, P.L. & Graham, B.S. Viral and host factors in human respiratory syncytial virus pathogenesis. J. Virol. 82, 2040–2055 (2008).

    Article  CAS  PubMed  Google Scholar 

  2. Wu, H., Pfarr, D.S., Losonsky, G.A. & Kiener, P.A. Immunoprophylaxis of RSV infection: advancing from RSV-IGIV to palivizumab and motavizumab. Curr. Top. Microbiol. Immunol. 317, 103–123 (2008).

    CAS  PubMed  Google Scholar 

  3. Welliver, R.C. Pharmacotherapy of respiratory syncytial virus infection. Curr. Opin. Pharmacol. 10, 289–293 (2010).

    Article  CAS  PubMed  Google Scholar 

  4. Olszewska, W. & Openshaw, P. Emerging drugs for respiratory syncytial virus infection. Expert Opin. Emerg. Drugs 14, 207–217 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Magro, M., Andreu, D., Gomez-Puertas, P., Melero, J.A. & Palomo, C. Neutralization of human respiratory syncytial virus infectivity by antibodies and low-molecular-weight compounds targeted against the fusion glycoprotein. J. Virol. 84, 7970–7982 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Adams, O. et al. Palivizumab-resistant human respiratory syncytial virus infection in infancy. Clin. Infect. Dis. 51, 185–188 (2010).

    Article  CAS  PubMed  Google Scholar 

  7. Lowry, F. FDA panel nixes licensing request for motavizumab. Medscape News (3 June 2010).

  8. DeVincenzo, J. et al. A randomized, double-blind, placebo-controlled study of an RNAi-based therapy directed against respiratory syncytial virus. Proc. Natl. Acad. Sci. USA 107, 8800–8805 (2010).

    Article  CAS  PubMed  Google Scholar 

  9. Olszewska, W. et al. Antiviral and lung protective activity of a novel RSV fusion inhibitor in a mouse model. Eur. Respir. J. 38, 401–408 (2011).

    Article  CAS  PubMed  Google Scholar 

  10. Behera, A.K. et al. Blocking intercellular adhesion molecule-1 on human epithelial cells decreases respiratory syncytial virus infection. Biochem. Biophys. Res. Commun. 280, 188–195 (2001).

    Article  CAS  PubMed  Google Scholar 

  11. Krusat, T. & Streckert, H.J. Heparin-dependent attachment of respiratory syncytial virus (RSV) to host cells. Arch. Virol. 142, 1247–1254 (1997).

    Article  CAS  PubMed  Google Scholar 

  12. Malhotra, R. et al. Isolation and characterisation of potential respiratory syncytial virus receptor(s) on epithelial cells. Microbes Infect. 5, 123–133 (2003).

    Article  CAS  PubMed  Google Scholar 

  13. Dimmock, N.J., Easton, A.J. & Leppard, K. Introduction to Modern Virology (Blackwell, Malden, Massachusetts, USA, 2007).

  14. Techaarpornkul, S., Barretto, N. & Peeples, M.E. Functional analysis of recombinant respiratory syncytial virus deletion mutants lacking the small hydrophobic and/or attachment glycoprotein gene. J. Virol. 75, 6825–6834 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Hallak, L.K., Kwilas, S.A. & Peeples, M.E. Interaction between respiratory syncytial virus and glycosaminoglycans, including heparan sulfate. Methods Mol. Biol. 379, 15–34 (2007).

    Article  CAS  PubMed  Google Scholar 

  16. Techaarpornkul, S., Collins, P.L. & Peeples, M.E. Respiratory syncytial virus with the fusion protein as its only viral glycoprotein is less dependent on cellular glycosaminoglycans for attachment than complete virus. Virology 294, 296–304 (2002).

    Article  CAS  PubMed  Google Scholar 

  17. Hallak, L.K., Spillmann, D., Collins, P.L. & Peeples, M.E. Glycosaminoglycan sulfation requirements for respiratory syncytial virus infection. J. Virol. 74, 10508–10513 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Collins, P.L. & Mottet, G. Post-translational processing and oligomerization of the fusion glycoprotein of human respiratory syncytial virus. J. Gen. Virol. 72, 3095–3101 (1991).

    Article  CAS  PubMed  Google Scholar 

  19. Crim, R.L., Audet, S.A., Feldman, S.A., Mostowski, H.S. & Beeler, J.A. Identification of linear heparin-binding peptides derived from human respiratory syncytial virus fusion glycoprotein that inhibit infectivity. J. Virol. 81, 261–271 (2007).

    Article  CAS  PubMed  Google Scholar 

  20. Martínez, I. & Melero, J.A. Binding of human respiratory syncytial virus to cells: implication of sulfated cell surface proteoglycans. J. Gen. Virol. 81, 2715–2722 (2000).

    Article  PubMed  Google Scholar 

  21. Cao, W. et al. Identification of alpha-dystroglycan as a receptor for lymphocytic choriomeningitis virus and Lassa fever virus. Science 282, 2079–2081 (1998).

