Egg to Embryo: Gene Regulatory Circuitry in Development

卵子到胚胎：发育中的基因调控回路

• 批准号: 7881821
• 负责人: ERIC H DAVIDSON
• 金额: $ 0.8万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2009-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7881821
• 关键词: Egg; Embryo; Gene; Regulatory; Circuitry

DESCRIPTION (provided by applicant): This is an application for continuation and broadening of a Program Project now in its 9* year. Here we propose to build in many directions on the large success we have had in solving and authenticating a gene regulatory network (GRN) for development. GRNs provide causal explanations for developmental processes in the terms of the genomic regulatory code, where all species-specific developmental processes are ultimately programmed. A developmental GRN serves as a conceptual, system-level logic map, of direct predictive power. Thus GRNs bridge between functional genomic DNA sequence of regulatory significance and the biology of embryogenesis and body plan formation. They do this by specifying the regulatory interactions which causally drive the progression of regulatory states in diverse cellular territories. During recent years, this Program has been responsible for the experimental solution of the most advanced developmental GRN yet available for any developing animal organism. This is the GRN underlying the specification of the endomesodermal territories of the sea urchin embryo. Recently proof of the principle that as a GRN approaches completion it indeed provides explanation of all the observed biological functions has been obtained in this work. We now intend to capitalize on the growing suite of successful technological approaches to GRN analysis that we have developed, to confront challenges that heretofore were inaccessible, or could not even have been defined. The current sea urchin embryo GRN concerns about half of the embryo, that ultimately forming the gut, the skeletogenic cell lineages and the non-skeletogenic mesoderm, from the earliest zygotic genomic activity (at the beginning of cleavage) to just before gastrulation. A start on the oral and aboral ectodermal GRNs has also been made. We intend to expand this GRN in qualitatively distinct ways which will represent radical advances if successful. In the DAVIDSON COMPONENT the GRN will be expanded to include all regulatory genes predicted by genomic analysis, and observed to be expressed specifically in the endomesodermal territories, as well as in the oral and aboral ectoderm through gastrulation, to the point where the mesodermal cell types have appeared, and endodermal and ectodermal subdivisions (e.g. mouth, stomach, hindgut) have been territorially specified. This will amount to a several fold expansion of the GRN, but also produce a qualitatively new departure. Thus, except for the apical neurogenic domain, it would represent for the first time ever a global, (almost) whole embryo GRN that includes the mechanism for specification of almost all embryonic territories: it would enable us to see the dimensions and the organization of the genomic regulatory program for most of a whole developing embryo, something no one could have imagined until very recently. In the McCLAY COMPONENT the GRN will be extended in a different direction: to the downstream differentiation and morphogenesis genes that do the actual work of building tissues and expressing their particular functions: this will enable us to see how the transcriptional spatial regulatory states established during specification are used to control the actual "jobs" of development, something of which we have but glimpses at present. We believe the conceptual and technological advances of the GRN analysis supported by the current POI are now sufficiently mature so that the time is ripe for transfer of these ideas and methods to developing amniote systems: thus in the BRONNER-FRASER COMPONENT a system-level GRN is to be constructed for chick neural crest. This is not only in its own right a fascinating, vertebrate specific, developmental feature, but due to extensive prior work, would appear ripe for application of many of the approaches emerging from the sea urchin GRN project. This Program has been designed in a heavily interdependent way so that each Component will enjoy and indeed will require close scientific interactions with other Components. These interactions are detailed in the following. No less crucial will be the general reliance of all components on two Core Units. These are the SPECIALIZED RESEARCH SUPPORT (SRC) CORE and the SCIENTIFIC AND ADMINISTRATIVE COORDINATION (SAC) CORE. The functions of the SRC CORE will be to provide special high tech services that will be essential to all three P01 Components, i.e., procedural and instrumental functions that would require a vast outlay of equipment and a vast effort to develop know-how were these functions to be duplicated in each independent lab. Among these, as detailed below, are: (1) construction of cis-regulatory expression vectors by BAC recombineering; (2) accurate amplification of ng quantity RNA preparations; (3) large scale arraying services; (4) bioinformatics services; (5) year round supply of sea urchins; (6) use of a newly on-line instrument, the NanoString nCounter System for direct measurement of large numbers of specific transcripts simultaneously and very quantitatively on small amounts of material. The SAC CORE will constitute the central managerial oversight of the whole project; will interface with individual University grants managers; will coordinate personnel, fiscal, and publication policies; will ensure maximally economic supply acquisitions; and will arrange for webcast conferences among Components as well as biannual meetings of POI personnel.

