ERAB, A BETA, NEUROTOXICITY AND ALZHEIMERS DISEASE

描述（来自申请人摘要）：越来越多的证据表明淀粉样蛋白- β肽（Abeta）在阿尔茨海默病（AD）发病机制中的致病作用。因此，β影响细胞特性的机制已经得到了深入的研究。较高浓度的β(1-40/42)在阿尔茨海默病的后期出现，形成聚集体和原纤维，非特异性地损害几乎任何细胞的膜。在阿尔茨海默病早期，当存在较低水平的Abeta时，Abeta与特定细胞以及敏感的亚细胞位点的特异性相互作用也可能介导细胞毒性。我们已经确定了一种新的内质网相关的β结合蛋白，称为ERAB，它具有b-羟基酰基辅酶a脱氢酶/短链醇脱氢酶的特性。重组人ERAB特异性结合β(1-40/42)，尽管它不与β淀粉样蛋白前体蛋白（betaAPP）或含有前体c端氨基酸的β形式相互作用。ERAB由神经母细胞瘤细胞组成，与β共免疫沉淀；在培养物暴露于外源性β后，ERAB迅速重新分布到质膜的内部。通过脂质体介导的阻断抗ERAB F（ab’）2进入细胞，可以阻止Abeta对神经母细胞瘤细胞的毒性，并通过在COS细胞中过表达ERAB而增强。我们认为ERAB是一个关键的细胞内靶标，增强了β介导的细胞扰动，并最终导致细胞毒性。我们假设，Abeta通过影响ERAB向浆/核膜的易位来影响ERAB，在浆/核膜中，细胞应激的诱导，以产生有毒醛和触发凋亡的事件为标志。我们的第一个目标是通过分析ERAB-Abeta诱导的毒性醛的产生和细胞凋亡，确定ERAB酶活性和ERAB与Abeta结合对细胞毒性的贡献。我们的第二个目标是用转基因小鼠分析ERAB对β诱导的细胞应激的贡献；我们假设靶向过表达ERAB的小鼠在富含abβ的环境中会表现出夸大的细胞毒性，并且这种毒性会在缺血应激下进一步加剧。本工作的长期目标是确定ERAB是否是Abeta细胞毒性的重要细胞辅助因子，以评估抑制ERAB是否可能成为未来药物开发的一种新的神经保护策略。

DESCRIPTION (from applicant's abstract): Increasing evidence points to a causative role for amyloid-beta peptide (Abeta) in the pathogenesis of Alzheimer's disease (AD). Thus, mechanisms through which Abeta affects cellular properties have come under intensive study. Higher concentrations of Abeta(1-40/42), present later in the course of AD, form aggregates and fibrils which nonspecifically damage membranes of virtually any cell. Earlier in AD, when lower levels of Abeta are present, specific interactions of Abeta with particular cells, as well as sensitive subcellular loci, could also mediate cellular toxicity. We have identified a novel endoplasmic reticulum-associated Abeta binding protein termed ERAB which has properties of a b-hydroxyacyl-Coenzyme A dehydrogenase/short-chain alcohol dehydrogenase. Recombinant human ERAB binds Abeta(1-40/42) specifically, though it does not interact with beta-amyloid precursor protein (betaAPP) or forms of Abeta containing C-terminal amino acids from the precursor. ERAB, constitutively produced by neuroblastoma cells, was co-immunoprecipitated with Abeta; following exposure of cultures to exogenous Abeta, ERAB was rapidly redistributed to the inner aspect of the plasma membrane. Toxicity of Abeta to neuroblastoma cells was prevented by liposome-mediated introduction of blocking anti-ERAB F(ab')2 into the cells, and was enhanced by overexpression of ERAB in COS cells. We propose that ERAB is a critical intracellular target potentiating Abeta-mediated cellular perturbation and, ultimately, cytotoxicity. We hypothesized that Abeta influences ERAB by effecting ERAB translocation to the plasma/nuclear membrane, where induction of cell stress, marked by generation of toxic aldehydes and events triggering apoptosis, occurs. Our first aim is to determine the contribution of ERAB enzymatic activity and ERAB binding to Abeta on cytotoxicity by analyzing ERAB-Abeta-induced generation of toxic aldehydes and apoptosis. Our second aim is to analyze the contribution of ERAB to Abeta-induced cell stress using transgenic mice; we hypothesize that mice with targeted overexpression of ERAB will display exaggerated cytotoxicity in an Abeta-rich environment, and that this will be further exacerbated by ischemic stress. The long-term goal of this wok is to determine if ERAB is an important cellular cofactor for Abeta cytotoxicity, in order to assess whether inhibition of ERAB might be a novel neuroprotective strategy for future drug development.