Single-cell transcriptomics uncovers an instructive T-cell receptor role in adult gamma delta T-cell lineage commitment

After entering the adult thymus, bipotent T-cell progenitors give rise to alpha beta or gamma delta T cells. To determine whether the gamma delta T-cell receptor (TCR) has an instructive role in gamma delta T-cell lineage commitment or only "confirms" a pre-established gamma delta T-cell lineage state, we exploited mice lacking expression of LAT, an adaptor required for gamma delta TCR signaling. Although these mice showed a T-cell development block at the CD4(-)CD8(-) double-negative third (DN3) stage, 0.3% of their DN3 cells expressed intermediate levels of gamma delta TCR (further referred to as gamma delta(int)) at their surface. Single-cell transcriptomics of LAT-deficient DN3 gamma delta(int) cells demonstrated no sign of commitment to the gamma delta T-cell lineage, apart from gamma delta TCR expression. Although the lack of LAT is thought to tightly block DN3 cell development, we unexpectedly found that 25% of LAT-deficient DN3 gamma delta(int) cells were actively proliferating and progressed up to the DN4 stage. However, even those cells failed to turn on the transcriptional program associated with the gamma delta T-cell lineage. Therefore, the gamma delta TCR-LAT signaling axis builds upon a yi5 T-cell uncommitted lineage state to fully instruct adult gamma delta T-cell lineage specification.