Clustering of the T-cell receptor complex may facilitate T-cell activation by stretching of CD3ζ cytoplasmic tails

Abstract

The vertebrate immune system relies on the ability of T cells to distinguish between dangerous and harmless antigens by sensing peptide–MHC (pMHC) complexes displayed in antigen-presenting cells. This recognition event is mediated by the T cell receptor TCR–CD3 complex, which converts extracellular binding to intracellular signaling through the phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAM) located within the cytoplasmic tails of CD3 subunits. Although cryo-electron microscopy studies have resolved the extracellular and trans membrane arrangement of the complex, the conformational dynamics of the CD3ζ cytoplasmic tails remain elusive due to their intrinsic disorder and flexibility. Biochemical and fluorescence-based studies show that CD3ζ is bound within the plasma membrane under resting conditions, sequestering ITAMs and preventing phosphorylation, while receptor engagement induces their release and exposure to kinase. These findings raise the possibility that conformational switching of CD3ζ is a central step in TCR activation. There are several theoretical models that describe the mechanism responsible for the release of the CD3ζ chains after the binding event. One of these models is the so-called aggregation model, which suggests that clustering of several complexes is the main mechanism responsible for T-cell activation. In this work, we investigate whether clustering of CD3ζ cytoplasmic domains alone is sufficient to noticeably increase the activation probability. Drawing inspiration from polymer brush theory, which predicts that tethered chains adopt extended conformations at sufficient graft densities, we hypothesized that steric repulsion between adjacent CD3ζ tails could promote membrane detachment and ITAM exposure. To test this, we used coarse-grained molecular dynamics simulations and different Monte Carlo methods to compare the behavior of single CD3ζ chains in confinement, single CD3 complexes and clusters of CD3 complexes at the membrane interface. Our results demonstrate that clustering substantially increases both the frequency and dwell time of ITAM dissociation events relative to single-chain configurations, providing a physical basis for enhanced phosphorylation efficiency. These findings support the idea that the aggregation model represent a robust mechanism for converting receptor engagement into intracellular signaling.