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| |  Lipid Droplets Are Necessary for Production of Infectious HCV: Presented at HEP-DART By Barbara J. Rutledge, PhD LAHAINA, HAWAII -- December 17, 2007 -- Association of hepatitis C virus (HCV) proteins around the lipid droplet is crucial for production of infectious HCV, reported in a presentation given here at the HEP-DART 2007 Frontiers in Drug Development for Viral Hepatitis symposium. This finding was presented here on December 12 by Kunitada Shimotohno, PhD, Professor at the Centre for Integrated Medical Research, Keio University, Tokyo, Japan. Lipid droplets are organelles that store neutral lipids and facilitate the transport of lipids and proteins to other organelles, such as endoplasmic reticulum. Earlier work showed that the HCV core protein is associated with lipid droplets, but the function of the lipid droplet in virus assembly has not been defined. To investigate how HCV particles are produced in the cells, Dr. Shimotohno and colleagues transfected cells with a series of genetically engineered variants of JFH-1, an infectious strain of HCV, and analysed the localisation of viral elements. In cells transfected with RNA from wild-type JFH-1, HCV core protein and NS proteins localised to lipid droplets. In addition, NS proteins were diffusely associated with endoplasmic reticulum in cells. Configuration of HCV proteins around the lipid droplet formed a concentric circle, with the first layer consisting of core proteins. Overproduction of lipid droplets was observed in cells transfected with RNA from wild-type JFH-1. "The high prevalence of steatosis in chronic HCV patients might be explained by the high level of lipid droplets in infected cells," commented Dr. Shimotohno. By contrast, in cells transfected with a JFH-1 derivative deleted for core protein sequences, production of lipid droplets was not increased, and NS proteins were not localised to lipid droplets. Electron microscopic analysis showed lipid droplet-associated membranes in wild-type JFH-1-transfected cells with replicating virus, with core protein seen on the edge of the lipid droplet. NS5A proteins were detected around the outside of the lipid droplet. "Taken together, these results clearly indicate that HCV core protein recruits both NS proteins and the HCV replication complex to the lipid droplet-associated membranes," said Dr. Shimotohno. In further analysis, cells were transfected with a JFH-1 derivative producing a variant of NS5A unable to associate with the lipid droplet. HCV genome replication continued, but the replication complex was not recruited to the lipid droplet. Virus-like particles were released into the cell medium, but infectious virus was not produced. "Association of HCV proteins with the lipid droplet is necessary for production of infectious HCV," concluded Dr. Shimotohno. Small molecular-weight compounds that prevent the association of HCV core protein with lipid droplets or that prevent activation of lipid synthesis by HCV core protein might have therapeutic potential as anti-HCV agents. Dr. Shimotohno reports no conflicts of interest.
[Presentation title: Lipid Droplet Is an Important Organelle for Production of Infectious Hepatitis C Virus. Abstract 55]
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