Human Neonatal MR1T Cells Have Diverse TCR Usage, are Less Cytotoxic and are Unable to Respond to Many Common Childhood Pathogens

Neonatal sepsis is a leading cause of childhood mortality. Understanding immune cell development can inform strategies to combat this. MR1-restricted T (MR1T) cells can be defined by their recognition of small molecules derived from microbes, self, and drug and drug-like molecules, presented by the MHC class 1-related molecule (MR1). In healthy adults, the majority of MR1T cells express an invariant α-chain; TRAV1-2/TRAJ33/12/20 and are referred to as mucosal-associated invariant T (MAIT) cells. Neonatal MR1T cells isolated from cord blood (CB) demonstrate more diversity in MR1T TCR usage, with the majority of MR1-5-OP-RU-tetramer(+) cells being TRAV1-2(-). To better understand this diversity, we performed single-cell-RNA-seq/TCR-seq (scRNA-seq/scTCR-seq) on MR1-5-OP-RU-tetramer(+) cells from CB (n=5) and adult participants (n=5). CB-derived MR1T cells demonstrate a less cytotoxic/pro-inflammatory phenotype, and a more diverse TCR repertoire. A panel of CB and adult MAIT and TRAV1-2(-) MR1T cell clones were generated, and CB-derived clones were unable to recognize several common riboflavin-producing childhood pathogens (<i>S. aureus, S. pneumoniae, M. tuberculosis</i>). Biochemical and structural investigation of one CB MAIT TCR (CB964 A2; TRAV1-2/TRBV6-2) showed a reduction in binding affinity toward the canonical MR1-antigen, 5-OP-RU, compared to adult MAIT TCRs that correlated with differences in β-chain contribution in the TCR-MR1 interface. Overall, this data shows that CB MAIT and TRAV1-2(-) MR1T cells, express a diverse TCR repertoire, a more restricted childhood pathogen recognition profile and diminished cytotoxic and pro-inflammatory capacity. Understanding this diversity, along with the functional ability of TRAV1-2(-) MR1T cells, could provide insight into increased neonatal susceptibility to infections.