Downloaded from www.microbiologyresearch.org by IP: 54.224.121.223 On: Mon, 18 Apr 2016 23:06:48 Antiviral effect of a-glucosidase inhibitors on viral morphogenesis and binding properties of hepatitis C virus-like particles Cynthia Chapel, 1 Ce ´ line Garcia, 1 Philippe Roingeard, 2 Nicole Zitzmann, 3 Jean Dubuisson, 4 Raymond A. Dwek, 3 Christian Tre ´ po, 1 Fabien Zoulim 1 and David Durantel 1 Correspondence David Durantel durantel@lyon.inserm.fr 1 INSERM U271, Laboratoire des Virus He ´ patiques et Pathologies Associe ´ es, 151 cours Albert Thomas, 69424 Lyon Cedex 03, France 2 INSERM ESPRI 3856, Universite ´ Franc ¸ ois Rabelais, Tours, France 3 Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford, UK 4 CNRS-UPR2511, Institut de Biologie de Lille et Institut Pasteur de Lille, Lille, France Received 6 September 2005 Accepted 29 November 2005 Hepatitis C virus (HCV) infections are a major public-health concern. New antiviral drugs are needed urgently to complement and improve the efficacy of current chemotherapies. The morphogenesis of HCV represents an interesting, and still unexploited, novel molecular target. a-Glucosidase inhibitors derived from the glucose analogue deoxynojirimycin (DNJ) inhibit viral morphogenesis in cellulo via perturbation of the N-glycosylation pathway and hence the misfolding of viral glycoproteins that depend on certain N-glycans for correct folding. Due to the heavy N-glycosylation of HCV glycoproteins, it was hypothesized that such inhibitors would also affect HCV morphogenesis. To study the effect of a-glucosidase inhibitors on viral morphogenesis and binding properties, HCV virus-like particles (VLPs) were produced by using baculovirus loaded with HCV structural-protein genes. Here, it is demonstrated that, in the presence of these a-glucosidase inhibitors, viral glycoproteins synthesized and retained in the endoplasmic reticulum (i) contain unprocessed, triglucosylated N-glycans, (ii) are impaired in their interaction with calnexin and (iii) are at least partially misfolded. Moreover, it is shown that, although the production of VLPs is not affected by a-glucosidase inhibitors, these VLPs contain unprocessed, triglucosylated N-glycans and potentially misfolded glycoproteins. Finally, it is demonstrated that VLPs produced in the presence of a-glucosidase inhibitors have impaired binding properties to hepatoma cells. The inhibitors of morphogenesis studied here target steps of the HCV viral cycle that may prevent or delay viral resistance. These a-glucosidase inhibitors may prove to be useful molecules to fight HCV infection in combination protocols. INTRODUCTION Hepatitis C virus (HCV), a member of the family Flavi- viridae, is a small, positive-sense, enveloped virus of approxi- mately 9600 nt (Choo et al., 1989, 1991). The viral genome, containing a unique open reading frame, is translated into a single polyprotein precursor of 3000 aa that is processed by host and viral proteases (Bartenschlager et al., 2004; Tellinghuisen & Rice, 2002). The amino-terminal part of the genome encodes structural proteins, including the core protein, two envelope glycoproteins (gpE1 and gpE2) and p7, a small ion-channel protein. These proteins are mainly involved in HCV morphogenesis, although some might have other regulatory functions (Bartenschlager et al., 2004; Tellinghuisen & Rice, 2002). gpE1 and gpE2 can either dimerize in a non-covalent interaction in the endoplasmic reticulum (ER) to form potentially functional pre-budding complexes, or derive disulfide-linked aggregates that contain misfolded, non-functional glycoproteins (Dubuisson, 2000). It has been shown that two subsets of host chaperone pro- teins participate in the formation of both of these types of complex. Calnexin interacts with glycoproteins engaged in the formation of functional non-covalent complexes, whilst calreticulin and Bip interact with misfolded glycoproteins destined to aggregate (Dubuisson, 2000). Worldwide, around 170 million people, representing 3 % of the population, have been infected with HCV (Davis et al., 2003; Poynard et al., 2000; Seeff, 2002). More than 70 % of HCV-infected patients become chronic carriers. Long-term exposure to persistent HCV replication predisposes patients to cirrhosis and hepatocellular carcinoma (Liang & Heller, 0008-1503 G 2006 SGM Printed in Great Britain 861 Journal of General Virology (2006), 87, 861–871 DOI 10.1099/vir.0.81503-0