Hepatitis C virus (HCV) is a positive strand RNA virus that propagates primarily in the liver. We show here that the liver-specific microRNA-122 (miR-122), a member of a class of small cellular RNAs that mediate posttranscriptional gene regulation usually by repressing the translation of mRNAs through interaction with their 3 0 -untranslated regions (UTRs), stimulates the translation of HCV. Sequestration of miR-122 in liver cell lines strongly reduces HCV translation, whereas addition of miR-122 stimulates HCV translation in liver cell lines as well as in the non-liver HeLa cells and in rabbit reticulocyte lysate. The stimulation is conferred by direct interaction of miR-122 with two target sites in the 5 0 -UTR of the HCV genome. With a replication-defective NS5B polymerase mutant genome, we show that the translation stimulation is independent of viral RNA synthesis. miR-122 stimulates HCV translation by enhancing the association of ribosomes with the viral RNA at an early initiation stage. In conclusion, the liver-specific miR-122 may contribute to HCV liver tropism at the level of translation.

Certain viral and cellular mRNAs initiate translation capindependently at internal ribosome entry site (IRES) elements. Picornavirus IRES elements are widely used in dicistronic or multicistronic vectors in gene therapy, virus replicon systems, and analysis of IRES function. In such vectors, expression of the upstream gene often serves as internal control to standardize the readings of IRES-driven downstream reporter activity. Picornaviral IRES elements translate optimally at up to 120 mM K ؉ concentration, whereas genes used as upstream reporters usually have lower salt optima when present in monocistronic mRNAs. However, here we show that such reporter genes are efficiently translated at higher K ؉ concentrations when placed upstream of a functional picornavirus IRES. This translation enhancement occurs in cis, is independent of the nature of the first reporter and of second reporter translation, and is conferred by the IRESs of picornaviruses but not of hepatitis C virus. A defective picornavirus IRES with a deletion killing IRES activity but leaving the binding site for initiation factor eIF4G intact retains translation enhancement activity. Translation enhancement on a capped mRNA is disabled by m 7 GDP. In addition, the C-terminal fragment of eIF4G can confer translation enhancement also on uncapped mRNA. We conclude that whenever eIF4F has been captured to a dicistronic mRNA by binding to a picornavirus IRES via its eIF4G moiety, it can be provided in cis to the 5-end of the RNA and there stimulate translation initiation, either by binding to the cap nucleotide using its eIF4E moiety or by binding to the RNA cap-independently using its eIF4G moiety. Internal ribosome entry site (IRES)3 elements are cis-acting RNA regions that confer the internal entry of ribosomes to the RNA translation start site independent of the process of capdependent scanning from the 5Ј-end of the RNA (1, 2). Such IRES elements were first discovered in picornaviruses like encephalomyocarditis virus (EMCV) (3), poliovirus (4), and foot-and-mouth disease virus (FMDV) (5). Also the translation of hepatitis C virus (HCV) (6) and of several cellular mRNAs is driven by IRES elements (2).The picornaviral IRES elements are classified in three groups according to their sequences and secondary structures, the type I elements of the entero-/rhinovirus group (including poliovirus), the type II elements of the cardio-/aphthovirus group (including EMCV and FMDV), and the type III element of hepatitis A virus (1). After infection of a susceptible cell, these IRES elements guide the small ribosomal subunit to an AUG triplet in a starting window at the 3Ј-border of the IRES (1, 2, 7). Ribosome binding to the picornavirus IRES elements is mediated by a number of cellular RNA-binding proteins (8) that fall into two groups. On one hand, all standard eukaryotic translation initiation factors (eIFs) are required (9), except the actual cap-binding protein eIF4E, a protein that associates with the large adaptor protein eIF4G (and the RNA-helicase eIF4A) i...