Architecture and Dynamics of a Gene Regulatory Network Controlling Cell Fate

控制细胞命运的基因调控网络的结构和动力学

• 批准号: 9908461
• 负责人: Rachel Maczis Shahan
• 金额: $ 6.49万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-01 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9908461
• 关键词: Arabidopsis; Architecture; Cell Differentiation process; Cell Fate Control; Cell Maintenance; Cell Maturation; Cell Proliferation; Cells; Complex; Coupled; Data Analyses; Development; Developmental Process; Ectopic Expression; Embryonic Development; Endoderm Cell; Environment; Epithelial; Epithelium; Event; Expression Profiling; Foundations; Future; Gene Expression; Gene Expression Profiling; Genes; Genetic; Genetic Screening; Genetic Transcription; Goals; Image; Imaging Techniques; Individual; Knowledge; Lead; Lighting; Logic; Maps; Mentors; Methodology; Microscopy; Modeling; Molecular; Organ; Organism; Overlapping Genes; Pathway interactions; Pattern; Peptides; Plant Roots; Play; Process; Proteins; Regulation; Regulator Genes; Reporter; Research; Resolution; Resources; Role; Signal Transduction; Signaling Molecule; Structure; Study models; System; Technology; Testing; Time; TimeLine; Tissues; Training; Transcriptional Regulation; Universities; Work; Zebrafish; cancer cell; cell type; collaborative environment; design; differential expression; experimental study; forward genetics; genetic approach; insight; mathematical model; molecular dynamics; multidimensional data; mutant; non-Native; novel; organ regeneration; pluripotency; programs; protein expression; single-cell RNA sequencing; stem cell differentiation; stem cell niche; stem cell proliferation; stem cells; tissue regeneration; transcription factor

Project Summary Cell fate acquisition is a fundamental developmental process in all multicellular organisms and a growing body of evidence indicates that gene regulatory networks (GRNs) play an important role. However, the molecular regulation of the entire pathway from stem cell to differentiation has never been defined for any tissue. The Arabidopsis root, with its simple structure and defined stem cell niche, is a tractable model for studying the transcriptional regulation of cell fate. Over two decades of work have outlined the GRN that orchestrates cell proliferation and specification of the endodermis, a tissue analogous to the mammalian epithelium. This GRN is mapped in sufficient detail to now mathematically model its dynamics. However, crucial questions remain regarding downstream differentiation events. These include what regulators control endodermal fate stabilization and differentiation? And how closely are such regulators connected to the transcriptional events controlling stem cell proliferation? Until recently, technological constraints made it very difficult to study the molecular dynamics underlying development of a single cell type in the context of an entire organ or organism. The research proposed here utilizes new advances in imaging and transcriptional profiling to study protein and gene expression dynamics at cellular resolution. The overall goal of this proposal is to expand the topology of the endodermal GRN and begin to quantify the dynamics underlying differentiation. To achieve this goal, the proposed specific aims include conducting a forward genetic screen in a sensitized genetic background to uncover novel regulators of endodermal identity (Aim 1). In parallel, single cell RNA-sequencing experiments will define gene cascades underlying cell maturation events, thus providing systems-level insight into regulation of the entire pathway from stem cell to differentiated endodermis (Aim 2). To experimentally quantify the dynamics of known and novel regulators in the context of differentiation, state-of-the-art imaging techniques will be employed to track changes in protein accumulation over time at a cellular resolution in living roots (Aim 3). Together, these aims should expand the architecture of the endodermal GRN and begin to define how information flows through it to orchestrate cell differentiation events. The intellectually stimulating and collaborative environment of Duke University, coupled with individualized mentoring and enabling technology in the sponsoring lab, provide a resource-rich environment in which to conduct the proposed experiments. This research plan will facilitate advanced training in genetics, systems-level transcriptional regulation, multi-dimensional data analysis, and time-lapse microscopy, thereby providing the foundation for a long-term research program to dissect and model the GRNs underlying fundamental developmental processes. Insights gained from this work will deepen our mechanistic understanding of how stem cell progeny traverse the pathway to differentiation, thereby producing methodological and conceptual advances to inform tissue regeneration.

项目摘要 细胞命运的获取是所有多细胞生物的基本发育过程，并且身体不断增长 证据表明基因调节网络（GRN）起着重要作用。但是，分子 从未针对任何组织定义从干细胞到分化的整个途径的调节。这 拟南芥根具有简单的结构和定义的干细胞生态位，是一个可拖动的模型 细胞命运的转录调节。超过二十年的工作概述了精心策划细胞的GRN 内胚层的增殖和规范，一种类似于哺乳动物上皮的组织。这个grn是 绘制足够的详细映射到现在数学上对其动态进行建模。但是，仍然存在关键问题 关于下游分化事件。其中包括哪些监管机构控制内皮层命运稳定 和分化？以及这种调节器连接到控制茎的转录事件的亲密关系 细胞增殖？直到最近，技术限制使研究分子动力学变得非常困难 在整个器官或生物体的背景下，单细胞类型的基础开发。研究提出了 这里利用成像和转录分析的新进步来研究蛋白质和基因表达 细胞分辨率的动力学。该提议的总体目标是扩大内胚层的拓扑结构 GRN并开始量化分化的动力学。为了实现这一目标，提议的具体 目的包括在敏感的遗传背景中进行正向遗传筛查以发现新的调节剂 内胚层身份（AIM 1）。同时，单细胞RNA测序实验将定义基因级联 潜在的细胞成熟事件，从而提供系统级别的洞察 干细胞分化的内胚层（AIM 2）。实验量化已知和新颖的动力学 在差异化的背景下，将采用最先进的成像技术来跟踪变化 在蛋白质在生物根中的细胞分辨率中随着时间的推移积累（AIM 3）。在一起，这些目标应该 扩展内胚层GRN的架构，并开始定义信息如何通过该信息流到 编排细胞分化事件。公爵的智力刺激和协作环境 大学，以及在赞助实验室中的个性化指导和启用技术，提供了一个 资源丰富的环境进行了提出的实验。该研究计划将有助于 遗传学，系统级转录调节，多维数据分析和 延时显微镜，从而为长期研究计划提供了基础，以剖析和建模 基本的基本发展过程。从这项工作中获得的见解将加深我们 对干细胞后代如何穿越分化途径的机械理解，从而产生 方法论和概念上的进步，以告知组织再生。