The activity-dependent regulation of Argonaute 2 function in neurons by PICK1.

PICK1 对神经元中 Argonaute 2 功能的活动依赖性调节。

• 批准号: BB/L021307/1
• 负责人: Jonathan Hanley
• 金额: $ 48.45万
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2015
• 资助国家: 英国
• 起止时间: 2015 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FL021307%2F1
• 关键词: activity; dependent; regulation; Argonaute; function

The aim of this research is to investigate a mechanism for how nerve cells in the brain control long-term changes in their structure and function in response to communication from other nerve cells.Nerve cells (neurons) in the brain communicate with one another at connections called synapses, which are located in small protrusions on the neuronal surface called dendritic spines. A chemical (neurotransmitter) is released from a neuron and travels across the synapse to activate receptors in the adjacent neuron. Synapses can change their strength (known as "synaptic plasticity") by altering the number of receptors found on the surface of the neuron in the synapse, and also by changing the size and shape of the dendritic spine that houses the synapse. This process is thought to underlie learning and memory, because the memory is likely to be stored in a circuit of interconnected neurons.In order to retain long-term memories, neurons need to synthesize additional protein components that are important for maintaining the changes in spine structure, or the changes in receptor number at the synapse. Proteins are made by translating genetic information encoded in DNA sequences (genes). An intermediate between DNA and protein is called messenger RNA (mRNA), and neurons can transport mRNA to the parts of the neuron close to synapses and locally control the synthesis of a particular protein that is important for those synapses at a particular time. Another type of molecule, called micro RNA (miRNA) can bind to mRNA and stop the translation of mRNA into protein. This process is very precisely regulated.We have found that a protein (called PICK1), which is known to be involved in synaptic plasticity over the timescale of hours, interacts with another protein (called Argonaute2), which is an important component of the cell machinery that promotes the association of miRNA with mRNA to block protein synthesis. Preliminary experiments suggest that by interacting with Argonaute2, PICK1 can relieve this block of protein synthesis and increase the production of specific proteins. Our main hypothesis is that PICK1 plays an important role in regulating protein synthesis close to active synapses via its interaction with Argonaute2.We aim to test this hypothesis by analysing the precise mechanism for how PICK1 regulates the function of Argonaute2. We will use established methods for analysing protein synthesis while manipulating the PICK1-Argonaute2 interaction under different conditions of synaptic activity. Our preliminary results suggest that PICK1 anchors Argonaute2 to membrane-bound structures inside the neuron called endosomes, which are involved in trafficking important receptors to the synapse during synaptic plasticity. One of our hypotheses is that these trafficking events are linked to the regulation of Argonaute2 by PICK1. One important function of miRNAs is controlling the local synthesis of proteins that determine the size or shape of dendritic spines, which is an important factor that correlates with the strength of a synaptic connection. We will investigate whether PICK1 is involved in local protein synthesis in dendrites and consequent regulation of dendritic spine size via its interaction with Argonaute2.Most of our experiments will be carried out using neurons obtained from the rat brain. These neurons can be isolated from the brain and then kept 'alive' in a petri dish. Using these cells we will be able to understand more about the mechanisms that regulate the local control of protein synthesis in neurons in response to synaptic activity, and hence further our knowledge of the mechanisms that underlie long-term memory.

本研究的目的是探讨大脑中的神经细胞如何通过与其他神经细胞的交流来控制自身结构和功能的长期变化。大脑中的神经细胞（神经元）通过位于神经元表面称为树突棘的小突起中的称为突触的连接来相互交流。一种化学物质（神经递质）从神经元中释放出来，穿过突触激活相邻神经元中的受体。突触可以改变其强度（称为“突触可塑性”），方法是改变突触中神经元表面的受体数量，以及改变容纳突触的树突棘的大小和形状。这一过程被认为是学习和记忆的基础，因为记忆很可能储存在一个由相互连接的神经元组成的回路中。为了保持长期记忆，神经元需要合成额外的蛋白质成分，这些蛋白质成分对维持脊柱结构的变化或突触受体数量的变化至关重要。蛋白质是通过翻译DNA序列（基因）中编码的遗传信息而产生的。DNA和蛋白质之间的中间体称为信使RNA（mRNA），神经元可以将mRNA运输到靠近突触的神经元部分，并在特定时间局部控制对这些突触重要的特定蛋白质的合成。另一种类型的分子，称为微RNA（miRNA），可以与mRNA结合并阻止mRNA翻译成蛋白质。我们发现一种蛋白质（PICK 1）与另一种蛋白质（Argonaute 2）相互作用，这种蛋白质被称为PICK 1，已知它在数小时的时间尺度内参与突触可塑性，Argonaute 2是促进miRNA与mRNA结合以阻止蛋白质合成的细胞机制的重要组成部分。初步实验表明，通过与Argonaute 2相互作用，PICK 1可以缓解蛋白质合成的这种阻滞，并增加特定蛋白质的产量。我们的主要假设是PICK 1通过与Argonaute 2的相互作用在调节靠近活跃突触的蛋白质合成中起重要作用。我们的目的是通过分析PICK 1如何调节Argonaute 2功能的精确机制来验证这一假设。我们将使用已建立的方法来分析蛋白质合成，同时在不同的突触活性条件下操纵PICK 1-Argonaute 2相互作用。我们的初步结果表明，PICK 1锚Argonaute 2的膜结合结构内的神经元称为内体，这是参与运输重要的受体突触突触在突触可塑性。我们的假设之一是，这些贩运事件与PICK 1对Argonaute 2的调节有关。miRNA的一个重要功能是控制蛋白质的局部合成，这些蛋白质决定树突棘的大小或形状，这是与突触连接强度相关的重要因素。我们将研究PICK 1是否参与树突局部蛋白质的合成，以及通过与Argonaute 2的相互作用来调节树突棘的大小。这些神经元可以从大脑中分离出来，然后在培养皿中保持“活力”。利用这些细胞，我们将能够更多地了解神经元中响应突触活动的蛋白质合成的局部控制机制，从而进一步了解长期记忆的基础机制。