MOLECULAR GENETICS OF FACTOR IX AND OTHER BLOOD PROTEINS

因子IX在外在和内在中都起着举足轻重的作用 胆汁凝结的途径。终审法院的设立 因此，因子IX(FIX)生物合成的潜在机制是 不仅对了解血友病B的分子基础至关重要， 而且FIX在凝血及其动态平衡中的作用也很重要。 这项建议的长期目标是建立全面的 负责修复基因转录的控制机制，第一步 在固定的生物合成中。这将通过实现三个目标来实现 具体目标：(一)建立精准的基础性机制 负责FIX基因的整体转录控制；(Ii) 建立导致青春期开始的具体机制 改善血友病B Leyden表型；及(Iii) 建立年龄相关调控的分子基础 修复基因的表达。目标1将通过建立一个详细的 基本结构元素图、它们的功能和相互关系 与FIX基因整体调控的关系，从而提供了一种 为其余拟议研究奠定了坚实的基础。体外系统研究 顺式作用DNA序列的鉴定及功能分析 将携带与它们相互作用的要素和交互作用因素 经定点突变、核酸酶超敏试验、DNA- 蛋白质和蛋白质-蛋白质相互作用分析以及 关键的反式作用因子的特征，如果它们是新奇的。在……里面 这些元件在固定微基因结构背景下的功能 将在与研究相关的转基因小鼠身上进行测试 目标2和目标3。目标2，血友病的机制 B Leyden表型，这是由一组单碱基突变引起的 包含在一个很小的5‘端区域，将通过扩展我们的 新发现包括生长激素可能参与其中 途径，通过密集的体外和体内分析，包括 用转基因小鼠结合脑下垂体切除术进行测试。我们有 最近发现了位于5‘端的Line 1序列的一个新的作用 在与年龄相关的FIX基因调控中。目标3将通过以下方式实现 建立序列的作用机制 基因结构的其余部分。在这项研究中，转基因小鼠将作为 重要的体内检测系统。拟议的研究将显著 有助于我们了解FIX生物合成的具体情况，并 高血压病患者凝血功能及其动态平衡的整体调节 一般来说，这对发展来说是改进的，或许是小说 治疗人类的出血和血栓性疾病。

Factor IX plays a pivotal role in both the extrinsic and intrinsic pathways of bolld coagulation. Establishment of the regualtory mechanisms underlying factor IX (FIX) biosynthesis, therefore, is critical for understanding not only the molecular basis of hemophilia B, but also the role of FIX in the blood coagulation and its homeostasis. The long-term goal of this proposal is to establish the comprehensive control mechanism responsible for FIX gene transcription, the first step in the FIX biosynthesis. This will be accomplished by achieving three specific aims, (I) to establish the precise fundamental mechanism responsible for the overall transcriptional control of the FIX gene; (ii) to establish specific mechanisms responsible for the puberty-onset amelioration of the hemophilia B Leyden phenotype; and (iii) to establish the molecular basis underlying the age-associated regulation of FIX gene expression. Aim 1 will achieved by establishing a detailed map of essential structural elements, their functions and mutual relationship n the overall regulation of the FIX gene, thus providing a solid base for the rest of the proposed studies. Systematic in vitro identification and functional analysis of cis-acting DNA sequence elements and trans-acting factors interacting with them will be carried out by site-directed mutagenesis, nuclease hypersensitivity assay, DNA- protein and protein-protein interaction analyses as well as characterization of crucial trans-acting factors, if they are novel. In vivo, functions of these elements in context of FIX minigene structures will tested in transgenic mice designed in conjunction with the studies for aims 2 and 3. Aim 2, mechanisms responsible for the hemophilia B Leyden phenotype, which are due to a set of single-base mutations contained in a small 5' end region, will be studied by extending our new findings including possible involvement of growth hormone pathway, with intensive in vitro as well as in vivo analyses including testing with transgenic mice combined with hypophysectomy. We have recently found a novel role of the Line 1 sequence present at the 5' end in the age-associated FIX gene regulation. Aim 3 will achieved by establishing the mechanism of action of the sequence in context of the rest of gene structure. In this study, transgenic mice will serve as an important in vivo assay system. Proposed studies will significantly contribute to our understanding of the FIX biosynthesis in specific, and the overall regulation of blood coagulation and its homeostasis in general, and this to the development of improved or perhaps novel teratments for bleeding and thrombotic disorders in humans.