Amino acid substitutions in norovirus VP1 dictate cell tropism via an attachment process dependent on membrane mobility

Viruses interact with receptors on the cell surface to initiate and co-ordinate infection. The distribution of receptors on host cells can be a key determinant of viral tropism and host infection. Unravelling the complex nature of virus-receptor interactions is, therefore, of fundamental importance to understanding viral pathogenesis. Noroviruses are non-enveloped, icosahedral, positive-sense RNA viruses of global importance to human health, with no approved vaccine or antiviral agent available. Here we use murine norovirus as a model for the study of molecular mechanisms of virus-receptor interactions. We show that variation at a single amino acid residue in the major viral capsid protein had a key impact on the interaction between virus and receptor. This variation did not affect virion production or virus growth kinetics, but a specific amino acid was rapidly selected through evolution experiments, and significantly improved cellular attachment when infecting immune cells in suspension. However, reducing plasma membrane mobility counteracted this phenotype, providing insight into for the role of membrane fluidity and receptor recruitment in norovirus cellular attachment. When the infectivity of a panel of recombinant viruses with single amino acid variations was compared in vivo , there were significant differences in the distribution of viruses in a murine model, demonstrating a role in cellular tropism in vivo . Overall, these results highlight the importance of lipid rafts and virus-induced receptor recruitment in viral infection, as well as how capsid evolution can greatly influence cellular tropism, within-host spread and pathogenicity. Importance All viruses initiate infection by utilising receptors to attach to target host cells. These virus-receptor interactions can therefore dictate viral replication and pathogenesis. Understanding the nature of virus-receptor interactions could also be important to developing novel therapies. Noroviruses are non-enveloped icosahedral viruses of medical importance. They are a common cause of acute gastroenteritis with no approved vaccine or therapy and are a tractable model for studying fundamental virus biology. In this study, we utilise the murine norovirus model system to show that variation in a single amino acid of the major capsid protein can alone can affect viral infectivity through improved attachment to suspension cells. Reducing plasma membrane mobility reduced infectivity, providing an insight into the importance of membrane mobility for receptor recruitment. Furthermore, variation at this site was able to change viral distribution in a murine model, illustrating how in-host capsid evolution can influence viral infectivity and immune evasion. ### Competing Interest Statement The authors have declared no competing interest.