Pharmacologically stabilizing RNA G-quadruplexes in coronavirus genome reduces infectivity

RNA G-quadruplexes (rG4s) are unusual RNA secondary structures formed by stacking arrays of guanine tetrads. Although thousands of potential rG4-forming motifs occur throughout the mammalian transcriptome, many single-stranded RNA (ssRNA) viruses are thought to be depleted of rG4-forming sequences. Using in silico methods, we examine rG4-forming potential in single-stranded RNA (ssRNA) viruses and observe that, while canonical rG4 motifs are depleted, noncanonical rG4 motifs occur at comparable or higher frequencies relative to the mammalian transcriptome. We ask if the noncanonical rG4's can be leveraged to block viral replication and control infection using OC43, the coronavirus believed to be responsible for the 1889 "Russian flu" pandemic. Profiling with "d-rG4-seq" confirms a dearth of folded rG4 in the OC43 RNA genome during natural infection. Intriguingly, rG4 ligands induce synthetic rG4 structures of a noncanonical nature. Significantly, induced rG4 inhibits viral replication and reduces infectivity. We show that the rG4 ligands act by disrupting the unique pattern of OC43 "discontinuous transcription." Thus, rG4-targeting compounds present a potential therapeutic approach for targeting ssRNA viruses.