Nuclear Pore Complexes As Scaffolds For Genome Architecture And Epigenetic Maintenance

摘要： 核孔复合物（NPC）是一种核孔嵌入的多组分复合物， 其介导细胞核和细胞质之间的分子运输。除了 它们在运输中的经典功能，NPC组分（Nups）已经被牵连到 通过与基因组结合进行转录调控。然而转录的调控步骤 以及Nups的染色质结合作用如何促进后生动物的发育 目前发展尚不清楚。我们确定了多种Nup与数百种 果蝇基因组中的启动子和增强子，并发现了一个以前未报道的 NPC在增强子-启动子环形成中的作用。具体来说，我们确定了Nup 98 是在被激活的基因上形成增强子-启动子环所必需的。 发育激素蜕皮激素在功能上，我们发现Nup 98介导的缺失 增强子-启动子环影响随后激活的引发反应， 转录记忆有趣的是，蜕皮激素调节基因与细胞核稳定相关， 孔之前和之后的活化，这表明后生动物NPC可以利用作为一个 为等待未来激活事件的基因组织支架。总之，这些发现 暗示Nups是一类新的增强子结构蛋白， 增强子-启动子环通过核孔结合的稳定化构成了 表观遗传维持我们预测核孔的这种功能将是高度相关的 在后生动物发育过程中的基因调控。为了能够研究基因组- Nups在发育基因调控中的组织作用，我们首先计划鉴定分子 Nup介导的增强子-启动子环的决定因素。因此，在目标1中，我们将定义 Nups和其他建筑蛋白质参与蜕皮激素的建立- 诱导的基因组环。此外，我们计划确定足以 系到NPC或环路稳定所必需的。在目标2中，我们建议定义 Nup介导的增强子-启动子环的形成与转录的关系 激活和记忆，通过识别作为特定结果发生的染色质变化， 的循环。此外，我们将研究Nups对维持基因组循环的影响， 苍蝇组织在发展过程中，并确定了一套全面的基因组接触， 由Nups监管。总之，这些实验有望扩大我们对 基因组结构、基因表达和核孔生物学的驱动力和原理。

Abstract: The Nuclear Pore Complex (NPC) is a nuclear envelope-embedded multi-component complex, which mediates transport of molecules between the nucleus and the cytoplasm. In addition to their classical function in transport, NPC components (Nups) have been implicated in transcriptional regulation via binding to the genome. Yet what regulatory steps of transcription are controlled by Nups and how chromatin-binding roles of Nups contribute to metazoan development is currently unclear. We identified the binding of multiple Nups to hundreds of promoters and enhancers in the Drosophila genome, and discovered a previously unreported role of the NPC in the formation of enhancer-promoter loops. Specifically, we identified Nup98 to be required for the formation of an enhancer-promoter loop at a gene activated by a developmental hormone ecdysone. Functionally, we found that the loss of Nup98-mediated enhancer-promoter loop affected the primed response to subsequent activation or transcriptional memory. Interestingly, ecdysone-regulated genes stably associated with nuclear pores before and after activation, suggesting that metazoan NPCs can be utilized as an organizing scaffold for genes awaiting future activation events. Together, these findings implicate Nups as a new class of architectural proteins for enhancers and suggest that stabilization of enhancer-promoter loops by nuclear pore binding constitutes a mechanism of epigenetic maintenance. We project that this function of the nuclear pore will be highly relevant to gene regulation during metazoan development. To be able to investigate the genome- organizing role of Nups in developmental gene regulation, we first plan to identify molecular determinants of Nup-mediated enhancer-promoter looping. Thus, in Aim 1, we will define which Nups and which other architectural proteins participate in the establishment of ecdysone- induced genomic loops. Additionally, we plan to identify DNA elements that are sufficient to tether to the NPC or that are necessary for loop stabilization. In Aim 2, we propose to define the relationship between formation of Nup-mediated enhancer-promoter loops and transcriptional activation and memory, by identifying chromatin changes that occur as a specific consequence of looping. Furthermore, we will examine the effect of Nups on maintenance of genomic loops in fly tissues during development and identify a comprehensive set of genomic contacts that are regulated by Nups. Together, these experiments are expected to expand our knowledge of the driving forces and principles of genome architecture, gene expression and nuclear pore biology.