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.

Human occludin is a hepatitis C virus entry factor required for infection of mouse cells

Abstract

Hepatitis C virus (HCV) is a leading cause of liver disease worldwide. The development of much needed specific antiviral therapies and an effective vaccine has been hampered by the lack of a convenient small animal model. The determinants restricting HCV tropism to human and chimpanzee hosts are unknown. Replication of the viral RNA has been demonstrated in mouse cells1,2, but these cells are not infectable with either lentiviral particles bearing HCV glycoproteins (HCVpp)3 or HCV produced in cell culture (HCVcc) (A.P., M.E. and C.M.R., unpublished observations), suggesting that there is a block at the level of entry. Here we show, using an iterative complementary DNA library screening approach, that human occludin (OCLN) is an essential HCV cell entry factor that is able to render murine cells infectable with HCVpp. Similarly, OCLN is required for the HCV-susceptibility of human cells, because its overexpression in uninfectable cells specifically enhanced HCVpp uptake, whereas its silencing in permissive cells impaired both HCVpp and HCVcc infection. In addition to OCLN, HCVpp infection of murine cells required expression of the previously identified HCV entry factors CD81 (ref. 4), scavenger receptor class B type I (SR-BI, also known as SCARB1)5 and claudin-1 (CLDN1)6. Although the mouse versions of SR-BI and CLDN1 function at least as well as the human proteins in promoting HCV entry, both OCLN and CD81 must be of human origin to allow efficient infection. The species-specific determinants of OCLN were mapped to its second extracellular loop. The identification of OCLN as a new HCV entry factor further highlights the importance of the tight junction complex in the viral entry process, and provides an important advance towards efforts to develop small animal models for HCV.

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: OCLN expression confers susceptibility to HCVpp.
Figure 2: OCLN silencing inhibits HCV entry.
Figure 3: Expression of human OCLN and CD81 determines HCV species tropism.

References

  1. Uprichard, S. L., Chung, J., Chisari, F. V. & Wakita, T. Replication of a hepatitis C virus replicon clone in mouse cells. Virol. J. 3, 89–97 (2006)

    Article  Google Scholar 

  2. Zhu, Q., Guo, J. T. & Seeger, C. Replication of hepatitis C virus subgenomes in nonhepatic epithelial and mouse hepatoma cells. J. Virol. 77, 9204–9210 (2003)

    Article  CAS  Google Scholar 

  3. Bartosch, B., Dubuisson, J. & Cosset, F. L. Infectious hepatitis C virus pseudo-particles containing functional E1–E2 envelope protein complexes. J. Exp. Med. 197, 633–642 (2003)

    Article  CAS  Google Scholar 

  4. Pileri, P. et al. Binding of hepatitis C virus to CD81. Science 282, 938–941 (1998)

    Article  ADS  CAS  Google Scholar 

  5. Scarselli, E. et al. The human scavenger receptor class B type I is a novel candidate receptor for the hepatitis C virus. EMBO J. 21, 5017–5025 (2002)

    Article  CAS  Google Scholar 

  6. Evans, M. J. et al. Claudin-1 is a hepatitis C virus co-receptor required for a late step in entry. Nature 446, 801–805 (2007)

    Article  ADS  CAS  Google Scholar 

  7. von Hahn, T. & Rice, C. M. Hepatitis C virus entry. J. Biol. Chem. 283, 3689–3693 (2008)

    Article  CAS  Google Scholar 

  8. Blight, K. J., McKeating, J. A. & Rice, C. M. Highly permissive cell lines for hepatitis C virus genomic and subgenomic RNA replication. J. Virol. 76, 13001–13014 (2002)

    Article  CAS  Google Scholar 

  9. Bartosch, B. et al. In vitro assay for neutralizing antibody to hepatitis C virus: evidence for broadly conserved neutralization epitopes. Proc. Natl Acad. Sci. USA 100, 14199–14204 (2003)

    Article  ADS  CAS  Google Scholar 

  10. Drummer, H. E., Maerz, A. & Poumbourios, P. Cell surface expression of functional hepatitis C virus E1 and E2 glycoproteins. FEBS Lett. 546, 385–390 (2003)

    Article  CAS  Google Scholar 

  11. Hsu, M. et al. Hepatitis C virus glycoproteins mediate pH-dependent cell entry of pseudotyped retroviral particles. Proc. Natl Acad. Sci. USA 100, 7271–7276 (2003)

