CREATINE KINASE CONTROL ELEMENTS & CARDIAC DETERMINATION

这个项目的长期目标是了解分子 控制心肌发育和基因表达的机制。AS 我们将继续描绘顺式和跨式监管的模式体系 小鼠肌肉肌酸激酶(MCK)基因的成分是 对其在心肌培养和心脏中的表达至关重要 转基因小鼠。我们认为，对这些组件的详细分析 将有助于了解心脏的早期发育过程 肌肉决定力。具体目标是： (1.)正负顺式作用主要成分的识别 它们在心肌发育过程中调节MCK的表达。这一目标 包括重构具有拷贝数的MCK微基因座 稳定表达载体中依赖和整合位点非依赖性表达 心肌细胞和转基因小鼠。 (2)结合主要受体的心脏反式作用因子的鉴定 MCK控制区。最初的关注将集中在增强子区域 它活跃在心肌中，并与骨骼肌结合 决定因素。 (3)全长cDNA和编码主要心脏功能基因的分离 特定的MCK反式作用因子，特别是那些结合DNA元件的因子 具有增强剂特性。 (4)心肌决定因素的鉴定。 (5)心肌测定的作用和机制分析 心脏发育过程中的因素。 在本项目中获得的信息应适用于 重建心脏治疗的新方法(其中非心脏 患者的中胚层细胞可以转化为心肌和 然后用作嫁接)。它也应该适用于设计 多种心脏肌肉疾病的遗传疗法(其中心脏- 具体的MCK监管要素将与 不同的心肌DNA来纠正遗传损伤)。

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).