Figure 3.

Therapeutic strategies for miRNA manipulation. (Top left) Schematic of miRNA gene regulation. miRNAs are transcribed in the nucleus as primary transcripts (pri-miRNAs) subject to 2 sequential cleavage steps, the first in the nucleus generating miRNA precursors (premiRNAs) and the second generating mature miRNAs in the cytoplasm. Mature miRNAs are bound to Argonaute family proteins, which assemble into miRISCs that recognize cellular mRNAs by base pairing to imperfectly complementary target sites, generally found in the 3′ UTR. miRISC recruitment results in repression of gene expression through mRNA destabilization and translational inhibition. (Top center) miRNA function may be inhibited in vivo through introduction of short oligonucleotides, chemically modified for enhanced stability and complementary to the mature miRNA sequence. (Top right) Similarly, miRNA mimics may be introduced that are processed by DICER into mature miRNAs that are identical to the endogenous miRNA. In this way, mature miRNA levels can be dramatically enhanced. (Bottom left) miRNA sponges are mRNAs bearing multiple target sites for an miRNA. They reduce miRNA activity by sequestering the mature miRNA. Sponge mRNAs may be employed as a gene therapy agent, introduced via cell-type-specific transfection approaches or viral gene transfer techniques. (Bottom right) In addition to synthetic miRNA mimics, increased miRNA expression can be accomplished via gene therapy approaches, with additional or replacement copies of miRNA genes introduced into the nucleus via viral gene transfer. This approach has the potential advantages of avoiding adverse host immune responses to oligonucleotides and permitting tunable, cell-type-specific expression through the use of appropriate gene promoters.