    Article  ADS  CAS  Google Scholar 

  12. Paris, L., Tonutti, L., Vannini, C. & Bazzoni, G. Structural organization of the tight junctions. Biochim. Biophys. Acta 1778, 646–659 (2008)

    Article  CAS  Google Scholar 

  13. Chiba, H. et al. Transmembrane proteins of tight junctions. Biochim. Biophys. Acta 1778, 588–600 (2008)

    Article  CAS  Google Scholar 

  14. Flint, M. et al. Diverse CD81 proteins support hepatitis C virus infection. J. Virol. 80, 11331–11342 (2006)

    Article  CAS  Google Scholar 

  15. Coyne, C. B. & Bergelson, J. M. Virus-induced Abl and Fyn kinase signals permit coxsackievirus entry through epithelial tight junctions. Cell 124, 119–131 (2006)

    Article  CAS  Google Scholar 

  16. Coyne, C. B., Shen, L., Turner, J. R. & Bergelson, J. M. Coxsackievirus entry across epithelial tight junctions requires occludin and the small GTPases Rab34 and Rab5. Cell Host Microbe 2, 181–192 (2007)

    Article  CAS  Google Scholar 

  17. Brazzoli, M. et al. CD81 is a central regulator of cellular events required for hepatitis C virus infection of human hepatocytes. J. Virol. 82, 8316–8329 (2008)

    Article  CAS  Google Scholar 

  18. Marukian, S. et al. Cell culture-produced hepatitis C virus does not infect peripheral blood mononuclear cells. Hepatology 48, 1843–1850 (2008)

    Article  Google Scholar 

  19. Lindenbach, B. D. et al. Complete replication of hepatitis C virus in cell culture. Science 309, 623–626 (2005)

    Article  ADS  CAS  Google Scholar 

Download references

Acknowledgements

We thank D. Bowman, K. Mu, M. Hunter, J. Tassello and M. Holz for excellent technical assistance, J. L. Law and N. Shohdy for advice on siRNA experiments, T. von Hahn and A. J. Syder for helpful discussions and C. Murray for editing the manuscript. S. Mazel, C. Bare and X. Fan provided outstanding technical support. This work was supported by the Greenberg Medical Research Institute, the Ellison Medical Foundation, the Starr Foundation, the Ronald A. Shellow Memorial Fund, the Richard Salomon Family Foundation and funded in part by a Grant from the Foundation for the National Institutes of Health through the Grand Challenges in Global Health initiative (to C.M.R.), and the National Institutes of Health (to M.J.E. and C.M.R.). C.M.R. is an Ellison Medical Foundation Senior Scholar in Global Infectious Diseases. A.P. and M.J.E. were supported by Kimberly Lawrence-Netter Cancer Research Discovery Fund Award Postdoctoral Fellowships. M.J.E. was also supported by the Charles H. Revson Postdoctoral Fellowship. We would like to thank P. D. Bieniasz and T. Hatziioannou for providing reagents, including the LMN8 retroviral plasmid, and expertise necessary for the cDNA library construction and screening. R. Tsien provided the mCherry fluorescent protein encoding plasmid, A. Miyawaki the Venus/YFP plasmid and D. W. Piston the Cerulean/CFP plasmid. Constructs expressing HCV glycoproteins of diverse genotypes and the JFH-1 subgenomic replicon cDNA were provided by J. Bukh and T. Wakita, respectively.

Author Contributions A.P., M.J.E. and C.M.R. designed the project, analysed results and wrote the manuscript. A.P., M.J.E., H.Y., M.P., V.A.G. and Y.P.d.J. performed the experimental work.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Charles M. Rice.

Ethics declarations

Competing interests

C.M.R. has equity in Apath, LLC, which holds commercial licenses for the Huh-7.5 cell line and the HCVcc cell culture system.

Supplementary information

Supplementary Information

This file contains Supplementary Data and Methods, Supplementary References and Supplementary Figures S1-S4 (PDF 454 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Ploss, A., Evans, M., Gaysinskaya, V. et al. Human occludin is a hepatitis C virus entry factor required for infection of mouse cells. Nature 457, 882–886 (2009). https://doi.org/10.1038/nature07684

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature07684

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

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