Unconventional miR-122 binding stabilizes the HCV genome by forming a trimolecular RNA structure

MicroRNAs (miRNAs) typically downregulate protein expression from target mRNAs through limited base-pairing interactions between the 5′ ‘seed’ region of the miRNA and the mRNA 3′ untranslated region (3′UTR). In contrast to this established mode of action, the liver-specific human miR-122 binds at two sites within the hepatitis C viral (HCV) 5′UTR, leading to increased production of infectious virions. We show here that two copies of miR-122 interact with the HCV 5′UTR at partially overlapping positions near the 5′ end of the viral transcript to form a stable ternary complex. Both miR-122 binding sites involve extensive base pairing outside of the seed sequence; yet, they have substantially different interaction affinities. Structural probing reveals changes in the architecture of the HCV 5′UTR that occur on interaction with miR-122. In contrast to previous reports, however, results using both the recombinant cytoplasmic exonuclease Xrn1 and liver cell extracts show that miR-122-mediated protection of the HCV RNA from degradation does not correlate with stimulation of viral propagation in vivo. Thus, the miR-122:HCV ternary complex likely functions at other steps critical to the viral life cycle.