CHROMATIN REMODELING DURING LIVER DEVELOPMENT

本提案的主要目的是审查各种程序， 发育调节转录的顺序结合 因子组织构成增强子的顺式作用序列 将血清白蛋白基因分成一系列的三个阶段 包含必需增强因子的核小体样颗粒 肝染色质中的结合位点。这将使用三个 不同的方法。首先，将白蛋白增强剂组装成 肝脏发育过程中的活性高级核蛋白复合物 随后将比较白蛋白水平上的核小体定相 在连续的发育阶段，肝脏染色质中的增强子。这些 观察结果将与最近完成的体内DMS相结合 保护性分析已经鉴定了转录因子 负责激活白蛋白增强剂。第二，能力 纯化的转录因子单独和组合以结合并 从白蛋白增强剂体外重构的扰动核小体 片段和核心组蛋白将被确定，以确定 转录因子指定核小体的机制 白蛋白增强剂的结构。最后，具体发挥的作用 白蛋白染色质重塑中的转录因子 增强子将通过研究活性和染色质来评估 含有突变的白蛋白增强子转基因的结构 转基因小鼠肝脏发育过程中选择的增强子结合位点 小鼠这些研究应该进一步加深我们对 通过染色质结构在转录调控中的作用， 深入了解调节因素决定正常 肝脏的生长和发育

The primary aim of this proposal is to examine the processes whereby the sequential binding of developmentally regulated transcription factors organizes the cis-acting sequences which make up the enhancer of the serum albumin gene into a series of three translationally phased nucleosome-like particles encompassing the essential enhancer factor binding sites in liver chromatin. This will be accomplished using three different approaches. First, the assembly of the albumin enhancer into an active higher-order nucleoprotein complex during liver development will be followed by comparison of nucleosome phasing at the albumin enhancer in liver chromatin at successive developmental stages. These observations will be combined with recently completed in vivo DMS protection analyses which have identified the transcription factors responsible for activating the albumin enhancer. Second, the ability of purified transcription factors alone and in combination to bind to and perturb nucleosomes reconstituted in vitro from albumin enhancer fragments and core histones will be determined, in order to determine the mechanisms whereby transcription factors specify the nucleosome structure of the albumin enhancer. Finally, the role played by specific transcription factors in the chromatin remodeling of the albumin enhancer will be assessed by studying the activity and chromatin structure of albumin enhancer transgenes containing mutations in selected enhancer binding sites during liver development in transgenic mice. These studies should further our understanding of the role played by chromatin architecture in transcriptional regulation as well as lend insight into the mechanisms whereby regulatory factors dictate normal liver growth and development.