Defining astrovirus-specific T cell responses

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PROJECT SUMMARY/ABSTRACT Human astroviruses (HAstVs) are a global cause of pediatric gastroenteritis, and can cause disseminated infection in immunocompromised hosts. Seroprevalence studies indicate almost universal HAstV infection during childhood. Despite their clinical importance, HAstVs are highly understudied in part due to the lack of a well- defined small animal model. Murine astrovirus (muAstV), which shares numerous genomic and phenotypic features with HAstVs, causes chronic infection in immunodeficient models and has begun to provide important insights into innate and adaptive immune regulation of AstV pathogenesis in vivo. Preliminary data indicates that CD8+ T cells are critical for clearance of chronic muAstV. However, to date functional and phenotypic features of the CD8+ T cell response remain uncharacterized for both HAstV and muAstV. Further, the specific viral epitopes targeted by CD8+ T cells remain to be delineated for both HAstV and muAstV. The definition of immunodominant CD8+ T cell epitopes will facilitate characterization of AstV-specific T cell functions in vivo and reveal key viral epitopes to target for future vaccines and cellular therapies. Overlapping peptide libraries will be used for ex vivo screening of immunodominant muAstV CD8+ T cell epitopes by well- established protocols, with the goal of developing MHC class I peptide tetramers to track muAstV-specific CD8+ T cells. Two phenotypically-distinct muAstV strains, with one causing acute self-limited infection and the other causing chronic infection in immunocompetent mice, will be leveraged to define effector and memory muAstV- specific CD8+ T cell phenotypes and functionality across multiple tissues. These studies, to be conducted in established in vivo mouse models, will reveal whether functionally suboptimal CD8+ T cell responses contribute to impaired clearance of some viral strains. Finally, human peripheral blood mononuclear cells from donors carrying an MHC class I allele associated with protection from HAstV will be used to identify immunodominant HAstV CD8+ T cell epitopes, permitting development of tetramers to characterize HAstV-specific CD8+ T cell phenotypes across a variety of donors with this common allele. These studies are highly appropriate for the R21 grant mechanism, as they involve development of critical tools for immunological studies using methods that have not yet been applied towards AstVs. Completion of this proposal will both provide key insights into the cellular immune response against AstVs and develop tetramer reagents to be used for future definition of T cell responses following natural infection and vaccination. Further, immunodominant epitopes identified will represent important targets for vaccines and cellular therapies against these clinically important enteric viral pathogens.