Role of AF-6 nuclear signaling in regulating dendritic spine morphology

AF-6核信号在调节树突棘形态中的作用

• 批准号: 7912348
• 负责人: JON-ERIC VANLEEUWEN
• 金额: $ 3.3万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7912348
• 关键词: Adhesions; Brain; Brain Diseases; Cell Nucleus; Data; Dendritic Spines; Disease; Estrogens; Fostering; Goals; Health; MLLT4 gene; Mediating; Morphogenesis; Morphology; Mutate; N-Cadherin; N-Methyl-D-Aspartate Receptors; Neurodegenerative Disorders; Neurologic; Neuronal Plasticity; Neurons; Nuclear; Nuclear Localization Signal; Pathway interactions; Process; Proteins; Role; Signal Pathway; Signal Transduction; Structure; Synapses; Synaptic plasticity; Techniques; Time; Vertebral column; Western Blotting; brain cell; immunocytochemistry; insight; neurodevelopment; overexpression; prevent; public health relevance; response

DESCRIPTION (provided by applicant): The majority of excitatory neuronal input occurs at dendritic spines; remodeling of these structures leads to concurrent changes in neuronal function essential for normal neural development and processing. Furthermore, dysregulation of this phenomenon underlies numerous neurological and neurodegenerative disorders. In addition to signaling at synapses, signaling pathways to the nucleus are thought to be important for changes in dendritic spine morphology and neuronal plasticity. However, the signal transduction networks that regulate spine modulation are not well understood. AF-6 is a PDZ-domain-containing protein abundantly expressed at adhesion junctions and is involved in dendritic spine remodeling. Pathways upstream of AF-6, including NMDA receptor (NMDAR), estrogen (E2) and N-cadherin-dependent signaling, have been associated with modulating dendritic spine morphology via an AF-6 dependent pathway. In our preliminary data, we demonsrate that these pathways can also induce AF-6 translocation to the nucleus. Thus AF-6 is an ideal candidate to mediate dendritic spine morphogenesis by signaling in distinct subcellular compartments. The aim of this proposal is to determine the role of differential AF-6 translocation in regulating dendritic spine morphogenesis in cultured cortical neurons. First, we will characterize the time-dependent localization of AF-6 in NMDAR, E2 and N-cadherin-mediated signaling by using immunocytochemistry and Western-blotting techniques. We will also assess the role of the various AF-6 domains in its translocation by overexpressing truncated or mutated AF-6 constructs. Next, we will use immunocytochemistry to determine changes in spine expression induced by NMDAR, E2 and N-cadherin-mediated signaling or the overexpression of altered AF-6 constructs. Lastly, we will intervene with AF-6 translocation and then assess alterations in spine morphology. To this end, we will prevent AF-6 from localizing to synapses or the nucleus using specifically modified AF-6 constructs and we will force AF-6 into the nucleus using a nuclear localization signal. Results from these studies will foster a better understanding of the signaling pathways regulating dendritic spine morphogenesis and provide insight into the mechanisms governing synaptic plasticity in health and disease. PUBLIC HEALTH RELEVANCE: The goal of this project is to understand how brain cells change in response to signals from other brain cells. These changes are essential for normal brain function but also become disrupted in various brain disorders. Findings from these studies will help us to better understand the cause of certain brain disorders and potentially identify new forms of treatment.

描述（由申请人提供）：大部分兴奋性神经元输入发生在树突棘；这些结构的重塑会导致神经元功能的同时变化，这对于正常的神经发育和处理至关重要。此外，这种现象的失调是许多神经系统和神经退行性疾病的基础。除了突触的信号传导之外，通往细胞核的信号传导途径被认为对于树突棘形态和神经元可塑性的变化也很重要。然而，调节脊柱调节的信号转导网络尚不清楚。 AF-6 是一种含有 PDZ 结构域的蛋白质，在粘附连接处大量表达，参与树突棘重塑。 AF-6 上游通路，包括 NMDA 受体 (NMDAR)、雌激素 (E2) 和 N-钙粘蛋白依赖性信号传导，已通过 AF-6 依赖性通路与调节树突棘形态相关。在我们的初步数据中，我们证明这些途径也可以诱导 AF-6 易位至细胞核。因此，AF-6 是通过不同亚细胞区室中的信号传导介导树突棘形态发生的理想候选者。该提案的目的是确定差异 AF-6 易位在调节培养的皮层神经元树突棘形态发生中的作用。首先，我们将通过使用免疫细胞化学和蛋白质印迹技术来表征 AF-6 在 NMDAR、E2 和 N-钙粘蛋白介导的信号传导中的时间依赖性定位。我们还将通过过表达截短或突变的 AF-6 构建体来评估各种 AF-6 结构域在其易位中的作用。接下来，我们将使用免疫细胞化学来确定 NMDAR、E2 和 N-钙粘蛋白介导的信号传导或改变的 AF-6 构建体的过表达诱导的脊柱表达变化。最后，我们将干预 AF-6 易位，然后评估脊柱形态的变化。为此，我们将使用专门修改的 AF-6 结构来防止 AF-6 定位到突触或细胞核，并且我们将使用核定位信号迫使 AF-6 进入细胞核。这些研究的结果将促进人们更好地了解调节树突棘形态发生的信号通路，并深入了解健康和疾病中控制突触可塑性的机制。 公共健康相关性：该项目的目标是了解脑细胞如何响应来自其他脑细胞的信号而发生变化。这些变化对于正常的大脑功能至关重要，但也会在各种大脑疾病中受到干扰。这些研究的结果将帮助我们更好地了解某些大脑疾病的原因，并有可能找到新的治疗方法。