Transcriptional Regulatory Networks in Living Cells

描述（由申请人提供）：我们建议开始绘制控制胚胎干细胞和选定分化细胞基因表达程序的转录调控电路。这项研究的短期目标是了解胚胎干细胞的调控回路如何促进多能性，而长期目标是利用这一回路的知识来促进操纵细胞命运的努力。转录调控电路的绘制可以通过识别基因组的活跃和沉默部分以及确定主调控体如何控制一组核心基因来启动。这些细胞类型特异性图谱将定义整个非重复基因组的活性和抑制染色质结构，并确定定义细胞表型的核心调控电路。为实现这一目标，建议的具体目标是：1)进一步发展实验和分析技术，以确定与脊椎动物基因组相关的蛋白质在体内的全基因组位置；2)定义人类胚胎干细胞中整个非重复基因组的活性和抑制染色质结构，并将其与两种分化细胞类型进行比较；3)确定主要调控因子如何促进人类胚胎干细胞和两种分化细胞的核心转录调控回路。4)鉴定人类和小鼠细胞中表观遗传状态和核心转录调控回路的保守成分，以方便基因检测和帮助鉴定细胞状态的关键控制。这些研究提高了对脊椎动物染色质结构和转录回路的理解，将导致对胚胎干细胞多能性的新认识，将生成调控回路图，可能有助于为再生医学操纵细胞命运的努力，并将为进一步绘制人类和其他脊椎动物细胞的调控回路奠定基础。

DESCRIPTION (provided by applicant): We propose to begin mapping the transcriptional regulatory circuitry that controls the gene expression programs of embryonic stem cells and selected differentiated cells. A short-term goal of this study is to learn how the regulatory circuitry of embryonic stem cells contributes to pluripotency, and a long-term goal is to use knowledge of this circuitry to facilitate efforts to manipulate cellular fates. Mapping transcriptional regulatory circuitry can be initiated by identifying the active and silent portions of the genome and determining how master regulators control a core set of genes. These cell-type specific maps will define active and repressed chromatin structure for the entire non-repeat genome and identify the core regulatory circuitry that defines cellular phenotype. To accomplish this, the specific aims of the proposal are: 1) further develop experimental and analytical technologies that identify the genome-wide location of proteins associated with vertebrate genomes in vivo, 2) define the active and repressed chromatin structure for the entire non-repeat genome in human embryonic stem cells and compare it to that of two differentiated cell types, 3) determine how master regulators contribute to the core transcriptional regulatory circuitry of human embryonic stem cells and two differentiated cells, and 4) identify conserved components of the epigenetic state and core transcriptional regulatory circuitry in human and mouse cells to facilitate genetic tests and help identify the key controls of cell state. Improved understanding of vertebrate chromatin structure and transcriptional circuitry from these studies should lead to new insights into embryonic stem cell pluripotency, will generate maps of regulatory circuitry that may facilitate efforts to manipulate cell fates for regenerative medicine, and will provide the foundation for further mapping regulatory circuitry in human and other vertebrate cells.