title: Proteomics of HCV virions reveals an essential role for the nucleoporin Nup98 in virus morphogenesis.
abstract: Hepatitis C virus (HCV) is a unique enveloped virus that assembles as a hybrid lipoviral particle by tightly interacting with host lipoproteins. As a result, HCV virions display a characteristic low buoyant density and a deceiving coat, with host-derived apolipoproteins masking viral epitopes. We previously described methods to produce high-titer preparations of HCV particles with tagged envelope glycoproteins that enabled ultrastructural analysis of affinity-purified virions. Here, we performed proteomics studies of HCV isolated from culture media of infected hepatoma cells to define viral and host-encoded proteins associated with mature virions. Using two different affinity purification protocols, we detected four viral and 46 human cellular proteins specifically copurifying with extracellular HCV virions. We determined the C terminus of the mature capsid protein and reproducibly detected low levels of the viral nonstructural protein, NS3. Functional characterization of virion-associated host factors by RNAi identified cellular proteins with either proviral or antiviral roles. In particular, we discovered a novel interaction between HCV capsid protein and the nucleoporin Nup98 at cytosolic lipid droplets that is important for HCV propagation. These results provide the first comprehensive view to our knowledge of the protein composition of HCV and new insights into the complex virus-host interactions underlying HCV infection.
authors.affiliation: Department of Infectious Diseases, King's College London, London SE1 9RT, United Kingdom;
authors.forename: Marion
authors.initials: M
authors.lastname: Lussignol
authors.affiliation: Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065;
authors.forename: Martina
authors.initials: M
authors.lastname: Kopp
authors.affiliation: Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY 10065;
authors.forename: Kelly
authors.initials: K
authors.lastname: Molloy
authors.affiliation: Centre for Ultrastructural Imaging, King's College London, London SE1 9RT, United Kingdom;
authors.forename: Gema
authors.initials: G
authors.lastname: Vizcay-Barrena
authors.affiliation: Centre for Ultrastructural Imaging, King's College London, London SE1 9RT, United Kingdom;
authors.forename: Roland A
authors.initials: RA
authors.lastname: Fleck
authors.affiliation: Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065; Section of Virology, Imperial College London, London W2 1PG, United Kingdom.
authors.forename: Marcus
authors.identifier: http://orcid.org/0000-0003-0321-4226
authors.initials: M
authors.lastname: Dorner
authors.affiliation: Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065;
authors.forename: Kierstin L
authors.initials: KL
authors.lastname: Bell
authors.affiliation: Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY 10065;
authors.forename: Brian T
authors.initials: BT
authors.lastname: Chait
authors.affiliation: Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065; maria.catanese@kcl.ac.uk ricec@mail.rockefeller.edu.
authors.forename: Charles M
authors.initials: CM
authors.lastname: Rice
authors.affiliation: Department of Infectious Diseases, King's College London, London SE1 9RT, United Kingdom; Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065; maria.catanese@kcl.ac.uk ricec@mail.rockefeller.edu.
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Catanese
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D016174:Hepacivirus / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009024:Morphogenesis; D028861:Nuclear Pore Complex Proteins / Q000502:physiology*; D040901:Proteomics; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*
chemical_list: D028861:Nuclear Pore Complex Proteins; D014764:Viral Proteins; C093350:nuclear pore complex protein 98
keywords: Flaviviridae; enveloped virus; nucleoporin; virion proteomics; virus–host interactions