Asthma is a highly complex disease because individual etiologies result from composite elements of genetic susceptibility, personal medical histories and a myriad of environmental factors. This Program will collect and evaluate data relevant to each component. Project II contributes experience and technologies to describe the molecular mechanisms by which virus infections, specifically by human rhinoviruses (RV), contribute and shape episodes of asthma. Canonically linked to the common cold, RV infections contribute 50-85% of asthma exacerbations and are the most frequently isolated viruses during the most severe respiratory infections and hospitalizations among children. More so than RV-A&B isolates, viruses in the RV-C species are particularly linked to illnesses in early childhood, and their infections are strongly linked to subsequent development of asthma. The RV-C require cadherin-related family member 3 (CDHR3), an unusual airway-specific protein, as their cell-entry receptor. The ?A? allele of this gene (Tyr529 protein variant) is among the strongest known genetic correlates for a type of childhood asthma marked by severe episodic wheezing. Conversely, Cys529, encoded by the ?G? allele, and of much higher prevalence in modern human lineages, is not an asthma correlate. Our fundamental hypothesis is that the CDHR3 Cys529 variant fails at the biochemical level to display this protein properly or extensively on cell surfaces, making their carriers more refractive to RV-C infections and virus-induced asthma-exacerbations. The three Aims of the Project will examine the structure, biochemistry and function of CDHR3 in recombinant, cell culture and native tissue formats. The expected information includes (a) a cryoEM structure determination of the receptor:RV-C complex, (b) cryoEM structure determinations of RV-C complexed with neutralizing and non-neutralizing antibodies, (c) biochemical descriptions of CDHR3 requirements for differential cell surface display (Cys/Tyr529), (d) biochemical descriptions of CDHR3 glycosylation sites, dimerization sites and virus interaction sites. We will also examine primary tissue, and tissue-derived differentiated cell cultures (e.g. ALI), to validate and localize the sites of CDHR3 display and determine how the genetics-determined allele (?A? vs ?G?) confers susceptibility to RV-C infections. Our techniques and assays will also document for the first time, if and how common environmental factors, such as an individual's microbiome content, may influence the extent and severity of RV infections by altering RV receptor expression or a cell's infection susceptibility in its native primary context.
The core processes of structure, biochemistry and function are what translate clinical findings into cogent mechanisms, which can then cycle back into the clinics for new types of interventions. This project will originate new technical underpinnings for the field of rhinovirus C biology (RV-C) by providing a structural, biochemical and functional description of an RV-C in complex with its asthma-associated receptor, cadherin related family member 3 (CDHR3).
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