Heparan sulfate proteoglycans in signaling and development

硫酸乙酰肝素蛋白多糖在信号传导和发育中的作用

• 批准号: 10393549
• 负责人: Hiroshi Nakato
• 金额: $ 33.92万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10393549
• 关键词: Affect; Animal Model; Animals; Biological; Biology; Bone Morphogenetic Proteins; Carbohydrates; Cell Communication; Cell Line; Cell surface; Cells; Chondroitin Sulfate A; Chondroitin Sulfate Proteoglycan; Development; Drosophila genus; Enzymes; Erinaceidae; Event; Goals; Growth Factor; Heparan Sulfate Proteoglycan; Integral Membrane Protein; Knowledge; Label; Leucine-Rich Repeat; Ligand Binding; Link; Mediating; Metabolic; Modeling; Molecular; Morphogenesis; Output; Pathway interactions; Pattern; Pattern Formation; Phenotype; Physiological; Play; Proteins; Proteoglycan; Proteomics; Radio; Research; Role; Signal Transduction; Structure; Structure-Activity Relationship; System; Trachea; genetic approach; in vitro Model; in vivo; in vivo Model; interdisciplinary approach; morphogens; mutant; novel; receptor; receptor function; stem cell niche

Project Summary The long-term goal of our research is to understand fundamental principles of cell communications mediated by heparan sulfate proteoglycans (HSPGs). HSPGs are a special type of carbohydrate-modified proteins that serve as co-receptors for various growth factors, including bone morphogenetic proteins, Wnt/Wingless, and Hedgehog. These HSPG co-receptors play critical roles in quantitative and robust control of signaling output. We study in vivo functions of HSPGs using the Drosophila model. Our earlier research has established critical roles of HSPGs in development, namely in morphogen signaling and gradient formation. In addition, we have demonstrated that HSPGs regulate the stem cell niche as a universal factor. Although proteoglycan biology has made significant progress, several major questions still remain to be elucidated. For example, the molecular mechanisms of co-receptor activities of HSPGs are poorly understood. It is also unknown how distinct HS structures regulate specific signaling and patterning events. It is now critical to develop an interdisciplinary approach, which can directly link detailed HS structural motifs, ligand binding, quantitative signaling output, and developmental/physiological phenotypes. Our previous studies suggested that HSPGs cooperate with other factors to exert co-receptor activity on the cell surface. To reveal the molecular basis for co-receptor function, we use proteomic and genetic approaches to identify these missing players. We recently found that Windpipe (Wdp), a transmembrane protein containing four leucine-rich repeats, acts as a novel negative co-receptor for Hh signaling. We also showed that Wdp is a chondroitin sulfate proteoglycan (CSPG). Thus, the Hh pathway is controlled by two classes of proteoglycan co-receptors: HSPGs (positive regulators) and Wdp (CSPG, a negative regulator). We will elucidate how such a "dual PG co-receptor system" achieves quantitatively controlled signaling output and precise pattern formation. To understand how a change in HS structure affects signaling and morphogenesis, we will establish an "in vitro" model using Drosophila cells. Despite the many strengths of the Drosophila model for in vivo studies, information on Drosophila HS structure is limited. This is mainly due to the difficulty of metabolic radio-labeling of HS in vivo using Drosophila animals. To fill this gap, we have recently generated novel Drosophila cell lines mutant for five HS modifying enzymes. Our studies using these cell lines will provide a direct link between detailed structural information of Drosophila HS and a wealth of knowledge on biological phenotypic information obtained over the last two decades using this animal model. Together, this application will advance our field by: (1) defining fundamental molecular mechanisms of co-receptor function and (2) comprehensive understanding of the structure-function relationship of HS.

项目概要 我们研究的长期目标是了解细胞通讯的基本原理 由硫酸乙酰肝素蛋白聚糖（HSPG）介导。 HSPG 是一种特殊类型的碳水化合物修饰物 作为各种生长因子的共同受体的蛋白质，包括骨形态发生蛋白， Wnt/无翼和刺猬。这些 HSPG 共受体在定量和鲁棒性方面发挥着关键作用。 控制信号输出。我们使用果蝇模型研究 HSPG 的体内功能。我们早期的 研究已经确定 HSPG 在发育中的关键作用，即在形态发生信号传导和 梯度形成。此外，我们还证明 HSPG 作为一种调节干细胞生态位的机制。 普遍因素。尽管蛋白多糖生物学已取得重大进展，但仍有几个主要问题 仍有待阐明。例如，HSPGs 共受体活性的分子机制是 不太了解。目前还不清楚不同的 HS 结构如何调节特定的信号传导和模式 事件。现在至关重要的是开发一种跨学科的方法，可以直接链接详细的 HS 结构基序、配体结合、定量信号输出和发育/生理表型。 我们之前的研究表明HSPGs与其他因素合作发挥共受体 细胞表面的活性。为了揭示共受体功能的分子基础，我们使用蛋白质组学和 基因方法来识别这些失踪的球员。我们最近发现 Windpipe (Wdp) 含有四个富含亮氨酸重复序列的跨膜蛋白，作为 Hh 的新型负共受体 发信号。我们还表明 Wdp 是一种硫酸软骨素蛋白聚糖 (CSPG)。因此，Hh途径是 由两类蛋白多糖共受体控制：HSPG（正调节因子）和 Wdp（CSPG，一种 负调节器）。我们将阐明这样的“双PG共受体系统”如何实现定量 受控的信号输出和精确的模式形成。 为了了解 HS 结构的变化如何影响信号传导和形态发生，我们将建立 使用果蝇细胞的“体外”模型。尽管果蝇体内模型具有许多优点 研究表明，有关果蝇 HS 结构的信息有限。这主要是由于新陈代谢困难造成的。 使用果蝇动物对 HS 进行体内放射性标记。为了填补这一空白，我们最近生成了小说 五种 HS 修饰酶的果蝇细胞系突变体。我们使用这些细胞系的研究将提供 果蝇 HS 的详细结构信息与丰富的生物学知识之间的直接联系 使用这种动物模型在过去二十年中获得的表型信息。 总之，该应用将通过以下方式推进我们的领域：（1）定义基本分子机制 (2) 全面了解 HS 的结构与功能关系。