Department of Regulatory Bioorganic Chemistry, SANKEN (The Institute of Scientific and Industrial Research), Osaka University, Mihogaoka 8-1, Ibaraki, Osaka 567-0047, Japan.
Medical Research Support Center, Kyoto University Graduate School of Medicine, Konoecho Yoshida, Sakyo-ku, Kyoto 606-8501, Japan.
ACS Chem Biol. 2022 Oct 21;17(10):2817-2827. doi: 10.1021/acschembio.2c00452. Epub 2022 Sep 23.
Non-coding RNAs are emerging targets for drug development because they are involved in various cellular processes. However, there are a few reliable design strategies for small molecules that can target RNAs. This paper reports a simple and efficient method to comprehensively analyze RNA motifs that can be bound by a specific small molecule. The method involves Dicer-mediated pre-miRNA cleavage and subsequent analysis of the reaction products by high-throughput sequencing. A pre-miRNA mutant library containing a randomized region at the Dicer cleavage site was used as the substrate for the reaction. Sequencing analysis of the products of the reaction carried out in the presence or absence of a synthetic small molecule identified the pre-miRNA mutants whose Dicer-mediated cleavage was significantly altered by the addition of the small molecule. The binding of the small molecule to the identified pre-miRNA mutants was confirmed by surface plasmon resonance, demonstrating the feasibility of our method.
非编码 RNA 是药物开发的新兴靶点，因为它们参与了各种细胞过程。然而，针对 RNA 的小分子具有可靠设计策略的并不多。本文报道了一种简单而有效的方法，可全面分析可被特定小分子结合的 RNA 基序。该方法涉及 Dicer 介导的前体 miRNA 切割，以及随后通过高通量测序分析反应产物。将包含 Dicer 切割位点随机区域的前体 miRNA 突变文库用作反应的底物。在存在或不存在合成小分子的情况下进行反应产物的测序分析，鉴定出小分子的添加显著改变了 Dicer 介导切割的前体 miRNA 突变体。通过表面等离子体共振确认了小分子与鉴定出的前体 miRNA 突变体的结合，证明了我们方法的可行性。
