A Molecular Signaling Pathway Underlying Differential Predisposition of ApoE4 Genotype to Alzheimer's Disease

项目摘要/摘要 载脂蛋白E(ApoE)基因变异ApoE-ϵ4的遗传易感性是高血压的最大遗传危险因素 阿尔茨海默病(AD)，这是老龄化的美国迅速增加的医疗保健负担。有一个 在理解编码载脂蛋白E亚型的载脂蛋白E-ϵ4的表达上存在着根本的差距， 影响神经功能，参与AD的发病。这一空白，直到填补，代表着我们的 对阿尔茨海默病的认识不足，我们无法为阿尔茨海默病和风险携带者提供治疗和预防。 AD发病机制中的一个中心事件是淀粉样蛋白-β(A-β)多肽的沉积，由一系列 淀粉样前体蛋白(APP)的蛋白酶裂解。使用从人类中提取的纯人类神经元 胚胎干细胞或IPSCs，申请人最近发现了一种信号通路，ApoE4 刺激APP转录，从而在神经元中诱导Aβ的产生，比 另外两种同工酶APOE2和ApoE3。这一途径依赖于双亮氨酸拉链激酶DLK和 下游的MAPK信号，并被三种不同的ApoE亚型激活，其效价顺序为 ApoE4；ApoE3；APOE2，与AD风险等级顺序平行。考虑到这种惊人的ApoE异构体特异性效应， 本应用程序的总体目标是确定ApoE4特异性激活DLK/MAPK是否以及如何激活 载脂蛋白E4‘S有害促进AD的作用可能与信号转导途径有关。我的中心假设是 ApoE4可能通过慢性刺激DLK/MAPK和增加神经元Aβ的产生而易患AD。 通过操纵DLK/MAPK通路干扰ApoE增强的APP转录可能 延缓AD发病机制。我计划测试我的中心假设，从而实现以下总体目标 本项目通过追求以下具体目标：1)确定DLK/MAPK通路是否介导 载脂蛋白E亚型诱导差异APP/Aβ，载脂蛋白E4和GT；载脂蛋白E3和GT；载脂蛋白2)鉴定转录调控 ApoE刺激所需的APP基因元件；3)决定DLK/MAPK的功能输出 体内途径。基因和药物操作的组合将被用来增强或抑制 APOE激活培养神经元和动物脑中的DLK/MAPK通路。分子机制和 这条途径的行为意义将得到严格的检验。总体方法是创新的。 因为它改变了现状，利用人类神经元，专注于那些被低估的人 从AD发病机制看APP转录机制。这项拟议的研究意义重大，因为它 有望定义载脂蛋白E在大脑中的新角色，并通过靶向探索新的治疗视野 DLK-MAPK通路作为替代策略。这些发现将极大地提高我们对AD的认识 其致病机制，并有很大潜力转化为迫切需要的治疗和预防。

PROJECT SUMMARY/ABSTRACT Inheritance of Apolipoprotein E (ApoE) gene variant APOE-ϵ4 is the strongest genetic risk factor for Alzheimer’s disease (AD), a rapidly growing burden of health care for the aging United States. There is a fundamental gap in understanding how expression of APOE-ϵ4, which encodes ApoE isoform ApoE4, influences neuronal function and contributes to AD pathogenesis. This gap, until filled, represents our insufficiency in understanding AD and our inability to provide treatment and prevention to AD and risk carriers. A central event in AD pathogenesis is deposition of amyloid-β (Aβ) peptide, generated from a series of protease cleavages of amyloid precursor protein (APP). Using pure human neurons derived from human embryonic stem cells or iPSCs, the applicant has recently identified a signaling pathway by which ApoE4 stimulates APP transcription and consequently induces Aβ production in neurons, more effectively than the other two isoforms ApoE2 and ApoE3. This pathway hinges on double leucine-zipper kinase DLK and downstream MAPK signaling, and is activated by three ApoE isoforms differentially in the potency rank order of ApoE4>ApoE3>ApoE2, paralleling AD risk rank order. Given this striking ApoE isoform-specific effect, the overall objective of this application is to determine whether and how ApoE4-specific activation of DLK/MAPK signaling pathway may account for ApoE4’s deleterious AD-promoting effect. My central hypothesis is that ApoE4 may predispose to AD by stimulating DLK/MAPK and increasing neuronal Aβ production chronically, and that interference with ApoE-enhanced APP transcription via manipulations of DLK/MAPK pathway may delay AD pathogenesis. I plan to test my central hypothesis, thereby accomplishing the overall objective for this project by pursuing these following specific aims: 1) Determine whether DLK/MAPK pathway mediates differential APP/Aβ induction by ApoE isoforms, ApoE4>ApoE3>ApoE2, 2) Identify the transcription control elements of APP gene required for ApoE stimulation, and 3) Determine the functional output of DLK/MAPK pathway in vivo. A combination of genetic and pharmacological manipulations will be used to enhance or inhibit ApoE-activated DLK/MAPK pathway in cultured neurons and in animal brains. The molecular mechanism and behavioral significance of this pathway will be rigorously examined. The overall approach is innovative because it departs from the status quo by utilizing human neurons and focusing on the underappreciated mechanisms of APP transcription in light of AD pathogenesis. The proposed research is significant, because it is expected to define a new role of ApoE in the brain, and also to explore new therapeutic horizons by targeting DLK-MAPK pathway as an alternative strategy. Findings will advance greatly our knowledge of AD pathogenesis, and have strong potential for future translation into urgently needed treatment and prevention.