GENE-TARGETED DELETION AND REPLACEMENT MUTATIONS OF THE T-CELL RECEPTOR BETA-CHAIN ENHANCER - THE ROLE OF ENHANCER ELEMENTS IN CONTROLLING V(D)J RECOMBINATION ACCESSIBILITY

To assess the role of transcriptional enhancers in regulating accessib ility of the T-cell receptor beta-chain (TCR beta) locus, we generated embryonic stem cell lines in which a single allelic copy of the endog enous TCR beta enhancer (E beta) was either deleted or replaced with t he immunoglobulin heavy-chain intronic enhancer. We assayed the effect s of these mutations on activation of the TCR beta locus in normal T- and B-lineage cells by RAG-2 (recombination-activating gene 2)-deficie nt blastocyst complementation. We found that E beta is required for re arrangement and germ-line transcription of the TCR beta locus in T-lin eage cells. In the absence of E beta, the heavy-chain intronic enhance r partially supported joining region beta-chain rearrangement in T- bu t not in B-lineage cells. However, ability of the heavy-chain intronic enhancer to induce rearrangements was blocked by linkage to an expres sed neomycin-resistance gene (neo(r)). These results demonstrate a cri tical role for E beta in promoting accessibility of the TCR beta locus and suggest that additional negative elements may cooperate to furthe r modulate this process.