microRNA-122 target sites in the hepatitis C virus RNA NS5B coding region and 3′ untranslated region: function in replication and influence of RNA secondary structure

Abstract

We have analyzed the binding of the liver-specific microRNA-122 (miR-122) to three conserved target sites of hepatitis C virus (HCV) RNA, two in the non-structural protein 5B (NS5B) coding region and one in the 3' untranslated region (3'UTR). miR-122 binding efficiency strongly depends on target site accessibility under conditions when the range of flanking sequences available for the formation of local RNA secondary structures changes. Our results indicate that the particular sequence feature that contributes most to the correlation between target site accessibility and binding strength varies between different target sites. This suggests that the dynamics of miRNA/Ago2 binding not only depends on the target site itself but also on flanking sequence context to a considerable extent, in particular in a small viral genome in which strong selection constraints act on coding sequence and overlapping cis-signals and model the accessibility of cis-signals. In full-length genomes, single and combination mutations in the miR-122 target sites reveal that site 5B.2 is positively involved in regulating overall genome replication efficiency, whereas mutation of site 5B.3 showed a weaker effect. Mutation of the 3'UTR site and double or triple mutants showed no significant overall effect on genome replication, whereas in a translation reporter RNA, the 3'UTR target site inhibits translation directed by the HCV 5'UTR. Thus, the miR-122 target sites in the 3'-region of the HCV genome are involved in a complex interplay in regulating different steps of the HCV replication cycle.

Hepatitis C Virus Translation Regulation

“…Cooperative binding of two miR-122 molecules to the two adjacent target sites in the HCV 5 UTR contributes to RNA stability by protecting against cellular nucleases [64,65] and has a positive effect on the efficiency of HCV translation [66][67][68][69][70][71][72]. Although some studies investigated the possible roles of the conserved potential miR-122 binding sites in the NS5B coding region and in the 3 UTR, it is not yet clear if physical binding of miR-122 to these sites results in effector functions [73][74][75][76], leaving some doubts why these sequences are conserved among HCV isolates.…”

miR-29a Promotes Lipid Droplet and Triglyceride Formation in HCV Infection by Inducing Expression of SREBP-1c and CAV1

“…This dual function of miRNAs has been previously observed in several cases, such as the liver-specific miR-122, where it resulted in an increase in HCV viral replication, while it was shown to act as a tumor suppressor miRNA in hepatocellular carcinoma, repressing liver cancer development and progression 48,49. Nonetheless, the contradicting role of miR-182 was also recently reported by our group; specifically, it was shown to promote HCV viral replication, while resulting in a significant augmentation of primary natural killer cell cytotoxicity 47…”

Identification and characterization of microRNA in the lung tissue of pigs with different susceptibilities to PCV2 infection

“…In humans, miR - 122 is reported to enhance the accumulation of HCV RNA by binding to the 5′ UTR of the HCV genome [ 34 ]. Moreover, the 3′ region of the HCV genome targeted by miR - 122 is involved in regulating different steps of the HCV replication cycle [ 35 ]. Our results also indicate that ssc - miR - 122 could decrease PCV2 viral DNA replication and protein synthesis in PK15 cells.…”