Probing the Structural Organization of Human Respiratory Syncytial Virus. (360G-Wellcome-086774_Z_08_Z)

Human respiratory syncytial virus (hRSV) is an enveloped, negative-sense, single-stranded RNA (ssRN virus responsible for severe paediatric respiratory disease worldwide. The virus infects epithelial cells both the upper and lower respiratory tract, and is a leading cause of severe bronchiolitis and pneumonia young childre. Most children will be infected by the virus by the age of two, and infection may require hospitalization a few months after birth. Additional groups at risk of infection include the elderly and immunocompromised. The high morbidity and mortality of hRSV-associated diseases, as well as the susceptibility of individuals to repeated infection has strongly motivated the development of a vaccination. Furthermore, hRSV represents an excellent model for understanding other negative-sense ssRNA virus including important human pathogens such as the Ebola, influenza, measles, mumps, parainfluenza, rabies and Rift valley fever viruses. The proposed research project will investigate the structural organization of hRSV at both extracellular and intracellular stages of the virus life-cycle. Electron cryo-microscopy (cryo-EM) will be used to achieve each of the following: 1. Virion morphogenesis of hRSV strains A and B w/11 be investigated by visualizing mature virus particles (virions) grown using different cell types. 2. The structure of the hRSV RNA-dependent-RNA-polymerase (RdRp) and related complexes will be determined by single-particle cryo-EM (SPEM). 3. Electron cryo-tomography (cryo-ET) will be developed to study the structure of hRSV extracellular and intracellular states. hRSV specimens will be visualized in a native and hydrated state for the first time. Three dimensional (30) reconstruction of the resultant data will provide novel insights into the organization of the virus, and interactions between virus and host factors. Such information will shed light on the structural basis for virus infection, replication and spread, informing strategies for rational antiviral and vaccine development against viruses of this class.