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，突触丧失被认为是阿尔茨海默病（AD）中认知能力下降的主要原因。突触功能受淀粉样蛋白前体蛋白（APP）和Reelin （APP和apoE受体VLDLR的配体）的影响，两者也参与AD病理。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，我们还发现了APP与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apoE受体，在认知表现和LTP中起着重要作用。然而，VLDLR是否能改变树枝状脊柱的结构，APP和VLDLR复合物对树枝状脊柱的形成是否具有协同作用或负面作用尚不清楚。为了解决这个问题，我们初步研究了VLDLR对树突棘形成的影响，发现VLDLR的过表达增加了脊柱密度，而内源性VLDLR的敲低降低了海马初级神经元的脊柱密度。此外，vldlr感染的神经元增加Ras的活性，已知Ras可促进树突棘的形成。此外，我们发现VLDLR的细胞外结构域是树突棘形成所必需的，这表明VLDLR通过细胞外配体相互作用改变树突棘密度。基于文献和我们的发现，我们假设VLDLR通过与Reelin和/或APP的关联促进树突棘的形成。在本应用中，我们将研究VLDLR是否会改变体内树突棘密度，并将研究VLDLR影响树突棘形成的分子机制。此外，我们将研究VLDLR和APP之间的相互作用是否独立地、竞争性地或协同地改变树突棘的形成。我们将进一步研究这些作用是否受到与APP和VLDLR相互作用的细胞外配体Reelin的调节。我们将利用分子和细胞生物学的方法来阐明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Aim 1中，我们将确定VLDLR是否调节树突棘的形成。在Aim 2中，我们将研究VLDLR是否通过Reelin和/或APP介导树突棘的形成。所提出的实验将进一步了解VLDLR和APP和/或细胞外配体Reelin之间相互作用在树突棘形成中的生理参与。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，在树突棘的形成中具有至关重要的功能，因此，在突触连接和学习和记忆中也是如此。在这些实验中研究的生理机制将促进对阿尔茨海默病认知能力下降和突触丧失的理解和治疗方法的发展。

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.