Presenilin Biology and the Mechanisms of Alzheimer's Disease

自1995年克隆以来，它被鉴定为史无前例的膜内 天冬氨酸蛋白酶1999年，早老素参与了一系列显著的信号和调控 所有后生动物中的事件。多发性硬化症是通过对阿尔茨海默氏症的研究发现的，但它很快就被发现了 被证明具有生命所必需的功能，尤指作为使Notch核信号发生作用的蛋白酶。 因此，继续破译PS的结构、功能以及蛋白质和血脂调节机制是 基础细胞生物学。与此同时，大脑中淀粉样β蛋白(AU)几十年的不变积累 在痴呆症状使PS/γ-分泌酶成为机制和治疗研究的关键靶点之前 在公元后。尽管它在生物学上起着多效性的作用，但该酶的结构只在12A下才被解析 这项拨款)，以及可以选择性地抑制其处理APP的小分子尚未得到验证。为所有人 出于这些原因，六位在早老素研究方面具有深厚经验的合作者希望应用一系列 细胞生物学、遗传学、化学、结构生物学和动物建模方面的技术，以解决一些 PS/γ-分泌酶生物学中最棘手的问题。我们能推导出这个19-的原子分辨率结构吗？ 跨膜复合体？协同α、β和伽马分泌酶的细胞生物学机制是什么？ 处理？某些突触蛋白和膜脂是如何调节神经元中PS活性的，从而影响 关键的A(14 2/4 0？人们能否设计出既有足够的效力又有选择性的药物来长期抑制伽马分泌酶？ 我们的小组已经仔细修改了这份申请，以解决所有深思熟虑的批评 SEP提供了。我们提出了许多相互关联的目标，其中包括三个交叉主题 团结我们的工作。首先，我们将进一步证实和扩大我们最近发现的内源性复合体 α/β/伽马分泌酶(一种“脱氧核糖体”)可能调节APP--大概还有所有伽马--底物的有效的、顺序的处理。其次，我们将应用这里开发的一种基于FRET的独特探针来测量活神经元中PS的构象，并了解我们最近通过蛋白质组学确定的某些突触集蛋白是否能够使PS快速且可逆地改变其构象 突触对钙离子内流的反应，解释了神经活动促进AB产生的原因。 第三，我们将研究我们为定义SARS而开发的新型伽马分泌酶调节剂和缺口保留抑制剂 对于APP和Notch裂解，识别同源结合位点，并评估它们在其他底物上的作用， 所有的目标都是将一个或多个推进到临床前开发。简而言之，我们致力于 应用新方法阐明伽马分泌酶在健康和疾病中的结构和功能。

Since its cloning in 1995 and its identification (under this grant) as an unprecedented intramembrane aspartyl protease in 1999, Presenilin has been implicated in a remarkable array of signaling and regulatory events in all metazoans. PS was discovered through research on Alzheimer's disease, but it was soon shown to confer functions necessary for life, especially as the protease that enables Notch nuclear signaling. Thus, continuing to decipher the structure, functions, and protein and lipid regulators of PS is a priority for basic cell biology. At the same time, the invariant cerebral accumulation of amyloid beta-protein (AU) decades before symptoms of dementia has made PS/gamma-secretase a key target for mechanistic and therapeutic study in AD. Despite its pleiotropic role in biology, the protease's structure has only been resolved at 12 A (under this grant), and small molecules that can selectively inhibit its processing of APP are not yet validated. For all these reasons, six collaborators with deep experience in the study of Presenilin wish to apply a range of techniques in cell biology, genetics, chemistry, structural biology and animal modeling to tackle some of the thorniest questions in PS/gamma-secretase biology. Can one derive an atomic resolution structure of this 19- transmembrane complex? What is the cell biological mechanism of coordinated alpha-, beta- and gamma-secretase processing? How do certain synaptic proteins and membrane lipids regulate PS activity in neurons, affecting the crucial A(l42/4o ratio? Can one design drugs that are sufficiently potent yet selective to chronically inhibit gamma-secretase? Our group has carefully revised this application to address all of the thoughtful critiques the SEP offered. We propose numerous interrelated aims that incorporate three cross-cutting themes which unite our work. First, we will further confirm and extend our recent discovery of an endogenous complex of the alpha/beta/gamma-secretases (a "sheddasome") that may mediate the efficient, sequential processing of APP - and presumably all gamma-substrates. Second, we will apply a unique FRET-based probe developed here to measure PS conformation in living neurons and learn if certain synaptotagmins we recently identified by proteomics as novel interactors of both PS1 and APP enable PS to change its conformation rapidly and reversibly in response to Ca2+ influx at the synapse, explaining the enhancement of AB production by neural activity. Third, we'll study novel gamma-secretase modulators and Notch-sparing inhibitors we've developed to define SARs for APP vs. Notch cleavage, identify the cognate binding sites, and assess their actions on other substrates, all with the goal of advancing one or more into preclinical development. In short, we are committed to applying novel approaches to elucidate the structure and function of gamma-secretase in health and disease.