Single cell RNA and Single cell TCR sequencing

The T cell receptor (TCR) is instrumental in initiating intracellular signaling pathways that are vital for T cell development, repertoire selection, and the fine-tuning of effector responses to foreign antigens. Even subtle variations in TCR signal intensity can significantly influence T cell effector functions and differentiation. Despite this, the intricate mechanisms through which TCR signaling elicits a spectrum of T cell functional responses are not yet completely elucidated. In this study, we present a gain-of-function mutation of CD3e (CD3eI173A) at the germline level, which mutation has been proved to neutralizes the inhibitory effect of negatively charged lipids, thereby amplifying TCR signaling in our previous study. Our aim is to investigate how precisely modulated TCR signaling governs T cell fate determination throughout their developmental and maturation processes. Our results were intriguing: the CD3eI173A mutation specifically influenced the TCR repertoire formation of double-positive (DP) T cells during thymic selection. Notably, these CD3eI173A T cells exhibited reduced TCR signaling strength in peripheral T cells. Mechanistically, the CD3eI173A mutation led to the downregulation of AC149090.1 in the DP subset, triggering systematic changes in lipid metabolism that over-compensated for the aberrant enhancement of TCR signaling. Moreover, these adaptive changes in lipid metabolism resulted in significant dysfunction of mature T cells, as evidenced by impaired activation, proliferation, and antitumor activity of CD3eI173A-CD8+ T cells. Our findings highlight the plasticity of thymic T cell signaling and emphasize that the critical role of lipid-mediated regulation during T cell development is essential for ensuring that TCR signaling is properly calibrated to sustain immune homeostasis.