Heterogeneity of CD8αα intraepithelial lymphocytes is transcriptionally conserved between TCRαβ and TCRγδ cells.

Intestinal intraepithelial lymphocytes (IELs) are a versatile population of immune cells with both effector and regulatory roles in gut immunity. Although this functional diversity is thought to arise from distinct IEL subpopulations, the heterogeneity of TCRαβ + and TCRγδ + IELs have not been well-characterized. Using scRNAseq, we identified CD8αα + T cell subsets with memory-like ( Tcf7 ⁺) and effector-like ( Prdm1 ⁺) profiles in both TCRαβ + and TCRγδ + IELs. Using CD160 and CD122 as markers of memory-like and effector-like cells, respectively, we found that while effector-like cells dominated the small intestine, memory-like IELs were more prevalent in the large intestine, suggesting a functional specialization of immune responses along the gut. Further transcriptional analysis revealed shared profiles between TCRαβ + and TCRγδ + small intestinal IEL subsets, suggesting conserved functional roles across these populations. Finally, our analysis indicated that TCRαβ + memory-like IELs arise from Tcf7⁺ double-negative (DN) precursors, and that effector-like IELs subsequently differentiate from the memory-like population. In contrast, TCRγδ + IELs appear to originate from two distinct precursor populations, one expressing Tcf7 and the other Zeb2 , indicating the presence of parallel developmental pathways within this lineage. Overall, our findings reveal that both TCRαβ + and TCRγδ + cells contain memory-like and effector-like subsets, which may contribute to the functional heterogeneity of IELs.