Investigation of Anaphase Promoting Complex function in synaptic transmission

突触传递后期促进复合体功能的研究

• 批准号: 8287906
• 负责人: Jennifer Kowalski
• 金额: $ 37.31万
• 依托单位: BUTLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8287906
• 关键词: Affect; Alzheimer' s Disease; Animals; Behavioral Assay; Behavioral Genetics; Biochemical; Biochemistry; Brain; Caenorhabditis elegans; Cell Cycle; Data; Defect; Development; Disease; Drug Delivery Systems; Enzymes; Epilepsy; Equilibrium; Excitatory Synapse; Family; Functional disorder; G-Protein-Coupled Receptors; Genes; Genetic; Glutamates; Glycopeptides; Goals; Hormones; Human; Huntington Disease; Image; Investigation; Lead; Learning; Link; Location; Memory; Mental Retardation; Modeling; Molecular; Muscle; Muscle Contraction; Mutation; Nematoda; Nervous System Physiology; Nervous system structure; Neurodegenerative Disorders; Neuromuscular Junction; Neurons; Parkinson Disease; Physiological; Positioning Attribute; Proteins; RNA Interference; Reagent; Regulation; Research; Role; Seizures; Signal Transduction; Site; Synapses; Synaptic Transmission; Syndrome; System; Techniques; Testing; Therapeutic; Ubiquitin; Ubiquitination; Work; anaphase-promoting complex; cell type; cholinergic; enzyme activity; enzyme pathway; enzyme substrate; experience; follow-up; in vivo; insight; loss of function; mutant; nervous system disorder; neurogenetics; neuron development; novel; polypeptide; postsynaptic; presynaptic; prevent; protein degradation; protein function; receptor; research study; synaptic function; transmission process; ubiquitin ligase

DESCRIPTION (provided by applicant): The ubiquitin signaling system, which promotes regulated protein degradation, is essential for synapse development and synaptic transmission. Hundreds of enzymes exist that have the ability to add or remove ubiquitin polypeptides from proteins. Less is known about the functions and relevant substrates of these enzymes in neurons and at synapses. The long-term goal is to determine how ubiquitin enzymes regulate synaptic transmission. Given that ubiquitin system dysfunction is linked to neurological and neurodegenerative diseases in which aberrant synaptic transmission occurs, such information may lead to the identification of drug targets to aid in treating these conditions. Recent work established that the Anaphase Promoting Complex (APC), an ubiquitin ligase that controls protein abundance in the cell cycle, also regulates neuronal development and glutamatergic synaptic transmission; yet much remains unknown regarding its mechanism of action and relevant substrates at these or other synapses. The objective of this proposal is to elucidate the molecular mechanisms by which the APC regulates synaptic transmission in vivo. This will be done using the model nematode, Caenorhabditis elegans, which shares significant conservation of nervous system proteins and function with humans. Preliminary experiments indicate that, in addition to regulating glutamatergic synapses, the APC negatively regulates transmission at the neuromuscular junction (NMJ), a tripartite synapse in C. elegans where contraction of postsynaptic muscle is controlled by a balance of inputs from excitatory cholinergic and inhibitory GABAergic motorneurons. The APC may act in any or all of these cell types to impact the amount of muscle activity. Additional preliminary localization studies and behavioral assays led to the central hypothesis that the APC acts in presynaptic motorneurons to promote ubiquitination and degradation of NMJ proteins important for increasing muscle excitation. In Aim 1, cell type-specific loss of function and rescue experiments, behavioral assays, and quantitative imaging will be used to determine the site and mechanism of APC action during NMJ signaling. Additional preliminary data led to the identification of one potential APC substrate at the NMJ, the G protein-coupled receptor, FSHR-1. In Aim 2, genetic and behavioral analyses, along with imaging and biochemistry, will be employed to characterize the role of FSHR-1 at the NMJ and the relationship between FSHR-1 and the APC. In Aim 3, RNA interference will be used to identify additional APC substrates. This research will provide new insight into how the balance of excitatory to inhibitory transmission is regulated-a balance lost i human seizure syndromes, such as epilepsy. These studies also will contribute to the global picture of how ubiquitin signaling impacts synaptic transmission, which is critical for learning an memory. Given that ubiquitin system misregulation occurs in Alzheimer's, Parkinson's, and Huntington's Diseases, and mutations in ubiquitin enzymes are linked to several neurogenetic disorders, elucidating how the APC and its substrates control synaptic transmission may facilitate the development of relevant therapeutics. PUBLIC HEALTH RELEVANCE: This project involves determining the molecular mechanism by which the Anaphase Promoting Complex (APC), an ubiquitin ligase enzyme, regulates synaptic transmission in vivo. Proper signaling at neuronal synapses is required for learning and memory and underlies all nervous system function. Misregulation of synaptic transmission and ubiquitin system dysfunction occur in many neurological disorders. Thus, a more complete understanding of the mechanism of action and relevant substrates of the APC ubiquitin ligase will contribute critical information to the global picture of synaptic transmission and ubiquitin enzyme activity.

