Next Generation Sequencing Facilitates Quantitative Analysis of FOXP3-YFP-Cre and FOXP3-YFP-Cre/USP21-floxp/floxp Treg Cells Transcriptomes

Regulatory T (Treg) cells harbor immune suppressive capacity and are crucial for the maintenance of peripheral tolerance. Treg cells are considered to be heterogenic, where compromised FOXP3 expression results in the generation of exTreg cells. Here we report that the E3 deubiquitinase USP21 prevents the depletion of FOXP3 protein and restricts tissue-resident exTreg cell generation. Mice lacking USP21 in Treg cells display immune disorders characterized by spontaneous T cell activation and excessive T helper type 1 (Th1) skewing. USP21 stabilizes FOXP3 protein by mediating its deubiquitination and therefore helps to maintain the expression of Treg signature genes. Moreover, at inflamed loci, tissue-resident USP21-deficient Treg cells display a Th1-like effector phenotype. Therefore, we demonstrate how USP21 controls the identity of tissue-resident Treg cells by preventing FOXP3 loss. We purified CD4+CD25hiYFP+ Treg cells from spleens and LNs of Foxp3-YFP-Cre mice or FOXP3-YFP-Cre/USP21-floxp/floxp mice and performed RNA-seq experiments subsequently. mRNA profiles were further analyzed and compared according to standard protocols.