Investigation of a neuromesendodermal progenitor population in the posterior avian endoderm

项目摘要/摘要 早期胚胎从看似杂乱无章的干细胞球转变为复杂的和 精确图案化的成人形态需要通过生物物理和生化来调节细胞行为的刻板印象 暗示。使用鸡胚胎，我的研究计划的长期目标是从一个高度 从量化的角度来看，这是如何安排的。实验室大约一半的人专注于机械生物学。 胚胎发育，试图了解塑造胚胎组织和器官的力量，以及 指定这些力的线索。实验室的其余部分研究细胞命运确定，或者在上下文中 早期大脑发育或发育中的肠道。本申请源于后者。 在胚胎发育期间，谱系分离的第一个主要步骤之一是原肠形成，当一个 单层全能细胞分为三个胚层：内胚层、中胚层和外胚层。而当 按照惯例，每个胚层的所有祖细胞都只在 原肠分裂，最近的研究对此提出了挑战，确定了一个持续存在的多能祖先种群 在尾芽中原肠发育结束后。在这个被称为神经中胚层祖细胞的群体中，单个 细胞可以产生后代，形成脊索(中胚层)和神经管(外胚层)。而当 在研究鸡胚胎的肠管形成时，我们最近发现了一组内胚层细胞 尾芽的腹面，似乎经历了广泛的和以前未报道的上皮到... 间充质转化(EMT)。这些细胞侵入邻近的尾芽组织，一直延伸到背部 尾芽的表面。重要的是，细胞散布在神经中胚层所在的区域。 已知祖细胞存在于神经管和脊索中。这表明这些 内胚层细胞是未被重视的神经中胚层前体细胞的来源，如果它们保留了 此外，还能产生内胚层组织，这将是非常了不起的。本申请结合了 经典的胚胎学方法，如命运图谱和基因错误表达和移植实验 单细胞RNA测序，试图解释内胚层简单但令人费解的观察 细胞经历局部的EMT，并在原肠分裂完成时侵入尾芽。这项建议 目的是确定这一潜在祖先群体贡献的组织，并构建一种 这一群体的详细单细胞转录图谱，其中包括身份、调节电路和建议的 可以推断出血统轨迹。当拟议的工作完成后，我们将在香港建立一个新的 体内模型来研究内皮内皮细胞转化，并潜在地发现了一种新的干细胞群体，居住在 内胚层，具有产生中胚层和外胚层细胞的能力。这与长期持有的 胚胎发育期间的血统分离的概念，因此有可能成为 对发育生物学和干细胞重新编程领域具有变革性。

PROJECT SUMMARY/ABSTRACT Transformation of the early embryo from a seemingly disorganized ball of stem cells to the complex and precisely patterned adult form requires stereotyped regulation cell behaviors via biophysical and biochemical cues. Using the chick embryo, the long term goals of my research program are to understand from a highly quantitative perspective how this is orchestrated. Approximately half of the lab focuses on mechanobiology of embryonic development, seeking to understand the forces that shape tissues and organs in the embryo and the cues that specify those forces. The remainder of the lab studies cell fate determination, either in the context of early brain development or in the developing gut. The present application stems from the latter. During embryonic development, one of the first major steps in lineage segregation is gastrulation, when a single totipotent layer of cells separates into three germ layers, endoderm, mesoderm, and ectoderm. While conventionally, it has long been accepted that all progenitors of each germ layer are generated only during gastrulation, recent studies have challenged this, identifying a multipotent progenitor population that persists after gastrulation ends in the tailbud. Within this population, known as neuromesodermal progenitors, a single cell can give rise to progeny that contribute to both notochord (mesoderm) and neural tube (ectoderm). While studying gut tube formation in the chick embryo, we recently identified a population of endoderm cells that line the ventral surface of the tailbud, and appear to undergo a widespread and previously unreported epithelial-to- mesenchymal transition (EMT). These cells invade the neighboring tailbud tissue, reaching as far as the dorsal surface of the tailbud. Importantly, cells can be seen dispersed throughout the region where neuromesodermal progenitors are known to reside, as well as in the neural tube and notochord. This suggests that these endodermal cells represent an unappreciated source of neuromesodermal progenitors, which, if they retain the ability to generate endodermal tissues as well, would be remarkable. The present application combines classical embryology approaches such as fate mapping and grafting experiments with gene misexpression and single-cell RNA Sequencing in an effort to make sense of the simple yet puzzling observation that endoderm cells undergo a localized EMT and invade the tailbud at a time when gastrulation is complete. The proposal aims to identify the tissues to which this prospective progenitor population contributes, and to construct a detailed single-cell transcriptomic atlas of this population, from which identity, regulatory circuits, and proposed lineage trajectories can be inferred. At the completion of the proposed work, we will have established a new in vivo model to study endodermal EMT, and potentially identified a novel stem cell population residing in the endoderm, with the ability to give rise to mesodermal and ectodermal cells. This stands in contrast to long-held notions of lineage segregation during embryonic development, and therefore has the potential to be transformative for the fields of developmental biology and stem cell reprogramming.