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PICK1 and cortactin as antagonistic regulators of Arp2/3-mediated actin polymerisation in GluA2-dependent AMPA receptor trafficking.

PICK1 和 cortactin 作为 GluA2 依赖性 AMPA 受体运输中 Arp2/3 介导的肌动蛋白聚合的拮抗调节剂。

基本信息

批准号：
BB/L007266/1
负责人：
Jonathan Hanley
金额：
$ 67.03万
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：
2014
资助国家：
英国
起止时间：
2014 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FL007266%2F1
关键词：
PICK1 cortactin antagonistic regulators Arp2

项目摘要

The aim of this research is to study specific aspects of how nerve cells in the brain communicate with each other, and how this communication can change during learning and also as a result of certain kinds of brain disorder. Nerve cells (neurons) in the brain communicate with one another at connections called synapses. 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. This process is thought to underlie learning and memory, because the memory is likely to be stored in a circuit of interconnected neurons.AMPA receptors are the neurotransmitter receptors involved in most synaptic excitation in the brain, and they are made up of distinct protein components, different combinations of which govern how the receptors work. The presence of a component called GluA2 is important for two reasons. First, a number of other proteins physically interact with GluA2 in order to move AMPA receptors to or from the synapse. Second, it influences the entry of calcium ions into the neuron. AMPA receptors lacking GluA2 are permeable to calcium, so they allow calcium to flow into the cell, but those that contain GluA2 are calcium impermeable. Calcium is very important for triggering a wide range of biochemical processes inside neurons that can lead to short or long-term changes to the function of the neuron. Previous work has suggested that some kinds of learning involve changing the total number of AMPA receptors localised to synapses, and others involve reducing the GluA2 content of the receptors at synapses for short periods of time. Both of these processes require the movement of receptors via GluA2, so it is crucial to understand how this happens. In certain disease states such as stroke, traumatic brain injury and motor neuron disease, longer-lasting removal of GluA2-containing AMPA receptors can lead to too much calcium entry, resulting in the dysfunction or death of the neurons affected.An important mechanism for moving receptors around neurons involves a protein called actin, which forms filaments that shrink and grow to physically manoeuvre parts of the cell or its constituents. A protein called PICK1 interacts with GluA2 and regulates the removal of AMPA receptors from synapses by inhibiting the formation of these actin filaments. Another protein, called cortactin, stimulates the formation of actin filaments, and we have found that it also binds directly to GluA2. Our preliminary experiments suggest that cortactin might function in an opposing manner to PICK1, and stabilise synaptic GluA2. We propose to study precise molecular mechanisms that control the GluA2-dependent movement of AMPA receptors at synapses, focussing on PICK1 and cortactin. 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 AMPA receptors at synapses in normal and disease states. We will use advanced forms of light microscopy as well as electron microscopy to visualise the precise location of these proteins relative to each other under conditions that lead to changes in synaptic levels of GluA2. We will use genetic techniques to manipulate interactions between relevant proteins and determine what effect this has on GluA2 movement under these conditions.This work will provide crucial mechanistic information about how neurons regulate the function of AMPA receptors, which are the most important neurotransmitter receptors in the brain. This will have wide-reaching implications for our understanding of learning and memory processes as well as a range of neurological diseases.

这项研究的目的是研究大脑中神经细胞如何相互交流的具体方面，以及这种交流在学习过程中以及由于某些类型的大脑障碍而发生的变化。大脑中的神经细胞(神经元)通过称为突触的连接相互通信。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。突触可以通过改变突触中神经元表面发现的受体的数量来改变它们的强度(称为“突触可塑性”)。这一过程被认为是学习和记忆的基础，因为记忆很可能存储在相互连接的神经元电路中。AMPA受体是大脑中参与大多数突触兴奋的神经递质受体，它们由不同的蛋白质成分组成，这些蛋白质的不同组合控制着受体的工作方式。名为GluA2的组件的存在非常重要，原因有两个。首先，许多其他蛋白质与GluA2物理上相互作用，以便将AMPA受体移动到突触或从突触移出。第二，它影响钙离子进入神经元。缺乏GluA2的AMPA受体对钙离子是通透的，因此它们允许钙流入细胞，但含有GluA2的AMPA受体是不透钙的。钙对于触发神经元内广泛的生化过程非常重要，这些过程可能导致神经元功能的短期或长期变化。以前的工作表明，有些学习涉及改变定位于突触的AMPA受体的总数，另一些涉及在短时间内减少突触受体的GluA2含量。这两个过程都需要受体通过GluA2进行移动，所以了解这一过程是如何发生的至关重要。在某些疾病状态下，如中风、创伤性脑损伤和运动神经元疾病，较长时间地移除含有GluA2的AMPA受体可能会导致过多的钙内流，导致受影响的神经元功能障碍或死亡。围绕神经元移动受体的一个重要机制涉及一种名为肌动蛋白的蛋白质，它形成细丝，收缩并生长，以物理方式操纵细胞的部分或其成分。一种名为PICK1的蛋白质与GluA2相互作用，通过抑制这些肌动蛋白细丝的形成来调节AMPA受体从突触中的移除。另一种名为Cortactin的蛋白质刺激肌动蛋白细丝的形成，我们发现它也直接与GluA2结合。我们的初步实验表明，Cortactin可能以与PICK1相反的方式发挥作用，并稳定突触GluA2。我们建议研究精确的分子机制，控制依赖于GluA2的AMPA受体在突触的运动，重点是PICK1和Cortactin。我们的大部分实验将使用从大鼠大脑中获得的神经元进行。这些神经元可以从大脑中分离出来，然后在培养皿中保持“活着”。使用这些细胞，我们将能够更多地了解在正常和疾病状态下调节突触AMPA受体的机制。我们将使用先进的光学显微镜以及电子显微镜来可视化这些蛋白质在导致GluA2突触水平变化的条件下相对于彼此的准确位置。我们将使用基因技术来操纵相关蛋白质之间的相互作用，并确定在这些条件下这对GluA2运动的影响。这项工作将为神经元如何调节AMPA受体的功能提供关键的机制信息，AMPA受体是大脑中最重要的神经递质受体。这将对我们理解学习和记忆过程以及一系列神经疾病产生广泛的影响。