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Single-cell profiling of goose thymus uncovers the developmental program and regulatory logic of T cell differentiation

ABSTRACT: The thymus is essential for T cell development and the establishment of central tolerance, yet avian thymic development remains poorly characterized. Using the Zhedong White goose as a model, we analyzed thymic development across four stagesby integrating histological analysis with single-cell RNA sequencing. The goose thymus exhibits a multilobulated architecture, undergoes rapid post-hatch maturation, and shows stage-dependent involution. Single-cell analysis identified the major thymic cell populations, with T cells predominating and γδ T cells present at higher proportions than in mammals. Pseudotime analysis revealed double-positive (DP) cells as a central differentiation hub. Double-negative (DN) thymocytes progressed from DN1 to DN3b through a stepwise program characterized by lineage commitment marked by upregulation of BCL11B, NOTCH1, and TCF7, followed by induction of β-selection–associated genes, including RAG1/2 and DNTT. Thymic epithelial cells functioned as a major signaling hub, with adhesion- and microenvironment-related pathways coordinating TEC–T cell interactions, while DP and mature T cells progressively emerged as key signal integration nodes. Cross-species comparison indicated an overall conserved developmental framework, with the most pronounced species-specific differences observed at the DP stage. Collectively, this study systematically delineates the developmental trajectories of thymic T cells and their microenvironmental regulation in the goose, providing a valuable resource for studies of avian immune development and comparative immunology.

ORGANISM(S): Anser cygnoides

PROVIDER: GSE315444 | GEO | 2026/02/28

REPOSITORIES: GEO

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Project description:The current model of hematopoiesis only resolves the thymus around the production and establishment of the peripheral T cell pool. However, the role and in situ development of other immune subsets has been overlooked. We aim to demonstrate that during TL1A and IL-18 induced inflammation, the thymus is capable of producing other cell types such as neutrophils, monocytes, and macrophages and that these potentially have other functions aside from supporting T cell development as scavengers. Here we have proven that ex vivo and in vivo treatment with TL1A and IL-18 results in acute thymic atrophy by a massive loss of DP T cells and shrinking of the thymic lobe. By electron microscopy, flow cytometry and single cell we demonstrate that neutrophils are able to maturate inside the thymus lobe in a NOTCH-independent manner. We used fate-mapping tools to elucidate the origin of thymic neutrophils since we observed a great expansion in our culture that is isolated from the influx of BM progenitors. The Rag1-Cre Rosa26-YFP fate mapping model revealed that neutrophils share a common progenitor with T cells, while monocytes/macrophages do not. Furthermore, we found that most of the thymic GMPs (defined as Lin-Sca-1-c-Kit+CD16/32+CD34+) show history of Rag1 expression in comparison with bone marrow GMPs. Additionally, by using Ms4a3-Cre Rosa26Tdtomato fate mapping model we observed that there was not difference in Tdt labeling between the thymus and bone marrow, suggesting that thymic neutrophils still undergo conventional neutrophils pathway arising from GMPs. Moreover, we show that thymic-derived neutrophils are functional and are capable of forming extracellular neutrophils traps (NETs) similarly to benchmark peritoneal neutrophils. We found that the expansion of thymic neutrophils is GM-CSF dependent by using Csf2rb KO mice. Additionally, we identified DR3+ and IL-18Rα+ expressing subsets of ILCs and gdT cells as the cellular source GM-CSF. Lastly, in vivo treatment with TL1A+IL-18 lead to emergency granulopoiesis and an increase of neutrophils in all the organs investigated, including the thymus.

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Single-cell profiling of goose thymus uncovers the developmental program and regulatory logic of T cell differentiation

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