To understand normal development and differentiation, it is necessary to determine the mechanisms by which cells initiate new programs of gene expression and promote formation of specific cell lineages. Typically, this involves activation of genes that are transcriptionally silent and that are likely incorporated into repressive chromatin structure. Evidence supports the idea that differentiation specific transcriptional regulators and enzymes that remodel chromatin structure cooperate to render genomic DNA more accessible to the transcriptional machinery. SWI/SNF enzymes alter nucleosome structure in an ATP dependent manner and facilitate transcription factor function in vitro and in vivo. Components of these enzymes are essential for embryonic development and some act as tumor suppressors. Additionally, SWI/SNF enzymes interact with other known tumor suppressors and are implicated in cell cycle control. Thus these enzymes are broadly required for normal cell function and for differentiation and development, and their mis-regulation is implicated in tumor formation. Skeletal muscle differentiation has long been a model for studying fundamental principles of tissue differentiation. Our recent studies mechanistically describe how chromatin remodeling enzymes facilitate the activation of specific myogenic genes using cell culture models. Via modification of existing methodologies, we are now also capable of examining changes in chromatin structure and regulatory protein interactions leading to gene activation during embryonic myogenesis, during adult myogenesis, and during maintenance of adult tissue, thereby giving our observations unprecedented biological relevance. In addition, we can assess the functional relevance of specific regulatory proteins in developing embryonic skeletal muscle tissue using a novel adaptation of a recently developed technique called in utero electroporation. This renewal application will focus on the contributions of SWI/SNF (Aim 1) and cooperating chromatin remodeling enzymes (Aim 2) at individual myogenic loci. We also demonstrate that SWI/SNF enzymes induce changes in higher order chromatin structure that result in rearrangement of myogenic genes during differentiation. We will pursue the mechanisms and consequences of these changes during myogenesis (Aim 3). Understanding embryonic and adult skeletal muscle differentiation and maintenance at a molecular level will have significant impact on studies of muscle regeneration and on the formation of rhabdomyosarcomas, which are tumors of myogenic derivation.
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