Investigating the role of genome folding in transcriptional regulation

项目摘要 在转录过程中，增强子需要跨越大的染色体结构域接触基因启动子。 染色体折叠可以通过将增强子-启动子在细胞内靠近在一起来辅助它们的通讯。 原子核改变基因组折叠的突变是包括发育障碍在内的许多人类疾病的基础 和一些癌症。然而，即使经过几十年的研究，我们仍然不明白 染色质结构和转录调控。具体来说，我们不明白为什么DNA循环可以 与基因活性正相关、负相关，有时甚至完全不相关。超越相关性， 因果关系，我们需要更好地了解耦合基因组折叠和基因的分子过程， 调控粘着蛋白复合物已经成为DNA循环中的关键角色，因为它可以容纳两个染色质 纤维在一起并在DNA上移位时挤出DNA环。粘着蛋白通过其辅因子负载在染色质上 NIPBL。然后，粘附素易位，直到它在CTCF转录因子的结合位点被阻断。我们最近 开发的工具允许通过操纵NIPBL控制粘附素环挤出的各个方面， 小鼠胚胎干细胞中的CTCF。这些工具提供了一种新的方法来调查的关系 和转录之间的联系。在这项拟议的研究中，我们将确定转录如何 和环挤出是分子耦合的，并阐明了解释为什么只有一些基因 依靠循环挤出来发挥作用。我们还将讨论如何凝聚环挤出有助于功能 CTCF在细胞分化过程中的作用此外，我们将确定新的途径，调节增强子-启动子 通过调制环挤出进行通信。为了实现这些目标，我们将联合收割机， 测定、生物化学测定和具有高通量报告测定的新型表观基因组工程模式 小鼠胚胎干细胞及其分化衍生物。这些调查将大大深化 我们对基因组折叠如何被粘附蛋白质影响基因转录的理解。完成本 该项目将开辟新的途径来探索这些过程如何在疾病中出错，我们的团队正在研究这个问题。 有兴趣在未来进行调查。

Project Summary During transcription, enhancers need to contact gene promoters across large chromosome domains. Chromosome folding can assist enhancer-promoter communication by bringing them close together inside the nucleus. Mutations that alter genome folding underlie many human diseases, including developmental disorders and some cancers. Yet, even after decades of research, we do not understand the causal links between chromatin architecture and transcription regulation. Specifically, we do not understand why DNA looping can correlate positively, negatively, or sometimes not at all with gene activity. To move beyond correlation to causation, we need to better understand the molecular processes that couple genome folding and gene regulation. The cohesin complex has emerged as a key player in DNA looping, because it can hold two chromatin fibers together and extrude DNA loops as it translocates on DNA. Cohesin is loaded on chromatin by its co-factor NIPBL. Cohesin then translocates until it is blocked at binding sites for the CTCF transcription factor. We recently developed tools that allow controlling various aspects of cohesin loop extrusion by manipulating NIPBL and CTCF in mouse embryonic stem cells. These tools provide a novel approach to investigate the relationship between loop extrusion by cohesin and transcription. In this proposed study, we will determine how transcription and loop extrusion are molecularly coupled and elucidate the mechanisms that explain why only some genes rely on loop extrusion to function. We will also address how cohesin loop extrusion contributes to the functions of CTCF during cell differentiation. In addition, we will identify novel pathways that regulate enhancer-promoter communication by modulating loop extrusion. To achieve these goals, we will combine gene editing, epigenomic assays, biochemical assays and novel epigenome engineering modalities with high-throughput reporter assays in mouse embryonic stem cells and their differentiated derivatives. These investigations will substantially deepen our understanding of how genome folding by cohesin proteins influences gene transcription. Completion of this project will open new avenues to explore how these processes go awry in disease, a question our group is interested to investigate in the future.