Transcriptional regulation in vivo involves both local and long-range interactions that occur within specialized """"""""open"""""""" chromosomal domains. Composite DNA elements which induce the formation of these structures are termed Locus Control Regions (LCRs). Transcriptional enhancers within LCRs initiate local changes in otherwise condensed higher-order chromatin structures, which can be propagated to more distal regions in the presence of nuclear matrix attachment regions (MARs). These LCR-induced changes in chromatin accessibility are required for functional interactions between widel separated enhancers and promoters. These effects have been described in transgenic mouse experiments and are, consequently difficult to study at the mechanistic level. To this end, the PI has recently developed a transfection method which reveals the functional contribution of each of the subelements within a well described LCR. This assa will expedite the identification of novel DNA regulatory elements and trans-acting factors that participate in long-range remodeling of chromatin. The immunoglobulin mu LCR consists of a classical enhancer element and flankin MARs. To understand how these elements collaborate to remodel large chromatin domains and permit interactions between the enhancer and distal promoters, the PI proposes the following 3 specific aims: 1) To identify the specific MAR sequences necessary for LCR function, 2) To identify MAR-binding protein factors which are a functional component of the active mu LCR, and 3) To investigate the mechanism by which the MARs and MAR-binding factors govern distal enhancer function and chromatin remodeling. LCRs, and the mechanisms by which they govern gene expression, are likely to b used at many, if not all, developmentally regulated loci. These studies will yield novel insights into LCR structure and function which should expand opportunities for regulated and targeted gene therapy.
