Trafficking of a Neuronal Glutamate Transporter, EAAC1

描述（由申请人提供）：谷氨酸是哺乳动物中枢神经系统中主要的快速兴奋性神经递质，参与发育和成人神经系统的几种形式的可塑性。在急性中枢神经系统损伤中，包括缺血和头部创伤，谷氨酸受体的过度激活会导致脑损伤。谷氨酸的细胞外浓度由钠依赖性转运蛋白家族控制。我们和其他人发现，其中一种转运体EAAC1的活性可以通过激活蛋白激酶C （PKC）或血小板衍生的生长因子受体而迅速增加。这些影响与EAAC1从亚细胞区室重新分布到细胞表面有关，并且与新转运体的合成无关。基于我们的初步研究，我们希望实现以下四个具体目标：1)我们建议研究pdgf依赖性EAAC1再分布所需的信号分子。2)我们最近发现EAAC1在质膜和细胞内区室之间本构循环，半衰期约为5-7分钟。我们建议确定EAAC1是否通过小窝蛋白依赖的内吞途径内化，并开始确定用于EAAC1回收的亚细胞区室。3)我们发现EAAC1与C激酶（PICK1）和PKC α亚型相互作用的蛋白形成复合物。我们建议确定这些相互作用所需的域，并确定这些相互作用如何影响EAAC1贩运。4)调节EAAC1转运和谷氨酸受体之一的机制似乎非常相似，增加EAAC1的信号会降低GluR2。我们建议使用多种方法来确定EAAC1和嗜离子性谷氨酸受体是否以这种明显互补的方式受到调节，并使用其他方法来化学诱导长期抑制（LTD）或长期增强（LTP）。我们对这种调控很感兴趣，因为EAAC1在突触传递具有高度可塑性的区域和对兴奋毒性损伤非常敏感的神经元中富集。EAAC1定位于突触后膜，位于突触周围的谷氨酸周围，是调节突触传递的理想位置。我们怀疑了解EAAC1的调控可能对理解谷氨酸毒性和/或突触可塑性有意义。

DESCRIPTION (provided by applicant): Glutamate is the predominant rapid excitatory neurotransmitter in the mammalian CNS and is involved in several forms of plasticity in the developing and adult nervous system. Excessive activation of glutamate receptors contributes to brain damage observed in acute insults to the CNS, including ischemia and head trauma. The extracellular concentrations of glutamate are controlled by a family of sodium-dependent transporters. We and others have found that the activity of one of these transporters, called EAAC1, can be rapidly increased by either activation of protein kinase C (PKC) or the platelet-derived growth factor receptor. These affects are associated with a redistribution of EAAC1 from a subcellular compartment to the cell surface and are independent of synthesis of new transporters. Based on our preliminary studies, we would like to pursue the following four specific aims: 1) We propose to examine the signaling molecules that are required for the PDGF-dependent redistribution of EAAC1. 2) We have recently found that EAAC1 constitutively recycles between the plasma membrane and intracellular compartments with a half-life of approximately 5-7 min. We propose to determine if EAAC1 is internalized by a caveolin-dependent endocytosis pathway and begin to identify the subcellular compartments used for recycling of EAAC1. 3) We have found that EAAC1 forms complexes with protein interacting with C kinase (PICK1) and the alpha subtype of PKC. We propose to identify domains required for these interactions and to determine how these interactions affect EAAC1 trafficking. 4) The mechanisms that regulate EAAC1 trafficking and one of the glutamate receptors appear to be quite similar, and a signal that increases EAAC1 decreases GluR2. We propose to use a variety of approaches to determine if EAAC1 and ionotropic glutamate receptors are regulated in this apparently complementary fashion using other approaches to chemically induce either long term depression (LTD) or long term potentiation (LTP). We are intrigued by this regulation because EAAC1 is enriched in areas where synaptic transmission is highly plastic and in neurons that are exquisitely sensitive to excitotoxic insults. EAAC1 is localized on the post-synaptic membrane and is perisynaptic surrounding GluRs where it is ideally situated to fine-tune synaptic transmission. We suspect that understanding the regulation of EAAC1 could have implications for understanding glutamate toxicity and/or synaptic plasticity.