REGULATION OF CONNEXIN-43 GENE EXPRESSION IN THE UTERUS

这项研究计划描述了旨在揭示 连接蛋白43基因在子宫肌层中的调节机制。 该基因编码形成细胞间差距连接蛋白 渠道这些通道负责传播电 通过肌肉的脉冲使子宫同步收缩 分娩时的肌肉与心肌相反，连接蛋白43在心肌中， 在任何时候都存在，如果它是宪法表达， 子宫肌层细胞-细胞通道仅在分娩开始时出现， 在分娩后不久消失。这种细胞间通道的诱导 它的形成受雌激素和孕激素的控制。拟议 将用大鼠连接蛋白43进行连接蛋白43基因调控的研究 含有6.7 kb上游调控序列的基因组克隆。它 包括6个半回文雌激素和3个孕激素反应元件。 转录蛋白与这些和其他调节蛋白的结合， 将通过DNA足迹分析和迁移率变化来研究元素 测定。启动子研究将用包含以下的构建体进行： 启动子的不同区段，包括上游调控序列， 与荧光素酶的报告基因相连。这些结构将是 在转基因小鼠中表达，不同激素的作用将是不同的。 测定单个启动子元件的贡献将是 通过诱变分析进行评估。 最后，反式作用转录 将从cDNA表达文库中分离因子并进行表征。 这项研究的长期目标是了解 子宫内膜缝隙连接蛋白43基因表达的调控 怀孕和分娩时。这项研究具有临床意义，因为 已经表明早产与早产有关， 子宫肌层间隙连接的出现。一旦相互作用的不同 具有连接蛋白43基因调控元件的激素更好 了解，更好的治疗和预防早产儿 劳动力是可以设计的。

This research proposal describes experiments designed to reveal the mechanism by which the connexin43 gene is regulated in the myometrium. This gene codes for the gap junction protein that forms cell-cell channels. These channels are responsible for the propagation of electrical pulses through the muscle to allow synchronous contraction of the uterine muscle during labor. In contrast to heart muscle, where connexin43 is present at all times and were it is expressed constitutively, the myometrial cell-cell channels appear only at the onset of labor and disappear shortly after parturition. This induction of cell-cell channel formation is under the control of estrogen and progesterone. The proposed study of connexin43 gene regulation will be done with a rat connexin43 genomic clone that contains 6.7 kb of upstream regulatory sequence. It includes 6 half-palindromic estrogen and 3 progesterone response elements. The binding of transcriptional proteins to these and other regulatory elements will be studied by DNA footprinting analysis and mobility shift assays. Promoter studies will be performed with constructs that contain various segments of the promoter, including upstream regulatory sequences, linked to the reporter gene for luciferase. These constructs will be expressed in transgenic mice, and the effect of different hormones will be determined. The contribution of individual promoter elements will be assessed by mutagenic analysis. Finally, trans-acting transcription factors will be isolated from a cDNA expression library and characterized. The long-term goal of this research is to gain an understanding of the regulation of connexin43 gene expression in the myometrium during pregnancy and at parturition. This research is clinically relevant because it has been shown that premature labor is associated with the premature appearance of myometrial gap junctions. Once the interaction of different hormones with the regulatory elements of the connexin43 gene is better understood, better regimens for the treatment and prevention of premature labor may be designed.