描述（由申请人提供）：这是一个延续和扩大计划项目，现在在其9* 年的申请。在这里，我们建议建立在许多方向上的巨大成功，我们已经在解决和验证一个基因调控网络（GRN）的发展。GRNs在基因组调控密码方面为发育过程提供了因果解释，所有物种特异性发育过程最终都是编程的。一个发展的GRN作为一个概念，系统级的逻辑地图，直接预测能力。因此，GRNs在具有调节意义的功能基因组DNA序列与胚胎发生和身体计划形成的生物学之间架起了桥梁。他们通过指定调节相互作用来实现这一点，这些相互作用因果地驱动不同细胞区域中调节状态的进展。近年来，该计划一直负责为任何发育中的动物生物体提供最先进的发育GRN的实验解决方案。这是GRN潜在的海胆胚胎的内中胚层领土的规范。最近的证明的原则，作为一个GRN接近完成，它确实提供了所有观察到的生物功能的解释已经在这项工作中获得。我们现在打算利用我们已经开发的越来越多的成功的GRN分析技术方法来应对迄今为止无法实现的挑战，甚至无法定义。目前的海胆胚胎GRN涉及大约一半的胚胎，最终形成肠道，骨骼发生细胞谱系和非骨骼发生中胚层，从最早的合子基因组活动（在卵裂开始时）到原肠形成之前。口腔和离口外胚层GRN也已开始。我们打算以不同的方式扩大GRN，如果成功的话，这将代表根本性的进步。在DAVIDSON COMPONENT中，GRN将扩展至包括通过基因组分析预测的所有调控基因，并观察到在内胚层区域中特异性表达，以及通过原肠胚形成在口腔和离口腔外胚层中特异性表达，直至中胚层细胞类型已经出现，并且内胚层和外胚层亚部（例如，口腔、胃、后肠）已经在区域上指定。这将相当于GRN的几倍扩展，但也产生了质的新出发。因此，除了顶端的神经源性结构域，它将首次代表一个全球性的（几乎）整个胚胎GRN，包括几乎所有胚胎区域的特化机制：它将使我们能够看到大部分发育中胚胎的基因组调控程序的尺寸和组织，直到最近才有人能想象到这一点。在麦克莱成分中，GRN将向一个不同的方向延伸：延伸到下游分化和形态发生基因，这些基因进行构建组织和表达其特定功能的实际工作：这将使我们能够看到，在特化过程中建立的转录空间调控状态是如何用来控制发育的实际“工作”的，我们目前只瞥见了一些东西。我们认为，目前POI支持的GRN分析的概念和技术进步现在已经足够成熟，因此将这些想法和方法转移到开发神经元系统的时机已经成熟：因此，在BRONNER-FRASER组件中，将为小鸡神经嵴构建系统级GRN。这不仅本身是一个迷人的，脊椎动物特有的，发展的特点，但由于大量的前期工作，将出现成熟的应用程序中出现的许多方法从海胆GRN项目。 该计划以高度相互依赖的方式设计，以便每个组件都将享受并确实需要与其他组件进行密切的科学互动。这些相互作用在下文中详述。同样重要的是，所有组成部分一般都依赖于两个核心单元。这些是专门研究支助（SRC）核心和科学和行政协调（SAC）核心。SRC CORE的功能将是提供对所有三个P01组件必不可少的特殊高技术服务，即，如果在每个独立的实验室重复这些功能，则需要大量的设备支出和巨大的努力来开发专门知识。其中，如下详述的是：（1）通过BAC重组工程构建顺式调节表达载体;（2）ng量RNA制备物的精确扩增;（3）大规模阵列服务;（4）生物信息学服务;（5）海胆的全年供应;（6）使用新的在线仪器，NanoString nCounter系统，用于同时直接测量大量特定转录物，并对少量进行非常定量的测量材料。SAC核心将构成整个项目的中央管理监督;将与个别大学赠款经理接口;将协调人事，财政和出版政策;将确保最大限度地经济供应收购;并将安排组件之间的网络广播会议以及POI人员的半年期会议。