A new protein-dependent riboswitch activates ribosomal frameshifting

Abstract Programmed –1 ribosomal frameshifting (PRF) is a ubiquitous translational control mechanism in RNA viruses, allowing them to change the relative abundance of proteins encoded in different reading frames. The cardioviruses exhibit the highest known viral PRF efficiency in nature, with up to 85 % of ribosomes diverted into the –1 frame. This unusual event requires an interaction between the viral 2A protein and a regulatory element in the RNA genome, thus PRF is conditionally activated by 2A accumulation during infection. Here we present the structure of the 2A-RNA complex, revealing a novel pseudoknot architecture and illustrating the mechanism of 2A recognition. By combining single-molecule Fӧrster resonant energy transfer (smFRET) and small-angle X-ray scattering (SAXS), we demonstrate that the RNA does not pre-exist as a pseudoknot, rather 2A binding induces a striking conformational change from a stem-loop into a pseudoknot. Together, this defines the conformational landscape of a protein-dependent riboswitch, revealing new mechanistic principles that underpin viral gene expression. We suggest that PRF stimulatory elements in other viruses may also be regulated in a similar way, highlighting their potential as targets for new antiviral drugs.