title: High diversity in the TCR repertoire of GAD65 autoantigen-specific human CD4+ T cells.
abstract: Autoreactive CD4(+) T cells are an essential feature of type 1 diabetes mellitus. We applied single-cell TCR α- and β-chain sequencing to peripheral blood GAD65-specific CD4(+) T cells, and TCR α-chain next-generation sequencing to bulk memory CD4(+) T cells to provide insight into TCR diversity in autoimmune diabetes mellitus. TCRs obtained for 1650 GAD65-specific CD4(+) T cells isolated from GAD65 proliferation assays and/or GAD65 557I tetramer staining in 6 patients and 10 islet autoantibody-positive children showed large diversity with 1003 different TCRs identified. TRAV and TRBV gene usage was broad, and the TRBV5.1 gene was most prominent within the GAD65 557I tetramer(+) cells. Limited overlap (<5%) was observed between TCRs of GAD65-proliferating and GAD65 557I tetramer(+) CD4(+) T cells. Few TCRs were repeatedly found in GAD65-specific cells at different time points from individual patients, and none was seen in more than one subject. However, single chains were often shared between patients and used in combination with different second chains. Next-generation sequencing revealed a wide frequency range (<0.00001-1.62%) of TCR α-chains corresponding to GAD65-specific T cells. The findings support minor selection of genes and TCRs for GAD65-specific T cells, but fail to provide strong support for TCR-targeted therapies.
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany; anne.eugster@crt-dresden.de.
authors: Anne
authors: A
authors: Eugster
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany;
authors: Annett
authors: A
authors: Lindner
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany;
authors: Mara
authors: M
authors: Catani
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany;
authors: Anne-Kristin
authors: AK
authors: Heninger
authors: Deep Sequencing Group, Collaborative Research Center 655/BIOTEC, Biotechnology Center, Dresden University of Technology, 01307 Dresden, Germany;
authors: Andreas
authors: A
authors: Dahl
authors: Deep Sequencing Group, Collaborative Research Center 655/BIOTEC, Biotechnology Center, Dresden University of Technology, 01307 Dresden, Germany;
authors: Sylvia
authors: S
authors: Klemroth
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany;
authors: Denise
authors: D
authors: Kühn
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany;
authors: Sevina
authors: S
authors: Dietz
authors: Technology Development Studio, Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany;
authors: Marc
authors: M
authors: Bickle
authors: Institute of Diabetes Mellitus Research, Helmholtz Zentrum München, 85764 Neuherberg, Germany; Forschergruppe Diabetes Mellitus, Klinikum Rechts der Isar, Technische Universität München, 81664 Munich, Germany;
authors: Anette-Gabrielle
authors: AG
authors: Ziegler
authors: German Research Foundation Center for Regenerative Therapies Dresden, Dresden University of Technology, 01307 Dresden, Germany; Paul Langerhans Institute Dresden, German Center for Diabetes Mellitus Research, Dresden University of Technology, 01307 Dresden, Germany; and Institute for Diabetes Mellitus and Obesity, Helmholtz Zentrum München, 85764 Neuherberg, Germany Germany.
authors: Ezio
authors: E
authors: Bonifacio
mesh_terms: D000293:Adolescent; D000328:Adult; D001323:Autoantibodies / Q000276:immunology; D001324:Autoantigens / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D002675:Child, Preschool; D022801:Complementarity Determining Regions / Q000235:genetics / Q000276:immunology; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics / Q000276:immunology; D005260:Female; D014644:Genetic Variation / Q000276:immunology*; D005968:Glutamate Decarboxylase / Q000276:immunology*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D007223:Infant; D008297:Male; D011236:Prediabetic State / Q000235:genetics / Q000276:immunology; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology*; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics / Q000276:immunology; D059010:Single-Cell Analysis / Q000379:methods; D055815:Young Adult
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D022801:Complementarity Determining Regions; D011948:Receptors, Antigen, T-Cell; D016693:Receptors, Antigen, T-Cell, alpha-beta; C024211:islet cell antibody; D005968:Glutamate Decarboxylase; C402565:glutamate decarboxylase 1