Tang Zhichao, Hegde Shalakha, Hao Siyuan, Selvaraju Manikandan, Qiu Jianming, Wang Jingxin
Department of Medicinal Chemistry, University of Kansas, Lawrence, USA.
Section of Genetic Medicine, Department of Medicine, Biological Sciences Division, University of Chicago, Chicago, USA.
Nat Commun. 2025 Jan 8;16(1):483. doi: 10.1038/s41467-024-55608-w.
One of the hallmarks of RNA viruses is highly structured untranslated regions (UTRs) which are often essential for viral replication, transcription, or translation. In this report, we discovered a series of coumarin derivatives that bind to a four-way RNA helix called SL5 in the 5' UTR of the SARS-CoV-2 RNA genome. To locate the binding site, we developed a sequencing-based method namely cgSHAPE-seq, in which an acylating probe was directed to crosslink with the 2'-OH group of ribose at the binding site to create read-through mutations during reverse transcription. cgSHAPE-seq unambiguously determined a bulged G in SL5 as the primary binding site, which was validated through mutagenesis and in vitro binding experiments. The coumarin derivatives were further used as a warhead in designing RNA-degrading chimeras to reduce viral RNA expression levels. The optimized RNA-degrading chimera C64 inhibited live virus replication in lung epithelial carcinoma cells.
RNA病毒的一个特征是具有高度结构化的非翻译区（UTR），这些区域通常对病毒复制、转录或翻译至关重要。在本报告中，我们发现了一系列香豆素衍生物，它们与严重急性呼吸综合征冠状病毒2（SARS-CoV-2）RNA基因组5'UTR中一个名为SL5的四链RNA螺旋结合。为了定位结合位点，我们开发了一种基于测序的方法，即cgSHAPE-seq，其中酰化探针被定向与结合位点处核糖的2'-OH基团交联，以在逆转录过程中产生通读突变。cgSHAPE-seq明确确定SL5中一个凸起的G作为主要结合位点，这通过诱变和体外结合实验得到了验证。香豆素衍生物进一步用作设计RNA降解嵌合体的弹头，以降低病毒RNA表达水平。优化后的RNA降解嵌合体C64抑制了肺上皮癌细胞中活病毒的复制。
