CREATINE KINASE CONTROL ELEMENTS & CARDIAC DETERMINATION

该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。 作为 一个模型系统，我们将继续划定顺式和反式监管 小鼠肌肉肌酸激酶（MCK）基因的组成部分， 对于其在心肌培养物和心脏中的表达至关重要 转基因小鼠。 我们相信对这些成分的详细分析 将有助于了解心脏的早期发育过程， 肌肉的决心 具体目标是： （1.） 主要正、负顺式作用元件的鉴定 其在心肌发育期间调节MCK表达。 这一目标 包括重建MCK微型基因座，所述MCK微型基因座表现出拷贝数- 依赖性和整合位点非依赖性表达 心肌细胞和转基因小鼠。 （二） 鉴定与心肌细胞主要膜结合的心脏反式作用因子 MCK控制区。 最初的注意力将集中在增强子区域 它在心肌中很活跃， 决定因素 （3.） 分离全长cDNA和编码主要心脏- 特异性MCK反式作用因子，尤其是结合DNA元件的那些 具有增强剂特性。 （4.） 心肌决定因子的鉴定。 （五） 心肌决定的作用及机制分析 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 来自患者的中胚层细胞可以转化为心肌， 然后用作移植物）。 它也应该适用于设计 用于各种心脏肌肉疾病的基因疗法（其中心脏- 特定的MCK调节元件将与 不同的心肌cDNA以矫正遗传损伤）。

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).