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Research Project: Intervention Strategies to Support the Global Control and Eradication of Foot-and-Mouth Disease Virus(FMDV)

Location: Foreign Animal Disease Research

Title: Characterization of binding specificities of Bovine Leucocyte class I molecules: Impacts for rational epitope discovery

item Hansen, Andreas - University Of Copenhagen
item Rasmussen, Michael - University Of Copenhagen
item Svitek, Nicholas - International Livestock Research Institute (ILRI) - Kenya
item Harndahl, Mikkel - University Of Copenhagen
item Golde, William
item Barlow, John - University Of Vermont
item Nene, Vishvanath - International Livestock Research Institute (ILRI) - Kenya
item Buus, Soren - University Of Copenhagen
item Nielsen, Morten - Technical University Of Denmark

Submitted to: Immunogenetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/25/2014
Publication Date: 9/4/2014
Citation: Hansen, A.M., Rasmussen, M., Svitek, N., Harndahl, M., Golde, W.T., Barlow, J., Nene, V., Buus, S., Nielsen, M. 2014. Characterization of binding specificities of Bovine Leucocyte class I molecules: Impacts for rational epitope discovery. Immunogenetics. 66(12):705-718. DOI: 10.1007/s00251-014-0802-5.

Interpretive Summary: The rational design of vaccines for livestock in general, and cattle in particular, requires a better knowledge of the immune response. In human vaccine research, the design of vaccine draws on the extensive data available analyzing control of the immune response mediated by the gene products of the major histocompatibility (MHC) locus. This highly variable gene complex is termed human leukocyte antigens (HLA) in human, and bovine leukocyte antigens (BoLA) in cattle. Databases available online contain information on over 1000 human HLA proteins and allow human vaccine researchers to quickly determine potential T cell epitopes across large populations of man. We have now added 14 BoLA proteins to this database and demonstrated the information available for HLA can be applied to the BoLA analysis. Here we demonstrate the use of BoLA bioinformatics and the application of high throughput assays to test these predictions. These data show we have established a system based on combined bioinformatics and rapid screening assay that allows for the rapid identification of epitopes for T cell mediated immune responses in cattle.

Technical Abstract: The binding of peptides to classical major histocompatibility complex (MHC) class-I proteins is the single most selective step in antigen presentation. However, the peptide binding specificity of cattle MHC (bovine leucocyte antigen, BoLA) class I (BoLA-I) molecules remains poorly characterized. Here, we demonstrate how a combination of high-throughput assays using positional scanning combinatorial peptide libraries, peptide dissociation and peptide binding affinity binding measurements can be combined with bioinformatics to effectively characterize the functionality of BoLA-I molecules. Using this strategy, we characterized 8 BoLA-I molecules, and found the peptide specificity to resemble that of human MHC-I molecules with primary anchors at P2 and P9, and occasional auxiliary P1 and P3 anchors. We analyzed 9 reported CTL epitopes from Theileria parva, and in 8 cases, stable and high affinity binding was confirmed. A set of peptides were tested for binding affinity to the 8 BoLA proteins and used to refine the predictors of peptide-MHC binding NetMHC and NetMHCpan. The inclusion of BoLA specific peptide binding data led to a significant improvement in prediction accuracy for reported T. parva CTL epitopes. For reported CTL epitopes with weak or no predicted binding, these refined prediction methods suggested presence of nested minimal epitopes with highpredicted binding affinity. The enhanced affinity of the alternative peptides was in all cases confirmed experimentally. This study demonstrates how biochemical high-throughput assays combined with immunoinformatics can be used to characterize the peptide binding-motifs of BoLA-I molecules, boosting performance of MHC-peptide binding prediction methods, and empowering rational epitope discovery in cattle.