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Molecular mechanisms of direct reprogramming of fibroblasts to trophoblast stem cells

成纤维细胞直接重编程为滋养层干细胞的分子机制

基本信息

批准号：
329123747
负责人：
Professor Dr. Hubert Schorle
金额：
--
依托单位：
Institut für Pathologie
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2017
资助国家：
德国
起止时间：
2016-12-31 至 2020-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/329123747?language=en
关键词：
Molecular mechanisms direct reprogramming fibroblasts

项目摘要

Lineage-identity of a differentiated somatic cell is being stabilized by specific chromatin configuration, DNA-methylation and networks of self-sustaining transcription factors. Experimental settings, such as somatic cell nuclear transfer or the overexpression of transcription factors are able to interfere with these stabilizing networks result in reprogramming other somatic lineages or to pluripotent cells. Recently, we demonstrated that four transcription factors (Tfap2c, Eomes, Gata3 and Ets2; abbreviated: TEGE) are sufficient for reprogramming of murine fibroblasts to trophoblast stem cells. Direct reprogramming is a promising approach for regenerative medicine and human disease modeling: die molecular mechanism of such direct reprogramming is only poorly understood. Here we propose, to take advantage of our recently published fibroblast-to-trophoblast reprogramming model and subject it to an highly sophisticated integrative genomic approach. At various time-points during reprogramming we propose to determine expression (by RNA-seq), DNA binding of the TEGE-factors (by ChIP-seq) and the nucleosome-coverage (by ATAC-seq). We will be able to define the so-called pioneer-factors, which are able to bind to nucleosome-covered (and silenced) DNA-elements, transactivate transcription and initiate the epigenetic reprogramming. Thus, we will be able to define direct targets of the TEGE-Factors in this system. These targets should be able to replace one or more of the TEGE-factors when overexpressed. Knowledge of such dynamics enables us to define the molecular steps in the reprogramming. The experiments will allow an insight about the close interconnection of transcription factor circuits, alterations in nucleosome coverage, histones and DNA-Methylation. We aim to establish a model, which precisely describes the dynamics of this reprogramming. With this in hand, we will be able to predict the efficiency of reprogramming of other cell types.

分化的体细胞的谱系特性通过特定的染色质配置、DNA甲基化和自我维持的转录因子网络来稳定。实验环境，如体细胞核移植或转录因子的过度表达，能够干扰这些稳定网络，导致对其他体细胞谱系或多能细胞的重新编程。最近，我们证明了四种转录因子(Tfap2c、Eome、GATA3和Ets2；缩写：TEGE)足以将小鼠成纤维细胞重编程为滋养层干细胞。直接重编程是再生医学和人类疾病建模的一种很有前途的方法：这种直接重编程的分子机制还知之甚少。在这里，我们建议利用我们最近发表的成纤维细胞到滋养层细胞重编程模型，并将其应用于高度复杂的整合基因组方法。在重新编程的不同时间点，我们建议确定表达(通过RNA-seq)、TEGE因子的DNA结合(通过芯片-seq)和核小体覆盖(通过ATAC-seq)。我们将能够定义所谓的先锋因子，它们能够与核小体覆盖的(和沉默的)DNA元件结合，反式激活转录，并启动表观遗传重新编程。因此，我们将能够在这个系统中定义TEGE因子的直接目标。当过度表达时，这些目标应该能够取代一个或多个TEGE-因子。对这种动力学的了解使我们能够定义重新编程中的分子步骤。这些实验将使人们能够深入了解转录因子电路的紧密联系、核小体覆盖的变化、组蛋白和DNA甲基化。我们的目标是建立一个模型，准确地描述这种重新编程的动态。有了这一点，我们将能够预测其他类型细胞的重新编程效率。