Core A - Establishing the regulatory mechanisms defining cellular function

项目摘要 染色质结构、转录因子结合和RNA的协同调控 表达是细胞分化的基础。不同层数的测量方法 单细胞内的基因调节可以用来确定细胞的基本调节因子 分化和功能是细胞特性和谱系潜力的敏感标记。对这件事 完了，我们(Buenrostro)最近开发了Share-seq(同步高吞吐量 ATAC和RNA表达与测序)，用于联合测量染色质的可及性 以及在同一单细胞内的基因表达，以低成本和巨大的吞吐量。vbl.使用 Share-Seq，我们已经证明了对来自同一细胞ATAC-和RNA-SEQ数据的联合分析 可用于将转录因子与其目标调节区和远端 对其靶基因的调控元件，使因果基因调控的定义成为可能 网络。此外，我们已经证明，染色质可及性的变化先于变化 在基因表达中，预示着世系承诺，并确定染色质的作用 启动染色质以激活基因的可及性。在此，我们建议设立 基因调控核心(GRC)，将由哈佛大学的Jason Buenrostro博士领导 干细胞与再生生物学系。作为P01提案的一部分，GRC将 使P01成员拥有Share-Seq和其他在 Buenrostro实验室，可用于在功能上注释监管区域，识别关键 转录因子和染色质调节器，定义基因调控网络并解析细胞 各州。GRC将与P01实验室密切合作，设计实验，组织和 采集造血样本(目标1)。GRC将使用SHARE-SEQ联合分析染色质 在单细胞水平上的可及性和RNA表达。这将包括扩展共享序号 到由Share-Seq唯一启用的功能，例如转录谱系的读出 条形码和克隆造血相关突变(目标2)。最后，GRC将是 负责数据的前期处理、数据管理、网页界面的开发 工具和组织P01实验室之间的数据讨论和讲习班(目标3)。总体而言， GRC将为P01社区的单细胞基因组需求提供完整的解决方案 通过一套独特和创新的基因组技术。

Project Summary The coordinated regulation of chromatin structure, transcription factor binding and RNA expression is fundamental to cellular differentiation. Methods for measuring different layers of gene regulation within single cells can be used to determine essential regulators of cell differentiation and function as sensitive markers of cellular identity and lineage potential. To this end, we (Buenrostro) have recently developed SHARE-seq (Simultaneous High-throughput ATAC and RNA Expression with sequencing), for joint measurement of chromatin accessibility and gene expression within the same single-cell, at low-cost and massive throughput. Using SHARE-seq, we have shown that co-analysis of ATAC- and RNA-seq data from the same cell can be used to associate transcription factors to their target regulatory regions and distal regulatory elements to their target genes, enabling the definition of causal gene regulatory networks. Furthermore, we have shown that changes in chromatin accessibility precedes changes in gene expression, foreshadowing lineage commitment, and identifying a role for chromatin accessibility in priming chromatin for gene activation. Here, we propose the establishment of the Gene Regulation Core (GRC), which will be led by Dr. Jason Buenrostro at Harvard University's Department of Stem Cell and Regenerative Biology. As part of this P01 proposal, the GRC will enable the P01 members with SHARE-seq and other genomic technologies established in the Buenrostro lab, which can be used to functionally annotate regulatory regions, identify key transcription factors and chromatin regulators, define gene regulatory networks and parse cellular states. The GRC will work closely with the P01 labs to design experiments, and organize and collect hematopoietic samples (Aim 1). The GRC will use SHARE-seq to jointly profile chromatin accessibility and RNA expression at the single cell level. This will include extending SHARE-seq to capabilities uniquely enabled by SHARE-seq, such as the readout of transcribed lineage barcodes and Clonal Hematopoiesis-associated mutations (Aim 2). Finally, the GRC will be responsible for the primary processing of the data, data management, developing web interface tools and organizing data discussions and workshops between the P01 labs (Aim 3). Overall, the GRC will offer a complete solution for the single-cell genomics needs of the P01 community through a unique and innovative suite of genomic technologies.