CREATINE KINASE CONTROL ELEMENTS & CARDIAC DETERMINATION

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The long-term objectives of this project are to understand the molecular mechanisms which control heart muscle development and gene expression. As a model system we will continue delineating the cis- and trans regulatory components of the mouse muscle creatine kinase (MCK) gene which are essential for its expression in cardiac muscle cultures and in the hearts of transgenic mice. We believe that detailed analyses of these components will lead to understanding the earlier developmental process of cardiac muscle determination. The specific aims are: (1.) Identification of major positive and negative cis-acting elements which regulate MCK expression during cardiac muscle development. This aim includes reconstructing an MCK mini-gene-locus which exhibits copy number- dependent and integration site-independent expression in stably transfected cardiac muscle cells and transgenic mice. (2.) Identification of cardiac trans-acting factors that bind to the major MCK control regions. Initial attention will focus on an enhancer region that is active in cardiac muscle and which binds skeletal muscle determination factors. (3.) Isolation of full-length cDNAs and genes encoding the major cardiac- specific MCK trans-acting factors, especially those which bind DNA elements with enhancer properties. (4.) Identification of cardiac muscle determination factors. (5.) Analyses of the role and mechanism of cardiac muscle determination factors during heart development. Information obtained in this project should be applicable to the design of new methods for reconstructive heart therapy (in which non-cardiac mesodermal cells from a patient could be transformed into heart muscle and then used as grafts). It should also be applicable to the design of genetic therapies for a variety of heart muscle diseases (in which cardiac- specific MCK regulatory elements would be used in conjunction with different cardiac muscle cDNAs to rectify genetic lesions).