Chromatin Domain Dynamics

描述（由申请人提供）：细胞核在不同结构和功能区域的组织对于正常的染色体过程（如转录和复制）是重要的，已知这种组织的破坏会导致癌症和发育缺陷等疾病状态。因此，了解细胞核是如何组织和对刺激作出反应的，应该有助于我们更好地操纵过程，以造福于研究和医学。本建议的总体目标是达到真核生物染色体结构域的结构和组织机制的理解。细胞核中的相邻结构域通常具有拮抗的结构或功能特性，确定不同结构域的精确结构和分子特性对于了解细胞核的组织方式是必要的。虽然DNA绝缘体元件已被确定为分离不同的功能域，但分离结构上确定的染色体结构域的元件尚未得到很好的定义。了解组织细胞核的功能元素和结构元素之间的关系是本提案的重点。分离不同染色质结构域的机制才刚刚开始被理解，可能涉及组蛋白的翻译后修饰以及染色质环与蛋白质上层结构的连接，许多机制问题仍然存在，也是本研究基金提出的研究领域。该研究将涉及使用分子遗传学和基因组学，使用定量染色质免疫沉淀等技术操纵染色质，染色体构象捕获和生化分析，以确定调节绝缘体元件和染色质环形成的因素。鉴于许多核蛋白和染色质的基本结构从酵母到人类都是保守的，这些研究将有助于辨别染色质结构域动力学的一般原理，这些原理可以外推到人类的基因调控中。由于已知染色质结构的改变会导致癌症和发育缺陷等状态，因此这项研究的结果将有助于开发治疗这些疾病状态的新策略，也有助于开发更好的基因治疗分子工具。

DESCRIPTION (provided by applicant): The organization of the nucleus into distinct structural and functional domains is important for normal chromosomal processes like transcription and replication and disruption of this organization is known to lead to disease states like cancer and developmental defects. Thus understanding how the nucleus is organized and responds to stimuli should help us better manipulate processes for benefit in research and medicine. The overall objective of this proposal is to reach a mechanistic understanding of the structure and organization of domains in eukaryotic chromosomes. Adjacent domains in the nucleus often have antagonistic structural or functional properties and determining the precise structural and molecular properties of different domains will be necessary to understand how the nucleus is organized. While DNA insulator elements have been identified that separate distinct functional domains, the elements that separate the structurally defined chromosomal domains are not well defined. Understanding the relationships between the functional and structural elements that organize the nucleus is a key focus this proposal. The mechanisms involved in separating distinct chromatin domains is just beginning to be understood and likely involve post-translational modifications of histones as well as attachment of chromatin loops to proteinaceous superstructures, many mechanistic questions remain and is also an area of inquiry proposed in this research grant. The research will involve use of molecular genetics and genomics, manipulation of chromatin using techniques like quantitative chromatin immunoprecipitations, and chromosome conformation capture and biochemical analysis to identify factors regulating insulator elements and chromatin loop formation. Given that many nuclear proteins and the basic structure of chromatin is conserved from yeast to humans, these studies will help discern general principles of chromatin domain dynamics that can be extrapolated to gene regulation in humans. Since alterations in chromatin architecture is known to cause states like cancer and developmental defects, the results from this study will be useful in developing new strategies for treating these disease states and also in developing better molecular tools for gene therapy.