Our long term goal is to elucidate mechanisms by which respiratory syncytial virus (RSV) causes lung dysfunction. In the USA, RSV hospitalizes >100K infants/year. Pulmonary mucus is a hallmark of RSV disease. Mucus mixed with epithelial cell debris blocks the airways. Mechanisms by which RSV infection induces airway mucus are not known. Goals of this proposal are to define the role of the viral fusion (F) proteins of mucus-inducing RSV strains in pathogenesis and to define the roles of neutrophils and basophils in RSV-induced mucus. In contrast to laboratory RSV strains, RSV strains line 19 and 2-20 induce mucus and bronchiolitis in mice. Using a RSV reverse genetics system, we showed that the F protein of line 19 plays a role in mucus induction. We identified five amino acids in the line 19 F protein as candidates for involvement in mucus induction. We will use F mutant viruses to define line 19 F residues that are important for mucus expression in vivo. RSV 2- 20 is virulent in mice. We hypothesize that RSV 2-20 F contributes to airway mucus and virulence. We will define the role of 2-20 F in pathogenesis using 2-20F-chimeric RSV. Line 19 and 2-20 cause greater epithelial cell cytopathic effect (CPE) and desquamation in vivo than lab RSV strains. We will use F mutants to define effects of RSV line 19 F and 2-20 F on epithelial cell CPE, syncytia, and desquamation in vivo and in vitro. We will define the fusogenicity of RSV line 19, 2-20, and lab strain F proteins in epithelial cells in vitro using a fusion assay. We hypothesize that line 19 F and 2-20 F induce greater syncytia and fusion in vivo and in vitro. The study will also determine the roles of neutrophils and basophils in RSV bronchiolitis. Concurrently wih epithelial cell damage, RSV line 19 and 2-20 induce neutrophilia in the lungs of mice. We hypothesize that neutrophils contribute to viral clearance and mucus expression in line 19 and 2-20 pathogenesis. Line 19- induced mucus is dependent on IL-13, a TH2 cytokine. IL 4 is the primary TH2-driving cytokine. We showed that basophils are the major IL 4-expressing cell type in RSV-infection. We hypothesize that basophils contribute to RSV line 19- and 2-20-induced IL-13 and mucus expression. These RSV bronchiolitis models will advance our understanding of RSV strain-dependent pathogenesis and may lead to much-needed therapies.

Public Health Relevance

RSV causes >100,000 infant hospitalizations in the US each year and is the leading cause of bronchiolitis and viral death in infants. Mucus production is a hallmark feature of RSV disease, leading to airway obstruction, hypoxia, and mechanical ventilation. Mechanisms of RSV-induced mucus are unknown and the focus of this proposal. Elucidation of these mechanisms will identify targets for therapies

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI087798-04
Application #
8462893
Study Section
Virology - B Study Section (VIRB)
Program Officer
Kim, Sonnie
Project Start
2010-05-15
Project End
2015-04-30
Budget Start
2013-05-01
Budget End
2014-04-30
Support Year
4
Fiscal Year
2013
Total Cost
$359,811
Indirect Cost
$121,210
Name
Emory University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
Rostad, Christina A; Stobart, Christopher C; Todd, Sean O et al. (2018) Enhancing the Thermostability and Immunogenicity of a Respiratory Syncytial Virus (RSV) Live-Attenuated Vaccine by Incorporating Unique RSV Line19F Protein Residues. J Virol 92:
Ke, Zunlong; Dillard, Rebecca S; Chirkova, Tatiana et al. (2018) The Morphology and Assembly of Respiratory Syncytial Virus Revealed by Cryo-Electron Tomography. Viruses 10:
Boyoglu-Barnum, S; Todd, S O; Meng, J et al. (2017) Mutating the CX3C Motif in the G Protein Should Make a Live Respiratory Syncytial Virus Vaccine Safer and More Effective. J Virol 91:
Stobart, Christopher C; Hotard, Anne L; Meng, Jia et al. (2017) BAC-Based Recovery of Recombinant Respiratory Syncytial Virus (RSV). Methods Mol Biol 1602:111-124
Chatterjee, Srirupa; Luthra, Priya; Esaulova, Ekaterina et al. (2017) Structural basis for human respiratory syncytial virus NS1-mediated modulation of host responses. Nat Microbiol 2:17101
Stobart, Christopher C; Hotard, Anne L; Meng, Jia et al. (2016) Reverse Genetics of Respiratory Syncytial Virus. Methods Mol Biol 1442:141-53
Currier, Michael G; Lee, Sujin; Stobart, Christopher C et al. (2016) EGFR Interacts with the Fusion Protein of Respiratory Syncytial Virus Strain 2-20 and Mediates Infection and Mucin Expression. PLoS Pathog 12:e1005622
Lee, Sujin; Nguyen, Minh Trang; Currier, Michael G et al. (2016) A polyvalent inactivated rhinovirus vaccine is broadly immunogenic in rhesus macaques. Nat Commun 7:12838
Meng, Jia; Hotard, Anne L; Currier, Michael G et al. (2016) Respiratory Syncytial Virus Attachment Glycoprotein Contribution to Infection Depends on the Specific Fusion Protein. J Virol 90:245-53
Stobart, Christopher C; Rostad, Christina A; Ke, Zunlong et al. (2016) A live RSV vaccine with engineered thermostability is immunogenic in cotton rats despite high attenuation. Nat Commun 7:13916

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