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Virology. 2011 Feb 20;410(2):336-44. doi: 10.1016/j.virol.2010.11.015. Epub 2010 Dec 24.

Competing roles of microRNA-122 recognition elements in hepatitis C virus RNA.

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1
Steacie Institute for Molecular Sciences, National Research Council of Canada, 100 Sussex Drive, Ottawa, Canada K1A 0R6.

Abstract

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.

PMID:
21185047
DOI:
10.1016/j.virol.2010.11.015
[Indexed for MEDLINE]
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