Comprehensive Characterization of Heterochromatin Domains

描述（由申请人提供）：异染色质是一种基因组结构，在所有真核生物中发挥作用，通过沉默基因来确定细胞身份并维持基因组稳定性。异染色质组装的要求是将具有组蛋白修饰活性的沉默蛋白复合物掺入染色质中，从而导致抑制的基因表达。这种一般机制在所有真核生物中是保守的，尽管存在多种异染色质结构域，即使在同一生物体中也是如此。我们对异染色质的形成因素了解很多，但对特定途径中的许多关键事件却知之甚少。研究异染色质的一个挑战是，在体内操纵中心组分往往会影响细胞中多种类型的异染色质或其他相关途径。本项目的目标将从三个方面解决这个问题：首先，通过使用功能性的体外重建异染色质系统，以更好地了解异染色质沉默在隔离环境中的机制。其次，使用预组装的异染色质结构域来鉴定细胞内有助于异染色质形成和维持的新因子。第三，通过建立一个系统，以解决特定的非编码RNA元件的异染色质的启动和维持的贡献。 异染色质酵母模型的初步研究将为染色质生物学领域做出重要贡献，也将作为过渡到独立研究者后在人类系统中进行后续研究的模型。该项目的主要目标是全面表征异染色质结构域，以比较对基因沉默和基因组稳定性至关重要的保守特征。人类异染色质结构域的多样性是显而易见的，但控制这些结构域组装的一般原则仍然与芽殖和裂殖酵母模型中的那些非常相似。这些基本的相似性使得将酵母研究的结果与人类进行比较成为可能，并且还可以使用在更简单的酵母模型中开发的方法作为研究人类异染色质的指南。这一主题是本提案的基础，也是我向独立过渡的方向。 公共卫生相关性：异染色质是在所有真核生物中起作用的基因组结构，其通过沉默基因来确定细胞身份并维持基因组稳定性。异染色质组装的一般要求是将具有组蛋白修饰活性的沉默蛋白复合物掺入染色质中，从而导致抑制的基因表达。在同一真核生物中存在多种异染色质结构域。

DESCRIPTION (provided by applicant): Heterochromatin is a genomic structure that functions in all eukaryotes to determine cell identity and maintain genome stability by silencing genes. The requirements for heterochromatin assembly are incorporation into chromatin of silencing protein complexes with histone modification activity that leads to repressed gene expression. This general mechanism is conserved in all eukaryotes, though multiple varieties of heterochromatic domains exist, even within the same organism. Much is known about the factors that contribute to heterochromatin, yet many key events in specific pathways are poorly understood. One challenge in studying heterochromatin is the fact that manipulation of the central components in vivo often affects multiple types of heterochromatin or other related pathways in the cell. The aims of this project will address this problem in three ways: firstly, by using a functional in vitro reconstituted heterochromatin system to better understand the mechanism of heterochromatic silencing in an isolated environment. Secondly, using a pre- assembled heterochromatin domain to identify new factors within the cell that contribute to heterochromatin formation and maintenance. Third, by establishing a system to address the contribution of specific non-coding RNA elements to initiation and maintenance of heterochromatin. Initial studies in yeast models of heterochromatin will make important contributions to the field of chromatin biology and will also serve as a model for subsequent studies in the human system to be carried out after the transition to independent investigator. The primary goal of the project is to comprehensively characterize heterochromatin domains in order to compare conserved features that are central to gene silencing and genome stability. The diversity of human heterochromatic domains is evident, yet the general principles that govern assembly of these domains remain quite similar to those in the budding and fission yeast models. These basic similarities make it feasible to compare results of yeast studies to humans and also to use the approaches that have been developed in the simpler yeast models as a guide to study human heterochromatin. This theme is the foundation of this proposal and the direction I will take as I transition to independence. PUBLIC HEALTH RELEVANCE: Heterochromatin is a genomic structure that functions in all eukaryotes to determine cell identity and maintain genome stability by silencing genes. The general requirements for heterochromatin assembly are the incorporation into chromatin of silencing protein complexes with histone modification activity that leads to repressed gene expression. Multiple varieties of heterochromatic domains exist within the same eukaryotic organism.