Transcriptional Control of Hemoglobin Synthesis

描述（由申请人提供）：由血红蛋白合成异常引起的人类疾病折磨着大量患者，而这些疾病的治疗方法远非最佳。结合使用的优点？-珠蛋白位点作为理解细胞类型特异性转录机制的模型，这一问题受到了相当大的关注。使用创新的方法，我们确定了控制成人的因素和机制？比如珠蛋白基因。GATA-1及其交互体通过分散在？-珠蛋白位点和编码红细胞生物学关键调节因子的位点，包括血红素生物合成酶。了解因子如何选择染色质位点、招募修饰/改造染色质的共调节因子、参与转录机制和赋予基因亚核定位是至关重要的。鉴于强大的技术，相关因素和独特的专业知识，我们将解决关键问题，开发一个综合模型来解释血红蛋白合成的转录控制。具体目标1 -测试模型来解释控制珠蛋白基因的机制和编码血红素生物合成酶的基因之间的关系。我们假设GATA-1与一组独特的反式作用因子和共调节因子一起发挥作用，将血红素生物合成中编码4-氨基乙酰丙酸合成酶（Alas2）的珠蛋白基因和基因从核外周排出。这将建立高水平转录和协调珠蛋白链生产和血红素合成所需的亚核定位。我们将测试模型来解释gata -1调节因子如何启动和/或维持这些位点在外围和内部位点的锚定或排斥。特异性目的2 -阐明gata -1依赖性转录的启动和维持机制。关于启动和维持转录差异的机制几乎一无所知，这对于控制转录的治疗结果的策略尤其重要。我们将使用一种强大的RNA干扰实验来验证这种假设，即成分调节？-globin和Alas2转录支持起始和维持的能力不同。特异性目标3 -确定GATA-1突变如何导致人类血液病中GATA因子依赖的遗传网络失调。关于疾病突变如何损害GATA-1活性的许多问题仍未得到解答。我们将确定锌指n端Arg216W和Arg216Gln突变是先天性红细胞生成性卟啉症、X连锁灰色血小板综合征和血小板减少症的基础。-地中海贫血影响GATA-1活性。这些研究将对控制血红蛋白合成、正常和恶性造血以及其他发育过程的机制产生广泛的见解。

DESCRIPTION (provided by applicant): Human diseases resulting from aberrant hemoglobin synthesis afflict large numbers of patients, and therapeutics for these disorders are far from optimal. Taken together with the merits of using the ?-globin locus as a model to understand cell type-specific transcriptional mechanisms, this problem has received considerable attention. Using innovative approaches, we identified factors and mechanisms controlling the adult ?-like globin genes. GATA-1 and its interactors function through dispersed sites at the ?-globin locus and loci encoding pivotal regulators of red cell biology, including heme biosynthetic enzymes. It is crucial to understand how factors select chromatin sites, recruit coregulators that modify/remodel chromatin, engage transcriptional machinery, and confer gene subnuclear localizations. Given powerful technologies, implicated factors, and unique expertise, we will address key issues to develop an integrative model to explain the transcriptional control of hemoglobin synthesis. Specific Aim 1 - To test models to explain the relationship between mechanisms controlling globin genes and genes encoding heme biosynthetic enzymes. We hypothesize that GATA-1 functions with a unique cohort of trans-acting factors and coregulators to expel globin genes and the gene encoding a rate-limiting enzyme in heme biosynthesis, 4- aminolevulinic acid synthase (Alas2), from the nuclear periphery. This would establish a subnuclear positioning required for high-level transcription and coordinate globin chain production and heme synthesis. We will test models to explain how GATA-1-regulating factors initiate and/or maintain anchoring or repulsion of these loci at peripheral and internal sites. Specific Aim 2 - To elucidate mechanisms underlying initiation and maintenance of GATA-1-dependent transcription. Nearly nothing is known about mechanisms that differentially initiate vs. maintain transcription, a problem particularly important for strategies to control transcription with a therapeutic outcome. We will use a powerful RNA interference asay to test the hypothesis that components regulating ?-globin and Alas2 transcription differ in their capacities to support initiation vs. maintenance. Specific Aim 3 - To establish how GATA-1 mutations underlying human hematologic disorders dysregulate GATA factor-dependent genetic networks. Many questions remain unanswered regarding how disease mutations impair GATA-1 activity. We will determine how Arg216W and Arg216Gln mutations within the N-terminal zinc finger that underlie congenital erythropoietic porphyria, X- linked grey platelet syndrome, and thrombocytopenia with ?-thalassemia afect GATA-1 activity. These studies will yield broad insights into mechanisms controling hemoglobin synthesis, normal and malignant hematopoiesis, and additional developmental processes.