The mechanism of RNAi. by Renchang. Guo S et al, a gene required for establishing polarity in C. elegans embryos, encoding a putative Ser/Thr kinase that is asymmetrically distributed, cell, 1995, 81:611. Fire Andrew, et al, Potent and specific genetic interference
The mechanism of RNAi
Guo S et al,
a gene required for establishing polarity in C. elegans embryos, encoding a putative Ser/Thr kinase that is asymmetrically distributed,
cell, 1995, 81:611
Fire Andrew, et al,
Potent and specific genetic interference
by double-strand RNA in Caenorhabditis
Nature, 1998, 391:806-811
RNAi is characteristic of:
Extreme efficiency: a few trigger dsRNA
molecules suffice to inactivate a continuously
transcribed target mRNA for long periods
Long-lasting: the inactivation persists through
cell division, spreads to untreated cells and
tissues of plants, and is even inhetited by
subsequent generations of nematodes.
Hamilton, A.J., et al,
A species of small antisense RNA in
posttranscriptional gene silencing in plants.
1999, Science, 286, 950-952.
Bernstein E, et al,
Role for a bidentate ribonuclease in the
initiation step of RNA interference.
Nature, 2001, 409:363-366
Dicer: a dsRNA-specific endonuclease
responsible for processing of long
targeting dsRNA into siRNAs.
Cogoni, et al, Gene silencing in Neurospora crassa reqiures a protein homologous to RNA-dependent RNA polymerase, Nature 399, 166-169
QDE-1 N. crassa
Smardon. A., et al, EGO-1 is related to RNA-directed RNA polymerase and functions in gern-line development and RNA interference in C. elegans. 2000, curr.Biol. 10, 169-178.
EGO-1, C. elegans
Cell, Vol. 101, 543-553
Material: Arabidopsis, transgenes
PVS:GFP: responsible for initiation of PTGS
GFP: reporter of silencing
Four genetic loci are requried for PTGS
Sde Silencing defective
Sde1, Sde2, Sde3, Sde4
This paper first proves that siRNA serves as
primers to transform the target mRNA into
dsRNA. The nascent dsRNA is degraded to
Eliminate the incorporated target mRNA
while generating new siRNAs in a cycle of
dsRNA synthesis and degradation.
Pp-Luc Photinus pyralis luciferase
Proved the nascent RNA is
siRNA’s antisense strand
The polarity of the RdRP reaction
How the ds siRNAs anneal to ss RNA?
Whether Target dsRNA can self replicate?
How to control the secondary RNAi?
How to explain the long-lasting of RNAi?