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可塑性，或突触数量和强度随经验的变化。长时程突触可塑性是一个转录和突触传递依赖的过程，可以限制在单个神经元内的突触子集。我们提出将基因表达限制在单个突触上的一种机制是通过mRNA定位和调节翻译。该模型系统为研究突触可塑性提供了一个很好的模型系统。Martin实验室的先前工作已经证明，在培养的Aussia感觉运动神经元的学习相关可塑性期间，蛋白质合成可以在空间上限制于受刺激的突触（Wang et al.，2009年）。在这个建议中，我专注于潜在的机制，突触定位的mRNA编码的感觉细胞特异性神经肽，sensorin。对神经元和其他不对称细胞类型的研究表明，定位转录本的非翻译区通常含有顺式作用定位元件 UTR通常由茎环结构编码。Martin实验室已经证明传感蛋白的5'和3个非翻译区（UTR）足以用于报告RNA的突触定位。虽然远端神经突定位需要3'UTR，但突触定位需要5'UTR。我已经鉴定了传感蛋白mRNA的5'UTR中的66个核苷酸的序列，当与传感蛋白3'UTR配对时，该序列是传感蛋白mRNA突触定位所需的并且足够。我的实验进一步表明该定位元件由茎环结构编码（梅尔et al.，2012年）。顺式作用RNA元件与反式作用RNA结合蛋白相互作用以介导RNA定位。除了表征神经炎RNA定位元件在sensorin 3'UTR，我建议实验，以确定顺式作用元件如何在RNA的稳定性和翻译的sensorin。最后，我将确定RN结合蛋白，本地化sensorin mRNA的神经突和突触，并研究这些蛋白质在突触形成的动力学。所提出的实验的结果将提供深入了解的细胞和分子生物学机制，神经元能够通过mRNA定位和调节翻译空间限制基因表达。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、焦虑症和药物成瘾。

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.