In vivo analysis of endodermal and dorsal forerunner cell migration

项目摘要 在发育中的胚胎中，器官和组织的形成来自许多人的集体行动。 个体细胞，其可以改变它们的形状、大小、粘附性和运动性。我们的长期目标是了解 这些细胞行为是如何在空间和时间上被调节以正确地构建每个组织和器官的。在 在这个项目中，我们将利用两种密切相关但表型不同的细胞类型，内胚层和背侧 为了更好地了解迁移间充质细胞之间的调节， 状态和连贯的上皮细胞。在原肠胚形成早期，内胚层细胞表现出间充质特征; 它们高度迁移，空间分散，避免相互接触。相反， 细胞彼此紧密相连，即使在迁移时也显示上皮特征 集体的值得注意的是，这两种细胞类型由相同的发育信号指定，并表达许多 相同的标记基因，尽管它们的迁移行为不同。在本项目的目标1中，我们将使用单- 细胞RNA测序和生物信息学分析，以比较内胚层和背 前导细胞，并确定差异表达的候选基因可能参与细胞的调控， 迁移模式在目标2中，我们将产生分裂荧光蛋白敲入斑马鱼系，以表征 关键内胚层和背侧前体基因的表达和定位。这种方法基于 技术开发的母赠款，以这种多样性补充申请。如果成功，这项研究将 在两种密切相关但不同的细胞类型中确定负责不同迁移模式的关键基因。我们 这些发现将揭示更广泛的问题，即转录状态的微小差异如何转化为 细胞行为的主要差异。

PROJECT SUMMARY In the developing embryos, the formation of organs and tissues arises from the collective actions of many individual cells which may change their shape, size, adhesion, and motility. Our long-term goal is to understand how these cellular behaviors are regulated in both space and time to correctly build each tissue and organ. In this project, we will leverage two closely related yet phenotypically distinct cell types, the endodermal and dorsal forerunner cells in zebrafish embryos, to better understand the regulation between migratory mesenchymal states and coherent epithelia. During early gastrulation, endodermal cells exhibit mesenchymal characteristics; they are highly migratory, spatially dispersed, and avoid contact with each other. In contrast the dorsal forerunner cells are tightly associated with each other and display epithelial characteristics, even as they migrate collectively. Remarkably, both cell types are specified by the same developmental signals and express many of the same marker genes, despite their divergent migration behaviors. In Aim 1 of this project, we will use single- cell RNA sequencing and bioinformatic analysis to compare the transcriptional profiles of endodermal and dorsal forerunner cells and identify differentially expressed candidate genes likely involved in the regulation of cell migration modes. In Aim 2, we will generate split fluorescent protein knock-in zebrafish lines to characterize the expression and localization of key endodermal and dorsal forerunner genes. This approach is based on technology developed in the parent grant to this diversity supplement application. If successful, this study will identify key genes responsible for different modes of migration in two closely related, yet distinct cell types. Our findings will shed light on the broader question of how small differences in transcriptional states translate into major differences in cellular behavior.