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

Recombination activating gene (RAG) expression increases as thymocytes transition from the CD4CD8 double-negative (DN) to the CD4CD8 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 repertoire due to inefficient secondary Vα-Jα rearrangement. These data indicate that RAG up-regulation is required for a replete 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 RORγt directs RAG up-regulation in DP thymocytes by binding to the DPASE and that RORγt influences the repertoire by binding to the enhancer. These data, together with prior work showing RORγt to control rearrangement by regulating DP thymocyte proliferation and survival, reveal RORγt to orchestrate multiple pathways that support formation of the repertoire.