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1 Centre for Infectious Diseases, University of Edinburgh, Summerhall, Edinburgh EH9 1QH, Scotland, UK
2 Department of Virology, University of Glasgow, Church Street, Glasgow G11 5JR, Scotland, UK
Correspondence
P. Simmonds
Peter.Simmonds{at}ed.ac.uk
There is accumulating evidence from bioinformatic studies that hepatitis C virus (HCV) possesses extensive RNA secondary structure in the core and NS5B-encoding regions of the genome. Recent functional studies have defined one such stem–loop structure in the NS5B region as an essential cis-acting replication element (CRE). A program was developed (STRUCTUR_DIST) that analyses multiple rna-folding patterns predicted by MFOLD to determine the evolutionary conservation of predicted stem–loop structures and, by a new method, to analyse frequencies of covariant sites in predicted RNA folding between HCV genotypes. These novel bioinformatic methods have been combined with enzymic mapping of RNA transcripts from the core and NS5B regions to precisely delineate the RNA structures that are present in these genomic regions. Together, these methods predict the existence of multiple, often juxtaposed stem–loops that are found in all HCV genotypes throughout both regions, as well as several strikingly conserved single-stranded regions, one of which coincides with a region of the genome to which ribosomal access is required for translation initiation. Despite the existence of marked sequence conservation between genotypes in the HCV CRE and single-stranded regions, there was no evidence for comparable suppression of variability at either synonymous or non-synonymous sites in the other predicted stem–loop structures. The configuration and genetic variability of many of these other NS5B and core structures is perhaps more consistent with their involvement in genome-scale ordered RNA structure, a structural configuration of the genomes of many positive-stranded RNA viruses that is associated with host persistence.
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