Mechanisms of Translation in the CNS

描述（由申请人提供）：本提案的主要目的是描述哺乳动物神经元，特别是树突中mRNA翻译的机制，这些机制改变了突触功效。CPEB是一种序列特异性mRNA结合蛋白，其促进细胞质聚腺苷酸化诱导的翻译，存在于哺乳动物神经元的突触树突。CPEB基因敲除小鼠在突触可塑性和学习记忆方面表现出缺陷，表明细胞质多聚腺苷酸化机制在复杂的脑功能中的重要性。CPEB使mRNA上的一组因子成核以促进多聚腺苷酸化，包括非典型的多聚（A）聚合酶Gld 2、去腺苷酸化酶PARN、eIF 4 E结合蛋白神经导向蛋白（Ngd）、支架蛋白symplekin等。这些蛋白质存在于哺乳动物神经元树突中的复合物中，在那里它们调节多聚腺苷酸化和几种mRNA的翻译。其中两个因子Gld 2和Ngd调节海马神经元的突触可塑性，但方向相反; Gld 2缺失可诱导长时程增强（LTP）功能的缺失，而Ngd缺失则可增强LTP功能。此外，Gld 2缺失可减少树突的翻译，而Ngd缺失可刺激LTP功能。这些数据表明，CPEB、Gld 2、和Ngd形成树突中翻译控制的连贯分子基础，其反过来调节突触功效。第一个具体目标的目标是鉴定可能修饰poly（A）长度和翻译变化的去腺苷酸化酶，以及评估它们是否影响突触功能。目的2是研究CPEB在脑中结合的mRNA的完整性，并确定它们是否在树突中进行活性依赖性聚腺苷酸化和翻译。目的3的目标是开发和使用一种新的深度测序方法来鉴定树突状mRNA，这些mRNA在体外响应于LTP诱导和体内学习而经历细胞质聚腺苷酸化和翻译。这些实验将增强我们对神经元中的局部mRNA翻译如何介导突触功能的理解，这对高级脑功能和神经病如自闭症，阿尔茨海默病，帕金森病等具有重要意义。

DESCRIPTION (provided by applicant): The broad objective of this proposal is to delineate mechanisms of mRNA translation in mammalian neurons and especially dendrites that modify synaptic efficacy. CPEB, a sequence-specific mRNA binding protein that promotes cytoplasmic polyadenylation- induced translation, is present at synapto-dendrites of mammalian neurons. CPEB knockout mice display defects in synaptic plasticity and learning and memory, indicating the importance of the cytoplasmic polyadenylation machinery in complex brain function. CPEB nucleates a set of factors on mRNA to promote polyadenylation including the non- canonical poly(A) polymerase Gld2, the deadenylating enzyme PARN, the eIF4E-binding protein neuroguidin (Ngd), the scaffold protein symplekin, and others. These proteins reside in a complex in dendrites of mammalian neurons where they modulate the polyadenylation and translation of several mRNAs. Two of these factors, Gld2 and Ngd, regulate synaptic plasticity in hippocampal neurons but do so in opposite directions; Gld2 depletion induces a deficit in long-term potentiation (LTP) while Ngd depletion enhances it. Moreover, Gld2 depletion reduces translation in dendrites while Ngd depletion stimulates it. These data indicate that the interplay among CPEB, Gld2, and Ngd form a coherent molecular foundation of translation control in dendrites that in turn modulates synaptic efficacy. The goals of the first specific aim are to identify deadenylating enzymes that are likely to modify poly(A) length and changes in translation as well as to assess whether they influence synaptic function. Aim 2 is to investigate the full panoply of mRNAs that are bound by CPEB in the brain and determine whether they undergo activity-dependent polyadenylation and translation in dendrites. The goal of aim 3 is to develop and use a new deep sequencing method to identify dendritic mRNAs that undergo cytoplasmic polyadenylation and translation in response to in LTP induction in vitro and learning in vivo. These experiments will enhance our understanding of how local mRNA translation in neurons mediates synapse function, which has important implications for higher brain function and neuropathies such as autism, Alzheimer's Disease, Parkinson's Disease, and others.