    Article  CAS  Google Scholar 

  22. Reyes-Reyes, E.M. & Akiyama, S.K. Cell-surface nucleolin is a signal transducing P-selectin binding protein for human colon carcinoma cells. Exp. Cell Res. 314, 2212–2223 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Losfeld, M.E. et al. The cell surface expressed nucleolin is a glycoprotein that triggers calcium entry into mammalian cells. Exp. Cell Res. 315, 357–369 (2009).

    Article  CAS  PubMed  Google Scholar 

  24. Chen, X., Kube, D.M., Cooper, M.J. & Davis, P.B. Cell surface nucleolin serves as receptor for DNA nanoparticles composed of pegylated polylysine and DNA. Mol. Ther. 16, 333–342 (2008).

    Article  CAS  PubMed  Google Scholar 

  25. Taylor, G., Stott, E.J., Hughes, M. & Collins, A.P. Respiratory syncytial virus infection in mice. Infect. Immun. 43, 649–655 (1984).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Hibbs, A.R., MacDonald, G. & Garsha, K. Handbook of Biological Confocal Microscopy (Springer, New York, 2006).

  27. Zhang, L., Peeples, M.E., Boucher, R.C., Collins, P.L. & Pickles, R.J. Respiratory syncytial virus infection of human airway epithelial cells is polarized, specific to ciliated cells, and without obvious cytopathology. J. Virol. 76, 5654–5666 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Srinivasakumar, N., Ogra, P.L. & Flanagan, T.D. Characteristics of fusion of respiratory syncytial virus with HEp-2 cells as measured by R18 fluorescence dequenching assay. J. Virol. 65, 4063–4069 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Osiowy, C. & Anderson, R. Neutralization of respiratory syncytial virus after cell attachment. J. Virol. 69, 1271–1274 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Mikhaylova, L.M., Boutanaev, A.M. & Nurminsky, D.I. Transcriptional regulation by Modulo integrates meiosis and spermatid differentiation in male germ line. Proc. Natl. Acad. Sci. USA 103, 11975–11980 (2006).

    Article  CAS  PubMed  Google Scholar 

  31. Said, E.A. et al. Pleiotrophin inhibits HIV infection by binding the cell surface-expressed nucleolin. FEBS J. 272, 4646–4659 (2005).

    Article  CAS  PubMed  Google Scholar 

  32. Bose, S., Basu, M. & Banerjee, A.K. Role of nucleolin in human parainfluenza virus type 3 infection of human lung epithelial cells. J. Virol. 78, 8146–8158 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Levin Perlman, S., Jordan, M., Brossmer, R., Greengard, O. & Moscona, A. The use of a quantitative fusion assay to evaluate HN-receptor interaction for human parainfluenza virus type 3. Virology 265, 57–65 (1999).

    Article  CAS  PubMed  Google Scholar 

  34. Tufaro, F. Virus entry: two receptors are better than one. Trends Microbiol. 5, 257–258, discussion 258–259 (1997).

    Article  CAS  PubMed  Google Scholar 

  35. Hovanessian, A.G. et al. The cell-surface-expressed nucleolin is associated with the actin cytoskeleton. Exp. Cell Res. 261, 312–328 (2000).

    Article  CAS  PubMed  Google Scholar 

  36. Leistner, C.M., Gruen-Bernhard, S. & Glebe, D. Role of glycosaminoglycans for binding and infection of hepatitis B virus. Cell. Microbiol. 10, 122–133 (2008).

    CAS  PubMed  Google Scholar 

  37. Nisole, S. et al. The anti-HIV pentameric pseudopeptide HB-19 binds the C-terminal end of nucleolin and prevents anchorage of virus particles in the plasma membrane of target cells. J. Biol. Chem. 277, 20877–20886 (2002).

    Article  CAS  PubMed  Google Scholar 

  38. Said, E.A. et al. The anti-HIV cytokine midkine binds the cell surface-expressed nucleolin as a low affinity receptor. J. Biol. Chem. 277, 37492–37502 (2002).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors thank R.A. Weiss, B. McManus and F. Jean for helpful comments, and L. Morikawa and Q. Xu for help with immunohistochemical staining of mouse lung. F.T. is the recipient of a Canadian Institutes of Health Research (CIHR) Institute of Genetics Grant for Short-term Visits, and D.M. is the recipient of CIHR IMPACT and Heart and Stroke Foundation of British Columbia Post-doctoral Fellowship awards.

Author information

Authors and Affiliations

Authors

Contributions

F.T. conducted VOPBAs, mass spectrometry analysis, neutralization and competition experiments in vitro. D.M. conducted immunoprecipitation, cell culture and virus experiments with mammalian and Sf9 cells. T.J.M. performed mouse dissections and lung fixation. W.D. generated virus for the mouse experiments and did quantitative plaque assays. P.M. assisted with cell culture and mouse experiments, analyzed the data, prepared figures and graphs, and coordinated manuscript writing. R.G.H. did mouse lung histological examination and supervised the project.

Corresponding author

Correspondence to Richard G Hegele.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–13, Supplementary Table 1 and Supplementary Methods (PDF 1429 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Tayyari, F., Marchant, D., Moraes, T. et al. Identification of nucleolin as a cellular receptor for human respiratory syncytial virus. Nat Med 17, 1132–1135 (2011). https://doi.org/10.1038/nm.2444

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm.2444

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing