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13:04
A Protocol for Analyzing Hepatitis C Virus Replication
Published on: June 26, 2014
Hepatitis C virus nonstructural protein 5B is involved in virus morphogenesis
Hamed Gouklani1, Rowena A Bull, Claudia Beyer
1The Macfarlane Burnet Institute for Medical Research and Public Health, Melbourne, Victoria, Australia.
Journal of Virology
|February 21, 2012
Summary
Hepatitis C virus (HCV) p7 protein is crucial for virus assembly. Mutations in NS2 and NS5B restored infectivity, revealing essential interactions for HCV morphogenesis.
Area of Science:
- Virology
- Molecular Biology
- Hepatitis C Research
Background:
- The hepatitis C virus (HCV) p7 protein, a viroporin, is essential for virus morphogenesis but not genome replication.
- Understanding the roles of viral proteins in HCV assembly and maturation is critical for developing antiviral strategies.
Purpose of the Study:
- To investigate the role of heterologous genotype 1b (GT1b) p7 in a JFH1-based chimeric HCV genome.
- To identify genetic determinants that restore infectivity to a nonviable chimeric HCV genome.
Main Methods:
- Generation of a JFH1-based chimeric HCV genome encoding GT1b p7.
- Serial passage of transfected cells to recover infectious virus.
- Sequence analysis of recovered virus and forward genetic analysis of mutations.
Main Results:
- A nonviable chimeric HCV genome regained infectivity after serial passage, acquiring mutations T23N in NS2 and K151R in NS5B.
- Each mutation independently restored infectivity, indicating their critical roles in virion assembly/maturation.
- The NS5B K151R mutation did not affect p7 and NS5B colocalization or NS5B polymerase activity.
Conclusions:
- Interactions between HCV p7, NS2, and NS5B are essential for viral morphogenesis.
- HCV nonstructural protein 5B (NS5B) plays a role in virus assembly and maturation, beyond its known polymerase function.
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