ABSTRACT
The Coronaviridae are a family of positive-strand RNA viruses that includes SARS-CoV-2, the etiologic agent of the COVID-19 pandemic. Bearing the largest single-stranded RNA genomes in nature, coronaviruses are critically dependent on long-distance RNA-RNA interactions to regulate the viral transcription and replication pathways. Here we experimentally mapped the in vivo RNA-RNA interactome of the full-length SARS-CoV-2 genome and its subgenomic mRNAs. We uncovered a network of RNA-RNA interactions spanning tens of thousands of nucleotides. These interactions reveal that the viral genome adopts alternative topologies inside cells and undergoes genome cyclization. In addition, the SARS-CoV-2 genome and subgenomic mRNAs engage in different interactions with host RNAs. Most importantly, we discovered a long-range RNA-RNA interaction - the FSE-arch - that encircles the programmed ribosomal frame-shifting element. The FSE-arch is conserved in the related MERS-CoV virus and is under purifying selection. Our findings illuminate RNA-based mechanisms governing replication, discontinuous transcription, and translation of coronaviruses, and will aid future efforts to develop antiviral strategies.