Regulatory RNAs as mediators and biomarkers in Alzheimer's Disease

描述(申请人提供)：我们和其他人的高通量努力提供了强有力的证据，与早期的理解形成鲜明对比的是，哺乳动物基因组的大部分被转录成非编码的RNA。我们现在试图了解非编码RNA调控网络是否与阿尔茨海默病(AD)的2-淀粉样假说有关。这项应用侧重于一个非编码的RNA网络，它似乎调节分泌酶-1(BACE1)的表达，BACE1是AD病理生理学中的关键酶，也是AD药物的主要靶点。我们最近报道了一个BACE1非编码反义转录本(BACE1-AS)能够有效地调节BACE1mRNA和蛋白的表达。重要的是，在人类AD患者和淀粉样前体蛋白转基因小鼠(AD动物模型)中，BACE1-AS浓度显著升高。BACE1-AS在细胞核中丰富，但当暴露于淀粉样蛋白-1-42(A？1-42)或其他细胞应激源时，这一非编码转录本易位到细胞质中，导致BACE1 mRNA的稳定性增加，以及BACE1 mRNA上的microRNA结合位点的“掩蔽”；这些似乎是协同机制，导致在一个“恶性循环”中产生额外的A？1-42。这种与核库释放BACE1-AS相关的前馈通路可能是AD相关病理的驱动因素。考虑到这些发现，本研究的应用集中在三个目标上。我们的第一个目标是从机制上详细了解BACE1-AS在体外对BACE1表达的调控作用，以及BACE1-AS对AD相关病理的潜在影响。其次，我们将试图验证BACE1-AS对BACE1表达的体外调控作用在体内也存在于小鼠脑中。在这里，我们将建立BACE1-AS转基因小鼠，以确定这种非编码RNA的过度表达是否足以引起或加速AD样病理。第三，我们将解决BACE1相关转录本，特别是BACE1-AS是否可用作AD研究的生物标志物的问题。为了补充我们对AD和对照脑样本的研究，我们将在大量的外周血单核细胞样本中调查外周血BACE1相关生物标记物是否可以用作生物标记物。总体而言，这些实验有望对BACE1表达的控制机制产生重要的见解。此外，我们认为非编码调控RNA，特别是BACE1-AS，必须被视为AD早期生物标记物的候选。公共卫生相关性：阿尔茨海默病(AD)是一种破坏性的与年龄相关的神经退行性疾病，其特征是进行性认知障碍和短期记忆丧失。虽然淀粉样蛋白通路与AD之间的联系以及导致AD的事件的先后顺序仍然存在争议，但A2 1-42沉积在老年斑中是AD神经病理的一个已被证实的特征。BACE1是一种淀粉样前体蛋白(APP)裂解酶，对A2 1-42的生物合成至关重要。因此，BACE1在AD中是一个热门的药物靶点。了解BACE1的调控机制可能会对AD的发病机制有重要的启示，并有助于开发新的治疗方法和AD生物标志物。

DESCRIPTION (provided by applicant): High-throughput efforts by us and others have provided strong evidence that, in sharp contrast to earlier understanding, much of the mammalian genome is transcribed into noncoding RNA. We now seek to understand if noncoding RNA regulatory networks may relate to the 2-amyloid hypothesis in Alzheimer's disease (AD). This application focuses on a noncoding RNA network that appears to regulate the expression of ¿-secretase-1 (BACE1), a critical enzyme in AD pathophysiology and a prime AD drug target. We have recently reported that a BACE1 noncoding antisense transcript (BACE1-AS) concordantly and potently regulates BACE1 mRNA and protein expression. Important, BACE1-AS concentrations were markedly elevated in human AD patients as well as in amyloid precursor protein transgenic mice, an AD animal model. BACE1-AS is enriched in the cell nucleus but upon exposure to amyloid-¿ 1-42 (A¿ 1-42), or other cellular stressors, this noncoding transcript translocates into the cytoplasm resulting in increased BACE1 mRNA stability, as well as "masking" of a microRNA binding site on BACE1 mRNA; these appear to be synergistic mechanisms leading to generation of additional A¿ 1-42 in a "vicious cycle". This putative feed-forward pathway, linked to the release of BACE1-AS from a nuclear reservoir, may be driving aspects of AD related pathologies. Considering these findings, the present research application focuses on three aims. Our first aim is to gain a detailed mechanistic understanding of the in vitro regulatory effect that BACE1-AS exerts on BACE1 expression as well as the potential impact of BACE1-AS on AD related pathologies. Second, we will seek to verify that the in vitro regulatory effects of BACE1-AS on BACE1 expression are also present in vivo in mouse brain. Here we will develop BACE1-AS transgenic mice to determine if the overexpression of this noncoding RNA is sufficient to cause or accelerate AD- like pathology. Third, we will address the issue as to whether BACE1-related transcripts, notably BACE1-AS, may be useful as biomarkers for studies on AD. To complement our studies on AD and control brain samples, we shall investigate, in a large number of peripheral blood mononuclear cell samples, whether peripheral BACE1-related biomarkers might be useful as biomarkers. Overall, these experiments promise to yield significant insights into mechanisms by which BACE1 expression is controlled. Furthermore we argue that noncoding regulatory RNAs, notably BACE1-AS, must be viewed as candidates for an early biomarker signature in AD. PUBLIC HEALTH RELEVANCE: Alzheimer's disease (AD) is a devastating age-related neurodegenerative disorder characterized by progressive impairment of cognition and short- term memory loss. Although controversy still exists in the link between amyloid pathway and AD and in the precedence of events leading to AD, deposition of A2 1-42 into senile plaques is a proven feature of AD neuropathology. BACE1 is a ¿-site amyloid precursor protein (APP) cleaving enzyme essential for A2 1-42 biosynthesis. BACE1 is therefore a hotly pursued drug target in AD. Understanding the mechanisms by which BACE1 is regulated may reveal important insights into the pathogenesis of AD, and lead to the development of novel therapeutics and AD biomarkers.