title: Functional role of T-cell receptor nanoclusters in signal initiation and antigen discrimination.
abstract: Antigen recognition by the T-cell receptor (TCR) is a hallmark of the adaptive immune system. When the TCR engages a peptide bound to the restricting major histocompatibility complex molecule (pMHC), it transmits a signal via the associated CD3 complex. How the extracellular antigen recognition event leads to intracellular phosphorylation remains unclear. Here, we used single-molecule localization microscopy to quantify the organization of TCR-CD3 complexes into nanoscale clusters and to distinguish between triggered and nontriggered TCR-CD3 complexes. We found that only TCR-CD3 complexes in dense clusters were phosphorylated and associated with downstream signaling proteins, demonstrating that the molecular density within clusters dictates signal initiation. Moreover, both pMHC dose and TCR-pMHC affinity determined the density of TCR-CD3 clusters, which scaled with overall phosphorylation levels. Thus, TCR-CD3 clustering translates antigen recognition by the TCR into signal initiation by the CD3 complex, and the formation of dense signaling-competent clusters is a process of antigen discrimination.
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Sophie V
authors: 0000-0003-1701-5551
authors: SV
authors: Pageon
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Thibault
authors: T
authors: Tabarin
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Yui
authors: Y
authors: Yamamoto
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Yuanqing
authors: Y
authors: Ma
authors: Philip R
authors: PR
authors: Nicovich
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom; Cellular Therapeutics Ltd., Manchester M13 9XX, United Kingdom;
authors: John S
authors: JS
authors: Bridgeman
authors: T Cell Signalling Laboratory, Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom;
authors: André
authors: A
authors: Cohnen
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Carola
authors: C
authors: Benzing
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Yijun
authors: Y
authors: Gao
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom;
authors: Michael D
authors: MD
authors: Crowther
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom;
authors: Katie
authors: K
authors: Tungatt
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom;
authors: Garry
authors: G
authors: Dolton
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom;
authors: Andrew K
authors: AK
authors: Sewell
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom; Human Immunology Section, Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892;
authors: David A
authors: DA
authors: Price
authors: T Cell Signalling Laboratory, Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom;
authors: Oreste
authors: O
authors: Acuto
authors: Institute for Molecular Bioscience, University of Queensland, St. Lucia, QLD 4072, Australia; Australia Centre for Microscopy and Microanalysis, University of Queensland, St. Lucia, QLD 4072, Australia; Australian Research Council Centre of Excellence in Convergent Bio-Nano Science and Technology, University of Queensland, St. Lucia, QLD 4072, Australia;
authors: Robert G
authors: RG
authors: Parton
authors: Australian Research Council Centre of Excellence in Convergent Bio-Nano Science and Technology, University of Queensland, St. Lucia, QLD 4072, Australia; School of Chemistry and Australian Centre of NanoMedicine, University of New South Wales, Sydney, NSW 2052, Australia;
authors: J Justin
authors: JJ
authors: Gooding
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia;
authors: Jérémie
authors: J
authors: Rossy
authors: Institute of Infection and Immunity, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom; Infection and Immunity Program, Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC 3800, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, VIC 3800, Australia.
authors: Jamie
authors: J
authors: Rossjohn
authors: European Molecular Biology Laboratory (EMBL) Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia; Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia; k.gaus@unsw.edu.au.
authors: Katharina
authors: K
authors: Gaus
mesh_terms: D056704:Adaptive Immunity / Q000235:genetics; D000818:Animals; D000941:Antigens / Q000235:genetics / Q000276:immunology*; D017252:CD3 Complex / Q000276:immunology*; D006801:Humans; D008285:Major Histocompatibility Complex / Q000276:immunology*; D051379:Mice; D010455:Peptides / Q000276:immunology; D010766:Phosphorylation / Q000276:immunology; D011948:Receptors, Antigen, T-Cell / Q000276:immunology*; D015398:Signal Transduction; D000072760:Single Molecule Imaging
chemical_list: D000941:Antigens; D017252:CD3 Complex; D010455:Peptides; D011948:Receptors, Antigen, T-Cell
keywords: TCR triggering; signal transduction; single-molecule localization microscopy