An integrative cellular blueprint of vertebrate tissue development

项目总结/摘要 每个动物都是从一个细胞开始的。这种细胞最终如何产生拥有数百万至数万亿个细胞的脊椎动物，每个细胞专门在各种器官系统中执行特定功能，仍然是一个未知数。 生物学的基本问题。到目前为止，在全面捕捉集体细胞行为，产生 脊椎动物的身体计划是不可能的。我们组建了一个具有互补专业知识的团队， 发育遗传学、组织模式、成像和定量生物学，这些将协同作用， 对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， 随着遗传命运绘图技术的最新进展，高分辨率，全动物4D显微镜， 计算方法来推导其完整的谱系图。我们提出的方法是精确的，能够进行前瞻性和回顾性谱系分析，并且它们是可视化的，能够直接识别和分析。 细胞行为的量化。我们已经开发出不可磨灭的标记系统，以激光靶向单细胞， 胚胎并永久激活报告基因表达。胚胎以外的克隆分析将涉及诱导型Cre重组酶和单/双Cre可释放报告转基因。我们的工具集将 允许对任何感兴趣的细胞及其后代进行终身跟踪。通过推导出一个包含数千个标记的 动物，我们将创造前所未有的获得每个器官的传记。为了补充我们的细胞 标记方法，我们将开发成像平台，以阐明标记克隆的整个后代集 及其组织分布。我们还将开发新的计算方法来跟踪发育过程中的克隆，并生成一个数字化地图，概括成年人身体的细胞动力学 计划克隆数据集将用于开发概率模型，以推断增殖和分化动力学如何引起观察到的谱系限制。总的来说，这项巨大的技术成就 将产生器官形态发生的模板，我们将使任何用户都可以轻松访问，可视化和操作。这些身体计划重建可以作为一个基线，用于解释细胞谱系的决定，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.