Identification and characterization of the class I and class II MHC molecules of the Syrian hamster to enable advanced studies of T cell immunity

PROJECT SUMMARY The Syrian hamster (Mesocricetus auratus) is a valuable model for the study of many RNA viruses, including SARS-CoV-2, as well as numerous other diseases relevant to human health. T cells are an important component of SARS-CoV-2-specific adaptive immunity, and they represent a key potential correlate of immunity or immunopathology in viral diseases in general. Remarkably, T cell immunity cannot be readily evaluated in Syrian hamsters at present, because their MHC molecules have not yet been identified. The lack of knowledge regarding the identity of the MHC molecules of Syrian hamsters has limited the ability of the model to support advanced investigations of cellular immunity in the response to human pathogens and vaccines. Filling this knowledge gap will be necessary to fully realize the promise of the Syrian hamster as an animal model for human diseases and is the primary objective of our proposed work. As a result of our analysis of gene structure, order, and position in the Syrian hamster genome, coupled with comparison to the mouse genome, we have identified plausible candidates for the genes encoding the classical class I and class II MHC molecules of the research Syrian hamster. We also discovered that select monoclonal antibodies (mAbs) raised to mouse MHC molecules cross-react with splenocytes of Syrian hamsters. In Aim 1, we will generate a group of cell lines, each of which will express a single candidate hamster MHC molecule. These "mono-allelic" cell lines will allow us to characterize the cross-reactivity of the anti-mouse mAbs in terms of which hamster MHC candidates they are recognizing, thus enabling the tentative identification of the candidates as genuine MHC molecules. We will verify that the cell lines can present peptides to stimulate T cells from infected or vaccinated hamsters, using SARS- CoV-2 as a model system. An important practical advance to arise from this work will be the ability to create peptide/MHC tetramers to follow T cell responses in hamsters. In Aim 2, we will define binding motifs for the hamster MHC molecules from the sequences of their natural ligands and create binding matrices to permit the identification of candidate T cell epitopes for pathogens of interest. This will permit the confirmation of the hamster candidates as true MHC molecules capable of presenting diverse arrays of peptides. Primary anchor residues and the most common peptide lengths will also be revealed. The sequences of the eluted peptides will be used to develop binding matrices that will permit prediction of candidate T cell antigens and epitopes for any current or emerging pathogen that can be modeled in the Syrian hamster. Our project will achieve the identification and functional characterization of the class I and class II MHC molecules of the Syrian hamster, enabling advanced studies of T cell immunity and greatly enhancing its utility as a model for diseases relevant to human health.