title: Paired TCRαβ analysis of virus-specific CD8(+) T cells exposes diversity in a previously defined 'narrow' repertoire.
abstract: T-cell receptor (TCR) usage has an important role in determining the outcome of CD8(+) cytotoxic T-lymphocyte responses to viruses and other pathogens. However, the characterization of TCR usage from which such conclusions are drawn is based on exclusive analysis of either the TCRα chain or, more commonly, the TCRβ chain. Here, we have used a multiplexed reverse transcription-PCR protocol to analyse the CDR3 regions of both TCRα and β chains from single naive or immune epitope-specific cells to provide a comprehensive picture of epitope-specific TCR usage and selection into the immune response. Analysis of TCR repertoires specific for three influenza-derived epitopes (D(b)NP(366), D(b)PA(224) and D(b)PB1-F2(62)) showed preferential usage of particular TCRαβ proteins in the immune repertoire relative to the naive repertoire, in some cases, resulting in a complete shift in TRBV preference or CDR3 length, and restricted repertoire diversity. The NP(366)-specific TCRαβ repertoire, previously defined as clonally restricted based on TCRβ analysis, was similarly diverse as the PA(224)- and PB1-F2(62)-specific repertoires. Intriguingly, preferred TCR characteristics (variable gene usage, CDR3 length and junctional gene usage) appeared to be able to confer specificity either independently or in concert with one another, depending on the epitope specificity. These data have implications for established correlations between the nature of the TCR repertoire and response outcomes after infection, and suggest that analysis of a subset of cells or a single TCR chain does not accurately depict the nature of the antigen-specific TCRαβ repertoire.
authors: Department of Microbiology and Immunology, University of Melbourne, Peter Doherty Institute of Infection and Immunity, Melbourne, VIC, Australia.
authors: Tania
authors: T
authors: Cukalac
authors: Department of Microbiology and Immunology, University of Melbourne, Peter Doherty Institute of Infection and Immunity, Melbourne, VIC, Australia.
authors: Wan-Ting
authors: WT
authors: Kan
authors: Department of Immunology, St Jude Children's Research Hospital, Memphis, TN, USA.
authors: Pradyot
authors: P
authors: Dash
authors: Department of Microbiology and Immunology, University of Melbourne, Peter Doherty Institute of Infection and Immunity, Melbourne, VIC, Australia.
authors: Jing
authors: J
authors: Guan
authors: Department of Microbiology and Immunology, University of Melbourne, Peter Doherty Institute of Infection and Immunity, Melbourne, VIC, Australia.
authors: Kylie M
authors: KM
authors: Quinn
authors: Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia.|Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, VIC, Australia.
authors: Stephanie
authors: S
authors: Gras
authors: Department of Immunology, St Jude Children's Research Hospital, Memphis, TN, USA.
authors: Paul G
authors: PG
authors: Thomas
authors: Department of Microbiology and Immunology, University of Melbourne, Peter Doherty Institute of Infection and Immunity, Melbourne, VIC, Australia.
authors: Nicole L
authors: NL
authors: La Gruta
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism / Q000821:virology; D002999:Clone Cells / Q000276:immunology / Q000378:metabolism; D022801:Complementarity Determining Regions / Q000235:genetics / Q000276:immunology; D018984:Epitopes, T-Lymphocyte / Q000276:immunology; D005260:Female; D005434:Flow Cytometry; D014644:Genetic Variation / Q000276:immunology; D053122:Influenza A Virus, H3N2 Subtype / Q000276:immunology* / Q000502:physiology; D008810:Mice, Inbred C57BL; D009976:Orthomyxoviridae Infections / Q000276:immunology* / Q000821:virology; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics / Q000276:immunology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000378:metabolism / Q000821:virology
chemical_list: D022801:Complementarity Determining Regions; D018984:Epitopes, T-Lymphocyte; D016693:Receptors, Antigen, T-Cell, alpha-beta