Transcription Factor Interactions with Nucleosomes

描述（由申请人提供）：染色质结构的调节在所有研究的细胞核过程中起着重要作用。染色质的主要组成部分是核小体。这个粒子包含四个核心组蛋白的两个拷贝；这些蛋白质组成了一个核心，组成了146个碱基对的DNA。核小体，曾经被认为是一个静态的构件，现在被认为是动态的。一大类复合体，即依赖ATP的染色质重塑复合体，利用ATP水解的能量来重塑核小体的结构和位置。该应用程序的一个主要焦点是探索重塑复合物在功能上的差异程度。对该复合体家族的全谱活动的机制理解对于理解染色质结构可以被调节的方式以及染色质结构的调节如何有助于核过程的调节是必要的。这将通过比较SWI/ snf家族重塑复合体和其他重塑复合体来完成，目的有三：目的1:atp酶结构域在重塑中的作用。我们将创建具有“交换”atp酶结构域的嵌合蛋白，以定义atp酶结构域在功能中的作用。目的2：描述组蛋白变异对重塑反应的影响。我们将测试几种不同的重塑蛋白在核小体上的功能，这些核小体由不同的组蛋白变体组装而成。目的3:atp依赖性重构和高阶染色质结构。我们将使用可量化的方法来检查连接体长度和压实状态的变化对重塑的影响，通过重塑蛋白的光谱。我们将开发一个组织培养系统来表征由不同重塑活动引起的染色质结构变化的幅度和范围。

DESCRIPTION (provided by applicant): Regulation of chromatin structure plays a major role in all studied processes in the nucleus. The primary building block of chromatin is the nucleosome. This particle contains two copies each of four core histones; these proteins which form a core which organizes 146 base pairs of DNA. The nucleosome, once thought of as a static building block, is now known to be dynamic. A large class of complexes, the ATP-dependent chromatin remodeling complexes, use the energy of ATP hydrolysis to remodel nucleosome structure and position. A major focus of this application is to explore the extent to which remodeling complexes differ in function. A mechanistic understanding of the full spectrum of activities of this family of complexes is necessary to understanding the ways in which chromatin structure can be modulated, and how modulation of chromatin structure might contribute to the regulation of nuclear processes. This will be done by comparing SWI/SNF-family remodeling complexes with other remodeling complexes in three Aims: Aim 1: The role of the ATPase domain in remodeling. We will create chimeric proteins with 'swapped' ATPase domains to define the role of the ATPase domain in function. Aim 2: Characterization of the effects of histone variants on the remodeling reaction. We will test the ability of several different remodeling proteins to function on nucleosomes that have been assembled with different histone variants. Aim 3: ATP-dependent remodeling and higher order chromatin structure. We will use quantifiable approaches to examine the effects of changes in linker length and compaction status on remodeling by a spectrum of remodeling proteins. We will develop a tissue culture system to characterize the magnitude and spread of changes in chromatin structure caused by different remodeling activities.