Molecular Regulation of Metabotropic Glutamate Receptors in Striatal Neurons

描述（由申请人提供）：蛋白磷酸化是谷氨酸受体翻译后调节的重要机制。通过磷酸化细胞内结构域中的特定氨基酸，蛋白激酶调节给定谷氨酸受体的锚定、运输和信号传导。I组代谢型谷氨酸受体（mGluR 1/5）密集表达于纹状体，纹状体是涉及精神刺激剂成瘾性质的脑区。长型mGluR 1/5剪接变体（1a、5a和5 b）具有大的细胞内C末端尾，这为直接蛋白质-蛋白质相互作用和磷酸化提供了基础。在我们最近的研究中，我们发现Ca ~（2+）/钙调素依赖性蛋白激酶II（CaMKII）直接结合到mGluR 5a C-末端的近端区域。这种结合将mGluR 5a转化为可能在选择性丝氨酸位点磷酸化的生化底物。这些发现提出了创新的问题，如CaMKII是否通过直接的蛋白质-蛋白质相互作用和磷酸化调节mGluR 1/5，以及这种调节在疾病模型中是否具有高度的临床相关性。在这个延续的提议中，提出了一系列从分子到行为的连贯实验，以证实体外CaMK II与mGluR 1/5的直接结合，并建立体内纹状体神经元中天然CaMK II和mGluR 1/5相互作用。我们将表征钙离子是否以及如何在体外和体内调节CaMKII和mGluR 1/5之间的相互作用。然后，我们将研究在纹状体神经元或异源细胞中，Ca 2+调节的CaMKII-mGluR 1/5相互作用是否调节1）mGluR 1/5的信号传导功效，2）受体的运输，以及3）mGluR 1/5与关键支架Homer蛋白的相互作用。最后，我们将进行神经行为实验，以确定CaMKII-mGluR 1/5相互作用在精神兴奋剂安非他明成瘾作用中的作用。我们的结果将为激酶调节mGluRs的新突触模型及其与精神疾病（物质成瘾）的联系提供证据和见解。他们还将通过靶向mGluRs和CaMKII，最终有助于开发新的药物疗法，用于治疗各种精神疾病，包括成瘾。

DESCRIPTION (provided by applicant): Protein phosphorylation is an important mechanism for post-translational regulation of glutamate receptors. Through phosphorylating a specific amino acid in the intracellular domain, protein kinases regulate anchoring, trafficking, and signaling of a given glutamate receptor. Group I metabotropic glutamate receptors (mGluR1/5) are densely expressed in the striatum, a brain area involved in addictive properties of psychostimlants. The long-form mGluR1/5 splice variants (1a, 5a, and 5b) have a large intracellular C-terminal tail, which provides a basis for direct protein-protein interactions and phosphorylation. In our recent studies, we found that Ca2+/calmodulin-dependent protein kinase II (CaMKII) binds directly to the proximal region of mGluR5a C-terminus. This binding converts mGluR5a into a biochemical substrate for phosphorylation likely at a selective serine site. These findings raise innovative questions as to if CaMKII regulates mGluR1/5 via a direct protein-protein interaction and phosphorylation and if this regulation has a high clinical relevance in a disease model. In this continuation proposal, a series of coherent experiments from molecule to behavior was proposed to confirm the direct binding of CaMKII to mGluR1/5 in vitro and to establish that native CaMKII and mGluR1/5 interact with each other in striatal neurons in vivo. We will characterize if and how Ca2+ regulates the interaction between CaMKII and mGluR1/5 in vitro and in vivo. We will then investigate whether Ca2+-regulated CaMKII-mGluR1/5 interactions regulate 1) signaling efficacy of mGluR1/5, 2) trafficking of the receptors, and 3) interactions of mGluR1/5 with key scaffold Homer proteins, in striatal neurons or heterologous cells. Finally, we will carry out neurobehavioral experiments to define the role of CaMKII-mGluR1/5 interactions in the addictive action of the psychostimulant amphetamine. Our results will provide evidence and insights for a new synaptic model of kinase-regulated mGluRs and for its linkage to a mental illness (substance addiction). They will also ultimately contribute to the development of novel pharmacotherapies, by targeting mGluRs and CaMKII, for treating various mental illnesses, including addiction.