Genome-wide mapping of SARS-CoV-2 RNA structures identifies therapeutically-relevant elements

SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of effective and durable therapeutic strategies a challenging and urgent need. As for other RNA viruses, genomic RNA structures are expected to play crucial roles in several steps of the coronavirus replication cycle. Despite this, only a handful of functionally-conserved coronavirus structural RNA elements have been identified to date. Here, we performed RNA structure probing to obtain single-base resolution secondary structure maps of the full SARS-CoV-2 coronavirus genome both in vitro and in living infected cells. Probing data recapitulate the previously described coronavirus RNA elements (5' UTR and s2m), and reveal new structures. Of these, ~10.2% show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. Secondary structure-restrained 3D modeling of these segments further allowed for the identification of putative druggable pockets. In addition, we identify a set of single-stranded segments in vivo, showing high sequence conservation, suitable for the development of antisense oligonucleotide therapeutics. Collectively, our work lays the foundation for the development of innovative RNA-targeted therapeutic strategies to fight SARS-related infections. Overall design: SHAPE-MaP, DMS-MaPseq

严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）属于β冠状病毒（betacoronavirus），其基因组为线性单股正链RNA，该病毒引发了持续至今的新冠肺炎（COVID-19）大流行。冠状病毒具备快速演化、适应环境并跨越物种屏障的能力，这使得开发高效且持久的治疗策略成为一项极具挑战性且亟待解决的需求。与其他RNA病毒类似，冠状病毒的基因组RNA结构被认为在其复制周期的多个关键环节中发挥重要作用。尽管如此，截至目前仅发现少量功能保守的冠状病毒结构RNA元件。本研究通过RNA结构探测技术，分别在体外与活感染细胞中获取了完整SARS-CoV-2基因组的单碱基分辨率二级结构图谱。探测数据不仅重现了此前已报道的冠状病毒RNA元件（5'非翻译区（5' UTR）与s2m元件），还揭示了全新的RNA结构。其中约10.2%的结构在SARS-CoV-2与其他冠状病毒间存在显著共变异现象，提示其具备功能保守性。基于二级结构约束的上述区段三维建模，进一步帮助我们鉴定出潜在的可药物结合口袋。此外，我们在活细胞中鉴定出一批序列保守性较高的单链RNA区段，可用于开发反义寡核苷酸治疗药物。综上，本研究为开发靶向RNA的创新型治疗策略以对抗SARS相关感染奠定了基础。实验整体设计：SHAPE-MaP、DMS-MaPseq