RUI: Examining Molecular Players Integrating Autophagy and Neuronal Development and Maintenance

在胚胎发育期间，神经系统细胞生长并分枝，与其他神经细胞(突触)建立专门的连接。突触对于神经系统正常运作所需的细胞间通信是必不可少的，突触是在被称为轴突的线状细胞投射到达另一个目标细胞后创建的。突触形成的一个步骤是通过一种称为自噬的过程从轴突中移除额外的物质。本研究项目考察了一种关键蛋白(UNC-33/CRMP-2)的确切作用，该蛋白是从以前的工作中已知的影响轴突生长和维持的。实验使用线虫，因为它为遗传操作和活细胞成像提供了优势。UNC-33/CRMP2的基因编码中将引入遗传损伤，该分子同时调节自噬和突触发育的假设将通过复杂的细胞成像和分析技术进行验证。这项研究有可能有助于更深入地了解神经元电路发育的分子机制，以及更好地了解突触生长和维持方面的缺陷，这些缺陷是包括自闭症、精神分裂症和焦虑症在内的一系列神经疾病的特征。此外，这项研究将在西南俄克拉荷马州立大学本科生的帮助和支持下进行。该项目包括针对当地小学和高中的各种外展计划，以及与公众就与大脑发育和功能相关的主题进行的教育参与。尽管围绕神经发育和神经元存活的信息丰富，但仍不清楚为什么在轴突生长和细胞骨架稳定的水平上工作的神经元组件与自噬机制的组件绑定。此外，自噬基因产物最初在酵母中被鉴定为Atg1、6、8、9、13、18，为什么会在神经系统中丰富，目前还不确定。为了阐明自噬与神经元发育和存活之间的联系，研究人员建议对UNC-33在神经元自噬中的作用进行表征。线虫UNC-33基因编码CRMP/TOAD/ULIP/DRP家族的保守成员。在线虫中，UNC-33亚型介导神经元的轴突引导和轴突发生。此外，初步调查表明，UNC-33突变体在达尔幼虫形成方面存在缺陷，达尔幼虫形成是一种受阻的发育阶段，由于细胞自噬增强，该阶段可在恶劣环境中存活。在这项研究中，研究人员将检验自噬和UNC-33有助于防止神经元发育和/或神经元稳定性缺陷的假设。在目标1中，他们将定义UNC-33在神经元自噬中的作用。在目标2中，他们将描述基础自噬在维持神经元等长寿命细胞中的作用。在目标3中，他们建议揭示UNC-33；daf-2双重突变体合成致死性的潜在机制。总之，这些研究将增进对自噬和神经元发育和维护的理解；将显著加强本科教育和课堂研究的整合；并将促进与当地社区的联系。

During embryonic development, nervous system cells grow and branch off to make specialized connections with other nerve cells (synapses). Synapses are essential for the cell-to-cell communication that nervous systems need to function properly, and are created after threadlike cellular projections, known as axons, reach another target cell. One step in synapse formation involves removing extra material from axons by a process called autophagy. The present research project examines the precise role of one key protein (UNC-33/CRMP-2) that is known from previous work to affect axon growth and maintenance. Experiments use the roundworm C. elegans because of the advantages it offers for genetic manipulation and live cell imaging. Genetic lesions will be introduced in the gene coding for UNC-33/CRMP2, and the hypothesis that this molecule simultaneously regulates autophagy and synapse development will be tested using sophisticated cellular imaging and analysis techniques. This study has the potential to contribute to a deeper understanding of the molecular mechanisms underlying the development of neuronal circuits, as well as to a better understanding of defects in the growth and maintenance of synapses that are characteristic of a range of neurological diseases including autism, schizophrenia, and anxiety disorders. Moreover, this research will be carried out with the assistance of, and support the training of, undergraduate students at Southwestern Oklahoma State University. This project includes a variety of outreach programs for local elementary and high schools, and educational engagement with the general public on topics related to brain development and function.Despite the wealth of information surrounding neurodevelopment and neuronal survival, it is still unclear why neuronal components shown to work at the level of axonal outgrowth and cytoskeleton stability bind to components of the autophagy machinery. Moreover, it is undetermined why autophagy gene products, first characterized in yeast as Atg1, 6, 8, 9, 13, 18, are enriched in the nervous system. To shed light on the link of autophagy and neuronal development and survival, the investigators propose to characterize the role of UNC-33 in neuronal autophagy. The nematode unc-33 gene encodes for conserved members of the CRMP/TOAD/Ulip/DRP family of proteins. In C. elegans, UNC-33 isoforms mediate axonal guidance and axonogenesis in neurons. Moreover, preliminary investigations show that unc-33 mutants have a defect in dauer larva formation, an arrested developmental stage that survives in harsh environments due to enhanced cellular autophagy. In this study, the investigators will test the hypothesis that autophagy and UNC-33 serve to protect against defects in neuronal development and/or neuronal stability. In aim 1, they will define the role of UNC-33 in autophagy in neurons. In aim 2, they will characterize the role of basal autophagy in the maintenance of long-lived cells such as neurons. In aim 3, they propose to uncover the mechanism underlying the synthetic lethality of unc-33; daf-2 double mutants. Together, these studies will advance understanding of both autophagy and neuronal development and maintenance; will significantly enhance undergraduate education and the integration of research in the classroom; and will promote outreach to local communities.