Mechanisms of mRNA localization to neuronal synapses

描述(申请人提供)：学习和记忆的最佳生物学相关性是突触的可塑性，即突触的数量和强度随经验的变化。长期突触可塑性是一个依赖转录和翻译的过程，可以限制在单个神经元内的突触亚群。我们认为，将基因表达限制在单个突触的一种机制是通过mRNA定位和调节翻译。海兔模型系统为研究突触可塑性提供了一个很好的模型系统。马丁实验室以前的工作已经证明，在培养的海兔感觉运动神经元与学习相关的可塑性过程中，蛋白质的合成可以在空间上限于受刺激的突触(Wang等人，2009年)。在这项提议中，我将重点放在编码海兔感觉细胞特异性神经肽Sensorin的突触定位的潜在机制上。对神经元和其他不对称细胞类型的研究表明，定位的转录本通常在其非翻译区包含顺式作用的定位元件 (UTRs)，通常由茎环结构编码。Martin实验室已经证明，Sensorin的5‘和3个非翻译区(UTRs)足以用于报告RNA的突触定位。远端轴突定位需要3‘非编码区，突触定位需要5’编码区。我已经在感觉蛋白mRNA的5‘非编码区中确定了一个66核苷酸序列，当与感觉蛋白3’非编码区配对时，它是必需的，并且足以进行感觉蛋白mRNA的突触定位。我的实验进一步表明，该定位元件由茎环结构编码(Meer等人，2012年)。顺式作用的RNA元件与反式作用的RNA结合蛋白相互作用，介导RNA定位。除了鉴定感觉蛋白3‘非编码区中的神经性RNA定位元件外，我还建议进行实验，以确定顺式作用元件如何在感觉蛋白的RNA稳定性和翻译中发挥作用。最后，我将确定RN结合蛋白，将感觉蛋白mRNA定位于神经突起和突触，并研究这些蛋白在突触形成过程中的动态。拟议的实验结果将为神经元通过mRNA定位和受调控的翻译在空间上限制基因表达的细胞和分子生物学机制提供深入的见解。它们对一系列长期突触可塑性受损的大脑疾病具有重要意义，包括自闭症、精神发育迟滞、焦虑症和毒瘾。 与公共健康相关：学习和记忆的能力取决于大脑中神经细胞之间的连接能力，这种能力随着经验的变化而变化。了解大脑回路这种经验依赖型变化的潜在机制，对于开发有效的药物和治疗各种认知障碍的方法至关重要，这些认知障碍包括智力低下、神经精神障碍和与年龄相关的记忆丧失。

DESCRIPTION (provided by applicant): The best biological correlate of learning and memory is synaptic plasticity, or changes in the number and strength of synapses with experience. Long-term synaptic plasticity is a transcription- and translation-dependent process that can be restricted to subsets of synapses within a single neuron. We propose that one mechanism for restricting gene expression to individual synapses is through mRNA localization and regulated translation. The model system Aplysia californica provides an excellent model system to study synaptic plasticity. Previous work in the Martin lab has demonstrated that protein synthesis can be spatially restricted to stimulated synapses during learning- related plasticity of cultured Aplysia sensory-motor neurons (Wang et al., 2009). In this proposal, I focus on the mechanisms underlying the synaptic localization of the mRNA encoding an Aplysia sensory cell-specific neuropeptide, sensorin. Studies in neurons and other asymmetric cell types have shown that localized transcripts often contain cis- acting localization elements in their untranslated regions (UTRs), which often are encoded by stem-loop structures. The Martin lab has demonstrated that the 5' and 3 untranslated regions (UTRs) of sensorin are sufficient for synaptic localization of reporter RNA. While the 3'UTR is required for distal neurite localization, the 5'UTR is required for synaptic localization. I have identified a 66 nucleotide sequence in the 5'UTR of sensorin mRNA that when paired with the sensorin 3'UTR is required and sufficient for sensorin mRNA synaptic localization. My experiments further indicate that this localization element is encoded by a stem-loop structure (Meer et al., 2012). Cis-acting RNA elements interact with trans-acting RNA binding proteins to mediate RNA localization. In addition to characterizing the neuritic RNA localization element in the sensorin 3'UTR, I propose experiments to determine how cis-acting elements function in the RNA stability and translation of sensorin. Finally, I will identify the RN binding proteins that localize sensorin mRNA to neurites and synapses and study the dynamics of these proteins during synapse formation. The results of the proposed experiments will provide insight into the cell and molecular biological mechanisms by which neurons are able to spatially restrict gene expression through mRNA localization and regulated translation. They are of significance to a range of brain disorders in which long-term synaptic plasticity is impaired, including autism, mental retardation, anxiety disorders and drug addiction. PUBLIC HEALTH RELEVANCE: The ability to learn and remember depends on the capacity of connections between nerve cells in the brain to change with experience. Knowledge of the underlying mechanisms that drive such experience-dependent changes in brain circuitry is critical to the development of effective drugs and therapies for a wide range of cognitive disorders, including mental retardation, neuropsychiatric disorders and age-related memory loss.