Thermodynamic Stabilities in the siRNA Seed and Non- seed Regions Regulate Off-target Effect in Opposite Direction | Chapter 13 | Research Aspects in Biological Science Vol. 2

 RNA interference (RNAi) is a phenomena in which a small interfering RNA (siRNA) with a perfect complementary sequence reduces the production of its target mRNA. However, siRNA also inhibits the production of undesired mRNAs with partial sequence complementarities, primarily in the siRNA seed region (nucleotides 2–8 from the guide strand's 5' end). This method is known as the siRNA-mediated off-target effect and is very similar to microRNA (miRNA)-mediated RNA silencing. The off-target effect, on the other hand, is an undesired side effect of RNAi that reduces the expression of genes other than the target, therefore thus should be avoided. We explored the exact mechanism of the off-target impact in order to effectively avoid it in this work. The off-target impact is caused by thermodynamically stable base-pairing between the siRNA seed area and off-target mRNAs, but not by unstable base-pairing, as previously shown. However, we recently discovered that the siRNA seed region has two functional domains: nucleotides 2–5, which are essential for off-target effects, and nucleotides 6–8, which are involved in both RNAi and off-target effects. In this work, we used a machine learning approach and a random sampling procedure to examine the thermodynamic characteristics of all possible siRNA subregions in order to find the most responsible area for the off-target impact. As a consequence, nucleotides 2–5 in the seed area had the largest positive relationship with the off-target impact, whereas nucleotides 8–14 in the non-seed region had the strongest negative relationship. As a result, the siRNA off-target impact is defined by the base-pairing stabilities of two distinct subregions with opposing effects.

Author(S) Details

Yoshiaki Kobayashi
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan.

Shen Tian
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan.

Kumiko Ui-Tei
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Tokyo 113-0033, Japan and Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8561, Japan.

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