RAG1/RAG2 (RAG) endonuclease-mediated assembly of diverse lymphocyte Ag receptor genes by V(D)J recombination is critical for the development and immune function of T and B cells. The RAG1 protein contains a ubiquitin ligase domain that stabilizes RAG1 and stimulates RAG endonuclease activity in vitro. We report in this study that mice with a mutation that inactivates the Rag1 ubiquitin ligase in vitro exhibit decreased rearrangements and altered repertoires of TCRβ and TCRα genes in thymocytes and impaired thymocyte developmental transitions that require the assembly and selection of functional TCRβ and/or TCRα genes. These Rag1 mutant mice present diminished positive selection and superantigen-mediated negative selection of conventional αβ T cells, decreased genesis of invariant NK T lineage αβ T cells, and mature CD4+ αβ T cells with elevated autoimmune potential. Our findings reveal that the Rag1 ubiquitin ligase domain functions in vivo to stimulate TCRβ and TCRα gene recombination and influence differentiation of αβ T lineage cells, thereby establishing replete diversity of αβ TCRs and populations of αβ T cells while restraining generation of potentially autoreactive conventional αβ T cells.
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