Graduate Student St. Jude Children�s Research Hospital Memphis, Tennessee, United States
Disclosure(s):
Logan Rice: No financial relationships to disclose
Introduction/Rationale: KIR+CD8+ T cells are present in diverse tissues and disease states, including autoimmunity, infection, and cancer. Interest in these cells has recently increased due to their detection by single-cell transcriptomics in these contexts. However, their development and function remain poorly understood. They show gene expression and T cell receptor (TCR) features associated with autoreactivity and agonist selection, a developmental pathway shared by intraepithelial lymphocytes (IELs), another unconventional T cell population thought to contribute to peripheral tolerance in the small intestine. This study investigates the ontogeny of Ly49+CD8+ T cells, the murine equivalent of human KIR+CD8+ T cells, through single cell transcriptomic analysis of IEL precursors, with the aim of identifying Klra6+ cells, a specific marker for Ly49+CD8+ T cells.
Methods: Mature IEL precursors were isolated from wild-type C57BL/6 mouse thymi using a defined antibody panel (CD1d-Tet- CD25- CD4- CD8- CD5+ TCRβ+ H-2kb+ CD122+). Hashtag antibodies identified the mouse of origin and CITE-seq antibodies distinguished type-A (PD1+ NK1.1- CD44- CXCR3-) from type-B (PD1- NK1.1+ CD44+ CXCR3+) precursors. Single-cell gene expression, CITE-seq, and TCR library data were generated with the 10x Genomics platform and analyzed using Seurat in R.
Results: IEL precursors separated into two major groups, forming 10 clusters. Previous findings have shown that IEL precursors can be separated into two groups named type A and type B. To determine if this distinction drove clustering, module scores based on PD1, NK1.1, CD44, and CXCR3 expression were used to assign clusters as A-like or B-like. This analysis confirmed that A/B identity explained the major separation.
Conclusion: Klra6+ cells were distributed across both A-like and B-like clusters, rather than clustering together, suggesting the existence of two distinct Ly49+CD8+ T cell populations: one potentially functioning within the thymus and another contributing to peripheral tolerance.
