Assembly of Picornaviral Replication Complexes

The picornaviruses are a family of small positive-sense single-stranded RNA viruses that cause a wide range of diseases at an annual cost well into the hundreds of million dollars. Members include paralyzing poliovirus and enterovirus D68, the heart disease causing coxsackie B3 virus, and rhinoviruses that cause more than half the occurrences of the common cold. These viruses share a life cycle where RNA replication and viral assembly occurs in large membrane anchored replication complexes assembled and the replication process is driven by a virally encoded RNA dependent RNA polymerase (3Dpol) that is responsible for the synthesis of all viral RNA. This research project is focused on the structure and assembly of the viral replication centers and on structure-function studies of the viral polymerases to understand mechanisms that control elongation rates and replication fidelity. We will use biochemical and structural biology approaches to study how the fidelity checkpoint used in 3Dpol was rearranged as these polymerases evolved to support large-genome coronavirus replication, providing novel information about functional constraints to help us understanding pathways for virus evolution. In a second aim we will elucidate the mechanisms whereby RNAs interact with and stimulate viral proteases, a finding that suggests viral RNA elements can regulated polyprotein processing the context of a viral replication center. Last, we will solve the structure of the picornaviral uridylylation complex that generates the VPg-pUpU primers used for all RNA synthesis by the viral 3Dpol polymerase.