Nizhnik, A.; Ulloa, B. A.; Jackson, K.; Zheng, D.; Bowman, T. V.

During embryogenesis, hematopoietic stem cells (HSC) and HSC-independent progenitor cells form concurrently and produce overlapping and distinct immune cell types. The full repertoire of immune cells generated during development and their origin across vertebrates is incompletely elucidated. Here, we performed temporal lineage tracing of the emerging hematopoietic system during zebrafish larval development followed by single cell RNA sequencing and identified a greater diversity of lymphoid cells than previously recognized. The cells are comprised of T-lymphocytes and Innate Lymphoid-like Cells (ILCs) that are Runx1-dependent. Both T-lymphocytes and ILC-like cells depended on Il2r-gamma; while only T-lymphocytes depended on Rag1. The larval ILC-like cells were detected in lymphoid and mucosal organs and were responsive to viral mimicry-induced stimulation, indicating their functionality in early vertebrate life. The work provides new fundamental knowledge on the early establishment of immune hierarchies and opens the zebrafish model to broader exploration of lymphoid immunity origination and function.