The Role of Tropomodulin in Dendritic Spine Development and Synapse Formation

原调节蛋白在树突棘发育和突触形成中的作用

• 批准号: 8912202
• 负责人: Omotola Folashade Omotade
• 金额: $ 4.31万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8912202
• 关键词: Actin-Binding Protein; Actins; Address; Adult; Affect; Alzheimer' s Disease; Autistic Disorder; Automobile Driving; Behavior; Behavioral; Brain; Cells; Communication; Complex; Cytoskeleton; Data; Defect; Dendritic Spines; Development; Disease; Electrophysiology (science); Excitatory Synapse; Exhibits; Filament; Filopodia; Fragile X Syndrome; Genetic; Goals; Hippocampus (Brain); Immunofluorescence Immunologic; Immunofluorescence Microscopy; Intellectual functioning disability; Knock-out; Life; Maintenance; Measures; Mediating; Mental disorders; Microfilaments; Minus End of the Actin Filament; Modification; Morphology; Mus; Mutant Strains Mice; Mutate; Nervous system structure; Neuraxis; Neurons; Pathway interactions; Phenotype; Physiological; Play; Process; Protein Family; Protein Isoforms; Proteins; Regulation; Role; Schizophrenia; Structure; Surface; Synapses; Synaptic Transmission; Time; Tissues; Vertebral column; Vertebrates; Work; base; cellular imaging; density; depolymerization; immunocytochemistry; insight; monomer; mutant; nervous system development; nervous system disorder; postnatal; postsynaptic; presynaptic; protein expression; public health relevance; research study; small hairpin RNA; spatiotemporal; synaptic function; synaptogenesis; tropomodulin

 DESCRIPTION (provided by applicant): The proper establishment of synapses during development and their maintenance throughout adulthood is critical for complex brain functions. Synaptic dysregulation is associated with many neurological diseases such as schizophrenia, Fragile X and Alzheimer's. An understanding of how synapses are altered in disease states will first require an understanding of the molecules and pathways responsible for the establishment of synapses in the central nervous system during brain development. In vertebrates, the majority of excitatory synapses are formed on dendritic spines, tiny, actin-rich structures that protrude from the dendritic surface and act as the platform for presynaptic input. The dynamic remodeling of the actin cytoskeleton drives dendritic spine development and synapse formation. However, the cellular mechanisms responsible for actin organization and remodeling during postsynaptic development are not fully understood. Tropomodulins (Tmods) are a multi-domain family of proteins that cap the minus end of actin filaments; thereby blocking monomer exchange at the minus ends, inhibiting depolymerization, and stabilizing actin-based structures. Tmods are highly expressed in the central nervous system and genetic knock-out of Tmod2 in mice leads to behavioral and synaptic defects. However, the mode of action and underlying cellular mechanisms responsible for such phenotypes are unknown. Likewise, the isoform specific functions as well as the actin-regulatory domains of Tmod responsible for proper neuronal function have not been investigated. Thus, if and how Tmods play a role in actin remodeling during dendritic spine development and synapse formation remain unknown. Our preliminary data reveal that Tmods are expressed in both developing and mature dendritic spines, where in mature spines, Tmods concentrate in a region that contains relatively stable actin filaments. We find that Tmod protein levels increase in the hippocampus during postnatal development, a time when extensive synaptogenesis and refinement occurs. We hypothesize that minus end capping of actin filaments by Tmod is essential for the development and stability of dendritic spines during postsynaptic development, thereby allowing the structural modification of dendritic spines that occurs during synapse formation. In Specific Aim 1, we will knockdown Tmods in developing primary hippocampal neurons and use immunocytochemistry, live cell imaging, and electrophysiology to determine the function of Tmods during spinogenesis and synapse formation. In Specific Aim 2, we will mutate the Tmod minus end capping domain in order to investigate the role of Tmod minus end capping during postsynaptic development. By understanding the mechanisms by which Tmod minus end capping regulates the actin cytoskeleton within dendritic spines, this work will aid in our long term goal of understanding the mechanisms by which excitatory synapses are established, maintained and dysregulated in disease states.

 描述（由申请人提供）：在发育过程中突触的正确建立及其在整个成年期的维持对于复杂的大脑功能至关重要。突触失调与许多神经系统疾病有关，例如精神分裂症、脆性 X 肌瘤和阿尔茨海默病。要了解疾病状态下突触如何改变，首先需要了解大脑发育过程中中枢神经系统中负责建立突触的分子和途径。在脊椎动物中，大多数兴奋性突触形成于树突棘上，树突棘是一种微小的富含肌动蛋白的结构，从树突表面突出，充当突触前输入的平台。肌动蛋白细胞骨架的动态重塑驱动树突棘发育和突触形成。然而，突触后发育过程中负责肌动蛋白组织和重塑的细胞机制尚未完全了解。原调节蛋白 (Tmod) 是一个多结构域蛋白质家族，覆盖肌动蛋白丝的负端；从而阻断负端的单体交换，抑制解聚，并稳定基于肌动蛋白的结构。 Tmods 在中枢神经系统中高度表达，小鼠中 Tmod2 基因敲除会导致行为和突触缺陷。然而，导致这种表型的作用模式和潜在细胞机制尚不清楚。同样，尚未研究负责适当神经元功能的 Tmod 的亚型特异性功能以及肌动蛋白调节域。因此，Tmod 是否以及如何在树突棘发育和突触形成过程中的肌动蛋白重塑中发挥作用仍然未知。我们的初步数据显示，Tmod 在发育中和成熟的树突棘中都有表达，其中在成熟的树突棘中，Tmod 集中在包含相对稳定的肌动蛋白丝的区域。我们发现，在出生后发育过程中，海马体中的 Tmod 蛋白水平会增加，此时突触发生和细化会发生广泛的变化。我们假设 Tmod 对肌动蛋白丝的负端加帽对于突触后发育过程中树突棘的发育和稳定性至关重要，从而允许在突触形成过程中发生树突棘的结构修饰。在具体目标 1 中，我们将敲低初级海马神经元发育过程中的 Tmod，并使用免疫细胞化学、活细胞成像和电生理学来确定 Tmod 在棘发生和突触形成过程中的功能。在具体目标 2 中，我们将突变 Tmod 负封端结构域，以研究 Tmod 负封端在突触后发育过程中的作用。通过了解 Tmod 减端帽调节树突棘内肌动蛋白细胞骨架的机制，这项工作将有助于我们了解 在疾病状态下兴奋性突触建立、维持和失调的机制。