Lentiviral vector-mediated expression of pre-miRNAs and antagomiRs

Abstract

Micro (mi)RNAs are highly conserved small regulatory RNAs, which regulate gene expression by hybridization to specific binding sites in the 3'untranslated region (UTR) of many mRNAs. Upon miRNA-guided recruitment of a multiprotein complex, target mRNAs are either degraded or their translation is blocked depending on the complementarity between the miRNAs and their binding sites in target mRNAs. Individual miRNAs have been shown to regulate the expression of hundreds of genes with corresponding miRNA binding sites in the 3'UTR in a dose-dependent manner. Although miRNA-target genes may be predicted by bioinformatic tools, each potential target needs to be confirmed experimentally. We describe here the expression of individual miRNAs or miRNA-specific antagomiRs by lentiviral gene transfer to induce stable gain- and loss-of-function phenotypes. These techniques provide some tools to analyze miRNA function in cell culture or animal models.