Phosphorylation of Runx protein controls helper CD4⁺ T cell versus cytotoxic CD8⁺ T cell lineage choice

MHC-I- and MHC-II-selected CD4⁺CD8⁺ precursor thymocytes differentiate into cytotoxic CD8⁺ and helper CD4⁺ lineage T cells, during which suppression of Cd4 and Thpok genes by Runx-dependent-silencers in those genes is crucial to segregate the two lineages. However, how TCR signals are linked to cytotoxic-lineage-specific Cd4-Thpok silencing remains unclear. Here we show that the terminal Y residue within the evolutionarily conserved C-terminal WRPY motif in Runx1, which is essential for interacting with TLE co-repressor proteins, was phosphorylated more in CD4^-CD8⁺ thymocytes than in CD4⁺CD8^- thymocytes, inducing an interaction with TLE co-repressors for cytotoxic-lineage specific Cd4-Thpok silencing. Non-receptor tyrosine kinases Lck and Zap70 interacted with Runx in the cytoplasm more in MHC-I-signaled CD4^-CD8⁺ thymocytes than in CD4⁺CD8^- thymocytes. Collectively, these findings reveal that differential phosphorylation states at the terminal tyrosine residue in Runx connect MHC restriction with the helper versus cytotoxic T cell lineage choice.