Systems Analysis of cell type differentiation in Xenopus development

摘要 参与胚胎发育的途径一直是了解疾病的丰富资源。 在追踪遗传损伤和环境毒物的影响方面也至关重要 在胎儿中。由于青蛙卵和胚胎的大小，青蛙胚胎特别有用。新的 我们开发了在单细胞水平上测量RNA表达的工具，以及蛋白质和 磷酸肽测量技术，为理解信号如何产生重大进展带来了希望 参与胚胎成熟的细胞指示个体细胞采用特定的命运。我们的第一个目标是定义 使用单细胞转录组的细胞类型，并使用HIGH定义导致特定细胞类型的谱系 解析时间映射。相关重要分子的靶向转录和蛋白质组学研究 指定细胞命运，如转录因子，将提供每个细胞中信号活性水平的指数 单个细胞。这将产生一个前所未有的详细的分子图像涉及的因素 产生表型，以及它们从卵裂早期到中期的相互转化 器官发生。 非洲爪哇的模型系统允许我们解剖出分化为外胚层的早期胚胎的部分。 如果不被打扰，就叫动物帽。在胚胎的情况下，动物帽中的细胞接受一个 发育信号的数量，包括Nodal、BMP和Wnt。这三个信号的组合(在 不同的比例)能够产生许多主要的组织。我们将把动物的帽子暴露在 这三种信号的矩阵，并使用单细胞RNA追踪导致的分化路径 测序。这种对分化决定的分子根源的研究将被用来开发一种 一种基于机器学习的数学方法，用于预测尝试扰动的结果 非洲爪哇的发展。我们会问，细胞类型是否由严格控制的 配体的组合或是否存在难以逃脱的缺省状态 因此形成了大多数胚胎细胞类型。这个问题的答案是 我们对非洲爪哇胚胎如何可靠地发育成青蛙的理解，并将加快努力 创建计算方法来预测其他生物路径的行为，例如参与 癌症。

Summary The pathways involved in embryonic development have been a rich resource for understanding disease in adults, as well as being critically important in tracing the effects of genetic lesions and environmental poisons in the fetus. Frog embryos have been particularly useful due to the large size of the frog egg and embryo. New tools we developed for measuring the expression of RNA at a single-cell level, and advances in protein and phosphopeptide measurement technologies, offer hope for dramatic progress in understanding how signals involved in the maturation of the embryo direct individual cells to adopt specific fates. Our first goal is to define cell types using single-cell transcriptomics, and to define the lineages that result in specific cell types using high resolution temporal mappings. Targeted transcriptomics and proteomics of important molecules involved in specifying cell fate, such as transcription factors, will provide an index of the levels of signaling activity in each individual cell. This will result in an unprecedentedly detailed molecular picture of the factors involved in producing the phenotypes, and their interconversions from the early cleavage stage to the middle of organogenesis. The Xenopus model system allows us to dissect out portions of the early embryo that differentiate to ectoderm if not disturbed, called the animal cap. In the context of the embryo the cells in the animal cap receive a number of developmental signals, including Nodal, BMP, and Wnt. Combinations of these three signals (in different proportions) are capable of generating many of the major tissues. We will expose animal caps to a matrix of these three signals and trace the differentiation pathways that result, using single-cell RNA sequencing. This study of the molecular roots of differentiation decisions will be used to develop a mathematical approach, based on machine learning, to predicting the results of an attempted perturbation of the development of Xenopus. We will ask whether cell types are carefully specified by tightly controlled combinations of ligands or whether there are default states that are hard to escape from ("basins of attraction"), that therefore form the majority of embryonic cell types. The answer to this question is central to our understanding of how the Xenopus embryo reliably develops into a frog, and will accelerate efforts to create computational methods to predict the behavior of other biological pathways such as those involved in cancer.