Genetic Analysis of Sequences in the 3′ Nontranslated Region of Hepatitis C Virus That Are Important for RNA Replication
- 1 June 2002
- journal article
- Published by American Society for Microbiology in Journal of Virology
- Vol. 76 (11), 5326-38
- https://doi.org/10.1128/jvi.76.11.5326-5338.2002
Abstract
The genome of the hepatitis C virus (HCV) is a plus-strand RNA molecule that carries a single long open reading frame. It is flanked at either end by highly conserved nontranslated regions (NTRs) that mediate crucial steps in the viral life cycle. The 3′ NTR of HCV has a tripartite structure composed of an about 40-nucleotide variable region, a poly(U/UC) tract that has a heterogeneous length, and a highly conserved 98-nucleotide 3′-terminal sequence designated the X tail or 3′X. Conflicting data as to the role the sequences in the 3′ NTR play in RNA replication have been reported. By using the HCV replicon system, which is based on the self-replication of subgenomic HCV RNAs in human hepatoma cell line Huh-7, we mapped in this study the sequences in the 3′ NTR required for RNA replication. We found that a mutant with a complete deletion of the variable region is viable but that replication is reduced significantly. Only replicons in which the poly(U/UC) tract was replaced by a homouridine stretch of at least 26 nucleotides were able to replicate, whereas RNAs with homopolymeric guanine, adenine, or cytosine sequences were inactive. Deletions of individual or all stem-loop structures in 3′X were not tolerated, demonstrating that this region is most crucial for efficient RNA replication. Finally, we found that none of these deletions or substitutions within the 3′ NTR affected RNA stability or translation, demonstrating that the primary effect of the mutations was on RNA replication. These data represent the first detailed mapping of sequences in the 3′ NTR assumed to act as a promoter for initiation of minus-strand RNA synthesis.This publication has 47 references indexed in Scilit:
- Sequences in the 5′ Nontranslated Region of Hepatitis C Virus Required for RNA ReplicationJournal of Virology, 2001
- Heterogeneous Nuclear Ribonucleoprotein A1 Binds to the 3′-Untranslated Region and Mediates Potential 5′-3′-End Cross Talks of Mouse Hepatitis Virus RNAJournal of Virology, 2001
- Specific Interaction of Hepatitis C Virus Protease/Helicase NS3 with the 3′-Terminal Sequences of Viral Positive- and Negative-Strand RNAJournal of Virology, 2001
- Hepatitis C Virus 3′X Region Interacts with Human Ribosomal ProteinsJournal of Virology, 2001
- The effects of the conserved extreme 3′ end sequence of hepatitis C virus (HCV) RNA on the in vitro stabilization and translation of the HCV RNA genomeJournal of Hepatology, 2000
- The effects of the conserved extreme 3' end sequence of hepatitis C virus (HCV) RNA on the in vitro stabilization and translation of the HCV RNA genomeJournal of Hepatology, 2000
- Heterogeneous Nuclear Ribonucleoprotein I (hnRNP-I/PTB) Selectively Binds the Conserved 3′ Terminus of Hepatitis C Viral RNABiochemical and Biophysical Research Communications, 1999
- Characterization of RNA Binding Activity and RNA Helicase Activity of the Hepatitis C Virus NS3 ProteinBiochemical and Biophysical Research Communications, 1996
- Poly(U) binding activity of hepatitis C virus NS3 protein, a putative RNA helicaseFEBS Letters, 1995
- A Novel Sequence Found at the 3′-Terminus of Hepatitis C Virus GenomeBiochemical and Biophysical Research Communications, 1995