TRANSCRIPTIONAL REGULATION OF MUSCLE-SPECIFIC GENES BY ELECTRICAL ACTIVITY

电活动对肌肉特异性基因的转录调控

• 批准号: 3756675
• 负责人: A BUONANNO
• 金额: --
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3756675
• 关键词: denervation; electrophysiology; gene induction /repression; genetic regulation; genetic regulatory element; genetic transcription; genetically modified animals; innervation; laboratory mouse; molecular cloning; muscle cells; muscle proteins; myogenesis; nicotinic receptors; nucleic acid sequence; striated muscles; tissue /cell culture; transcription factor; troponin

Skeletal muscle plasticity is dramatically influenced by motoneuron innervation. We have identified muscle genes that are either repressed or stimulated by specific patterns of nerve-derived electrical activity; our goal is to elucidate the molecular mechanisms underlying activity-dependent transcriptional regulation. To this end, we previously demonstrated that myogenin, a muscle-specific factor that regulates transcription of nicotininc acetylcholine receptor (nAChR) genes in cultured cells, is repressed by innervation and de-repressed by denervation. Changes in myogenin levels in muscle preceded receptor regulation during development and after denervation, suggesting that regulation of myogenin may lie upstream of receptor. Analyses of a 3.7 and 1.5 kb myogenin upstream region in transgenic mice have shown that these sequences confer muscle-, developmental-, and denervation-specific regulation. We have developed a modification of a myoblast implantation technique to map transcription regulatory sequences in mature skeletal muscle. Using this technique, we have delineated a 450 bp region that confers the denervation response. To begin mapping the DNA sequences that stimulate the expression of the troponin I slow (TnIs) gene in response to specific patterns of electrical activity, we have begun to map the promoter in cultured C2C12 muscle cells and transgenic mice. Delineation of TnIs upstream sequences in cultured myocytes, demonstrated that 200 bp are necessary and sufficient to confer muscle- and developmental-specific transcription; MyoD and MEF-2 binding sites were shown to be required for activity. However, this region does not direct transcription of the gene in the adult slow muscles of transgenic mice; there is a requirement for sequences further upstream. We have begun to clone and characterize putative trans-acting factors that may regulate myogenin and TnIs transcription. We have isolated cDNAs coding for a novel member of the ets-family and a rat homolog of MEF-2. The functional properties of these factors are currently under investigation. Analysis of the interactions of these families of trans-acting factors should help elucidate the molecular mechanisms underlying fiber-type specific regulation of muscle genes by innervation.

运动神经元对骨骼肌的可塑性有显著影响 神经支配我们已经确定了肌肉基因， 由特定的神经电活动模式刺激;我们的 目的是阐明潜在的分子机制 活性依赖性转录调节。为此，我们此前 证明了肌细胞生成素，一种肌肉特异性因子， 烟碱乙酰胆碱受体基因转录 培养的细胞，被神经支配抑制， 去神经肌前受体中肌细胞生成素水平的变化 在发育过程中和去神经支配后的调节，这表明， 肌细胞生成素的调节可能位于受体的上游。分析3.7 和1.5kb肌细胞生成素上游区的研究表明， 这些序列赋予肌肉、发育和去神经特异性 调控我们开发了一种改良的成肌细胞移植 在成熟骨骼肌中定位转录调节序列技术 肌肉.使用这种技术，我们已经划定了一个450 bp的区域， 赋予去神经支配反应。 为了开始绘制刺激细胞表达的DNA序列， 肌钙蛋白I减慢（TnIs）基因对特定模式的反应 电活性，我们已经开始在培养的C2 C12中定位启动子 肌肉细胞和转基因小鼠。TnIs上游序列的描述 在培养的心肌细胞中，证明200 bp是必需的， 足以赋予肌肉和发育特异性转录; 显示MyoD和MEF-2结合位点是活性所需的。 然而，该区域并不指导基因在细胞中的转录。 成年转基因小鼠的慢肌;需要 更上游的序列。我们已经开始克隆和描述 可能调节肌细胞生成素和肌钙蛋白I的反式作用因子 转录。我们已经分离出编码一个新成员的cDNA， ets家族和MEF-2的大鼠同系物。这些的功能特性 目前正在调查这些因素。相互作用分析 这些家族的反式作用因子应该有助于阐明 肌纤维型特异性调节的分子机制 神经支配的基因。