Tolerated wobble mutations in siRNAs decrease specificity, but can enhance activity in vivo.

Torgeir Holen; Svein Erik Moe; Jan Gunnar Sørbø; Trine J Meza; Ole Petter Ottersen; Arne Klungland

doi:10.1093/nar/gki785

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Tolerated wobble mutations in siRNAs decrease specificity, but can enhance activity in vivo.

Torgeir Holen·Svein Erik Moe·Jan Gunnar Sørbø·Trine J Meza·Ole Petter Ottersen·Arne Klungland

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Abstract

RNA interference (RNAi) has become an invaluable tool for functional genomics. A critical use of this tool depends on an understanding of the factors that determine the specificity and activity of the active agent, small interfering RNA (siRNA). Several studies have concluded that tolerance of mutations can be considerable and hence lead to off-target effects. In this study, we have investigated in vivo the toleration of wobble (G:U) mutations in high activity siRNAs against Flap Endonuclease 1 (Fen1) and Aquaporin-4 (Aqp4). Mutations in the central part of the antisense strand caused a pronounced decrease in activity, while mutations in the 5' and 3'ends were tolerated very well. Furthermore, based on analysis of nine different mutated siRNAs with widely differing intrinsic activities, we conclude that siRNA activity can be significantly enhanced by wobble mutations (relative to mRNA), in the 5' terminal of the antisense strand. These findings should facilitate design of active siRNAs where the target mRNA offers limited choice of siRNA positions.

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Aquaporin 4 Aquaporins Base Pairing Blotting, Western Flap Endonucleases HeLa Cells Humans Mutation RNA Interference RNA, Messenger RNA, Small Interfering Journal Article Research Support, Non-U.S. Gov't

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Torgeir Holen, Svein Erik Moe, Jan Gunnar Sørbø, Trine J Meza, Ole Petter Ottersen, Arne Klungland. 2005-08-19. Tolerated wobble mutations in siRNAs decrease specificity, but can enhance activity in vivo.. https://doi.org/10.1093/nar%2Fgki785

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