Université Paris 7, Inserm, UMR 1162, 75013 Paris, France.
ICCVS, University of Gdańsk, Science, ul. Wita Stwosza 63, 80-308 Gdańsk, Poland.
Molecules. 2018 Nov 29;23(12):3124. doi: 10.3390/molecules23123124.
Protein-RNA interactions (PRIs) control pivotal steps in RNA biogenesis, regulate multiple physiological and pathological cellular networks, and are emerging as important drug targets. However, targeting of specific protein-RNA interactions for therapeutic developments is still poorly advanced. Studies and manipulation of these interactions are technically challenging and in vitro drug screening assays are often hampered due to the complexity of RNA structures. The binding of nucleolin (NCL) to a G-quadruplex (G4) structure in the messenger RNA (mRNA) of the Epstein-Barr virus (EBV)-encoded EBNA1 has emerged as an interesting therapeutic target to interfere with immune evasion of EBV-associated cancers. Using the NCL-EBNA1 mRNA interaction as a model, we describe a quantitative proximity ligation assay (PLA)-based in cellulo approach to determine the structure activity relationship of small chemical G4 ligands. Our results show how different G4 ligands have different effects on NCL binding to G4 of the mRNA and highlight the importance of in-cellulo screening assays for targeting RNA structure-dependent interactions.
蛋白质-RNA 相互作用（PRIs）控制着 RNA 生物发生的关键步骤，调节多种生理和病理细胞网络，并且正在成为重要的药物靶点。然而，针对特定的蛋白质-RNA 相互作用进行治疗开发的工作仍进展不佳。这些相互作用的研究和操作在技术上具有挑战性，并且由于 RNA 结构的复杂性，体外药物筛选测定常常受到阻碍。核仁素（NCL）与 Epstein-Barr 病毒（EBV）编码的 EBNA1 的信使 RNA（mRNA）中的 G-四链体（G4）结构的结合已成为一种有趣的治疗靶点，可干扰 EBV 相关癌症的免疫逃逸。我们使用 NCL-EBNA1 mRNA 相互作用作为模型，描述了一种基于定量邻近连接测定（PLA）的细胞内方法，用于确定小分子 G4 配体的结构活性关系。我们的结果表明，不同的 G4 配体对 NCL 与 mRNA 中 G4 的结合有不同的影响，并强调了针对 RNA 结构依赖性相互作用的细胞内筛选测定的重要性。
