Cutting Edge: Evidence for a Dynamically Driven T Cell Signaling Mechanism

Abstract

T cells use the αβ T cell receptor (TCR) to bind peptides presented by major histocompatibility complex proteins (pMHC) on antigen presenting cells. Formation of a TCR-pMHC complex initiates T cell signaling via a poorly understood process, potentially involving changes in oligomeric state, altered interactions with CD3 subunits, and mechanical stress. These mechanisms could be facilitated by binding-induced changes in the TCR, but the nature and extent of any such alterations are unclear. Using hydrogen/deuterium exchange, we demonstrate that ligation globally rigidifies the TCR, which via entropic and packing effects will promote associations with neighboring proteins and enhance the stability of existing complexes. TCR regions implicated in lateral associations and signaling are particularly affected. Computational modeling demonstrated a high degree of dynamic coupling between the TCR constant and variable domains that is dampened upon ligation. These results raise the possibility that TCR triggering could involve a dynamically driven, allosteric mechanism.

Molecular architecture of the αβ T cell receptor–CD3 complex

“…Our data provide a structural framework that can potentially begin to reconcile disparate observations about how the TCR relays a ligation event to signaling. Such observations include studies showing that monomeric pMHC possesses little to no ability to induce signaling regardless of affinity (49), there exist signaling-permissive and -nonpermissive pMHC-TCR docking geometries (48), conformational changes occur in the TCR upon pMHC ligation (46,(50)(51)(52), the CD4 and CD8 coreceptors are required to place Lck relative to the CD3 ITAMs (53, 54), phosphatases are occluded from the T-cell Ag-presenting cell synapse (18,26,55), and mechanotransduction appears to be important for TCR signaling (19,36,(56)(57)(58)(59) (Fig. 4).…”

Peptide Modulation of Class I Major Histocompatibility Complex Protein Molecular Flexibility and the Implications for Immune Recognition*

“…The identities of pepsin-generated fragments of HLA-A2 were previously established in our laboratory by LC/MS/MS (18). The analysis of the exchange behavior of these fragments was performed by MALDI mass spectrometry as described previously (18).…”

“…The analysis of the exchange behavior of these fragments was performed by MALDI mass spectrometry as described previously (18). Briefly, a matrix solution of 2,5-dihydroxybenzoic acid in 50:50 0.1% TFA (pH 2.4) and acetonitrile was mixed 1:1 with the acidified protein digest on a chilled MALDI plate.…”

“…Hydrogen/Deuterium Exchange-Mass Spectrometry-Hydrogen/deuterium exchange-mass spectrometry (HDX-MS) was performed as described previously (18). Briefly, exchange was initiated by diluting protein samples at 25°C in 25 mM HEPES, 50 mM NaCl (pH 7.4) 10-fold with the same buffer made with 99.9% 2 H 2 O. pMHC concentrations were 30 M after dilution.…”

Hydrogen/Deuterium Exchange Kinetics Demonstrate Long Range Allosteric Effects of Thumb Site 2 Inhibitors of Hepatitis C Viral RNA-dependent RNA Polymerase

“…For proteins the size of NS5B, HDX-MS provides a useful approach for probing conformational dynamics and perturbation by ligands. Proteins for which HDX-MS has revealed the dynamical basis of allostery include nuclear receptors (12), G protein-coupled receptors (13), the viral polymerase HIV-1 reverse transcriptase (14), and many others (15)(16)(17)(18)(19).…”