Role of CPEB in dendritic CaMKlla mRNA transport and translation

CPEB 在树突状 CaMKlla mRNA 转运和翻译中的作用

• 批准号: 7932166
• 负责人: Sharon Ann Swanger
• 金额: $ 3.02万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-12-01 至 2011-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7932166
• 关键词: 3' Untranslated Regions; Address; Alzheimer' s Disease; Autistic Disorder; Binding; Binding Proteins; Biochemical; Biological Assay; Biological Process; CPE-binding protein; Cells; Co-Immunoprecipitations; Complex; D Aspartate; Data; Defect; Dendrites; Development; Diffusion; Disease; Dominant-Negative Mutation; Elements; Event; Fluorescent in Situ Hybridization; Genetic Translation; Germ Lines; Hippocampus (Brain); Image; Immunofluorescence Immunologic; In Situ Hybridization; Knockout Mice; Knowledge; Lead; Learning; Life; Mediating; Memory; Mental Retardation; Messenger RNA; Mission; Modification; Molecular; National Institute of Neurological Disorders and Stroke; Neurologic; Neurons; Pathology; Pattern; Phosphorylation; Poly(A)-specific ribonuclease; Polyadenylation; Polyadenylation Pathway; Polynucleotide Adenylyltransferase; Polyribosomes; Process; Protein Biosynthesis; Proteins; RNA; RNA-Binding Proteins; Regulation; Reporter; Resolution; Role; Scaffolding Protein; Site; Structure; Symptoms; Synapses; Synaptic plasticity; Synaptosomes; System; Testing; Translational Regulation; Translations; Traumatic Brain Injury; Work; Xenopus oocyte; aspartate receptor; calmodulin-dependent protein kinase II; cellular imaging; digital imaging; experience; lentiviral-mediated; mRNA tagging; particle; protein complex; small hairpin RNA; synaptic function

DESCRIPTION (provided by applicant): Dendritic protein synthesis is essential for several forms of long-term synaptic plasticity that are thought to underlie learning and memory. However, the molecular mechanisms that regulate activity-induced dendritic translation remain unclear. One hypothesis is that activity may regulate mRNA binding proteins (RBP) that control the dendritic localization and translation of specific mRNAs. This proposal investigates the role of the RBP cytoplasmic polyadenylation element binding protein 1 (CPEB1) in dendritic mRNA targeting and local translation. The overall hypothesis is that CPEB1 associates with a multi-protein dendritic complex that regulates activity-induced transport and translation of known CPEB1 target mRNA CaMKIIa. The first aim of this proposal examines 1) dendritic localization of the multi-protein complex in CPEB1 null neurons compared to wild type and 2) the molecular mechanisms mediating activity regulation of the CPEB1 complex at synapses. The two proposed approaches for this aim are high-resolution quantitative immunofluorescence in cultured hippocampal neurons and biochemical analysis of synaptic fractions. The second aim addresses whether the CPEB1 complex regulates basal and activity-induced dendritic localization of CaMKIIa mRNA. CaMKIIa mRNA localization will be assayed in CPEB1 null neurons and following lentiviral-mediated knockdown of the CPEB1 complex proteins using in situ hybridization and live imaging of a tagged mRNA construct. The third aim evaluates whether the CPEB1 complex regulates dendritic CamKlla mRNA translation using live-cell imaging of a fluorescent mRNA reporter combined with lentiviral-mediated manipulation of the CPEB1 complex components. Consistent with the mission of NINDS to "pursue an understanding of the normal activities" of neurons, this proposal examines a potential mechanism by which activity might regulate the local synthesis of new proteins. This work contributes to the basic knowledge that founds the development of useful therapies for disorders of synaptic function. Lay summary: Deficits in learning and memory are symptoms of varied neurological conditions such as mental retardation, autism, traumatic brain injury and Alzheimer's disease. The proposed study investigates the regulation of experience-dependent modification of neuronal connections - the biological process that is thought to underlie learning and memory. Acquiring this basic knowledge is critical for understanding the pathology of these wide-spread disorders as well as the establishment of useful therapies.

描述（由申请人提供）：树突状蛋白的合成对于被认为是学习和记忆基础的几种形式的长期突触可塑性是必不可少的。然而，调节活性诱导的树突翻译的分子机制仍不清楚。一种假设是活性可以调节控制树突定位和特定mRNA翻译的mRNA结合蛋白（RBP）。本研究旨在探讨RBP胞质多聚腺苷酸化元件结合蛋白1（CPEB 1）在树突状mRNA靶向和局部翻译中的作用。总体假设是CPEB 1与多蛋白树突状复合物相关，该复合物调节已知CPEB 1靶mRNA CaMKIIa的活性诱导的转运和翻译。本提案的第一个目的是检查1）与野生型相比，CPEB 1无效神经元中多蛋白复合物的树突定位，以及2）介导CPEB 1复合物在突触处的活性调节的分子机制。为此目的提出的两种方法是培养海马神经元的高分辨率定量免疫荧光和突触组分的生化分析。第二个目的是解决CPEB 1复合物是否调节CaMKIIa mRNA的基础和活性诱导的树突状定位。将在CPEB 1缺失神经元中以及在慢病毒介导的CPEB 1复合物蛋白敲低后，使用标记的mRNA构建体的原位杂交和实时成像测定CaMKIIa mRNA定位。第三个目标评估是否CPEB 1复合物调节树突状CamKlla mRNA翻译使用活细胞成像的荧光mRNA报告相结合的慢病毒介导的操纵的CPEB 1复合物的组件。与NINDS的使命“追求对神经元正常活动的理解”相一致，该提议研究了一种潜在的机制，通过该机制，活动可能调节新蛋白质的局部合成。这项工作有助于基础知识的发展，建立有用的治疗疾病的突触功能。敷设总结：学习和记忆缺陷是各种神经系统疾病的症状，如精神发育迟滞、自闭症、创伤性脑损伤和阿尔茨海默病。这项拟议中的研究调查了神经元连接的经验依赖性修饰的调节-这一生物学过程被认为是学习和记忆的基础。获得这些基本知识对于理解这些广泛传播的疾病的病理学以及建立有用的治疗方法至关重要。