An integrative cellular blueprint of vertebrate tissue development

项目摘要/摘要 每种动物都是从单个细胞开始的。这种细胞最终是如何产生拥有数百万到数万亿个细胞的脊椎动物，每个细胞都专门在各种器官系统中执行特定的功能，仍然是一个 生物学中的基本问题。到目前为止，为了全面捕捉产生 脊椎动物的身体计划是不可能的。我们组建了一支在以下领域具有互补专业知识的团队 发育遗传学、组织图案化、成像学和数量生物学将协同产生血统 脊椎动物的分析达到了一个全新的水平。我们将利用斑马鱼的独特优势，结合 随着遗传命运图谱技术的最新进展，高分辨率、全动物4D显微镜和 计算方法，以得出其完整的谱系图。我们建议的方法是精确的，能够进行前瞻性和回溯性的谱系分析，并且它们是可视的，能够直接识别和 细胞行为的量化。我们已经开发出不可磨灭的标记系统来激光瞄准单个细胞 胚胎并永久激活报告基因的表达。胚胎以外的克隆分析将涉及可诱导的Cre重组酶和单色/多色Cre释放的报告转基因。我们的工具集将 允许任何感兴趣的细胞及其后代被终身跟踪。通过派生出一个包含数千个标签的库 动物，我们将史无前例地获取每个器官的传记。为了补充我们的蜂窝 标记方法，我们将开发成像平台来阐明标记克隆的整个后代集 以及它们的组织分布。我们还将开发新的计算方法来跟踪克隆人的整个发育过程，并生成一张数字化的地图，重述构成成人身体的细胞动力学 计划。克隆数据集将被用来开发概率模型，以推断增殖和分化动态如何引起观察到的谱系限制。总体而言，这一巨大的技术成就 将产生器官形态发生的模板，我们将使任何用户都可以方便地访问、可视化和操作。这些身体平面重建可以用作解释细胞谱系决定的基线，以响应组织损伤、遗传操作或恶性肿瘤。

Project Summary / Abstract Every animal begins as a single cell. How this cell ultimately gives rise to a vertebrate animal possessing millions to trillions of cells, each specialized to perform specific functions in a variety of organ systems, remains a fundamental question in biology. Until now, in toto capture of the collective cellular behaviors that generate the vertebrate body plan has not been possible. We have assembled a team with complementary expertise in developmental genetics, tissue patterning, imaging, and quantitative biology that will synergize to take lineage analysis in vertebrates to an entirely new level. We will utilize the unique strengths of the zebrafish, combined with recent advances in genetic fate mapping techniques, high-resolution, whole-animal 4D microscopy, and computational methodologies to derive its complete lineage map. Our proposed approaches are precise, enabling prospective and retrospective lineage analyses, and they are visual, enabling direct identification and quantification of cell behaviors. We have developed indelible marking systems to laser-target single cells in the embryo and permanently activate reporter gene expression. Clonal analysis beyond the embryo will involve inducible Cre recombinases and single/multicolor Cre-releasable reporter transgenes. Our toolset will allow any cell of interest and its progeny to be followed for life. By deriving a library of thousands of labeled animals, we will create unprecedented access to the biography of each organ. To complement our cellular tagging approaches, we will develop imaging platforms to elucidate the entire progeny sets of labeled clones and their tissue distribution. We will also develop novel computational methods to track clones across development and to generate a digitized map that recapitulates the cellular dynamics that underlie the adult body plan. Clonal data sets will be utilized to develop probabilistic models to infer how proliferation and differentiation dynamics give rise to observed lineage restriction. Collectively, this massive technological achievement will yield templates of organ morphogenesis that we will make facile for any user to access, visualize, and manipulate. These body plan reconstructions can be employed as a baseline for interpreting cell lineage decisions in response to tissue damage, genetic manipulation, or malignancy.