Journal
VIROLOGY
Volume 410, Issue 2, Pages 336-344Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.virol.2010.11.015
Keywords
MicroRNA; Hepatitis C; Cooperativity; Translation; RNA replication; Subgenomic replicon; Antiviral immunity; Hybridization kinetics
Categories
Funding
- Canadian Institutes of Health Research (CIHR)
- Natural Sciences and Engineering Research Council of Canada (NSERC)
- National CIHR
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MicroRNA-122 positively modulates hepatitis C virus (HCV) through direct interactions with viral RNA. Three microRNA-122 recognition elements (MREs) have been previously identified: two in the 5'UTR and one in the 3'UTR. Herein, we report the relative affinity of microRNA-122 to these sites using viral RNA-coated magnetic beads, with mutagenesis and probes to disrupt interactions of microRNA-122 at specific sites. We demonstrate cooperativity in binding between the closely spaced MREs within the 5'UTR in vitro. We also identified a well conserved fourth site in the coding region and showed that it is the highest affinity MRE site. Site-directed mutagenesis of the MREs in HCV subgenomic replicons expressed in Huh-7.5 cells demonstrated competing roles of the stimulatory MREs in the 5'UTR with the inhibitory role of an MRE in the open reading frame (ORF). These data have important implications in elucidating the mechanism of interaction between microRNA-122 and HCV RNA. Crown Copyright (C) 2010 Published by Elsevier Inc. All rights reserved.
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