TCR-peptide-MHC interactions in situ show accelerated kinetics and increased affinity.
The recognition of foreign antigens by T lymphocytes is essential to most adaptive immune responses. It is driven by specific T-cell antigen receptors (TCRs) binding to antigenic peptide-major histocompatibility complex (pMHC) molecules on other cells. If productive, these interactions promote the formation of an immunological synapse. Here we show that synaptic TCR-pMHC binding dynamics differ significantly from TCR-pMHC binding in solution. We used single-molecule microscopy and fluorescence resonance energy transfer (FRET) between fluorescently tagged TCRs and their cognate pMHC ligands to measure the kinetics of TCR-pMHC binding in situ. When compared with solution measurements, the dissociation of this complex was increased significantly (4-12-fold). Disruption of actin polymers reversed this effect, indicating that cytoskeletal dynamics destabilize this interaction directly or indirectly. Nevertheless, TCR affinity for pMHC was significantly elevated as the result of a large (about 100-fold) increase in the association rate, a likely consequence of complementary molecular orientation and clustering. In helper T cells, the CD4 molecule has been proposed to bind cooperatively with the TCR to the same pMHC complex. However, CD4 blockade had no effect on the synaptic TCR affinity, nor did it destabilize TCR-pMHC complexes, indicating that the TCR binds pMHC independently of CD4.
Huppa
Johannes B
JB
Department of Microbiology and Immunology, Stanford School of Medicine, California 94305-5323, USA.
Axmann
Markus
M
Mörtelmaier
Manuel A
MA
Lillemeier
Björn F
BF
Newell
Evan W
EW
Brameshuber
Mario
M
Klein
Lawrence O
LO
Schütz
Gerhard J
GJ
Davis
Mark M
MM
D000199:Actins / Q000378:metabolism; D000818:Animals; D015704:CD4 Antigens / Q000187:drug effects / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D004331:Drosophila melanogaster; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes; D015395:Histocompatibility Antigens Class I / Q000276:immunology / Q000378:metabolism*; D054992:Immunological Synapses / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D051379:Mice; D008822:Mice, Transgenic; D010455:Peptides / Q000276:immunology* / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism*; D015398:Signal Transduction; D020349:Surface Plasmon Resonance; D006377:T-Lymphocytes, Helper-Inducer / Q000187:drug effects / Q000276:immunology / Q000378:metabolism
D000199:Actins; D015704:CD4 Antigens; D005456:Fluorescent Dyes; D015395:Histocompatibility Antigens Class I; D008024:Ligands; D010455:Peptides; D011948:Receptors, Antigen, T-Cell