The Role of Highwire/DFsn Ubiquitination Complex During Synaptic Development

Highwire/DFsn 泛素化复合物在突触发育过程中的作用

基本信息

批准号：
8672700
负责人：
Chunlai Wu
金额：
$ 30.14万
依托单位：
LSU HEALTH SCIENCES CENTER
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2015-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8672700
关键词：
Address Affinity Affinity Chromatography Alzheimer&apos s Disease Architecture Autistic Disorder Back Binding Proteins Brain Child Complex Cullin Proteins Development Drosophila genus Enhancers Epilepsy F-Box Proteins Figs - dietary Future Genes Genetic Growth Lead MAPK8 gene Mediating Mental Retardation Modeling Modification Molecular Morphology Muscle Cells Nervous system structure Neurodegenerative Disorders Neuromuscular Junction Neurons Parkinson Disease Pathway interactions Peptides Phenotype Play Process Proteins Regulation Role Scaffolding Protein Shapes Signal Transduction Specificity Spinal cord injury Structure Synapses Testing Time Ubiquitin Ubiquitination Work axon guidance cell type fly improved in vivo loss of function mutant nervous system disorder novel synaptic function synaptogenesis ubiquitin ligase ubiquitin-protein ligase

项目摘要

DESCRIPTION (provided by applicant): Ubiquitination is a key regulatory mechanism for synaptic development, signaling, and plasticity. The covalent attachment of the 76 aa peptide ubiquitin to target proteins is a rapid and reversible modification that regulates protein stability, activity and localization. As such, it is a potent mechanism for sculpting the synapse. We have uncovered a ubiquitination complex composed of the E3 ubiquitin ligase Highwire (Hiw) and the F-Box protein DFsn that plays a central role in controlling synaptic growth and function at the Drosophila neuromuscular junction (NMJ). A highly homologous ubiquitination complex has also been identified in the mammalian brain, where it plays a critical role in regulating axon guidance and synaptogenesis. However, the molecular architecture and molecular action of this ubiquitination complex is not well understood. We propose that Hiw and DFsn form a non-SCF ubiquitin complex where Hiw functions as an E3 ligase and a scaffolding protein to facilitate multi-subunit interaction, and the combination of different co-factors and ubiquitin substrates confers time- and cell type-specific regulation of neuronal functions. Thus identifying other components and novel ubiquitin targets of this ubiquitination complex is key to understanding how the hiw-mediated ubiquitin pathway specifically regulates synaptic development. We have taken two independent approaches to address this question. Biochemically, we identified Hiw/DFsn interacting proteins through tandem affinity purification using fly brains that express affinity-tagged Hiw and DFsn proteins, respectively. Studying the role of two of the Hiw- binding proteins, NSF and Rae1, in synaptic development (aim1), and how they work together with Hiw and DFsn to modulate the ubiquitin ligase activity (aim2) will define an essential ubiquitination machinery that controls synaptic growth. Genetically, we identified 5 hiw enhancer complementation groups through a hiw enhancer screen. Studying these genetic hiw interactors will allow us to identify other molecular pathways that work together with the Hiw/DFsn ubiquitin pathway to shape the structure and strength of synaptic connections formed during development (aim3).

描述（由申请人提供）：泛素化是突触发育，信号传导和可塑性的关键调节机制。 76 AA肽泛素对靶蛋白的共价附着是一种快速且可逆的修饰，可调节蛋白质稳定性，活性和定位。因此，它是雕刻突触的有效机制。我们发现了由E3泛素连接酶Highwire（HIW）和F-box蛋白DFSN组成的泛素化复合物，该蛋白DFSN在控制果蝇神经肌肉肌肉发达（NMJ）的突触生长和功能方面起着核心作用。在哺乳动物大脑中也发现了高度同源的泛素化复合物，在调节轴突引导和突触发生中起着关键作用。但是，这种泛素化复合物的分子结构和分子作用尚不清楚。我们建议HIW和DFSN形成一种非SCF泛素复合蛋白复合蛋白，其中HIW充当E3连接酶和脚手架蛋白，可促进多生成相互作用，并且不同的co剂因子和泛素底物的组合赋予了时间和细胞类型的神经元功能的特异性调节。因此，识别这种泛素化复合物的其他成分和新型的泛素靶标是了解HIW介导的泛素途径如何专门调节突触发育的关键。我们采用了两种独立的方法来解决这个问题。从生物化学上讲，我们通过使用串联亲和力纯化鉴定了HIW/DFSN相互作用的蛋白质，该蛋白质分别使用蝇脑纯化，分别表达获得亲和力标记的HIW和DFSN蛋白。研究两种HIW结合蛋白NSF和RAE1在突触发育中的作用（AIM1），以及它们如何与HIW和DFSN一起调节泛素连接酶活性（AIM2）将定义一种控制突触生长的基本泛素化机制。从遗传上讲，我们通过HIW增强子屏幕确定了5个HIW增强子互补组。研究这些遗传HIW相互作用的人将使我们能够鉴定出与HIW/DFSN泛素途径一起使用的其他分子途径，以塑造开发过程中形成的突触连接的结构和强度（AIM3）。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A Cullin1-based SCF E3 ubiquitin ligase targets the InR/PI3K/TOR pathway to regulate neuronal pruning.

DOI：
10.1371/journal.pbio.1001657
发表时间：
2013-09
期刊：
PLoS biology
影响因子：
9.8
作者：
Wong JJ;Li S;Lim EK;Wang Y;Wang C;Zhang H;Kirilly D;Wu C;Liou YC;Wang H;Yu F
通讯作者：
Yu F

Size matters: formation and function of giant synapses.

大小很重要：巨型突触的形成和功能。