High dose antigen stimulation leads to binding defects caused by persistent spatial segregation of TCR and CD8 in cytotoxic T cells (178.9)

Abstract

Abstract The lateral organization of surface receptors controls ligand binding and activation. Our group has shown that activated T Cells exhibit persistent clustering of T Cell Receptor (TCR) several days after stimulation, leading to enhanced binding of soluble Major Histocompatibility Complex (MHC) dimers at low concentrations. In contrast, other groups have shown a complete loss of binding in activated T Cells during the same time period. Here, we resolve this contradiction by demonstrating that persistent binding behavior is controlled by the dose of antigen during primary activation. T Cells from TCR transgenic mice stimulated with high but not low dose of peptide are unable to specifically bind soluble Kb-SIY or Db-GP100 MHC dimer four days after stimulation. The behavior is not due to differences in the surface expression of TCR, CD8alpha, or CD8beta. Cytokine production mirrors the loss of binding. However, both high and low dose stimulated cells are able to bind the so-called ‘CD8 independent’ Ld-QL9 dimer, implicating the coreceptor in the binding defect. Using biophysical binding assays, k-space Image Correlation Spectroscopy, and Forster Resonance Energy Transfer between TCR and CD8, we demonstrate that TCR and CD8 are segregated, and TCR unclustered, on high dose activated T Cells, impairing CD8-dependent binding. Thus, membrane organization derived binding defects represent a mechanism for controlling T Cell responses to supraoptimal antigen.