The mechanism of RNAi

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The mechanism of RNAi

• by Renchang

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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, 81611
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Fire Andrew, et al, Potent and specific genetic
interference by double-strand RNA in
Caenorhabditis elegans. Nature, 1998,
391806-811
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RNAi is characteristic of Extreme efficiency a
few trigger dsRNA molecules suffice to
inactivate a continuously transcribed target
mRNA for long periods of time 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.
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Bernstein, 2001
Hamilton, 1999
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Cogoni, 1999
Smardon, 2000
Dalmay, 2000
Lipardi, 2001
Nykanen, 2001
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Sijen, 2001
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Hamilton, A.J., et al, A species of small
antisense RNA in posttranscriptional gene
silencing in plants. 1999, Science, 286, 950-952.
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Bernstein E, et al, Role for a bidentate
ribonuclease in the initiation step of RNA
interference. Nature, 2001, 409363-366
Dicer a dsRNA-specific endonuclease
responsible for processing of long
targeting dsRNA into siRNAs.
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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

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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

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Cell, Vol. 101, 543-553
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• Material Arabidopsis, transgenes
• PVSGFP responsible for initiation of PTGS
• GFP reporter of silencing
• Four genetic loci are requried for PTGS
• Sde Silencing defective
• Sde1, Sde2, Sde3, Sde4

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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.
Drosophila GFP Pp-Luc Photinus pyralis
luciferase
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Proved the nascent RNA is double stranded..
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The polarity of the RdRP reaction
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Puzzlement

• 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?

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Good night