Newly synthesized hepatitis C virus replicon RNA is protected from nuclease activity by a protease-sensitive factor(s).
Biochemical characterization of hepatitis C virus (HCV) replication using purified, membrane-associated replication complexes is hampered by the presence of endogenous nuclease activity that copurifies with the replication complex. In this study, pulse-chase analyses were used to demonstrate that newly synthesized replicon RNA was protected from nuclease activity by a factor(s) that was sensitive to 0.5% NP-40 or protease treatment. Nuclease susceptibility was not related to disruption of lipid membranes, since NP-40 did not significantly affect the buoyant density of HCV replication complexes or protease susceptibility of HCV NS3 and NS5A proteins. These results suggest that a protease-sensitive factor(s) protects newly synthesized RNA from nuclease degradation.
ViroPharma Incorporated, Exton, Pennsylvania, USA.
Guang
G
Yang
Daniel C
DC
Pevear
Marc S
MS
Collett
Srinivas
S
Chunduru
Dorothy C
DC
Young
Christopher
C
Benetatos
Robert
R
Jordan
D002460:Cell Line; D010450:Endopeptidases / Q000378:metabolism; D016174:Hepacivirus / Q000235:genetics* / Q000502:physiology; D006801:Humans; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D012367:RNA, Viral / Q000235:genetics* / Q000378:metabolism*; D012093:Replicon; D012260:Ribonucleases / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000378:metabolism; D014779:Virus Replication; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; D012324:RNA-Dependent RNA Polymerase
D010450:Endopeptidases; D017830:Octoxynol; D011092:Polyethylene Glycols; D012367:RNA, Viral; D012260:Ribonucleases; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C010615:Nonidet P-40; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase