Nucleic Acids Research Advance Access originally published online on September 9, 2008
Nucleic Acids Research 2008 36(18):5812-5821; doi:10.1093/nar/gkn584
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Nucleic Acids Research, 2008, Vol. 36, No. 18 5812-5821
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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Effect of asymmetric terminal structures of short RNA duplexes on the RNA interference activity and strand selection
1Biotherapeutic Research Laboratory, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan, 2Centre of Molecular and Macromolecular Studies, Polish Academy of Sciences, Department of Bioorganic Chemistry, Sienkiewicza 112, Lodz 90-363, Poland, 3Department of Gene Diagnostics and Therapeutics, Research Institute, International Medical Center of Japan, 1-21-1 Toyama, Tokyo, 4iGENE Therapeutics Inc., 5-1 Kobune-cho, Nihonbashi, Tokyo 103-8355 and 5School of Pharmacy, Shujitsu University, 1-6-1 Nishigawara, Okayama 703-8516, Japan
*To whom correspondence should be addressed. Tel: +81 29 861 3040; Fax: +81 29 861 2798; Email: m.sano{at}aist.go.jp
Received May 2, 2008. Revised August 26, 2008. Accepted August 27, 2008.
Short interfering RNAs (siRNAs) are valuable reagents for sequence-specific inhibition of gene expression via the RNA interference (RNAi) pathway. Although it has been proposed that the relative thermodynamic stability at the 5'-ends of siRNAs plays a crucial role in siRNA strand selection, we demonstrate here that a character of the 2-nt 3'-overhang of siRNAs is the predominant determinant of which strand participates in the RNAi pathway. We show that siRNAs with a unilateral 2-nt 3'-overhang on the antisense strand are more effective than siRNAs with 3'-overhangs at both ends, due to preferential loading of the antisense strand into the RNA-induced silencing complex (RISC). Regardless of the relative thermodynamic stabilities at the ends of siRNAs, overhang-containing strands are predominantly selected as the guide strand; whereas, relative stability markedly influences opposite strand selection. Moreover, we show that sense strand modifications, such as deletions or DNA substitutions, of siRNAs with unilateral overhang on the antisense strand have no negative effect on the antisense strand selection, but may improve RNAi potency. Our findings provide useful guidelines for the design of potent siRNAs and contribute to understanding the crucial factors in determining strand selection in mammalian cells.
The author wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors.
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