INVOLVEMENT OF RAP AND RAPGAP IN NMDA RECEPTOR SIGNALING

n -甲基-d -天冬氨酸受体（NMDAR）因其在活动依赖性突触可塑性和兴奋毒性中起着关键作用而被广泛研究。定义由NMDAR调控的信号级联对于了解NMDAR功能的机制至关重要，但由NMDAR启动的细胞内信号尚未得到很好的表征。我们的实验室对与NMDAR细胞质尾部相关的蛋白质进行了系统的分析，其基本原理是这些蛋白质将通过NMDAR通道将钙内流传导到下游信号通路。多结构域蛋白PSD-95似乎是这些转导元件的组织中心，它与NMDARs以及两种GTPase激活蛋白（gap）等信号分子结合。其中之一，SynGAP，是最近描述的ras特有的GAP；另一种是SpanGAP，是一种可能调节Rap小gtp酶的新型GAP。本提案将重点关注SpanGAP和Rap在NMDAR信号传导中的潜在作用，研究该途径（与Ras途径相比）在NMDAR信号传导级联中的相对贡献。显性负突变体和反义策略将用于解剖培养的原代神经元的特定转导机制。将通过使用各种转录报告以及针对MAP激酶和CREB的磷酸化特异性抗体来分析效果。本研究结果可能揭示nmdar介导的突触可塑性和兴奋毒性的新机制。

The N-methyl-D-aspartate receptor (NMDAR) has been extensively studied due to its critical role in activity-dependent synaptic plasticity and excitotoxicity. Defining the signaling cascades regulated by the NMDAR is essential for a mechanistic understanding of NMDAR function, but the intracellular signals initiated by NMDARs are not well characterized. Our laboratory has undertaken a systematic analysis of the proteins associated with the cytoplasmic tails of NMDARs with the rationale that such proteins would be ideally positioned to transduce calcium influx through the NMDAR channel to downstream signaling pathways. The multidomain protein PSD-95 appears to be an organizing center for these transducing elements, binding to NMDARs as well as to signaling molecules such as two GTPase activating proteins (GAPs). One of these, SynGAP, is a recently described Ras-specific GAP; the other, SpanGAP, is a novel GAP likely to regulate Rap small GTPases. This proposal will focus on the potential role of SpanGAP and Rap in NMDAR signaling, examining the relative contribution of this pathway (compared with the Ras pathway) in the NMDAR signaling cascade. Dominant negative mutants and antisense strategies will be employed to dissect specific transduction mechanisms in cultured primary neurons. Effects will be analyzed by using a variety of transcriptional reporters as well as phospho-specific antibodies against MAP kinases and CREB. The results of this study may shed light on novel mechanisms contributing to NMDAR-mediated synaptic plasticity and excitotoxicity.