Chromatin accessibility analysis of CNS4-sufficient and defincient regulatory T cells, Treg cell precursors, and mature and semimature CD4 single positives

We have discovered that deletion of a conserved regulatory element upstream of the Foxp3 gene, termed CNS4, results in the abrogation of IL-2 mediated induction of Foxp3 expression during regulatory T cell development. However, the mice still harbor a reduced but functional population of Treg cells. To understand potential compensatory mechanisms driving the differentiation and expansion of regulatory T cells in the absence of CNS4, we profiled CNS4-deficient and -sufficient regulatory T cells, as well as related cell types, from the thymuses of male mice by ATAC-sequencing. Overall design: Cells from 2 mice per genotype were analyzed, of the following genotypes: CNS4FL/YFoxp3GFP/Y; CNS4KO/YFoxp3GFP/Y. Cells were sorted from mechanically disrupted thymuses based on the following markers: Treg cells: Foxp3-GFP+CD4+CD8-TCRß+; preTreg cells: CD25+Foxp3-GFP-CD4+CD8-TCRß+; mSP cells: CD25-Foxp3-GFP-CD4+CD8-TCRßhiCD24lo; smSP cells: CD25-Foxp3-GFP-CD4+CD8-TCRßloCD24hi. 40,000 - 50,000 cells per replicate were used.