Characterization of the B Cell Receptor Repertoire and Discovery of New Immunoglobulin Genes in the Domestic Ferret

Abstract

Abstract Mustela putorius furo uniquely reflects physiological and immunological changes in human aging such as immune imprinting from influenza infection, yet key ferret immunobiology knowledge gaps limit the scope of molecular analyses. First, ferret germline B cell receptor genes require identification. Additionally, the V, D, and J gene repertoire expressed by B cell receptors in antigen-experienced cells needs characterization. Illumina NGS was used to generate B-cell receptor mRNA data sets. A candidate list of gene segments was assembled including ferret genomic accessions homologous to known immunoglobulin genes and previously-known ferret immunoglobulin genes. NGS reads were queried against the candidate list using IgBLAST. IgBLAST hits were focused into consensus sequences using principal components analysis and K-means clustering. To identify IgG subclasses, putative ferret genome sequences sharing human IgG C genes were used to query Oxford Nanopore data that covered the FR3 through C gene area of transcripts. We report two new functional IGHV, two IGKV, and one IGLV gene segments in the ferret, with more sequences likely to surface in downstream analyses. We also corroborate most already-proposed functional V, D, and J gene segment sequences. Ferret orthologues to human IGHV subgroups commonly involved in stem-directed, broadly neutralizing antibody responses were identified. For the first time, productive IgG3 was described in the ferret model and distinguished from productive IgG1. Our report of novel ferret immunoglobulin genes will disinhibit the ferret as a model organism for the study of infectious disease. Specifically, it will inform the immunogenicity of candidate HA antigens in preclinical vaccine studies. Research reported in this abstract was supported by NIH NIAID contract 75N93019C00050.