Differential hepatitis C virus RNA target site selection and host factor activities of naturally occurring miR-122 3' variants

Abstract

In addition to suppressing cellular gene expression, certain miRNAs potently facilitate replication of specific positive-strand RNA viruses. miR-122, a proviral hepatitis C virus (HCV) host factor, binds and recruits Ago2 to tandem sites (S1 and S2) near the 5' end of the HCV genome, stabilizing it and promoting its synthesis. HCV target site selection follows canonical miRNA rules, but how non-templated 3' miR-122 modifications impact this unconventional miRNA action is unknown. High-throughput sequencing revealed that a 22 nt miRNA with 3'G ('22-3'G') comprised <63% of total miR-122 in human liver, whereas other variants (23-3'A, 23-3'U, 21-3'U) represented 11-17%. All loaded equivalently into Ago2, and when tested individually functioned comparably in suppressing gene expression. In contrast, 23-3'A and 23-3'U were more active than 22- 3'G in stabilizing HCV RNA and promoting its replication, whereas 21-3'U was almost completely inactive. This lack of 21-3'U HCV host factor activity correlated with reduced recruitment of Ago2 to the HCV S1 site. Additional experiments demonstrated strong preference for guanosine at nt 22 of miR-122. Our findings reveal the importance of non-templated 3' miR-122 modifications to its HCV host factor activity, and identify unexpected differences in miRNA requirements for host gene suppression versus RNA virus replication.