RNA Interference

Three classes of small RNAs accomplish RNA interference: microRNAs (miRNAs) are encoded by a cell's genome; small interfering RNAs (siRNAs) are derived from exogenous viral double-stranded RNAs; and PIWI-interacting RNAs (piRNAs) are specific to the germline. Each class targets complementary mRNA sequences to prevent protein translation.

Dicer, an enzyme in the RNAi pathway, cleaves double-stranded RNA into approximately 20-25 nucleotide-long siRNA fragments. These shorter siRNAs then bind to the RNA-induced silencing complex (RISC), where the guide strand is retained and used to target complementary mRNA sequences for silencing.

The guide strand is the siRNA strand that remains attached to RISC after the complementary strand is removed. It binds to its complementary sequence in mRNA through base pairing, bringing the RISC complex to the target mRNA. This binding is highly specific because the siRNA is designed to be completely complementary to the targeted mRNA.

Argonaute is an enzyme within the RNA-induced silencing complex that cleaves and degrades the target mRNA after the guide strand binds to it. This cleavage prevents the mRNA from being translated into protein, effectively silencing the targeted gene and blocking its expression.

Short hairpin RNA (shRNA) is a single RNA strand folded over to create a double-stranded structure with a hairpin loop. It serves as a precursor to siRNA. Once introduced into cells, RNase enzymes like Dicer cleave shRNA to produce the shorter siRNA, removing the hairpin loop and generating the active form for RISC binding.

Researchers can synthesize double-stranded RNA with sequences complementary to disease-causing genes and introduce them into cells via injection or viral vectors. This allows targeted suppression of overactive genes in conditions like cancer. The synthetic RNAs hijack the cell's natural RNAi mechanism to silence specific genes without affecting others.

After siRNA binds to the RNA-induced silencing complex, the two strands separate. The complementary strand floats away and is discarded, while the guide strand remains attached to RISC. Only the guide strand participates in targeting and binding the mRNA, making it the functional component of the silencing complex.