RNA interference (RNAi) is a natural process in cells to silence unwanted or harmful genes. The initial discovery of this phenomenon goes back to 1991 by scientists who were trying to deepen the colour of petunias. Surprisingly, by introducing a gene for colour, they turned the gene for colour off.
Several years after the petunia experiments, the mechanism of RNA interference was revealed: it is triggered by double-stranded RNA (dsRNA), not usually found in healthy cells, but needed to turn genes off, if the cell is threatened or damaged by invading viruses.
In natural RNA interference, dsRNA in the cell’s cytoplasm is cut by an enzyme called Dicer into double stranded small interfering RNA (siRNA) molecules which are 20-25 nucleotides long. This siRNA binds to an RNA-Induced Silencing Complex (RISC) which separates the two strands into the passenger and guide strand. The passenger strand is degraded while the RISC takes the guide strand to a specific mRNA site, cleaving it so that the unwanted target protein isn’t made. This is how the gene is ‘silenced’.
These are the stages of the process as it pertains to disease control:
If the level of mRNA and the protein in the cell both decrease by at least 70%, it is likely that a successful gene silencing molecule has been developed. However, delivery to the target cells and organs can be difficult.
The RNAi pathway is initiated when dsRNA enters the cytoplasm. The source of the dsRNA may be:
In the synthetic approach, the molecule is introduced into the cytoplasm of the cell where it is usually quickly degraded. To gain effects lasting longer than a few days, repeat ongoing administration is needed.
In the ddRNAi approach, the shRNA is produced by the cell itself, as a result of the operation of the DNA construct which is delivered to the nucleus, and becomes part of the cell’s own DNA. The gene silencing effect can last up to years, because the cell continues to manufacture its own shRNA, which enters the RNAi pathway.