Presenilin Biology and the Mechanisms of Alzheimer's Disease

自1995年克隆并鉴定（根据该资助）为前所未有的膜内 早老素在1999年被发现参与了一系列重要的信号传导和调节作用， 所有后生动物的事件。PS是通过对阿尔茨海默病的研究发现的，但很快就被发现了 显示出赋予生命所必需的功能，特别是作为使Notch核信号传导成为可能的蛋白酶。 因此，继续破译PS的结构、功能以及蛋白质和脂质调节剂是研究的优先事项。 基础细胞生物学与此同时，淀粉样β蛋白（Au）在大脑中的恒定积累， 在痴呆症状出现之前，PS/γ-分泌酶已成为机制和治疗研究的关键目标 在AD中。尽管其在生物学中的多效性作用，但蛋白酶的结构仅在12 A（低于10 A）处被解析。 这项授权），和小分子，可以选择性地抑制其加工的APP尚未得到验证。为所有 由于这些原因，六位在早老素研究方面有丰富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- 跨膜复合物？什么是协调的α，β和γ分泌酶的细胞生物学机制 处理？某些突触蛋白和膜脂如何调节神经元中的PS活性， 关键的A（142/40）比？我们能设计出既有效又有选择性的药物来长期抑制γ-分泌酶吗？ 我们的团队已经仔细修改了这个应用程序，以解决所有深思熟虑的批评 SEP提供。我们提出了许多相互关联的目标，其中包括三个贯穿各领域的主题， 团结我们的工作。首先，我们将进一步证实和扩展我们最近发现的内源性复合物， α/β/γ-分泌酶（“脱落体”），其可介导APP -和可能所有γ-底物的有效、顺序加工。其次，我们将应用一种独特的基于FRET的探针来测量活神经元中的PS构象，并了解我们最近通过蛋白质组学鉴定的某些突触结合蛋白作为PS1和APP的新型相互作用物是否能够使PS在神经元中快速可逆地改变其构象。 突触对Ca 2+内流的反应，解释了神经活动对AB产生的增强。 第三，我们将研究新的γ-分泌酶调节剂和Notch-sparing抑制剂，我们已经开发出定义SAR 对于APP与Notch切割，鉴定同源结合位点，并评估其对其他底物的作用， 所有这些都是为了将一种或多种药物推进临床前开发。简而言之，我们致力于 应用新的方法来阐明γ-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.