Definition of a T-cell receptor beta gene core enhancer of V(D)J recombination by transgenic mapping

V(D)J recombination in differentiating lymphocytes is a highly regulated pr ocess in terms of both cell lineage and the stage of cell development. Tran sgenic and knockout mouse studies have demonstrated that transcriptional en hancers from antigen receptor genes play an important role in this regulati on by activating cis-recombination events. A striking example is the T-cell receptor beta-chain (TCR beta) gene enhancer (E beta), which in the mouse consists of at least seven nuclear factor binding motifs (beta E1 to beta E 7), Here, using a well-characterized transgenic recombination substrate app roach, we define the sequences within E beta required for recombination enh ancer activity. The E beta core is comprised of a limited set of motifs (be ta E3 and beta E4) and an additional previously uncharacterized 20-bp seque nce 3' of the beta E4 motif. This core element confers cell lineage- and st age-specific recombination within the transgenic substrates, although it ca nnot bypass the suppressive effects resulting from transgene integration in heterochromatic centromeres. Strikingly, the core enhancer is heavily occu pied by nuclear factors in immature thymocytes, as shown by in vivo footpri nting analyses. A larger enhancer fragment including the beta E1 through be ta E4 motifs but not the 3' sequences, although active in inducing germ lin e transcription within the transgenic array, did not retain the E beta reco mbinational activity, Our results emphasize the multifunctionality of the T CR beta enhancer and shed some light on the molecular mechanisms by which t ranscriptional enhancers and associated nuclear factors may impact on cis r ecombination, gene expression, and lymphoid cell differentiation.