Specificity and functionality of microRNA inhibitors

In this paper, we present a first systematic study to evaluate the effect of mismatches in the target site on synthetic inhibitor activity. Panels of miRNA inhibitors containing two-nucleotide mismatches across the target site were tested against three miRNAs (miR-21, miR-22 and miR-122). The results showed that the function of inhibitors vary as mismatch positions in the inhibitors change.The data indicate that features important for natural miRNA target recognition (such as seed region complementarity) are also important for inhibitor functionality. In addition, base pairing at a second, more 3' region appears to be equally important in determining the efficacy of synthetic inhibitors. Considering the importance of these inhibitor regions and the expression of closely related miRNA sequences will enable researchers to interpret results more accurately in future experiments.Micro (mi)RNAs are small (17 to 27 nucleotides), non-coding RNAs that act in association with Argonaute (Ago) proteins to modulate gene expression via an effector nucleic acid-protein complex (microribonucleoprotein (RNP) or miRNA-induced silencing complex (RISC)). In animals, miRNA-based gene modulation occurs predominantly by the mature miRNA binding to an mRNA target site through partial base pairing, resulting in translational attenuation (for recent reviews, see [1-6]). Computational and experimental techniques for identifying target sites [7-10] have found that complementarity to the seed region (nucleotide positions 2 to 7 or 2 to 8 of the mature miRNA) is often an important determinant of target sites. In some cases of incomplete seed-pairing, pairing at '3'-compensatory' sites of the mature miRNA creates a functional target site [11,12]. The large number of potential target sites per miRNA, combined with the hundreds of putative miRNAs, has led to the prediction that a large fraction of human genes could be modulated by miRNAs.The functional roles of miRNAs can be investigated using inh