Synapse formation in the C. elegans nervous system

描述(申请人提供)：谷氨酸是大脑中含量最丰富的兴奋性神经递质，谷氨酸能突触在中枢神经系统的学习、记忆和发育可塑性中发挥关键作用。了解谷氨酸能突触是如何在中枢神经系统中形成和调节的，以开发新的应用于神经疾病的诊断、治疗和预防是至关重要的。有几个基本问题仍未得到解答。谷氨酸受体和信号分子是如何定位于谷氨酸能突触的？这些神经元-神经元突触是如何在发育过程中形成和修改的？线虫一直是研究体内谷氨酸信号的一个很好的模型系统，重点是谷氨酸能突触在发育过程中是如何形成和修饰的。AMPA型谷氨酸受体(GluR)亚单位GLR-1是谷氨酸能信号转导所必需的，定位于线虫神经元之间的突触簇。我们感兴趣的是确定GLR-1的定位(以及谷氨酸受体的定位)是如何进行和调控的。通过正向遗传筛选，我们已经发现了几个突变导致GLR-1定位缺陷的基因。这项提案旨在确定通过这些筛查识别的三个基因的特征。首先，我们将对UNC-43的结构/功能进行分析，UNC-43是GLR-1本地化所需的CaMKll同源物。我们将在UNC-43的特定结构域中引入突变，以测试突变蛋白定位到突触的能力，并拯救针对GLR-1定位缺陷的UNC-43突变。其次，我们将鉴定两个新发现的基因，glo-2和glo-11，它们是GLR-1定位所必需的。我们将克隆GLO-2和GLO-11，确定哪些细胞表达GLO-2和GLO-11蛋白，并确定GLO-2和GLO-11蛋白在细胞内的定位。第三，我们将完成将glo-2和glo-11标识为饱和的屏幕。候选基因将受到严格的标准，以确定哪些基因值得进一步研究。有趣的是，UNC-43/CaMKII的功能在系统发育过程中是保守的。因此，我们希望我们未来对这一系统的实验将揭示关于中枢神经系统的形成和功能的普遍原理，并可能为人类神经疾病的治疗提出新的策略。

DESCRIPTION (provided by applicant): Glutamate is the most abundant excitatory neurotransmitter in the brain, and glutamatergic synapses play a critical role in learning, memory, and developmental plasticity of the central nervous system. It is critical to understand how glutamatergic synapses are formed and regulated in the CNS in order to develop novel applications for the diagnosis, treatment, and prevention of neurological disorders. Several fundamental questions remain unanswered. How are glutamate receptors and signaling molecules localized specifically to glutamatergic synapses? How are these neuron-neuron synapses made and modified during development? C. elegans has been an excellent model system for studying glutamate signaling in vivo with an emphasis on how glutamatergic synapses are formed and modified during development. The AMPA-type glutamate receptor (GluR) subunit GLR-1 is required for glutamatergic signaling and is localized to synaptic clusters between C. elegans neurons. We are interested in determining how GLR-1 localization (and the localization of glutamate receptors in general) is conducted and regulated. By forward genetic screening, we have identified several genes that when mutated result in defects in GLR-1 localization. This proposal aims to characterize three genes identified through those screens. First, we will conduct a structure/function analysis of UNC-43, a CaMKll homolog required for GLR-1 localization. We will introduce mutations into specific domains of UNC-43 to test the ability of the mutant proteins to localize to synapses and to rescue unc-43 mutants for GLR-1 localization defects. Second, we will characterize two newly-identified genes, glo-2 and glo-11, that are required for GLR-1 localization. We will clone glo-2 and glo-11, determine in which cells GLO-2 and GLO-11 proteins are expressed, and determine where GLO-2 and GLO-11 proteins are localized within the cell. Third, we will complete the screen that identified glo-2 and glo-11 to saturation. Candidate genes will be subjected to rigorous criteria to determine which genes merit further study. Interestingly, the function of UNC-43/CaMKII is conserved across phylogeny. Thus, we expect our future experiments with this system to reveal universal principles about the formation and function of the central nervous system, and perhaps suggest new strategies for the treatment of human neurological disorders.