Transcriptional regulation by TEF-1 in cardiac myocytes

描述(申请人提供)：心血管疾病是发达国家的主要死亡原因，晚期患者预后非常差。充血性心力衰竭是许多心血管问题的最终结果，也是美国最昂贵的健康问题之一。虽然心血管疾病的治疗有了很大的改善，但预后仍然很差，因此有必要开发新的治疗方法。在心脏发育、正常生命和心血管疾病期间，多种转录因子及其辅助因子(Nkx2、MEF2、NFAT、SRF、GATA、HDACs、HATS和TEF-1)控制基因表达的复杂变化。已知这些因子中的一些但不是全部的活性是如何在总水平上被调节的，例如TEF-1。这项提案中的研究将集中于确定TEF-1基因家族在心脏转录调控中的作用。这些研究将使用新生大鼠心肌细胞，这是一个成熟的正常和疾病心脏的体外模型系统。这项研究将解决这样一个假设，即MCAT位点、TEF-1蛋白、它们的辅因子和其他心脏转录因子之间的相互作用是在正常和肥大心肌细胞中产生复杂的基因表达模式的关键。这些实验将(1)确定TEF-1是否调节正常和肥厚心肌细胞(显性负蛋白，siRNA)的内源性心脏基因表达；(2)确定MCAT核心序列是否通过影响TEF-1及其辅因子的相互作用来影响MCAT位点的活性；(3)通过MS/MS分离新的TEF-1辅因子；(4)研究TEF-1辅因子(YAP65、TAZ、CK2)和TEF-1如何在正常和肥厚条件下相互作用以及与其他心脏转录因子相互作用来调节心肌细胞的基因表达。将使用的方法包括使用腺病毒载体表达显性负TEF-1蛋白。这些分析将详细说明正常和疾病心肌细胞中启动子对特定信号的反应机制，从而改进心脏病的治疗方法。

DESCRIPTION (provided by applicant): Cardiovascular diseases are the leading cause of death in developed countries, with a very poor prognosis for patients in advanced stages. Congestive heart failure, the end result of many cardiovascular problems is one of the most expensive health problems in the United States. Although there have been dramatic improvements in the treatment of cardiovascular diseases their continued poor prognosis makes the development of novel therapies necessary. During cardiac development, during normal life and during cardiovascular disease there are complex changes in gene expression controlled by multiple transcription factors and their cofactors (Nkx2, MEF2, NFAT, SRF, GATA, HDACs, HATs and TEF-1). It is known how the activity of some but not all of these factors, such as TEF-1 is regulated at a gross level. The research in this proposal will concentrate on determining the role of the TEF-1 gene family in transcriptional regulation in the heart. The studies will use rat neonatal cardiac myocytes, a well established in vitro model system of normal and diseased heart. The research in this grant will address the hypothesis that interactions between MCAT sites, TEF-1 proteins, their cofactors, and other cardiac transcription factors are key in generating the complex patterns of gene expression in normal and hypertrophic cardiac myocytes. The experiments will (1) determine if TEF-1 regulates endogenous cardiac gene expression in normal and hypertrophic cardiac myocytes (dominant negative proteins, siRNA), (2) determine if MCAT core sequence affects the activity of MCAT sites by influencing the interaction of TEF-1 with its cofactors, (3) isolate novel TEF-1 cofactors by state of the art MS/MS, and (4) study how the TEF-1 cofactors (YAP65, TAZ, CK2) and TEF-1 interact with each other and with other cardiac transcription factors to regulate gene expression in cardiac myocytes under normal and hypertrophic conditions. The approaches that will be used include expression of dominant negative TEF-1 proteins using adenoviral vectors. These analyses will detail the mechanisms of promoter response to specific signals in normal and diseased cardiac myocytes, leading to improved treatments for heart disease.