Common regulation of BACE1 and y-secretase substrate processing

淀粉样前体蛋白（APR）依次被BACE 1和PS/γ-分泌酶切割，产生A（3。 APLP 2经历类似的加工，导致产生Ap-like肽。我们最近 鉴定了BACE 1和PS/γ-分泌酶活性的第三种底物，即电压门控的β 32-亚基（（32） 钠通道（Nav 1）。Nav 1几乎完全负责动作电位的上升阶段。 我们的初步数据表明BACE 1-/γ-分泌酶介导的β 2加工调节钠通道 NaJ α亚基Nav1.1在神经元细胞和BACE 1中的功能以及mRNA和蛋白水平 转基因小鼠在BACE 1水平升高的AD患者脑中，我们还发现BACE 1水平升高（32 C-末端 片段和Nav1.1水平。有趣的是，癫痫发作的发病率增加与 AD，更常见于PS1突变引起的家族性AD。a-和（3- Najs的亚基与癫痫症状有关，这些症状是由Najs的减少和增加引起的。 钠通道活性导致钠通道功能失衡。因此，BACE 1/ysecretase- 介导的裂解可能导致两个独立的AD相关的致病事件，A（3 淀粉样蛋白生成（毒性淀粉样蛋白形成）和钠通道功能障碍（癫痫发作）。来定义 调节这些裂解的因素，在这里，我们提出解决的假设，新的结合蛋白 调节β 2的加工和代谢，并且这些可以由APP和/或APLP 2共享。在这些 研究中，我们将主要表征（32加工和代谢，因为这种蛋白质是不是以及 我们将利用APLP 2作为第三BACE 1/γ-分泌酶底物。在特定 目的1我们鉴定了β 2的新的结合伴侣，并评估了它们在蛋白水解中的功能意义。 处理（32.如果这些相互作用的蛋白质被APP或APLP 2所共享，我们也将研究它们是否 影响处理APP或APLP 2包括A（3代。特定目标2将表征代谢 β 2调节钠通道功能的通路，以及这些通路是否由APP和APLP 2共享。 由于所有三种蛋白质都定位于脂筏，我们将分析这些除了所有主要的 BACE 1/γ-分泌酶底物存在的亚细胞区室。我们将评估 新鉴定的β 2代谢途径上的结合配偶体和基于细胞的钠通道功能上的结合配偶体 测定和体内。糖尿病相关的FAD突变对PS1钠通道水平的影响 还将在细胞和体内确定功能。这些实验的结果将揭示新的 可能导致旨在减少A（3代）的治疗策略的加工或代谢途径 以及使具有癫痫症状的AD患者的膜兴奋性正常化。Lay language：我们 我建议了解淀粉样蛋白是如何在阿尔茨海默氏症患者的大脑中形成的，以及为什么一些 患者会出现癫痫发作。这些信息可以帮助找到阿尔茨海默病的治疗方法。

The amyloid precursor protein (APR) is sequentially cleaved by BACE1 and PS/y-secretase to yield A(3. APLP2 undergoes similar processing, resulting in the production of an Ap-like peptide. We have recently identified a third substrate for BACE1 and PS/y-secretase activities, the (32-subunit ((32) of the voltage-gated sodium channel (Nav1). Nav1s are almost exclusively responsible for the rising phase of action potentials. Our preliminary data indicate that BACE1-/y-secretase-mediated processing of (32 regulates sodium channel function and both mRNA and protein levels of a NaJ a-subunit, Nav1.1, in neuronal cells and BACE1 transgenic mice. In AD patient brains with elevated BACE1 levels, we also found elevated (32 C-terminal fragments and Nav1.1 levels. Interestingly, increased incidence of epileptic seizures has been associated with AD, more frequently with familial forms AD caused by mutations in PS1. Mutations in both a- and (3- subunits of the Najs have been linked to epileptic symptoms, induced by both decreased and increased sodium channel activity resulting in an imbalance in sodium channel function. Therefore, altered BACE1/ysecretase- mediated cleavages may result in two separate AD-associated pathogenic events, changes in A(3 generation (toxic amyloid formation) and sodium channel malfunction (epileptic seizures). To define the factors regulating these cleavages, here we propose to address the hypothesis that novel binding proteins regulate the processing and metabolism of (32 and these may be shared by APP and/or APLP2. In these studies, we will primarily characterize (32 processing and metabolism since this protein is not as well characterized as APP. We will take advantage of APLP2 as a third BACE1/y-secretase substrate. In Specific Aim 1 we identify novel binding partners for (32 and assess their functional significance in proteolytic processing of (32. If these interacting proteins are shared by APP or APLP2, we will also study whether they affect processing APP or APLP2 including A(3 generation. Specific Aim 2 will characterize metabolic pathways of (32 regulating sodium channel function, and whether these are shared by APP and APLP2. Since all three proteins share localization to lipid rafts, we will analyze these in addition to all major subcellular compartments for the presence of BACE1/y-secretase substrates. We will assess the effect of newly identified binding partners on metabolic pathways of (32 and on sodium channel function in cell-based assays and in vivo. The effects of seizure-associated FAD mutations in PS1 on sodium channel levels and function will also be determined in cells and in vivo. Results from these experiments will uncover novel processing or metabolic pathways that may lead to therapeutic strategies aimed at reducing A(3 generation as well as normalizing membrane excitability in AD patients with epileptic symptoms. Lay language: We propose to understand aspects of how amyloid is formed in the brains of Alzheimer's patients and why some patients develop seizures. This information can help find therapies for Alzheimer's disease.