Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor

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ID: 307587
2002
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Abstract
Chemically synthesised 21–23 bp double-stranded short interfering RNAs (siRNA) can induce sequence-specific post-transcriptional gene silencing, in a process termed RNA interference (RNAi). In the present study, several siRNAs synthesised against different sites on the same target mRNA (human Tissue Factor) demonstrated striking differences in silencing efficiency. Only a few of the siRNAs resulted in a significant reduction in expression, suggesting that accessible siRNA target sites may be rare in some human mRNAs. Blocking of the 3′-OH with FITC did not reduce the effect on target mRNA. Mutations in the siRNAs relative to target mRNA sequence gradually reduced, but did not abolish mRNA depletion. Inactive siRNAs competed reversibly with active siRNAs in a sequence-independent manner. Several lines of evidence suggest the existence of a near equilibrium kinetic balance between mRNA production and siRNA-mediated mRNA depletion. The silencing effect was transient, with the level of mRNA recovering fully within 4 – 5 days, suggesting absence of a propagative system for RNAi in humans. Finally, we observed 3′ mRNA cleavage fragments resulting from the action of the most effective siRNAs. The depletion rate-dependent appearance of these fragments argues for the existence of a two-step mRNA degradation mechanism.
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Authors Torgeir Holen
Journal Nucleic Acids Research
Year 2002
DOI
10.1093/nar/30.8.1757
URL
https://academic.oup.com/nar/article-pdf/30/8/1757/7063191/gkf272.pdf
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