Canonical T cell receptor docking on peptide–MHC is essential for T cell signaling

Zareie, Pirooz, Szeto, Christopher, Farenc, Carine, Gunasinghe, Sachith D., Kolawole, Elizabeth M., Nguyen, Angela, Blyth, Chantelle, Sng, Xavier Y. X., Li, Jasmine, Jones, Claerwen M., Fulcher, Alex J., Jacobs, Jesica R., Wei, Qianru, Wojciech, Lukasz, Petersen, Jan, Gascoigne, Nicholas R.J., Evavold, Brian D., Gaus, Katharina, Gras, Stephanie, Rossjohn, Jamie ORCID: https://orcid.org/0000-0002-2020-7522 and La Gruta, Nicole L. 2021. Canonical T cell receptor docking on peptide–MHC is essential for T cell signaling. Science 372 (6546) , eabe9124. 10.1126/science.abe9124

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Abstract

INTRODUCTION T cell receptor (TCR) recognition of peptide–major histocompatibility complexes (pMHCs) is one of the most diverse receptor–ligand interactions in biology. Nevertheless, these interactions exhibit a highly conserved, or canonical, TCR–pMHC docking polarity in both mice and humans. Whether this canonical docking polarity is driven by evolutionarily conserved, germline-encoded complementarity between the TCR and MHC or by signaling constraints imposed by coreceptors has been a question of enduring debate. Here, we demonstrate that although reversed-polarity TCR–pMHC recognition is prevalent within a naïve, viral epitope–specific T cell repertoire and may exhibit relatively high pMHCI affinity, such TCRs are unable to support TCR signaling in the presence of CD8 coreceptor because of mislocalization of Lck. These data support a paradigm in which the highly conserved TCR–pMHCI docking polarity is driven by structural constraints on TCR signaling. RATIONALE Evidence suggests that the canonical TCR–pMHC docking polarity is driven by evolutionary hardwiring of complementary germline-encoded motifs at the TCR and MHC interface. An alternate model suggests that TCR recognition of pMHC is driven during thymic selection by the need for the CD4 or CD8 coreceptors to bind MHC and deliver coreceptor-associated Lck to the CD3 signaling complex. We previously identified reversed-polarity TRBV17+ TCRs from the preimmune influenza A virus (IAV)–specific repertoire that bound pMHCI (H-2DbNP366) with a moderate affinity but were unable to support robust T cell recruitment. Here, using a range of canonical and reversed TCRs specific for the same cognate pMHCI, we tested the hypothesis that the TCR–pMHCI docking polarity precedes binding strength as a key determinant of T cell activation. We hypothesized that the underlying driver of the canonical docking polarity is the colocalization of signaling molecules central to the TCR signal transduction pathway. RESULTS In this study, we demonstrate that reversed TCRs are prevalent in a naïve virus–specific repertoire but are poorly represented in the immune response after virus challenge. We identified antigen-specific TCRαβ clonotypes that were either poorly recruited or clonally expanded and found an overriding association between immune prevalence and canonical TCR–pMHCI docking. This was irrespective of pMHCI affinity, catch or slip bond formation, or TCR clustering, demonstrating that a canonical docking polarity is required for T cell activation. This finding was verified after viral challenge of adoptively transferred retrogenic T cells expressing reversed or canonical docking TCRs of varying affinities. The inability of T cells expressing reversed-docking TCRs to be recruited into the antiviral immune response demonstrates that TCR–pMHCI docking topology supersedes TCR–pMHCI affinity as the primary determinant for effective in vivo immune recruitment. Using fluorescence lifetime imaging microscopy (FLIM)–Förster resonance energy transfer (FRET) analyses, we show that canonical TCR–pMHCI docking is essential for the colocalization of CD8–Lck with CD3ζ, which is impaired when the TCR engages pMHCI with reversed polarity. The requirement for canonical TCR–pMHCI docking can be circumvented by the removal of the CD8 coreceptor or by dissociation of Lck from CD8, suggesting that sequestration of Lck by the CD8 coreceptor has a dual role: potentiating signaling arising from canonical TCR–pMHCI interactions and impeding reversed-polarity TCR–pMHCI signaling. CONCLUSION The inability of reversed-polarity TCRs to participate in the immune response occurs independently of TCR–pMHCI binding affinity and instead is a direct consequence of reversed TCR–pMHCI engagement. Most TCR–pMHC complexes that have been solved to date, upon which the canonical TCR–pMHCI docking paradigm has been established, were derived from expanded immune repertoires. Thus, we conclude that the highly conserved docking polarity is driven predominantly by the structural constraints imposed on TCR signaling and recruitment into an immune response. In addition to the well-recognized augmentation of signaling resulting from canonical TCR–pMHCI engagement, our findings suggest a role for coreceptor–Lck association in preventing signaling by noncanonical TCR–pMHC recognition. Such negative regulation would serve to limit the extent of functional TCR cross-reactivity and constrain the number of signaling-competent TCR-binding modalities.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Schools > Medicine
Publisher:	American Association for the Advancement of Science
ISSN:	0036-8075
Date of First Compliant Deposit:	14 March 2025
Date of Acceptance:	23 April 2021
Last Modified:	18 Mar 2025 12:30
URI:	https://orca.cardiff.ac.uk/id/eprint/176886

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