Molecular actions of VLDLR in dendritic spine formation

描述（申请人提供）：突触和树突棘在学习和记忆中很重要。事实上，突触丢失被认为是阿尔茨海默病（AD）中认知能力下降的主要原因。突触功能受淀粉样前体蛋白（APP）和Reelin（APP的配体）和apoE受体VLDLR的影响，两者也参与AD病理学。最近，我们和其他人发现APP和Reelin对树突棘的形成都很重要。此外，我们确定了APP和极低密度脂蛋白受体（VLDLR）之间的一种新的相互作用，VLDLR是一种在认知能力和LTP中起重要作用的apoE受体。然而，VLDLR是否可以改变树突棘结构，以及APP和VLDLR复合物是否可以对树突棘形成产生协同或负面影响尚不清楚。为了解决这一问题，我们首先研究了VLDLR对树突棘形成的影响，发现VLDLR的过表达增加了原代海马神经元的棘密度，内源性VLDLR的敲低降低了原代海马神经元的棘密度。此外，VLDLR感染的神经元增加了Ras的活性，这是已知的促进树突棘的形成。此外，我们发现VLDLR的胞外结构域是树突棘形成所必需的，这表明VLDLR通过胞外配体相互作用改变树突棘密度。基于文献和我们的研究结果，我们假设，VLDLR促进树突棘形成通过与Reelin和/或APP。在此应用程序中，我们将研究是否VLDLR改变树突棘密度在体内，并将调查的分子机制，VLDLR影响树突棘的形成。此外，我们将研究VLDLR和APP之间的相互作用是否独立，竞争或协同改变树突棘的形成。我们将进一步研究这些作用是否受到细胞外配体Reelin的调节，Reelin与APP和VLDLR相互作用。我们将利用分子和细胞生物学方法来阐明VLDLR与APP和/或Reelin合作或竞争调节树突棘形成的分子机制。因此，在目标1中，我们将确定VLDLR是否调节树突棘的形成。在目的2中，我们将调查是否VLDLR介导树突棘形成通过Reelin和/或APP。拟议的实验将进一步我们的理解的生理参与的相互作用之间的VLDLR和APP和/或细胞外配体，Reelin，在树突棘的形成。我们的研究结果将有助于越来越多的证据表明，这些蛋白质，VLDLR和APP，在树突棘的形成，因此，也在突触连接和学习和记忆中具有至关重要的功能。在这些实验中研究的生理机制将促进对阿尔茨海默病中观察到的认知下降和突触丧失的治疗的理解和开发。

DESCRIPTION (provided by applicant): Synapses and dendritic spines are important in learning and memory. Indeed, synapse loss is thought to be a main contributor to the cognitive decline seen in Alzheimer's disease (AD). Synapse functions are affected by amyloid precursor protein (APP) and Reelin, a ligand for APP and the apoE receptor VLDLR, both of which also are involved in AD pathology. Recently, we and others found that both APP and Reelin are important for dendritic spine formation. Additionally, we indentified a novel interaction between APP and the very low density lipoprotein receptor (VLDLR), an apoE receptor that plays an important role in cognitive performance as well as LTP. However, whether VLDLR can alters dendritic spine structure and whether an APP and VLDLR complex can have synergistic or negative effects on dendritic spine formation is unknown. To address this, we initially examined the effects of VLDLR on dendritic spine formation and found that overexpression of VLDLR increased spine density, and knockdown of endogenous VLDLR decreased spine density in primary hippocampal neurons. Also, VLDLR-infected neurons increased the activity of Ras, which is known to promote dendritic spine formation. Moreover, we found that extracellular domain of VLDLR is required for dendritic spine formation, suggesting that VLDLR alters dendritic spine density through extracellular ligand interactions. Based on the literature and our findings, we hypothesize that VLDLR promotes dendritic spine formation through association with Reelin and/or APP. In this application, we will examine whether VLDLR alters dendritic spine density in vivo, and will investigate the molecular mechanism by which VLDLR affects dendritic spine formation. Additionally, we will examine whether the interactions between VLDLR and APP work independently, competitively or synergistically to alter dendritic spine formation. We will further examine whether these effects are regulated by the extracellular ligand, Reelin, which interacts with both APP and VLDLR. We will utilize molecular and cell biology approaches to elucidate the molecular mechanisms by which VLDLR cooperates or competes with APP and/or Reelin to regulate dendritic spine formation. Therefore, in Aim 1, we will determine whether VLDLR regulates dendritic spine formation. In Aim 2, we will investigate whether VLDLR mediates dendritic spine formation through Reelin and/or APP. The proposed experiments will further our understanding of the physiological involvement of the interaction between VLDLR and APP and/or the extracellular ligand, Reelin, in dendritic spine formation. Our results will contribute to the growing evidence that these proteins, VLDLR and APP, have crucial functions in dendritic spine formation, and therefore, also in synapse connection and in learning and memory. The physiological mechanisms investigated in these experiments will advance the understanding of, and the development of therapeutics for the cognitive decline and synapse loss observed in Alzheimer's disease.