Defining the epitope in antiNMDA receptor encephalitis

描述（申请人提供）：谷氨酸是中枢神经系统中主要的兴奋性递质，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神经元的长期变化也至关重要。然而，过度释放谷氨酸（在病理情况下发生）可导致细胞死亡和NMDA受体功能低下与精神分裂症有关。最近，在与Josep Dalmau博士的合作中，我们发现NMDA受体的抗体介导与人格和行为改变，急性精神病和短期记忆缺陷相关的免疫性脑炎。患有这种综合征的个体产生的抗体会选择性地与海马体（一个与记忆有关的区域）发生反应，这种抗原是NMDAR的NR1亚基。利用分子生物学方法，我们确定了表位在NR1上的前380个氨基酸内。此外，抗原表位的形成被tunicamycin阻断，NR1中特异性N-连接的糖基化/脱酰胺位点的破坏消除了免疫反应性，这表明位点特异性N-连接的糖基化和脱酰胺特异性参与了抗原表位的产生。为了更好地理解这两个关键生化过程在NMDAR特性中的作用，我们将对该表位的特征进行表征。我们将确定脱酰胺化和差异糖基化的NMDA受体在大脑中的分布，并将其与抗NMDA受体脑炎中发现的患者相关抗NMDA免疫反应的独特模式进行比较。此外，我们将用患者血清对NMDAR进行免疫标记，并评估标记的NMDAR在特定位点是否被糖基化或脱酰胺化。在我们的第二个目标中，我们将NMDA受体引入缺乏特异性糖基化酶的细胞系。然后，我们将评估对NMDAR的糖基化模式、细胞运输和生理特性的影响。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。这些目标将为抗NMDA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认识，并为研究NMDA的生物化学提供新的策略。公共卫生相关性：该提案解决了在抗n -甲基- d -天冬氨酸受体脑炎中产生抗体的机制。通过对这一过程的了解，该建议可能有助于预防这种疾病的损害的治疗，并对其他神经系统疾病（如中风）有基本的了解。

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.