Comprehensive Mapping of Long-Range Chromatin Interactions in Humanand Mouse Genomes

项目摘要/摘要 努力绘制人类基因组的功能元素图，其中包括绝缘体等元素， 增强子、启动子和转录起始点历来将基因组视为线性。然而，它 现在人们很好地认识到基因组有一个三维(3D)组织，这对于调节 调控元件和基因编码序列之间的功能关联。因此，线性地图提供了 基因组的不完整图景，通常很难或不可能推断出 没有提供背景的拓扑框架的监管要素。我们已经开发并发展了一种 用于提供这种拓扑框架的强大的高分辨率方法--染色质相互作用分析 使用成对末端标签测序(CHIA-PET)。利用CHIA-PET，我们已经证明了特定的DNA基序 CCTC结合因子(CTCF)是定义拓扑域和排列基因的关键 RNA聚合酶II(RNAPII)介导的协调转录的位置。因此，这一组合 CTCF和RNAPII CHIA-PET将有效地全面绘制主要结构编码和 支撑RNAPII相关转录调控的拓扑组织。我们将捐献佳亚-PET 为ENCODE项目提供技术，既加强现有的ENCODE数据集，又确定新的“结构” 编码“元件及其与基因编码序列的相互作用将有助于理解转录 基因组的景观。 我们已经建立了一条强大的CHIA-PET流水线，从库生产到数据处理，供人类和 小鼠细胞。在这里，我们建议应用该平台来检测大量的细胞系和原代细胞，这些细胞 代表了广泛的细胞空间，具有重要的生物学意义。根据我们目前的生产情况 规模和预算分配，我们的目标是从CTCF和RNAPII产生1024个高质量的数据集 CHIA-PET实验，每个实验有两个生物重复，用于256个生物样本。这条管道的运力将 应用于ENCODE联盟选择的样本，我们的目标是为这个样本库做出贡献 收集高价值生物样本，这些样本可能是财团和 更大的研究社区。这些样本包括原代和体外分化的人类血细胞， 健康和疾病诱导的多能干细胞(IPSC)，从IPSC分化的成熟神经元，以及 来自健康和疾病模型小鼠的几种主要细胞和组织类型。这些样本的选择是为了 扩展ENCODE项目探索的“细胞空间”，还因为CHIA-PET分析将被 尤其与揭示基础生物学有关。

PROJECT SUMMARY/ABSTRACT Efforts to map the functional elements of the human genome, which include elements such as insulators, enhancers, promoters and transcriptional start sites, have historically treated the genome as linear. However, it is now well appreciated that the genome has a three-dimensional (3D) organization that is important for mediating functional associations between regulatory elements and gene-coding sequences. Thus a linear map provides an incomplete picture of the genome, and it is often difficult or impossible to infer functional associations between regulatory elements without a topological framework to provide context. We have developed and advanced a powerful, high-resolution method for providing such a topological framework, Chromatin Interaction Analysis using Paired-End Tag sequencing (ChIA-PET). Using ChIA-PET, we have demonstrated that specific DNA motifs bound by CCTC-binding Factor (CTCF) are critical in defining topological domains and arranging the gene positions for coordinated transcription mediated by RNA Polymerase II (RNAPII). Therefore, the combination of CTCF and RNAPII ChIA-PET will be effective for comprehensively mapping the major structure codes and topological organization that scaffold RNAPII associated transcriptional regulation. We will contribute ChIA-PET technology to the ENCODE Project to both strengthen existing ENCODE datasets and identify new “structure code” elements and their interplays with gene-coding sequences that will aid in understanding the transcriptional landscape of the genome. We have established a robust ChIA-PET pipeline from library production to data processing for human and mouse cells. Here, we propose to apply this platform to assay large numbers of cell lines and primary cells that represent a wide-range of cellular space with important biological significance. Based on our current production scale and estimated budget allocation, we aim to produce 1024 high quality datasets from CTCF and RNAPII ChIA-PET experiments, each with two biological replicates for 256 biological samples. This pipeline capacity will be applied to samples selected by the ENCODE Consortium, and to this sample pool we aim to contribute a collection of high value biological samples that are likely of common interest to both the Consortium and the greater research community. These samples include both primary and in vitro–differentiated human blood cells, healthy and diseased induced pluripotent stem cells (iPSC), mature neurons differentiated from the iPSCs, and several major cell and tissue types from healthy and disease-model mice. These samples were selected to expand the “cell space” explored by the ENCODE Project and also because ChIA-PET analyses will be particularly relevant for revealing fundamental biology.