A1-ADRENERGIC REGULATION OF CARDIAC GENE EXPRESSION

这项工作的重点是心肌肥厚的临床问题， 特别是调节这一过程的分子机制。 一个新 建立了一个研究心肌肥厚的模型系统， 无血清培养的心肌细胞。 有人提出了一个新的意见， 刺激这些细胞上的α 1-肾上腺素能受体， 儿茶酚胺去甲肾上腺素诱导细胞增大或肥大， 没有DNA合成。 α 1肾上腺素能受体是第一个 受体显示调节心肌细胞肥大。 最近的研究表明，α 1受体调节mRNA的表达 在肥大期间。 特别是，α 1刺激诱导两个 早期心脏发育特征性收缩蛋白iso-mRNAs 和体内压力负荷肥大，骨骼α-肌动蛋白同工酶mRNA和 B-肌球蛋白重链同功mRNA。 这种α 1受体效应是选择性的 并在同基因转录水平上介导。 因此 关键的问题是诱导转录的机制， α 1受体 目前的实验将测试骨骼α- 肌动蛋白同工基因和B-肌球蛋白重链同工基因含有一个或多个 α 1响应元件，即，DNA序列需要专门为 这些基因的转录响应α 1-肾上腺素能 刺激. 两种互补的功能鉴定实验方法 提出了α 1反应元件：（1）杂交基因的转染 在瞬时测定系统中;和（2）DNA酶I超敏反应作图。 必要的克隆基因可用于开始这些实验， 初步研究证明这项工作是可行的。 长期的目标是从α-淀粉样蛋白的表达来确定细胞内的通路。 1受体对基因转录的影响。 整体 一种假说是受体刺激激活了一种预先存在的蛋白质 或与α 1反应元件结合的蛋白质， RNA聚合酶II的转录复合物。标识 本工作中的α 1响应元件或元件将提供 为随后鉴定一种或多种蛋白质奠定基础。

This work is focused on the clinical problem of myocardial hypertrophy, specifically the molecular mechanisms which regulate this process. A new model system was developed to study hypertrophy, employing neonatal heart myocytes in serum-free culture. The novel observation was made that stimulation of the alpha 1-adrenergic receptor on these cells by the catecholamine norepinephrine induces cell enlargement or hypertrophy, without DNA synthesis. The alpha 1-adrenergic receptor is the first receptor shown to regulate cardiac myocyte hypertrophy. Recent work has shown that the alpha 1 receptor regulates mRNA expression during hypertrophy. In particular, alpha 1 stimulation induces two contractile protein iso-mRNAs characteristic of early cardiac development and pressure-load hypertrophy in vivo, skeletal alpha-actin iso-mRNA and B-myosin heavy chain iso-mRNA. This alpha 1 receptor effect is selective and is mediated at the level of iso-gene transcription. Thus, the critical question is the mechanism for induction of transcription by the alpha 1 receptor. The present experiments will test the hypothesis that the skeletal alpha- actin iso-gene and the B-myosin heavy chain iso-gene contain one or more alpha 1 response elements, i.e., DNA sequences required specifically for transcription of these genes in response to alpha 1-adrenergic stimulation. Two complementary experimental approaches for functional identification alpha 1 response elements are proposed: (1) transfection of hybrid genes in a transient assay system; and (2) DNase I hypersensitivity mapping. The necessary cloned genes are available to begin these experiments and preliminary studies provide evidence that the work is feasible. The long-range goal is to define the intracellular pathway from the alpha 1 receptor at the cell surface to gene transcription. The overall hypothesis is that receptor stimulation activates a pre-existing protein or proteins that binds to the alpha 1 response element in a transcriptional complex with RNA polymerase II. Identification of an alpha 1 response element or elements in the present work will provide the foundation for subsequent identification of the protein or proteins.