The medulla controls effector primed γδT-cell development in the adult mouse thymus.

γδT cells are produced in the thymus throughout life and provide immunity at epithelial-rich sites. Unlike conventional αβT cells, γδT-cell development involves intrathymic acquisition of effector function, with priming for either IL17 or IFN-γ production occurring during embryonic or adult life, respectively. How the thymus controls effector-primed γδT-cell generation in adulthood is poorly understood. Here, we distinguished de novo γδT cells from those undergoing thymus recirculation and/or retention using Rag2GFP mice alongside markers of maturation/effector priming including CD24, CD25, CD73, and IFN-γ, the latter by crossing with IFN-γ GREAT mice. We categorize newly developing γδT-cells into an ordered sequence where CD25 CD73 IFN-γ precursors are followed sequentially by CD25 CD73 IFN-γ intermediates and CD25 CD73 IFN-γ effectors. To determine intrathymic requirements controlling this sequence, we examined γδT-cell development in Relb thymus grafts that lack medullary microenvironments. Interestingly, medulla deficiency did not alter CD25 γδT-cell precursor generation, but significantly impaired development of effector primed stages. This impact on γδT-cell priming was mirrored in plt/plt mice lacking the medullary chemoattractants CCL19 and CCL21, and also Ccl21a but not Ccl19 mice. Collectively, we identify the medulla as an important site for effector priming during adult γδT-cell development and demonstrate a specific role for the medullary epithelial product CCL21 in this process.