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

SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome of ~30 kb, whose outbreak caused the still 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 structural elements within coronavirus RNA genomes have been identified to date. Here, we performed RNA structure probing by SHAPE-MaP and DMS-MaPseq to obtain single-base resolution secondary structure maps of the full SARS-CoV-2 coronavirus genome both in vitro and in living infected cells. The probing data recapitulate the previously described coronavirus RNA elements (5′ UTR and 3’ UTR), and reveal new structures. Of these, ~11.3% show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. Secondary structure-restrained 3D modeling of these highly-structured conserved regions further allowed for the identification of several putative druggable pockets. In addition, we identify a set of persistently single-stranded regions 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.