Epigenetic activation of the human growth hormone gene

描述(由申请人提供)： 这一K01奖项的总体目标是扩大候选人的科学培训和能力，并为他的独立研究生涯做好准备。这将在对人类生长激素基因(HGH)表达的详细研究的背景下完成。该实验室以前的研究已经确定了一个组织特异性基因座控制区(HGH LCR)，它调节人类生长激素(HGH)基因簇。HGH LCR的元件位于距目的hGH-N基因启动子很远的位置(15-32kb)5‘。因此，这个模型系统中的一个中心问题是hGH LCR和hGH-N启动子之间隐含的长程基因激活是如何介导的。在目前的K01指导培训期间，候选人发现hGH LCR本身是由POL II主动转录的，并且转录的区域也被特定和独特的组蛋白改变模式修改。在目前的提案中，候选人的目标是确定LCR结构中的这些表观遗传变化如何有助于高阶染色质相互作用，最终导致hGH基因激活。该方案的假设是hGH LCR通过招募和传播表观遗传修饰物来发挥作用。这些修饰促进了LCR内部以及LCR与远端hGH-N启动子之间的直接相互作用，并介导了hGH基因定位到特定的、转录活性较高的生长细胞核亚区。目的研究促生长激素染色质中hGH LCR和hGH-N基因之间的长程相互作用。这一目标将通过应用染色质构象捕获(3C)分析来解决LCR内部以及LCR和hGH-N启动子之间的直接相互作用。要研究的染色质将从携带有信息的转基因小鼠的脑下垂体和对照组织中分离出来。目的利用高清晰度显微技术，建立人生长激素LCR活性与生长激素基因亚核定位之间的功能关系。新的协作将为候选人提供成功实现第二个目标所需的技术方面的额外培训。这些研究解决了基因控制的基本机制，并为了解垂体激素表达的正常和病理状态奠定了必要的知识基础。这种认识为内分泌疾病的诊断和治疗的进展奠定了基础。

DESCRIPTION (provided by applicant): The overall goal of this K01 award is to extend the scientific training and capabilities of the candidate and prepare him for an independent research career. This will be accomplished in the context of a detailed study of human growth hormone gene (hGH) expression. Prior studies from this laboratory have identified a tissue-specific locus control region (hGH LCR) that regulates the human growth hormone (hGH) gene cluster. The components of the hGH LCR are located quite far (15-32 kb) 5' to the target hGH-N gene promoter. Thus a central question in this model system is how the implied long-range gene activation between the hGH LCR and hGH-N promoter is mediated. During the current K01 mentored training period, the candidate has discovered that the hGH LCR is itself actively transcribed by Pol II and that the regions that are transcribed are also modified by specific and distinct patterns of histone alterations. The candidate's goal in the current proposal is to determine how these epigenetic alterations in LCR structure contribute to higher-order chromatin interactions that eventuate in hGH gene activation. The Hypothesis of the proposal is that the hGH LCR acts by recruitment and spreading of epigenetic modifiers. These modifications facilitate direct interactions within the LCR and between the LCR and the remote hGH-N promoter and they mediate relocation of the hGH locus to specific, transcriptionally-active sub-regions of the somatotrope nucleus. Aim I is to define the long-range interactions between the hGH LCR and hGH-N gene in somatotrope chromatin. This aim will address direct interactions within the LCR and between the LCR and the hGH-N promoter by applying Chromatin Conformation Capture (3C) analysis. The chromatin to be studied will be isolated from pituitaries and control tissues of mouse lines carrying informative transgenes. Aim II will establish functional relationships between hGH LCR activities and sub-nuclear localization of the hGH locus in somatotrope nuclei using high definition microscopic technology. A new collaboration will provide the candidate with additional training in the technology necessary to successfully implement this second aim. These studies address fundamental mechanisms of gene control and contribute to a foundation of knowledge necessary for understanding normal and pathological states of pituitary hormone expression. Such understanding establishes a basis for advances in diagnosis and treatment of endocrine disorders.