CARDIAC GROWTH REGULATION

心脏通过以下方式对各种血液动力学和体液刺激作出反应 心肌细胞大小和功能的改变。左心室肥厚 如由高血压或瓣膜疾病引起的是一种敏感 心肌功能障碍的发病率和死亡率的预测因子。 在本申请中，我们建议检验假设 心肌细胞收缩活动本身调节转录 (α)-肌球蛋白重链(αMHC)基因。我们已经确定了一个 转录起始点-47bp处的顺式作用元件， 指定的血流动力学反应元件(HME)，即两者 赋予收缩响应性的充要条件 在培养中自发收缩心肌细胞。HME 包含一个E-box基序，其结合蛋白(指定为HRP)是 与USF(上游刺激因子)抗原性相关。我们 从新生大鼠心肌细胞中克隆HRP的建议 文库通过其与HME序列的特异性结合的能力。 克隆的HRP在培养心肌细胞中的表达研究 确认其在收缩介导的(α)-MHC转录中的作用。 体外研究将确定它的二聚化、DNA结合和 转录激活特性，以及是否磷酸化 HRP是对机械性刺激的反应。角色 PKC(Z)在信号转导途径中的作用及其潜力 对原子核的定位将被确定。我们建议研究 甲状腺激素(TH)与收缩的联合作用 (α)-MHC基因转录调控活性的研究 理解这种基因在出生时的快速诱导 CARDIA血流动力学正在发生变化。这些激活剂的相互作用 与通用转录因子(TATA盒结合蛋白和 TAFs)将进行研究，以确定工作量的分子基础- 和激素调节(α)-MHC转录。要验证 HRP在体内的转录作用我们建议在体内过表达HRP 直接评估其对心脏血流动力学的影响 (Alpha)-MHC启动子活性并确定PKC(Z) HRP的磷酸化是其转录激活所必需的。 这些研究将促进我们对分子的理解。 工作负荷调节心脏生长和功能的途径。

The heart responds to various hemodynamic and humoral stimuli by changes in myocyte size and function. Left ventricular hypertrophy such as arises from hypertension or valvular disease is a sensitive predictor for morbidity and mortality due to myocardial dysfunction. In the present application we propose to test the hypothesis that myocyte contractile activity per se regulates the transcription of the (alpha)-myosin heavy chain (alpha-MHC) gene. We have identified a cis-acting element at -47 bp from the transcription initiation site, designated hemodynamic response element (HME), that is both necessary and sufficient to confer contractile responsiveness in spontaneously contracting cardiomyocytes in culture. The HME contains an E-box motif whose binding protein (designated HRP) is antigenically related to USF (upstream stimulatory factor). We propose to clone HRP from a neonatal rat cardiomyocyte expression library by its ability to bind specifically to the HME sequence. Expression studies of the cloned HRP is cultured cardiac myocytes will confirm its role in contractile-mediated (alpha-)-MHC transcription. In vitro studies will determine its dimerization, DNA binding and transcription activation properties, and whether phosphorylation of HRP occurs in response to the mechanogenic stimulus. The role PKC(Z) in the signal transduction pathway and its potential localization to the nucleus will be determined. We propose to study the combinatorial effects of thyroid hormone (TH) and contractile activity on transcriptional regulation of the (alpha)-MHC gene to understand the rapid induction of this gene at birth when both TH and cardia hemodynamics are changing. The interaction of these activators with general transcription factors (TATA box binding protein and TAFs) will be studied to determine the molecular basis of workload- and hormone-regulated (alpha)-MHC transcription. To verify the transcriptional role of HRP in vivo we propose to overexpress HRP in the hemodynamically unloaded heart to directly assess its effect on (alpha)-MHC promoter activity and to determine whether PKC(Z) phosphorylation of HRP is required for its transcriptional activation. These studies will advance our understanding of the molecular pathways of workload regulated cardiac growth and function.