Formation and function of hepatitis C virus replication complexes require residues in the carboxy-terminal domain of NS4B protein.
During replication, hepatitis C virus (HCV) NS4B protein rearranges intracellular membranes to form foci, or the web, the putative site for HCV replication. To understand the role of the C-terminal domain (CTD) in NS4B function, mutations were introduced into NS4B alone or in the context of HCV polyprotein. First, we show that the CTD is required for NS4B-induced web structure, but it is not sufficient to form the web nor is it required for NS4B membrane association. Interestingly, all the mutations introduced into the CTD impeded HCV genome replication, but only two resulted in a disruption of NS4B foci. Further, we found that NS4B interacts with NS3 and NS5A, and that mutations causing NS4B mislocalization have a similar effect on these proteins. Finally, we show that the redistribution of Rab5 to NS4B foci requires an intact CTD, suggesting that Rab5 facilitates NS4B foci formation through interaction with the CTD.
Aligo
Jason
J
Department of Biochemistry and Molecular Biology, The Pennsylvania State University, 308 Althouse Laboratory, University Park, PA 16802, USA.
Jia
Shuaizheng
S
Manna
David
D
Konan
Kouacou V
KV
D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure / Q000821:virology; D004252:DNA Mutational Analysis; D016174:Hepacivirus / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D054884:Host-Pathogen Interactions; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000235:genetics / Q000502:physiology; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D016415:Sequence Alignment; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014779:Virus Replication; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; D012324:RNA-Dependent RNA Polymerase
D050505:Mutant Proteins; D017361:Viral Nonstructural Proteins; D020696:rab5 GTP-Binding Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C080107:NS4B protein, flavivirus; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase
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