To understand how ligand affinity affects TCR signaling, Voisinne and Locard-Paulet et al. probed molecular-level responses of OT-1 T cells to SIINFEKL (N4, high-affinity) versus SIITFEKL (medium affinity) or SIIGFEKL (low affinity). N4 stimulation led to the greatest T cell proliferation, cytokine production, and global protein phosphorylation. Specific Zap70 and downstream Lat phosphorylation and Slp76 complex formation were reduced at low affinity, although Cd3 ITAM phosphorylation and Cd3ζ-Zap70 complex formation were unchanged. Inactivating Cd6 improved IFNγ response to low-affinity peptide stimulation, perhaps by reducing its negative signals.
Contributed by Alex Najibi
ABSTRACT: T cells recognize a few high-affinity antigens among a vast array of lower affinity antigens. According to the kinetic proofreading model, antigen discrimination properties could be explained by the gradual amplification of small differences in binding affinities as the signal is transduced downstream of the T cell receptor. Which early molecular events are affected by ligand affinity, and how, has not been fully resolved. Here, we used time-resolved high-throughput proteomic analyses to identify and quantify the phosphorylation events and protein-protein interactions encoding T cell ligand discrimination in antigen-experienced T cells. Although low-affinity ligands induced phosphorylation of the Cd3 chains of the T cell receptor and the interaction of Cd3 with the Zap70 kinase as strongly as high-affinity ligands, they failed to activate Zap70 to the same extent. As a result, formation of the signalosome of the Lat adaptor was severely impaired with low- compared with high-affinity ligands, whereas formation of the signalosome of the Cd6 receptor was affected only partially. Overall, this study provides a comprehensive map of molecular events associated with T cell ligand discrimination.