11.10: siRNA - Small Interfering RNAs
Small interfering RNAs, or siRNAs, are short regulatory RNA molecules that can silence genes post-transcriptionally, as well as the transcriptional level in some cases. siRNAs are important for protecting cells against viral infections and silencing transposable genetic elements.
In the cytoplasm, siRNA is processed from a double-stranded RNA, which comes from either endogenous DNA transcription or exogenous sources like a virus. This double-stranded RNA is then cleaved by the ATP-dependent riboendonuclease, Dicer, into 21-23 nucleotide long fragments with two nucleotide overhangs at both ends. This siRNA is then loaded onto another protein, Argonaute. Argonaute has four different domains – N-terminal, PAZ, Mid, and PIWI. Its PIWI domain has an RNase activity that enables Argonaute to cleave target mRNA. The Argonaute-siRNA complex then binds with a helicase and other proteins to form the RNA induced silencing complex (RISC). In RISC, the sense strand is separated from the antisense, or guide strand, which is thought to be catalyzed by the helicase. The sense strand is degraded in the cytoplasm, and the guide strand directs RISC towards a complementary target mRNA.
The fate of the target mRNA is determined by whether the guide mRNA shows optimal or suboptimal base-pairing with the target mRNA. If the guide strand shows optimal base-pairing with the target mRNA, then the target mRNA is cleaved by Argonaute. The RISC complex then is reused again to target another mRNA. In contrast, if the guide strand shows suboptimal base-pairing with the target mRNA strand, Argonaute will not cleave the mRNA. Instead, it will lead to translational arrest since the RISC complex will obstruct the ribosome binding and translocation. These mRNAs are then directed to the processing bodies (P-bodies) where they are gradually degraded. In the nucleus, siRNA can silence transposable DNA elements and thereby prevent their unwanted and dangerous random insertions in the genome.
As siRNA silences specific genes, it has important applications in both molecular biology research and therapeutic applications. In research, they can be used to study specific gene functions in vivo and in vitro by silencing that gene. They can also be used to silence genes from deadly viruses and can be employed as an effective anti-viral agent. siRNAs are being explored as a potential treatment for several diseases including neurological disorders such as Alzheimer’s and cancers by targeting respective disease-causing genes. The siRNAs can be used in personalized gene therapy as they are highly specific and can be easily designed for different target genes. Also, therapeutic siRNAs are programmed to target mRNA rather than DNA and therefore there is a significantly reduced risk of permanent DNA modification.