RNA interference (RNAi) is a process in which a small non-coding RNA molecule blocks the post-transcriptional expression of a gene by binding to its messenger RNA (mRNA) and preventing the protein from being translated.
This process occurs naturally in cells, often through the activity of genomically-encoded microRNAs. Researchers can take advantage of this mechanism by introducing synthetic RNAs to deactivate specific genes for research or therapeutic purposes. For example, RNAi could be used to suppress genes that are overactive in diseases such as cancer.
First, researchers synthesize double-stranded RNA with a sequence complementary to the targeted gene. Different types of double-stranded RNA can be used, including small interfering RNA (siRNA) and short hairpin RNA (shRNA). shRNA is one strand of RNA that is folded over—creating a double-stranded RNA with a hairpin loop on one side—and is a precursor of siRNA. The double-stranded RNA is then introduced into cells by methods such as injection or delivery by vectors, such as modified viruses. If shRNA is used, RNase enzymes in the cell, such as Dicer, cleave it down to the shorter siRNA, removing the hairpin loop.
The siRNA then binds to an enzyme Argonaute, which is part of a complex called the RNA-induced silencing complex (RISC). Here, the two strands of the siRNA separate. One floats away while the other—called the guide strand—remains attached to the RISC. It is known as the "guide strand" because it is the strand that binds the mRNA through complementary base pairing, bringing the RISC to the mRNA. This binding is very specific because the siRNA is usually designed to be completely complementary to the targeted mRNA. Argonaute then cleaves and degrades the mRNA, preventing it from being translated into protein— effectively silencing the gene.
Copyright © 2024 MyJoVE Corporation. All rights reserved