GSE137699
Single cell sequencing defines a branched progenitor population of stable medullary thymic epithelial cells
Medullary thymic epithelial cells (mTECs) play a critical role in central immune tolerance by mediating negative selection of autoreactive T cells through the collective expression of the peripheral self-antigen compartment, including tissue-specific antigens (TSAs). Recent work has shown that gene expression patterns within the mTEC compartment are remarkably heterogenous and include multiple differentiated cell states. To further define mTEC development and medullary epithelial lineage relationships, we combined lineage tracing and recovery from transient in vivo mTEC ablation with single cell RNA-sequencing in Mus musculus. The combination of bioinformatic and experimental approaches revealed a non-stem transit-amplifying population of cycling mTECs that preceded Aire expression. Based on our findings, we propose a branching model of mTEC development  wherein a heterogeneous pool of transit-amplifying cells gives rise to Aire- and Ccl21a-expressing mTEC subsets. We further use experimental techniques to show that within the Aire-expressing developmental branch, TSA expression peaked as Aire expression decreased, implying Aire expression must be established before TSA expression can occur. Collectively, these data provide a higher order roadmap of mTEC development and demonstrate the power of combinatorial approaches leveraging both in vivo models and high-dimensional datasets. Overall design: Lineage trace and ablation-recovery of medullary thymic epithelial cells to determine lineage relationships