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BioArray Backgrounder: How RNAi and siRNA Work


The technology of RNAi triggers a natural gene silencing process that occurs when a virus invades a cell. As many viruses have double-stranded RNA, the cell has a built-in mechanism to attack this RNA: As double-stranded RNA enters the cell, an enzyme called Dicer chops it up into short 21-25 base-pair segments (called siRNA, for short interfering RNA or silencing RNA), which then combine with another group of enzymes to form a complex called RISC, or RNA-induced silencing complex. This complex unwinds the short double-stranded siRNAs into single strands, then uses them in its hunt for RNA which is complementary to these strands (potential viral RNA). It can copy these bait siRNAs numerous times, using RNA polymerases. When these single strands hybridize to a complementary target mRNA, one of RISC’s enzymes chops this sequence up into meaningless bits.

In 1998, Andy Fire of the Carnegie Institution and Craig Mello of the University of Massachusetts discovered that they could trigger this silencing process for a select strand of target mRNA by introducing a double-stranded RNA of a complementary sequence into the cell of a model organism. In organisms such as C. elegans and Drosophila, introducing this RNAi completely or partially shuts down the expression of the target gene, and the resulting phenotypic changes reveal the function of that gene. While this process does not work the same way for mammals, in 2001, Tom Tuschl from the Max Planck Institute for Biophysical Chemistry demonstrated that introducing siRNAs of 21 base pairs, with two-nucleotide overhangs, to human cells could silence genes in the cell. (Philip Zamore of the University of Massachusetts, Philip Sharp and David Bartel of the Whitehead Institute also share credit for experiments that led to this discovery. See US Patent Application 20020086356, and WIPO application WO 0244321.) For the full story on RNAi, see the cover article in the May 2003 issue of Genome Technology magazine.



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