Mechanisms regulating immunoglobulin gene transcription

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DESCRIPTION (provided by applicant): One of the first steps in successful B cell development is establishing an accessible, or open, immunoglobulin mu (IgM) chromatin structure. Increased accessibility is critical for converting the quiescent IgM locus of the hematopoietic precursor into a biologically functional, or activated, locus in the first committed member of the B lineage. However, the mechanisms regulating IgM accessibility are unknown. The proposed work will focus on determining how an inaccessible, or closed, mu chromatin structure becomes accessible during B cell development then remains accessible in the mature B cell. This work will answer the question: how is the mu enhancer activated in the context of chromatin? Completing the proposed work represents a first step towards the long-term goal of characterizing mechanisms driving tissue-specific regulation of IgM transcription. Work from multiple investigators spanning 20 years suggests that alterations in chromatin packaging regulate mu enhancer activation during B cell development. Increased mu enhancer accessibility, a prelude to full activation, correlates with transcription factor binding and modification of histone proteins packaging the mu enhancer. However, a direct "cause and effect" relationship showing transcription factors directing changes in chromatin structure has not been established. Overall, we will test the hypothesis that the mu enhancer, and hence B cell development is regulated by transcription factor-directed histone modifications through three approaches: 1. We will define the combination of proteins required for establishing maximal accessibility from a naturally chromatinized mu locus by ectopically expressing transcription factors in cells; 2. We will destroy DNA binding activity of the accessibility factors then measure stability of the open mu chromatin structure; 3. We will test how transcription factors that induce mu accessibility affect acetylation and methylation of the histones packaging the mu enhancer by chromatin immunoprecipitation. Understanding the details of mu locus accessibility during B cell development will provide logical targets for pharmaceutically controlling B cell generation in disease states by either blocking or enhancing this developmentally critical process. In addition understanding the rules for tissue-specific gene expression will allow exquisitely regulated delivery of treatments for a variety of single tissue syndromes from cancer to diabetes.