Non-agonist peptide-MHC engagement triggers TCR proximal signaling distinct from agonists.

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Abstract The maintenance of naïve and memory T cell populations, is regulated by signals from cytokines including IL-7 as well as those ensuing from the tonic TCR recognition of endogenous peptide-MHC (pMHC) complexes. These TCR signals are thought to be distinct from typical agonist antigen-driven signals, but the precise biochemistry is poorly understood. In mice, the 5C.C7 TCR transgenic model has well characterized interactions with agonist and non-agonist foreign and self-peptides, respectively. Using these cells, we find that agonist pMHC engagement elicits strong phosphorylation of ERK, actin polymerization, increase of cell size as well as upregulation of surface markers such as CD69. Interestingly, co-culturing 5C.C7 T cells with both the agonist and non-agonist pMHC complexes amplifies these phenotypes of T cell activation when compared to the agonist alone. Additionally, we found that endogenous peptides alone do not directly elicit these downstream cellular responses. In contrast, examination of more TCR proximal signaling suggests that endogenous peptide engagement results in strong LAT phosphorylation comparable to levels triggered by the agonist. These non-agonist-induced phosphorylated LAT levels were present from 30 minutes until 4 hours at a relatively sustained level, compared to the synergistic effects of the downstream signaling (pERK, CD69, etc) which only began to emerge at the 2 hour time point and onwards. These data suggest that TCR signaling after engaging with endogenous self-peptides is uncoupled from agonistic signaling at the level of LAT. This bifurcation may allow T cells to use agonist vs non-agonistic signals for distinct biologic outcomes.