Cooperation between transcription factors and its role in the regulation of rhythmic enhancer activity by the circadian clock

摘要 转录激活依赖于结合顺式基因的多个转录因子(TF)的协同活性。 靶基因附近的调控元件(CRE)。尽管像染色质免疫沉淀这样的技术 与测序相结合，有助于揭示不同物种中许多转录因子结合位点的全基因组位置。 CIES和组织，目前仍不清楚转录因子如何与组蛋白竞争、结合DNA、调节Cre 活性，并驱动转录激活。在本提案中，我们将通过利用 转录因子时钟BMal1对基因表达的昼夜节律调控。我们实验室和文学院最近的报告- TRUE表明时钟对转录的昼夜节律调节：BMal1依赖于 时钟：BMal1和其他转录因子，而不仅仅是核心时钟基因。时钟：BMal1有节奏的DNA结合 促进核小体在其CRES的有节奏地移除，强烈表明时钟：BMal1基因- ATES是一种允许的染色质景观，有助于在其CRES有节奏地招募其他TF。这 CLOCK：BMal1和其他TF之间的合作可以解释为什么只有不到三分之一的CLOCK：BMal1 TAR- GET在正常情况下是有节奏的表达，以及它们的有节奏的转录是如何复制的。 当动物暴露在一个新的环境中时，这是一种语法。这项建议建立在这些发现和 拟议的实验将确定CLOCK：BMal1是否促进部分未包裹的核- SOMES(目标1)，时钟的方式：BMal1与其他TF采用单摩尔法合作- CULE足迹(目标2)，以及如果高脂饮食改变了Clock：BMal1和其他TF之间的协同作用，则 重新编程Cre转录活性(目标3)。这些实验的结果有望发现新的 以及控制数千个基因有节奏表达的重要机制，并提供了一种新的 生物功能如何在组织间协调的概念框架。预计他们还将 提供对Tf协作性在控制Cre活性中所起作用的机制的见解 可以广泛地与昼夜节律场以外的场相关。

SUMMARY Transcription activation relies on the cooperative activity of multiple transcription factors (TFs) that bind cis- regulatory elements (CREs) near their target genes. Although techniques like chromatin immunoprecipitation coupled to sequencing helped uncover the genome-wide location of binding sites for many TFs in different spe- cies and tissues, it still remains unclear how TFs cooperate to compete with histones, bind DNA, regulate CRE activity, and drive transcription activation. In this proposal, we will address this knowledge gap by exploiting the circadian regulation of gene expression by the TF CLOCK:BMAL1. Recent reports from our lab and the litera- ture indicate that the circadian regulation of transcription by CLOCK:BMAL1 relies on the interaction between CLOCK:BMAL1 and other TFs rather than on core clock genes only. CLOCK:BMAL1 rhythmic DNA binding promotes the rhythmic removal of nucleosomes at its CREs, strongly suggesting that CLOCK:BMAL1 gener- ates a permissive chromatin landscape that facilitates the rhythmic recruitment of other TFs at its CREs. This cooperation between CLOCK:BMAL1 and other TFs may explain why less than a third of CLOCK:BMAL1 tar- gets are rhythmically expressed under normal condition, and how their rhythmic transcription can be repro- grammed when animals are exposed to a new environment. This proposal builds upon these findings and the proposed experiments will determine if CLOCK:BMAL1 promotes the formation of partially unwrapped nucleo- somes (Aim 1), the modalities of CLOCK:BMAL1 cooperation with other TFs by using a method of single-mole- cule footprinting (Aim 2), and if high-fat diet alters the cooperativity between CLOCK:BMAL1 and other TFs to reprogram CRE transcriptional activity (Aim 3). Results from these experiments are expected to uncover novel and important mechanisms that control the rhythmic expression of thousands of genes, and to provide a new conceptual framework for how biological functions are coordinated between tissues. They are also expected to provide insights into the mechanisms that underlie the role of TF cooperativity in the control of CRE activity that can be widely relevant to fields beyond the circadian field.