IN PLANTS AND FUNGI
RNAi was actually first
observed in plants, where it was named posttranscriptional gene silencing
(PTGS). The phenomenon was first noted when some early transgenic experiments
in plants gave strange results. When an extra copy of a gene was inserted to
increase production of a particular protein, both the inserted gene (i.e., the
transgene) and the resident gene copy were silenced. The result was a plant
that made less of the target protein rather than more.
For example, in 1990,
researchers inserted a gene to make petunia flowers a darker purple. Instead,
the plant made white flowers. Both the transgene and the endogenous gene were
suppressed, leaving the flower without any pigment. A similar phenomenon was
seen in Neurospora, where it was called quelling. After the discovery of RNAi
in Caenorhabditis elegans, it was recognized that RNAi, PTGS, and quelling all
operate via the same mechanism. Initially, none of these processes affected the
level of transcription. There was plenty of mRNA produced from the transgene.
After some time, the mRNA for the transgene was found in two fragments,
suggesting an endonuclease cleaved it in two. Later, the target mRNA was found
in smaller and smaller fragments, suggesting that exonucleases were digesting
the large mRNA segments. Finally, the genes were converted into
How does an extra copy of a
gene induce a system that is triggered by dsRNA? One theory is that
overproduction of certain mRNAs triggers RdRP to make dsRNA from the excess.
This dsRNA activates Dicer to create siRNAs that quench mRNA, both from the
transgene and from any closely related endogenous gene. Alternatively, when
certain transgenes are expressed, some regions of the mRNA may fold back on
themselves to form hairpins. These
double-stranded segments may also activate Dicer. Genetic analysis of the model
plant, Arabidopsis, has shown that
the RdRP encoded by the SDE1 gene is necessary for transgene silencing but is
not needed for antiviral RNAi. (In the latter case, the virus RNA polymerase
would make dsRNA and the plant RdRP enzyme would therefore not be necessary.)
This favors the first model for transgene-triggered silencing.
The most interesting trait of
PTGS is the ability of silencing to propagate from one part of the plant to the
next. Plants can be grafted, that is, a leaf or stem can be attached to a
different plant. If the graft has a transgene silenced by PTGS, the scion
(piece of grafted plant) will then silence the corresponding endogenous gene.
The effect of RNAi travels through the vascular system of the plant, and
affects regions without the transgene. RNAi in C. elegans also has the ability to spread, not only from tissue to
tissue, but also from parent to progeny. It does not appear that mammalian
systems have the ability to spread the RNAi signal.
The ability to spread may not
rely solely on siRNA. In plants, the potyviruses produce an inhibitor of RNAi
called helper component proteinase
(Hc-Pro). This protein blocks the accumulation of siRNA. Despite this, the
RNAi signal still spreads to other parts of the plant and triggers methylation
of DNA, thus turning it into heterochromatin. Other viral genes that inhibit
different steps of the RNAi process will, it is hoped, illuminate the mechanism
Other terms used to describe
variants of RNAi are transcriptional
gene silencing, co-suppression,
and virus-induced silencing.
Virus-induced silencing occurs when the viral
genome has a double-stranded RNA intermediate, which triggers Dicer and RISC.
Co-suppression is an early name for PTGS. Transcriptional gene silencing refers
to the silencing of gene expression by converting the gene into
heterochromatin. A comprehensive term, GENE
impedance (GENEi), has been proposed to encompass all these phenomena but
is rarely used.