Mechanisms of Tissue Morphogenesis in C. elegans

发育生物学中的一个主要挑战是理解复杂的细胞相互作用 构成复杂器官形态发生的基础的组织。剖析其形态发生的分子基础 S有利于研究简单的模型组织或器官。这项提议的长期目标是 线虫表皮胚胎形态发生的分子和细胞解释。 在胚胎发育过程中，表皮覆盖在底物细胞上，即腹侧神经母细胞。我们以前的 研究表明，通过Eph受体酪氨酸激酶及其Ephin配体的信号转导是 底物腹侧神经母细胞的运动。一种特殊形式的Eph信号，涉及受体和 配体激活(双向信号)可能促进腹侧神经母细胞之间的黏附或吸引。 在这里，我们建议研究Eph信号和平行通路如何调节神经母细胞黏附和 动静。1.我们将结合共聚焦分析腹侧神经母细胞运动的细胞基础 时间流逝分析、遗传学、药物干预和激光显微外科。结果将允许 我们来解释信号通路是如何影响运动的。2.GPI连接的eparins的反向信号是 人们对此知之甚少。在RNAi增强子筛选中，我们确定G蛋白亚基是一个潜在的成分 伊夫林反向信号。我们将用基因和基因来确定G蛋白在ePhin信号传递中的作用 生化测试。3.Eph信号部分冗余地与其他几条途径一起发挥作用 细胞黏附在胚胎和轴突的引导下。为了解决这种冗余的性质，我们将分析 其中一个涉及KAL-1的平行通路，即Anosmin-1的线虫直系同源物。我们已经向KAL-1展示了 与细胞表面乙酰肝素、硫酸蛋白多聚糖、Syndecan和葡聚糖相互作用。我们将定义 KAL-1和HSPGs的调控途径以及它们是否需要在同一细胞中。HSPG是 线虫胚胎形态发生所必需的。我们将确定其他核心蛋白 这一基本功能是由候选基因检测和生化纯化相结合实现的。4. 表皮细胞在腹侧神经母细胞底物上迁移。表皮中涉及的分子- 基材粘附性是未知的。我们的初步数据表明，netrin信号在 神经母细胞迁移和表皮基质附着。我们将定义Netrin信号在 形态发生，并检验网状蛋白在表皮基质黏附中短程作用的假设。 相关性：对形态发生的理解与人类出生缺陷的治疗有关，对于 人工器官和组织修复的发展，以及肿瘤的发展。尤其是Eph信号具有 与人类遗传病、癌症和维持神经干细胞的利基有关。

A major challenge in developmental biology is to understand the intricate cellular interactions between tissues that underlie morphogenesis of complex organs. To dissect the molecular basis of morphogenesis it s advantageous to study simple model tissues or organs. The long term objective of this proposal is a molecular and cellular explanation of embryonic morphogenesis of epidermis of the nematode C. elegans. In embryonic development the epidermis spreads over substrate cells, the ventral neuroblasts. Our previous work showed that signaling via the Eph receptor tyrosine kinase and its ephrin ligands is required for movements of substrate ventral neuroblasts. A specific form of Eph signaling involving both receptor and igand activation (bidirectional signaling) likely promotes adhesion or attraction among ventral neuroblasts. Here we propose to investigate how Eph signaling and parallel pathways regulate neuroblast adhesion and movements. 1. We will analyze the cellular basis of ventral neuroblast motility by a combination of confocal timelapse analysis, genetics, pharmacological interventions, and laser microsurgery. The results will allow us to interpret how the signaling pathways affect motility. 2. Reverse signaling by GPI-linked ephrins is poorly understood. In RNAi enhancer screens we identified a G protein subunit as a potential component of ephrin reverse signaling. We will define the role of this G protein in ephrin signaling using genetic and biochemical tests. 3. Eph signaling functions partly redundantly with several other pathways that provide cell adhesion in the embryo and in axon guidance. To address the nature of this redundancy we will analyze one such parallel pathway involving KAL-1, the C. elegans ortholog of anosmin-1. We have shown KAL-1 interacts with the cell surface heparan sulfated proteoglycans syndecan and glypican. We will define the regulatory pathway of KAL-1 and HSPGs and whether they are required in the same cells. HSPGs are essential for C. elegans embryonic morphogenesis. We will identify the other core proteins that account for this essential function by a combination of candidate gene testing and biochemical purification. 4. Epidermal cells migrate over the ventral neuroblast substrate. The molecules involved in epidermal- substrate adhesion are not known. Our preliminary data suggest netrin signaling has functions both in neuroblast migration and in epidermal substrate attachment. We will define the roles of netrin signaling in morphogenesis and test the hypothesis that netrin acts over short range in epidermal substrate adhesion. Relevance: An understanding of morphogenesis is relevant to treatment of human birth defects, to the development of artificial organs and tissue repair, and to tumor development. Eph signaling in particular has been implicated in human genetic disease, cancer, and in maintenance of neural stem cell niches.