BACKGROUND & AIMS: The process and the site of hepatitis C virus (HCV) particle formation in cells after infection remain unknown. The aim of this study was to create an in vitro model for the study of HCV particle formation. METHODS: HeLa G cells were transfected with the full-length sequence of the HCV genome. Viral protein expression was analyzed using immunoblotting. The cells were examined using immunoelectron and conventional electron microscopy. RESULTS: Core, E2, NS3, NS5a, and NS5b proteins were identified using immunoblotting. Immunoelectron microscopy showed that the core antigen was located along the membrane of the endoplasmic reticulum (ER) and occasionally in its cisternae. Core antigen-positive particles of 30 nm in diameter were found in the cytosol and in the cisternae of the ER. The particles in the cisternae were coated with an outer membrane that was connected to the ER membrane. Conventional electron microscopy revealed particles of 45 nm in diameter with electron-dense cores in the cisternae of the ER. The outer membrane of the particles was occasionally connected to the ER membrane. CONCLUSIONS: The findings suggest that HCV core proteins are synthesized and assembled into particles in the cytosol and that they bud into the cisternae of the ER to form coated particles.
BACKGROUND & AIMS: The process and the site of hepatitis C virus (HCV) particle formation in cells after infection remain unknown. The aim of this study was to create an in vitro model for the study of HCV particle formation. METHODS: HeLa G cells were transfected with the full-length sequence of the HCV genome. Viral protein expression was analyzed using immunoblotting. The cells were examined using immunoelectron and conventional electron microscopy. RESULTS: Core, E2, NS3, NS5a, and NS5b proteins were identified using immunoblotting. Immunoelectron microscopy showed that the core antigen was located along the membrane of the endoplasmic reticulum (ER) and occasionally in its cisternae. Core antigen-positive particles of 30 nm in diameter were found in the cytosol and in the cisternae of the ER. The particles in the cisternae were coated with an outer membrane that was connected to the ER membrane. Conventional electron microscopy revealed particles of 45 nm in diameter with electron-dense cores in the cisternae of the ER. The outer membrane of the particles was occasionally connected to the ER membrane. CONCLUSIONS: The findings suggest that HCV core proteins are synthesized and assembled into particles in the cytosol and that they bud into the cisternae of the ER to form coated particles.
Authors: Pablo Gastaminza; Kelly A Dryden; Bryan Boyd; Malcolm R Wood; Mansun Law; Mark Yeager; Francis V Chisari Journal: J Virol Date: 2010-08-04 Impact factor: 5.103
Authors: Pablo Gastaminza; Guofeng Cheng; Stefan Wieland; Jin Zhong; Wei Liao; Francis V Chisari Journal: J Virol Date: 2007-12-12 Impact factor: 5.103