Molecular and Structural Basis of Cell Entry by Emerging and Zoonotic RNA Viruses

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Emerging zoonotic viruses cross the species barrier from animals to humans or vice-versa. Their ability to propagate in human cells is dependent on a variety of factors, most importantly, their ability to target particular cell types by binding to receptor molecules displayed on the surface of the host cell. Despite the severity of disease caused in both animals and humans and threat these viruses pose to global health and economy, little is known of how they infect their host, information which has proven to be extremely valuable for the development of antiviral drugs or antibody-based medicines for better-studied viruses such as flu and HIV-1. Using techniques in structural biology, immunology, and cell biology, I aim to show in atomic detail the mechanism by which these viruses attach to and infect human cells. This will involve the combination of high-resolution structural studies on isolated molecules with lower resolution analyses of intact viral prototypes. The information derived from these combined techniques will ultimately aid in our ability to combat these pathogens. I will elucidate the molecular mechanism by which by emergent single-stranded viruses undergo attachment and fusion with host cells. This will be addressed in the context of both intact virus and the individual structural glycoproteins responsible for viral entry. This investigation is limited to emergent viruses from three families: Paramyxoviridae (e.g. African Henipaviruses), Arenaviridae (e.g. Venezuelan Hemorrhagic fever virus), and Bunyaviridae (e.g. Hantavirus Pulmonary Syndrome associated diseases; a list of targets is provided in the Case for Support, Table 1).There are four complementary goals:1. Recombinant expression and purification of viral attachment glycoproteins, preparation of model non-pathogenic viruses, and the acquisition/development of neutralising antibodies (nAbs) against these glycoproteins.2. Crystallographic analysis of viral glycoproteins alone and in complex with cellular receptors and nAbs to provide atomic descriptions in pre- and post-fusion assemblies, and illustrate the morphological changes required for viral entry.3. Cryo-electron microscopic analysis of intact, non-pathogenic viral orthologues to visualise the ultrastructure of these viruses, the assembly of their subunit glycoproteins, and to provide a structural basis for -receptor and -antibody interactions for entire virions.4. Mutagenesis of the viral genome to generate modified viruses to complement structure predicted determinants for virus host cell entry.