RORγt up-regulates RAG gene expression in DP thymocytes to expand the Tcra repertoire

Recombination activating gene (RAG) expression increases as thymocytes transition from the CD4-CD8- double-negative (DN) to the CD4+CD8+ double-positive (DP) stage, but the physiological importance and mechanism of transcriptional up-regulation are unknown. Here, we show that a DP-specific component of the recombination activating genes antisilencer (DPASE) provokes elevated RAG expression in DP thymocytes. Mouse DP thymocytes lacking the DPASE display RAG expression equivalent to that in DN thymocytes, but this supports only a partial Tcra repertoire due to inefficient secondary V alpha-J alpha rearrangement. These data indicate that RAG up-regulation is required for a replete Tcra repertoire and that RAG expression is fine-tuned during lymphocyte development to meet the requirements of distinct antigen receptor loci. We further show that transcription factor ROR gamma t directs RAG up-regulation in DP thymocytes by binding to the DPASE and that ROR gamma t influences the Tcra repertoire by binding to the Tcra enhancer. These data, together with prior work showing ROR gamma t to control Tcra rearrangement by regulating DP thymocyte proliferation and survival, reveal ROR gamma t to orchestrate multiple pathways that support formation of the Tcra repertoire.