Immunoglobulin gene expression requires rearrangement of heavy and light chain gene segments during B cell development, that brings promoter elements into the proximity of enhancer elements. The focus of this proposal is to continue to study events that control Ig gene rearrangement and expression early in B cell ontogeny, in both mouse and human. This includes characterization of elements which regulate germline transcription of the kappa locus, and their potential role in targeting gene segments for rearrangement. The developmental activation of the kappa intron and 3' enhancers will be examined by transgenic mouse studies. The specific role of the intron enhancer will be determined by examining the effects of enhancer deletion in A-MuLV transformed cell lines that normally rearrange kappa genes, and ES cells used to generate genetically altered mice. The structural and functional components that comprise the kappa intron enhancer will be studied by defining sequences and protein-DNA interactions that contribute to transcriptional activation and suppression in model cell culture systems. This includes cloning a novel, inducible factor that binds a region in the enhancer, as well as defining requirements for spatial arrangements of all the sequence motifs that contribute to full enhancer activity. Sequences that flank the core intron enhancer and act as silencers of enhancer activity will be characterized. A number of these studies will involve direct comparisons of mouse and human enhancer organization and function. Finally, an ongoing collaborative study will be maintained to study the T cell dependent, heavy chain enhancer-mediated suppression of IgE expression in murine hybridomas.
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