fig3

Figure 3. Mechanism of miRNA, siRNA, and shRNA action. The Center (blue arrows) represents the canonical miRNA maturation and mechanism of action pathway. Transcription leads to a pri-miRNA, which folds into a small hairpin RNA with one or several mismatches. The pri-miRNA is processed by Drosha and DGCR8 to a pre-miRNA. After association to exportin 5, the pre-miRNA is transported through the nuclear envelope by a process dependent upon the small GTPase ran-GTP. The pre-miRNA is finally processed by the Dicer endoribonuclease (a class III RNase), which deletes the hairpin loop. This generates an RNA duplex made of a passenger and a guide strand. After miRNA homoduplex association with the RISC complex composed of protein TRBP and the argonaute 2 nuclease, the passenger strand is eliminated. The RISC/guide strand complex binds to the target mRNA, in the 5’ non-translated region, inducing a steric blocking of ribosome entry and translation. In some instances, miRNA binding can also lead to mRNA cleavage and degradation, but the mismatch generally make this less likely. Left side: siRNA mechanism. (green arrows), a double-strand siRNA with no mismatch is administered to the cells. All action takes place in the cytosol. After siRNA binding to the RISC complex composed of protein TRBP and the argonaute 2 nuclease, the passenger strand is eliminated. The RISC/guide strand complex binds to the target mRNA. The target sequence is generally chosen within the translated region. Because of the complete siRNA matching with the mRNA target sequence, argonaute 2 is now able to cleave the target mRNA with great efficiency. After mRNA cleavage and elimination, the RISC/guide siRNA complex can bind and cleave another mRNA (grey arrow). Right side: (green arrows): shRNA mechanism. A gene expression cassette coding a small hairpin RNA is administered either as a plasmid or by a viral gene delivery vector. The shRNA transcript is then processed as for the miRNA. However, no mismatch is introduced, which leads to fully efficient mRNA degradation and silencing.