The role of a novel viral-like signalling pathway in synaptic plasticity and neurological disorders

我们发现了一种与细胞外小泡(EV)相关的新的病毒样信号通路。我们 发现果蝇ARC(肌动蛋白调节细胞骨架相关蛋白)(Darc1)的同源物，是 在EVS中以信使核糖核酸和蛋白质形式存在。ARC是神经突触可塑性的主要调节器 它是哺乳动物的系统，对学习和记忆至关重要。DArc1包含一个类似于 逆转录病毒/反转录转座子Gag样蛋白，多聚体形成包装病毒RNA的衣壳。我们的 研究表明，darc1形成衣壳，与自己的rna结合，然后运输darc1转录本 穿过突触。DArc1的转移是依赖活性的可塑性所必需的。 神经肌肉接头(NMJ)。除了dArc1，目前还不清楚这种病毒样中是否有其他基因 路径。我们在目标1中解决了这种不确定性。在这里，我们描述了我们识别其他类似恶作剧的计划 EVS中的蛋白质，我们已经发现了EVS中富含的另一种Gag蛋白质，它由 反转录转座子Copia。我们已经发现，Copia通过突触环转移。当Copia是 在NMJ上被击倒，这显着地导致了可塑性的增加。这是darc1的反义词，其中 我们报告了可塑性的下降。在目标2中，我们将重点放在哪些货物与dArc1和 科皮亚。我们已经通过免疫共沉淀鉴定了与dArc1相关的mRNAs和蛋白质 还有科皮亚。关于Arc的转移是如何发生的，我们发现dArc1的3‘非翻译区(UTR)。 是dArc1跨突触突触转移的必要条件和充分条件。我们现在正在测试 DArc1 3‘非编码区指导将dArc1加载到EVS。此外，我们还建议进行实验，以了解 DARC1和COPIA介导突触可塑性。我们用免疫共沉淀dArc1和Copia来鉴定 潜在的相互作用因素，我们将采取候选方法来寻找基因相互作用因素。在前期工作中 我们发现，在NMJ，dArc1是正确的WNT信号通路所必需的。此外，我们观察到 Copia和dArc1与一些相同的蛋白质和mRNAs结合，这表明它们可能是 相互对立，从而潜在地解释了它们在调节可塑性方面似乎相反的角色。 通过这笔赠款，我们将扩大对电动汽车运输和突触可塑性的理解，同时 描述了反转录转座子在神经元通讯中的一种新的生理功能。

We have discovered a novel viral-like signaling pathway associated with extracellular vesicles (EV). We found the Drosophila homolog of ARC (Actin-Regulated Cytoskeleton-associated protein) (darc1), is present in EVs as both an mRNA and protein. ARC is a master regulator of synaptic plasticity in the nervous system of mammals and is crucial for learning and memory. dArc1 bears a domain resembling retroviral/retrotransposon Gag-like proteins that multimerizes into a capsid that packages viral RNA. Our work shows dARC1 forms a capsid, associates with its own RNA, and then transports the darc1 transcript across the synapse. The transfer of dArc1 is needed for activity-dependent plasticity at the fly neuromuscular junction (NMJ). Besides dArc1, it is unknown whether other genes are in this viral-like pathway. We address this uncertainty in Aim 1. Here we describe our plan to identify other Gag-like proteins in EVs, and we have already found another Gag protein enriched in EVs, that is encoded by the retrotransposon Copia. We have found that Copia transfers across the synaptic bouton. When copia is knocked down at the NMJ this strikingly leads to increased plasticity. This is the opposite of darc1, where we reported a decrease in plasticity. In Aim 2 we focus on what cargoes are co-transferring with dArc1 and Copia. We have identified through co-immunoprecipitation, mRNAs and proteins that associate with dArc1 and Copia. As to how the transfer of Arc occurs, we have found that the dArc1 3’untranslated region (UTR) is necessary and sufficient for the transfer of dArc1 across synaptic boutons. We are now testing if the dArc1 3’UTR directs the loading of dArc1 into EVs. As well, we propose experiments to understand how darc1 and copia mediate synaptic plasticity. We have co-immunoprecipitated dArc1 and Copia to identify potential interactors, and we will take a candidate approach to find genetic interactors. In preliminary work we found that dArc1 is needed for proper WNT pathway signaling at the NMJ. Additionally, we observe that Copia and dArc1 bind to some of the same proteins and mRNAs, suggesting that they may be antagonistic to each other, thus potentially explaining their seemingly opposite roles in mediating plasticity. Through this grant we will expand our understanding of EV trafficking and synaptic plasticity, while describing a novel physiological function of a retrotransposon in neuronal communication.