PGDF-Dependent Regulation of EAAC1

EAAC1 的 PGDF 依赖性调节

• 批准号: 7116352
• 负责人: AMANDA L SHELDON
• 金额: $ 4.11万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7116352
• 关键词: actins; binding proteins; biological signal transduction; chimeric proteins; cytoskeleton; enzyme activity; gene expression; glioma; glutamate transporter; hormone regulation /control mechanism; laboratory rat; neoplastic cell culture for noncancer research; neural transmission; neurons; neuroprotectants; neurotransmitter transport; phosphatidylinositol 3 kinase; phosphorylation; platelet derived growth factor; predoctoral investigator; protein localization; protein structure function; protooncogene; tissue /cell culture; transfection

DESCRIPTION (provided by applicant): The excitatory amino acid, glutamate, is cleared from the synapse by a family of glutamate transporters. EAAC1, a neuronal subtype, is enriched in the hippocampus and cortex, areas that are extremely sensitive to excitotoxic insults. EAAC1 subcellular localization is regulated by stimulation in C6 glioma cells. PDGF has been shown to be neuroprotective, and regulation of EAAC1 may represent one way in which PDGF exerts its neuroprotective effects. The goal of this proposal is to understand the mechanisms underlying PDGF-dependent regulation of EAAC1. Aim I is to determine if PDGF-dependent phosphorylation of the proto-oncogene product, Cbl, via an interaction with the Cbl adapter protein (CAP) is required. This will be examined by expressing a dominant interfering CAP construct and measuring cell surface EAAC1 expression after PDGF treatment. Aim II is to test whether intact actin filaments are required for stimulated trafficking, by pre-treating cells with an actin depolymerizer and measuring glutamate uptake and cell surface EAAC1. Aim Ill is to identify structural motifs of EAAC1 required for PDGF-dependent trafficking, using chimeras and mutant variants of EAAC1.

描述（由申请人提供）：兴奋性氨基酸谷氨酸被谷氨酸转运蛋白家族从突触中清除。 EAAC1 是一种神经元亚型，富含于海马体和皮质，这些区域对兴奋性毒性损伤极其敏感。 EAAC1 亚细胞定位受 C6 神经胶质瘤细胞刺激的调节。 PDGF已被证明具有神经保护作用，EAAC1的调节可能代表PDGF发挥其神经保护作用的一种方式。该提案的目标是了解 EAAC1 的 PDGF 依赖性调节的潜在机制。目的 I 是确定原癌基因产物 Cbl 是否需要通过与 Cbl 衔接蛋白 (CAP) 相互作用进行 PDGF 依赖性磷酸化。这将通过表达显性干扰 CAP 构建体并测量 PDGF 处理后细胞表面 EAAC1 表达来检查。目标 II 是通过用肌动蛋白解聚剂预处理细胞并测量谷氨酸摄取和细胞表面 EAAC1，来测试刺激运输是否需要完整的肌动蛋白丝。目的III是使用EAAC1的嵌合体和突变变体来鉴定PDGF依赖性运输所需的EAAC1的结构基序。