Early T cell receptor signals globally modulate ligand:receptor affinities during antigen discrimination

Rafal M. Pielak, Geoff P. O’Donoghue, Jenny J. Lin, Katherine N. Alfieri, Nicole C. Fay, Shalini T. Low-Nam, and Jay T. Groves
PNAS published ahead of print October 30, 2017 https://doi.org/10.1073/pnas.1613140114

  1. Edited by Ronald N. Germain, National Institutes of Health, Bethesda, MD, and approved August 28, 2017 (received for review August 7, 2016)

Significance

Antigen discrimination by T cells is based on subtle differences in binding of the T cell receptor (TCR) for its peptide major histocompatibility complex (pMHC) ligand. While such binding characteristics are readily mapped with great precision in reconstituted biochemical systems, it is less clear how these interactions are affected in the live cell environment. Here we utilize single-molecule imaging to individually resolve all of the pMHC:TCR binding events in live T cells. The quantitative measurements reveal an active feedback mechanism that globally modulates the probability of pMHC:TCR binding throughout the cell–cell interface, without affecting the unbinding rate. The result is to increase the efficiency with which TCRs scan for antigen pMHC after the first few molecular encounters have occurred.

Abstract

Antigen discrimination by T cells occurs at the junction between a T cell and an antigen-presenting cell. Juxtacrine binding between numerous adhesion, signaling, and costimulatory molecules defines both the topographical and lateral geometry of this cell–cell interface, within which T cell receptor (TCR) and peptide major histocompatibility complex (pMHC) interact. These physical constraints on receptor and ligand movement have significant potential to modulate their molecular binding properties. Here, we monitor individual ligand:receptor binding and unbinding events in space and time by single-molecule imaging in live primary T cells for a range of different pMHC ligands and surface densities. Direct observations of pMHC:TCR and CD80:CD28 binding events reveal that the in situ affinity of both pMHC and CD80 ligands for their respective receptors is modulated by the steady-state number of agonist pMHC:TCR interactions experienced by the cell. By resolving every single pMHC:TCR interaction it is evident that this cooperativity is accomplished by increasing the kinetic on-rate without altering the off-rate and has a component that is not spatially localized. Furthermore, positive cooperativity is observed under conditions where the T cell activation probability is low. This TCR-mediated feedback is a global effect on the intercellular junction. It is triggered by the first few individual pMHC:TCR binding events and effectively increases the efficiency of TCR scanning for antigen before the T cell is committed to activation.

Footnotes

  • ↵1R.M.P., G.P.O., and J.J.L. contributed equally to this work.

  • ↵2Present address: L'Oréal Tech Incubator, San Francisco, CA 94105.

  • ↵3Present address: Department of Cellular and Molecular Pharmacology, University of San Francisco, San Francisco, CA 94158.

  • ↵4Present address: Stanford ChEM-H, Stanford University, Stanford, CA 94305.

  • ↵5Present address: Applied Molecular Transport, South San Francisco, CA 94080.

  • ↵6To whom correspondence should be addressed. Email: jtgroves{at}lbl.gov.

  • Author contributions: R.M.P., G.P.O., J.J.L., N.C.F., S.T.L.-N., and J.T.G. designed research; R.M.P., G.P.O., J.J.L., K.N.A., and S.T.L.-N. performed research; R.M.P., G.P.O., J.J.L., K.N.A., N.C.F., and S.T.L.-N. contributed new reagents/analytic tools; R.M.P., G.P.O., J.J.L., K.N.A., S.T.L.-N., and J.T.G. analyzed data; and R.M.P., G.P.O., J.J.L., and J.T.G. wrote the paper.

  • The authors declare no conflict of interest.

  • This article is a PNAS Direct Submission.

  • This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1613140114/-/DCSupplemental.

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TCR signaling feedback
Rafal M. Pielak, Geoff P. O’Donoghue, Jenny J. Lin, Katherine N. Alfieri, Nicole C. Fay, Shalini T. Low-Nam, Jay T. Groves
Proceedings of the National Academy of Sciences Oct 2017, 201613140; DOI: 10.1073/pnas.1613140114
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