描述（由申请人提供）：泛素信号系统促进调节蛋白质降解，对突触发育和突触传递至关重要。存在数百种具有从蛋白质添加或去除泛素多肽的能力的酶。对这些酶在神经元和突触中的功能和相关底物知之甚少。长期目标是确定泛素酶如何调节突触传递。鉴于泛素系统功能障碍与发生异常突触传递的神经系统和神经退行性疾病有关，这些信息可能导致鉴定药物靶点以帮助治疗这些病症。最近的研究表明，后期促进复合物（APC），一种控制细胞周期中蛋白质丰度的泛素连接酶，也调节神经元发育和神经元突触传递;但关于其作用机制和这些或其他突触的相关底物仍有很多未知。本研究的目的是阐明APC在体内调控突触传递的分子机制。这将使用模型线虫秀丽隐杆线虫来完成，秀丽隐杆线虫与人类共享神经系统蛋白质和功能的重要保守性。初步的实验表明，除了调节突触外，APC还负调节神经肌肉接头（NMJ）的传递，NMJ是C.其中突触后肌肉的收缩由来自兴奋性胆碱能和抑制性GABA能运动神经元的输入的平衡控制。APC可以在任何或所有这些细胞类型中起作用，以影响肌肉活动的量。额外的初步定位研究和行为测定导致了中心假设，即APC在突触前运动神经元中起作用，以促进对增加肌肉兴奋重要的NMJ蛋白的泛素化和降解。在目标1中，将使用细胞类型特异性功能丧失和拯救实验、行为测定和定量成像来确定NMJ信号传导期间APC作用的位点和机制。额外的初步数据导致在NMJ鉴定出一种潜在的APC底物，G蛋白偶联受体FSHR-1。在目标2中，遗传和行为分析，沿着成像和生物化学，将用于表征FSHR-1在NMJ中的作用以及FSHR-1和APC之间的关系。在目标3中，RNA干扰将用于鉴定其他APC底物。这项研究将提供新的见解兴奋性抑制性传输的平衡是如何调节的平衡失去了人类癫痫综合征，如癫痫。这些研究还将有助于了解泛素信号如何影响突触传递的全球图景，这对学习记忆至关重要。鉴于泛素系统失调发生在阿尔茨海默氏症、帕金森氏症和亨廷顿氏症中，并且泛素酶的突变与几种神经遗传性疾病有关，阐明APC及其底物如何控制突触传递可能有助于相关疗法的开发。 公共卫生关系：该项目涉及确定后期促进复合物（APC）（一种泛素连接酶）调节体内突触传递的分子机制。神经元突触处的适当信号传导是学习和记忆所必需的，并且是所有神经系统功能的基础。突触传递失调和泛素系统功能障碍发生在许多神经系统疾病中。因此，更全面地了解APC泛素连接酶的作用机制和相关底物将为突触传递和泛素酶活性的全局图像提供关键信息。

项目成果

The Anaphase-Promoting Complex (APC) ubiquitin ligase affects chemosensory behavior in C. elegans.

• DOI: 10.7717/peerj.2013
• 发表时间: 2016
• 期刊: PeerJ
• 影响因子: 2.7
• 作者: Wang J;Jennings AK;Kowalski JR
• 通讯作者: Kowalski JR