Regulation of IgH gene assembly:: Role of the intronic enhancer and 5′DQ52 region in targeting DHJH recombination

The assembly of Ag receptor genes by V(D)J recombination is regulated by transcriptional promoters and enhancers which control chromatin accessibility at Ig and TCR gene segments to the RAG-1/RAG-2 recombinase complex. Paradoxically, germline deletions of the IgH enhancer (E mu) only modestly reduce D-H -> J(H) rearrangements when assessed in peripheral B cells. However, deletion of E mu severely impairs recombination of V-H gene segments, which are located over 100 kb away. We now test two alternative explanations for the minimal effect of E mu deletions on primary D-H -> J(H) rearrangement: 1) Accessibility at the D(H)J(H) cluster is controlled by a redundant cis-element in the absence of E mu. One candidate for this element lies 5' to D-Q52 (PDQ52) and exhibits promoter/enhancer activity in pre-B cells. 2) In contrast to endpoint B cells, D-H -> J(H) recombination may be significantly impaired in pro-B cells from enhancer-deficient mice. To elucidate the roles of PDQ52 and E-mu in the regulation of IgH locus accessibility, we generated mice with targeted deletions of these elements. We report that the defined PDQ52 promoter is dispensable for germline transcription and recombination of the D(H)J(H) cluster. In contrast, we demonstrate that E mu directly regulates accessibility of the D(H)J(H) region. These findings reveal a significant role for E mu in the control mechanisms that activate IgH gene assembly and suggest that impaired V-H -> D(H)J(H) rearrangement in enhancer-deficient cells may be a downstream consequence of the primary block in D-H -> J(H) recombination.