Abstract
It has been suggested that cellular proteins are involved in hepatitis C virus (HCV) RNA replication. By using the yeast two-hybrid system, we isolated seven cDNA clones encoding proteins interacting with HCV RNA polymerase (NS5B) from a human liver cDNA library. For one of these, alpha-actinin, we confirmed the interaction by coimmunoprecipitation, immunofluorescent staining and confocal microscopic analysis. Experiments with deletion mutants showed that domains NS5B(84-95), NS5B(466-478), and alpha-actinin(621-733) are responsible for the interaction. Studies of the HCV subgenomic replicon system with small interference RNA indicate that alpha-actinin is essential for HCV RNA replication. Our results suggest alpha-actinin may be a component of the HCV replication complex.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Actinin / chemistry
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Actinin / genetics
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Actinin / metabolism*
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Animals
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COS Cells
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Cell Line, Tumor
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Chlorocebus aethiops
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DNA-Directed RNA Polymerases / chemistry
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DNA-Directed RNA Polymerases / genetics
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DNA-Directed RNA Polymerases / metabolism*
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Fluorescent Antibody Technique
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Gene Expression Regulation / drug effects
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Gene Expression Regulation / physiology
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Gene Library
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Hepacivirus / genetics
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Hepacivirus / metabolism*
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Hepacivirus / ultrastructure
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Humans
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Microscopy, Confocal
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Precipitin Tests
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Protein Structure, Tertiary
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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RNA, Small Interfering / pharmacology
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RNA, Viral / biosynthesis
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RNA, Viral / genetics
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RNA, Viral / metabolism
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Replicon / genetics
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Sequence Deletion
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Two-Hybrid System Techniques
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Virus Replication / drug effects
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Virus Replication / physiology
Substances
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RNA, Small Interfering
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RNA, Viral
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Actinin
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DNA-Directed RNA Polymerases