Mechanisms of Tissue Morphogenesis in C. elegans

发育生物学的一个主要挑战是理解细胞之间复杂的相互作用， 构成复杂器官形态发生基础的组织。为了剖析形态发生的分子基础， 这有利于研究简单的模型组织或器官。这项建议的长远目标是 线虫表皮胚胎形态发生的分子和细胞学解释。优雅 在胚胎发育中，表皮覆盖在基质细胞（腹侧成神经细胞）上。我们以前的 研究表明，通过Eph受体酪氨酸激酶及其肝配蛋白配体的信号传导是必需的， 基底腹侧成神经细胞的运动。Eph信号传导的一种特殊形式涉及受体和 配体激活（双向信号传导）可能促进腹侧成神经细胞之间的粘附或吸引。 在这里，我们打算研究Eph信号和平行通路如何调节成神经细胞粘附， 动作1.我们将结合共聚焦显微镜和透射电镜分析腹侧成神经细胞运动的细胞基础。 时间推移分析、遗传学、药物干预和激光显微手术。结果将允许 我们来解释信号通路如何影响运动。2. GPI连接的肝配蛋白的反向信号传导是 不太了解。在RNAi增强子筛选中，我们鉴定了一个G蛋白亚基作为RNAi的潜在组分。 肝配蛋白反向信号传导。我们将使用遗传学和分子生物学来定义这种G蛋白在肝配蛋白信号传导中的作用。 生化测试3. Eph信号传导功能部分冗余与其他几个途径， 胚胎中的细胞粘附和轴突引导。为了解决这种冗余的性质，我们将分析 一个这样的平行途径涉及KAL-1，C. anosmin-1的elegans直系同源物。我们已经展示了KAL-1 与细胞表面乙酰肝素硫酸化蛋白聚糖多配体聚糖和磷脂酰肌醇蛋白聚糖相互作用。我们将定义 KAL-1和HSPGs的调节途径以及它们是否在相同的细胞中被需要。HSPG是 对C.线虫胚胎形态发生我们将确定其他核心蛋白， 通过候选基因测试和生化纯化的组合来实现这一基本功能。4. 表皮细胞迁移到腹侧成神经细胞基底上。参与表皮的分子- 衬底粘附性是未知。我们的初步数据表明，netrin信号传导既有功能， 成神经细胞迁移和表皮基质附着。我们将定义netrin信号传导在 形态发生和测试netrin在表皮基质粘附中短范围内起作用的假设。 相关性：对形态发生的理解与人类出生缺陷的治疗， 人工器官和组织修复的发展，以及肿瘤的发展。Eph信号尤其具有 与人类遗传疾病、癌症和维持神经干细胞生态位有关。

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.