Transcriptional Networks Controlling Erythroid Differentiation

控制红细胞分化的转录网络

基本信息

批准号：
10597204
负责人：
Gerd A Blobel
金额：
$ 75.22万
依托单位：
CHILDREN'S HOSP OF PHILADELPHIA
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-03-01 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10597204
关键词：
Acceleration Acute Adaptor Signaling Protein Architecture Auxins Binding Binding Sites Biological Assay Categories Cell Cycle Cell Cycle Progression Cell Differentiation process Cell Line Cell Maturation Cell Nucleus Cells ChIP-seq Chromatin Chromatin Loop Chromatin Modeling Chromatin Structure Chromosome Structures Chromosomes Collaborations Complement Complex Coupled Data Data Set Defect Dependence Development Disease Engineering Enhancers Erythroblasts Erythrocytes Erythroid Erythroid Cells Excision Funding Agency G1 Phase G1/S Transition GATA1 gene Gene Activation Gene Expression Gene Expression Regulation Genes Genetic Transcription Genome Hematopoietic stem cells Hi-C Influentials Interphase Knowledge Laboratories Life Maintenance Malignant Neoplasms Maps Measurable Measurement Measures Mechanics Metaphase Mitosis Monitor Mus Nuclear Nucleic Acid Regulatory Sequences Outcome Pattern Population Process Proteins Research Resolution Role Series Structure Sum System TAL1 gene Time Tissues Untranslated RNA Work YY1 Transcription Factor beta Globin cofactor cohesin connectome cost effective developmental disease developmental genetics erythroid differentiation experimental study follow-up forging gene repression genome-wide infancy insight novel programs promoter recruit superresolution microscopy synergism tool transcription factor virtual

项目摘要

Abstract The spatial organization of chromosomes and gene expression are mutually influential processes. Perturbations of either can lead to developmental defects and disease. Studies on chromatin structure have been dominated by those focusing on the architectural transcription factors CTCF, YY1 and the cohesin complex. The central hypothesis of this application, supported by preliminary data, is that the non-DNA binding adapter protein Ldb1 presents a separate, equally important category of nuclear chromatin organizers. In prior work we demonstrated that artificial recruitment of Ldb1 to chromatin can be sufficient to forge long range enhancer- promoter contacts at the b-globin locus. This nominated Ldb1 as a key architectural protein in erythroid cells. Yet a global view of Ldb1 chromatin occupancy patterns during cell differentiation or cell cycle progression is lacking, and genome wide direct Ldb1 dependent chromatin contacts and corresponding transcriptional patterns are largely unknown. More broadly, the cause-effect relationships of architectural features and gene expression are still hotly debated in the field. Here we propose to carry out the following aims in collaboration with the laboratory of Dr. Ross Hardison. In Aim 1 we will assess in erythroblasts undergoing cell maturation the dynamic Ldb1 chromatin landscapes in relation to high resolution Hi-C chromosomal connectivity maps, and nascent transcription measurements (PRO- seq). In Aim 2 we will use an auxin-inducible acute Ldb1 degradation system to interrogate direct architectural and transcriptional Ldb1 dependencies. In Aim 3 we will exploit the massive and swift changes in chromatin architecture during the mitosis to G1-phase progression to assess the chromatin occupancy dynamics of Ldb1 and its role in establishing the multi-layered hierarchy of chromatin organization and gene expression. All aims are accompanied by perturbative and mechanistic experiments. We believe that the power of the proposed studies lies in the complementarity of highly dynamic natural transition states (cell maturation and cell cycle progression) as well as acute experimental perturbation (auxin degron), which to our knowledge has never been done before within the same cell system. Gains in knowledge are further enhanced by parallel studies in the same cellular system on CTCF, YY1 and cohesin with support from different funding sources. The forward looking view is that the end result of our studies is, for the first time, a unified view of the dynamics of major architectural components and gene expression. Therefore, the studies proposed here coupled with parallel studies in the lab generate a cost-effective synergy, such that the outcome will be greater than the sum of its parts.

抽象的染色体的空间组织和基因表达是相互影响的过程。任何一个的干扰都会导致发育缺陷和疾病。对染色质结构的研究那些关注结构转录因子 CTCF、YY1 和粘连蛋白复合物的研究占主导地位。本申请的中心假设（得到初步数据的支持）是非 DNA 结合接头蛋白质 Ldb1 代表了一个单独的、同样重要的核染色质组织者类别。在之前的工作中我们证明人工招募 Ldb1 到染色质足以形成长程增强子- b-珠蛋白基因座处的启动子接触点。这将 Ldb1 指定为红系细胞中的关键结构蛋白。然而，细胞分化或细胞周期进展过程中 Ldb1 染色质占据模式的总体观点是缺乏全基因组直接 Ldb1 依赖的染色质接触和相应的转录模式很大程度上是未知的。更广泛地说，建筑特征和基因表达的因果关系至今仍引起业界的热烈讨论。在此，我们建议与罗斯·哈迪森博士的实验室合作实现以下目标。在目标 1 我们将评估正在经历细胞成熟的成红细胞中的动态 Ldb1 染色质景观与高分辨率 Hi-C 染色体连接图和新生转录测量（PRO- 序列）。在目标 2 中，我们将使用生长素诱导的急性 Ldb1 降解系统来询问直接的结构和转录 Ldb1 依赖性。在目标 3 中，我们将利用染色质的巨大而快速的变化有丝分裂到 G1 期进展期间的结构，以评估 Ldb1 的染色质占用动态及其在建立染色质组织和基因表达的多层体系中的作用。所有目标都伴随着微扰和机械实验。我们相信，所提出的研究的力量在于高度动态的自然环境的互补性。过渡态（细胞成熟和细胞周期进展）以及急性实验扰动（生长素 degron），据我们所知，以前从未在同一细胞系统中完成过这项工作。知识的收获通过在同一细胞系统中对 CTCF、YY1 和粘连蛋白进行平行研究，进一步增强了支持来自不同的资金来源。前瞻性的观点是，我们研究的最终结果是，第一次，主要建筑成分和基因表达动态的统一视图。因此，研究这里提出的建议与实验室的平行研究相结合，产生了具有成本效益的协同作用，这样的结果将大于其各部分之和。