Defining the epitope in antiNMDA receptor encephalitis

描述(申请人提供)：谷氨酸是中枢神经系统的主要兴奋性递质，不仅对突触传递至关重要，而且对突触可塑性和基因表达控制等长期神经元变化也是至关重要的。然而，谷氨酸的过度释放(在病理情况下发生)会导致细胞死亡，NMDA受体功能低下与精神分裂症有关。最近，我们与Josep Dalmau博士合作，发现NMDA受体抗体介导了与个性和行为变化、急性精神病和短期记忆缺陷相关的免疫性脑炎。患有这种综合征的人会产生抗体，选择性地与海马体发生反应，海马体是与记忆有关的区域，抗原是NMDAR的NR1亚单位。利用分子生物学方法，我们将NR1的表位定义在前380个氨基酸范围内。此外，衣霉素阻断了表位的产生，破坏了NR1上特定的N-连接的糖基化/去酰胺化位点就会去除免疫反应性，这表明特定的N-连接的糖基化和脱酰胺作用特定地参与了表位的产生。为了更好地理解这两个关键的生化过程在NMDAR特性中的作用，我们将对该表位的特征进行表征。我们将确定去胺化和差异糖基化的NMDAR受体在大脑中的分布，并将其与患者相关的抗NMDAR免疫反应的独特模式进行比较，这种免疫反应在抗NMDA受体脑炎中发现。此外，我们将用患者血清免疫标记NMDAR，并评估标记的NMDAR是否在特定位置糖基化或去酰胺化。在我们的第二个目标中，我们将NMDA受体引入缺乏特定糖基化酶的细胞系中。然后，我们将评估对NMDAR糖基化模式的影响，对它们的细胞运输和它们的生理特性的影响。这些将与NMDAR在特定大脑区域和细胞类型中的独特特性进行比较。总之，这些目标将为抗NMDAR脑炎的机制提供新的数据，为糖基化和脱酰胺在神经元化学中的作用提供新的理解，并为研究NMDA的生物化学设计新的策略。与公共卫生相关：该提案阐述了在被称为抗N-甲基-D-天冬氨酸受体脑炎的疾病中产生抗体的机制。通过对这一过程的理解，该提案可能有助于预防这种疾病的损害的治疗，以及对其他神经疾病(如中风)的基本了解。

DESCRIPTION (provided by applicant): Glutamate, the major excitatory transmitter in the central nervous system, is crucial not only for synaptic transmission but also for long-term neuronal changes such as synaptic plasticity and control of gene expression. However, excessive release of glutamate (as occurs in pathological situations) can result in cell death and NMDA receptor hypofunction has been implicated in schizophrenia. Recently, in collaboration with Dr. Josep Dalmau, we have found that antibodies to the NMDA receptor mediate an immune encephalitis associated with personality and behavioral changes, acute psychosis, and short-term memory deficits. Individuals with this syndrome make antibodies that selectively react with the hippocampus, an area involved in memory, and the antigen is the NR1 subunit of the NMDAR. Using molecular biological approaches we have defined the epitope to be within the first 380 amino acids on NR1. Furthermore, creation of the epitope is blocked by tunicamycin and disruption of a specific N- linked glycosylation/deamidation site in NR1 removes immunoreactivity, suggesting that site specific N-linked glycosylation and deamidation are specifically involved in the production of the epitope. We will characterize the features of this epitope in order to better understand the role of these 2 crucial biochemical processes in NMDAR properties. We will ascertain the distribution of deamidated and differentially glycosylated NMDA receptors in the brain and comparing this to the unique pattern of patients' related anti-NMDAR immunoreactivity that is found in anti NMDA receptor encephalitis. In addition, we will immunolabel NMDAR with patient serum and assess whether labeled NMDAR are glycosylated or deamidated at specific sites. In our second aim, we introduce NMDA receptors into cell lines lacking specific glycosylation enzymes. We will then assess the effect on the glycosylation pattern of NMDAR, on their cellular trafficking, and their physiological properties. These will be compared to the unique properties of NMDAR in specific brain regions and cell types. Together, these aims will supply new data on the mechanisms involved in antiNMDAR encephalitis, provide new understanding of the role of glycosylation and deamidation in neuronal chemistry, and devise new strategies for studying the biochemistry of NMDA. PUBLIC HEALTH RELEVANCE: The proposal addresses the mechanisms by which antibodies are generated in the disorder known as anti-N-methyl-D-aspartate Receptor encephalitis. Through understanding of this process, the proposal may facilitate therapies for preventing damage in this disorder and a basic understanding of other neurologic disorders such as stroke.