Higher-order structures of the foot-and-mouth disease virus RNA-dependent RNA polymerase required for dynamic inter-molecular interactions involved in viral genome replication

Replication of many positive-sense RNA viruses occurs within intracellular membrane-associated compartments. These are believed to provide a favourable environment for replication to occur, concentrating essential viral structural and non-structural components, as well as protecting these components from host-cell pathogen recognition and innate immune responses. However, the details of the molecular interactions and dynamics within these structures is very limited. One of the key components of the replication machinery is the RNA-dependent RNA polymerase, RdRp. This enzyme has been shown to form higher-order fibrils in vitro. Here, using the RdRp from foot-and-mouth disease virus (termed 3Dpol), we report fibril structures, solved at ~7-9 [A] resolution by cryo-EM, revealing multiple conformations of a flexible assembly. Fitting high-resolution coordinates led to the definition of potential intermolecular interactions. We employed mutagenesis using a sub-genomic replicon system to probe the importance of these interactions for replication. We use these data to propose models for the role of higher order 3Dpol complexes as a dynamic scaffold within which RNA replication can occur.