Hormonal Control of Histone Modifications in ES Cells

超出所提供的空间。胚胎干细胞（ES）分化的最早阶段的分子方面仍然知之甚少。我们的初步数据表明，全球组蛋白乙酰化可能是分化的关键第一步。本提案的目标是确定在ES细胞分化的多细胞诱导方法期间是否发生全局组蛋白乙酰化和/或甲基化，并使用组蛋白修饰的标准测定法建立这些组蛋白修饰发生的时间范围。我们的模型预测，这些组蛋白修饰的大部分可能发生在启动子区域，我们将利用新的CpG岛阵列来证实这一点。研究旨在确定在退出未分化ES细胞状态期间发生的全局组蛋白修饰是否与由激素信号传导至高度分化细胞诱导的基因特异性组蛋白修饰独特不同。第二个具体的目的是探索可能的机制，导致这些独特的全球组蛋白修饰早期在ES细胞分化通过筛选ES细胞提取物的各种酶活性负责共价修饰的历史。第三个具体目标是测试组蛋白修饰在直接调节ES细胞分化中的功能意义。组蛋白去乙酰化酶抑制剂，阿司他丁A（TSA），将与特定的生长因子结合使用，以增加细胞定向到特定的定向细胞命运的速率和比例。此外，将评估旨在抑制ES细胞中组蛋白乙酰转移酶活性或增加组蛋白脱乙酰酶活性的特定基因产物的过表达对ES细胞分化速率和总体细胞命运定型的影响。这些研究旨在了解早期胚胎干细胞调控的ES细胞分化，并有可能显著提高体外谱系特异性分化的产量。这将极大地促进ES细胞技术的发展，用于将“纯”细胞群移植到患有诸如帕金森病、再生障碍性贫血等疾病的患者中。本申请中提出的模型预测组蛋白脱乙酰酶抑制剂如丙戊酸（一种已知的致畸剂）和TSA可能通过改变正常谱系所需的非常早期的组蛋白乙酰化模式，大大增强环境化合物的致畸作用。具体区别。性能现场=

EXCEED THE SPACE PROVIDED. The molecular aspects of the earliest steps in embryonic stem (ES) cell differentiation remain poorly understood. Our preliminary data suggest that global histone acetylation may be a critical first step in differentiation. The goals of this proposal are to establish whether global histone acetylation and/or methytation occurs during multipie hormonally induced methods of ES cell differentiation and establish the time frame in which these histone modifications occur using standard assays for histone modifications. Our model predicts that the bulk of these histone modifications probably occur in promoter regions and we will make use of novel CpG island arrays to confirm this. Studies are designed to determine whether the global histone modifications that occur during exit from the undifferentiated ES cell state are uniquely different from the gene-specific histone modifications induced by hormonal signallingto highly differentiated cells. The second specific aim is to explore the possible mechanisms leading to these unique global histone modifications early in ES cell differentiation by screening ES cell extracts for various enzymatic activities responsible for covalent modification of histories. The third specific aim is designed to test the functional significance of histone modifications in directly regulating ES cell differentiation. The histone deacetylase inhibitor, trichostatin A (TSA), will be used in conjunction with specific growth factors to increase the rate and proportion of cells directed to a specific committed cell fate. In addition, overexpression of specific gene products designed to inhibit histone acetyltransferase activity or increase histone deacetylase activity in ES cells will be assessed for effects on the rate of ES cell differentiation and overall cell fate commitment. The proposed studies are designed to understand early hormonally regulated ES cell differentiation with potential application for significantly improving the yield of lineage-specific differentiation in vitro. This would greatly facilitate the development of ES cell technology for potential transplantation of "pure" cell populations into patients with diseases such as Parkinson disease, aplastic anemia, etc. In addition, the model proposed within this application predicts that histone deacetylase inhibitors such as valproic acid (a known teratogen) and TSA may greatly potentiate the teratogenic effects of environmental compounds by altering the very early histone acetylation pattern required for normal lineage-specific differentiation. PERFORMANCE SITE ========================================Section End===========================================