Uncovering the roles of dynamic RNA interactions during assembly and assortment of multi-segmented viral genomes (360G-Wellcome-213437_Z_18_Z)

Rotaviruses (RV) comprise a diverse and rapidly evolving group of segmented double-stranded (ds)RNA viruses of the Reoviridae family that cause deadly gastroenteritis in children worldwide. A hallmark of rotavirus morphogenesis is the stoichiometric assembly of eleven distinct genome ssRNA segment precursors prior to packaging. Elucidating the mechanism of accurate ‘molecular counting’ of RNAs is essential to understanding RV replication and their evolution. Our recent studies suggest that segmented genome assembly in RVs, and potentially other Reoviridae, requires the formation of an RNA assortment complex, whose assembly is facilitated by the non-structural proteins that accumulate in viroplasms. The key goals of this proposal are: a) to investigate how the dynamic RNA structures control accurate assembly of a multi-segmented genome; b) to directly visualize the assembly of distinct ssRNAs and to identify pathways for disrupting this process. To achieve these goals, I will combine structural probing of ssRNAs, tools for detecting inter-molecular RNA-RNA contacts and structural modeling. To visualize multiple interacting ssRNAs in situ, I will employ recently developed quantitative super-resolution multiplexed RNA imaging approaches. These fundamental studies will pave the way for identification of routes for controlling RV assembly, having potential uses in the development of antivirals and rationally designed vaccines.