Compensatory mutations in NS3 and NS5A proteins enhance the virus production capability of hepatitis C reporter virus.
In this study, an infectious HCV monocistronic reporter virus was constructed by inserting an EGFP gene into the C-terminus of NS5A in the JFH-1 genome. A robust adaptive mutant, which could produce infectious virions as robustly as the JFH-1 wild type in Huh7.5.1 cells, was subsequently isolated by monitoring EGFP fluorescence. Full genomic sequencing revealed five amino acid substitutions, three located in the helicase domain of NS3 and two positioned in the C-terminus of NS5A. Reverse genetics studies suggested that the NS3 and NS5A mutations acted synergistically to enhance virus production capability possibly by accelerating the virion assembly efficiency but did not affect the replication competence of the adaptive reporter virus. Further analysis revealed that the M260K and T462I substitutions in NS3 and NS5A, respectively, were the key mutations. These adaptive mutations were also effective in the context of the JFH-1 genome.
Han
Qingxia
Q
State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, People's Republic of China.
Xu
Chunlian
C
Wu
Chunchen
C
Zhu
Wenbo
W
Yang
Rongge
R
Chen
Xinwen
X
D019943:Amino Acid Substitution; D002460:Cell Line; D016679:Genome, Viral; D049452:Green Fluorescent Proteins; D016174:Hepacivirus / Q000502:physiology*; D006526:Hepatitis C / Q000821:virology; D006801:Humans; D017354:Point Mutation; D021381:Protein Transport; D012367:RNA, Viral / Q000032:analysis / Q000235:genetics; D017423:Sequence Analysis, RNA; D017361:Viral Nonstructural Proteins / Q000235:genetics* / Q000378:metabolism; D019065:Virus Assembly; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; D012324:RNA-Dependent RNA Polymerase
D049452:Green Fluorescent Proteins; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase