RNA interference (RNAi) is a phenomenon in which the introduction of double-stranded RNA (dsRNA) into certain organisms and cell types causes degradation of the homologous mRNA (Figure 1; step 1). In the cell, long dsRNAs are cleaved into short 21-25 nucleotide small interfering RNAs, or siRNAs, by a ribonuclease known as Dicer (step 2). The siRNAs subsequently assemble with protein components into an RNA-induced silencing complex (RISC). An ATP-generated unwinding of the siRNA activates the RISC (step 3), which in turn binds to the homologous transcript by base pairing interactions and cleaves the mRNA (step 4). This sequence specific degradation of mRNA (step 5) results in gene silencing (reviewed in 1-3).
Figure 1. RNAi Mechanism.
RNAi has been used as a tool by scientists to understand gene function in Caenorhabditis elegans and Drosophila. In these organisms, RNAi can be induced by introducing long dsRNA complementary to the target mRNA to be degraded. In mammalian cells and organisms, however, introducing dsRNA longer than 30 bp activates a potent antiviral response. To circumvent this, siRNAs are used to induce RNAi in mammalian cells and organisms. These siRNAs can be prepared by chemical synthesis, in vitro transcription, or digestion of long dsRNA by RNase III or Dicer, and can then be introduced into the cell by transfection, electroporation, or other methods. Alternatively, siRNAs can be expressed within cells after transfection of an siRNA expression vector.