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


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lentiviral particles bearing HCV glycoproteins (HCVpp)3 or HCV produced in cell culture (HCVcc) (A.P., M.E. and C.M.R., unpublished observations), suggesting that there is a block at the level of entry.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.

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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.


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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.

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Correspondence to Charles M. Rice.

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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.

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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).

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