Gene Specific
Silencing by RNAi
RNA interference (RNAi) is a phenomenon
in which the introduction of double-stranded RNA (dsRNA) into
a diverse range of organisms and cell types causes degradation
of the complementary mRNA (Figure 1; step 1). In the cell, long
dsRNAs are cleaved into short 21-25 nucleotide small interfering
RNAs, or siRNAs, by a ribonuclease known as Dicer (step 2). The
siRNAs subsequently assemble with protein components
into an RNA-induced silencing complex (RISC), unwinding in the
process (step 3). Activated RISC then binds to complementary
transcript by base pairing interactions between the siRNA antisense
strand and the mRNA. The bound mRNA is cleaved (step 4) and sequence
specific degradation of mRNA (step 5) results in gene silencing
(reviewed in 1-3).
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| Figure 1. RNAi
Mechanism. |
RNAi has been used as a tool by scientists
to understand gene function in Caenorhabditis elegans and Drosophila.
In these organisms, RNAi can be induced by introducing long
dsRNA complementary to the target mRNA to be degraded. In
mammalian cells and organisms, however, introducing dsRNA
longer than 30 bp activates a potent antiviral response.
To circumvent this, siRNAs are used to induce RNAi in mammalian
cells and organisms.
In the last few years, siRNAs have been
used in a number of different experimental settings to silence
gene expression. In some, chemically synthesized or in vitro
transcribed siRNAs have been transfected into cells, injected
into mice, or introduced into plants (e.g. by a particle
gun). In others, siRNAs have been expressed endogenously
from siRNA expression vectors or PCR products in cells or
in transgenic animals.
In addition to their role in gene silencing,
siRNAs have been determined to play diverse biological functions
in vivo -- roles that include antiviral defense, transposon
silencing, gene regulation, centromeric silencing, and genomic
rearrangements. This functional diversity has exemplified
the importance of siRNAs within cells and has also stirred
interest in their detection across species and tissues.
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