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Journal of Virology, July 2006, p. 6964-6972, Vol. 80, No. 14
0022-538X/06/$08.00+0     doi:10.1128/JVI.00024-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Hepatitis C Virus Entry Depends on Clathrin-Mediated Endocytosis

Emmanuelle Blanchard,1 Sandrine Belouzard,1 Lucie Goueslain,1 Takaji Wakita,2 Jean Dubuisson,1 Czeslaw Wychowski,1 and Yves Rouillé1*

CNRS-UMR8161, Institut de Biologie de Lille and Institut Pasteur de Lille, Lille, France,1 Department of Virology II, National Institute for Infectious Diseases, Tokyo, Japan2

Received 4 January 2006/ Accepted 3 May 2006

Due to difficulties in cell culture propagation, the mechanisms of hepatitis C virus (HCV) entry are poorly understood. Here, postbinding cellular mechanisms of HCV entry were studied using both retroviral particles pseudotyped with HCV envelope glycoproteins (HCVpp) and the HCV clone JFH-1 propagated in cell culture (HCVcc). HCVpp entry was measured by quantitative real-time PCR after 3 h of contact with target cells, and HCVcc infection was quantified by immunoblot analysis and immunofluorescence detection of HCV proteins expressed in infected cells. The functional role of clathrin-mediated endocytosis in HCV entry was assessed by small interfering RNA-mediated clathrin heavy chain depletion and with chlorpromazine, an inhibitor of clathrin-coated pit formation at the plasma membrane. In both conditions, HCVpp entry and HCVcc infection were inhibited. HCVcc infection was also inhibited by pretreating target cells with bafilomycin A1 or chloroquine, two drugs known to interfere with endosome acidification. These data indicate that HCV enters target cells by clathrin-mediated endocytosis, followed by a fusion step from within an acidic endosomal compartment.

* Corresponding author. Mailing address: Équipe Hépatite C, CNRS-UMR8161, Institut de Biologie de Lille, 1 rue du Professeur Calmette, BP447, 59021 Lille cedex, France. Phone: (33) 3 20 87 10 27. Fax: (33) 3 20 87 12 01. E-mail: yves.rouille{at}ibl.fr.

Journal of Virology, July 2006, p. 6964-6972, Vol. 80, No. 14
0022-538X/06/$08.00+0     doi:10.1128/JVI.00024-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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