GH ACTION--REGULATION OF IGF-I GENE TRANSCRIPTION

生长激素(GH)和胰岛素样生长因子(IGF-I)都 对脊椎动物的生长和分化至关重要。 GH是IGF-I产生的主要调节因子，主要作用是 通过激活IGF-I基因转录。描述这些机制 负责这种转录激活的人应该提供新的 药物设计的分子靶点并最终导致更好的 代谢和生长障碍的治疗。我们的实验室有 在大鼠IGF-I基因中发现了一个染色质区域，称为HS7 同时经历生长激素调节的结构变化 激素诱导的转录调节。这样做的目的是 项目的目的是刻画分子机制， HS7在GH激活IGF-I基因中的作用 转录控制。 第一组实验将检验HS7病毒的假设 介导生长激素诱导的IGF-I基因转录的刺激。几个 已知表达IGF-I的细胞系将被GH转染 受体和各种报告基因与IGF-I或 含或不含HS7位点的异源启动子。平行的一组 一系列实验将描述与之结合的核蛋白 HS7站点的IGF-I。体外凝胶漂移试验和足部 打印实验将与现场特定的实验结合使用 诱变以确定所需的特定核苷酸 并足以与高亲和力蛋白结合。几个 将使用实验方法来识别和克隆 与HS7相互作用的核蛋白。一系列实验 将检验克隆的蛋白质是特定的这一假设 生长激素调节IGF-I基因转录的介体。

Growth (GH) and insulin-like growth factor (IGF-I) are both critically important for growth and differentiation in vertebrates. GH is the principal regulator of IGF-I production and acts primarily by activating IGF-I gene transcription. Characterizing the mechanisms responsible for this transcriptional activation should provide new molecular targets for drug design and ultimately lead to better treatment of metabolic and growth disorders. Our laboratory has identified a chromatin region in the rat IGF-I gene termed HS7 that undergoes GH-regulated structural alteration in parallel with hormone-induced transcriptional regulation. The goals of this project are to characterize the molecular mechanisms that mediate activation of IGF-I gene by GH by evaluating the role of HS7 in transcriptional control. The first set of experiments will test the hypothesis that HS7 mediates GH-induced stimulation of IGF-I gene transcription. Several cell lines known to express IGF-I will be transfected with GH receptor and various reporter genes fused to the IGF-I or heterologous promoters with or without the HS7 site. A parallel set of experiments will characterize the nuclear proteins that bind to the IGF-I at the HS7 site. In vitro gel shift assays and foot printing experiments will be used in conjunction with site-specific mutagenesis to identify the specific nucleotides that are necessary and sufficient for high affinity protein binding. Several experimental approaches will be employed to identify and clone the nuclear proteins that interact with HS7. A series of experiments will test the hypothesis that the cloned proteins are specific mediators of GH-regulated IGF-I gene transcription.