Deciphering Molecular Mechanisms of Calcium Homeostasis

项目摘要/摘要 钙是许多细胞过程的关键媒介，包括肌肉收缩， 线粒体活性、转录、细胞分裂和突触小泡释放。矛盾的是， 钙还会引发细胞死亡和细胞坏死。因此，钙的调节失调 信号可以破坏细胞的动态平衡。因此，钙水平需要严格控制。 控制住了。事实上，钙信号缺陷与许多神经退行性变有关。 疾病、肌肉营养不良和心脏病。要了解其调控机制 钙信号以及这个过程中的缺陷如何导致细胞功能障碍，我们正在 利用模型系统秀丽隐杆线虫识别 参与钙离子转运的调节。我们最近证明了SEL-12，即 线虫早老素同源基因在调节内质网-线粒体中的作用 钙稳态。早老素功能受阻导致线粒体增多 钙水平和改变线粒体代谢促进蛋白质稳态崩溃和 神经退行性变。早老素是一种高度保守的蛋白质，从植物到人类都是如此 广泛存在于沙门氏菌的内膜结构(如内质网和溶酶体)上 大多数细胞类型。然而，早老素在血管内膜系统中的作用尚不清楚。 重要的是，人类早老素基因突变是早发性家族性疾病最常见的原因 阿尔茨海默氏症。尽管早老素与阿尔茨海默病的关系已被证实 20多年前，早老素突变导致的功能后果 阿尔茨海默氏症目前还不清楚。为了进一步了解早老素在 线粒体钙动态平衡与神经元适合性，我们开发了一种新颖而高效的 选择性RNA干扰筛查用于鉴定沉思提升的基因产物 在sel-12突变体中观察到内质网对线粒体钙信号的影响。由此 筛选，我们已经确定了几种已知的介导内质网钙离子的蛋白质 释放和线粒体钙摄取，但我们也鉴定了几个基因产物与一个 在内质网和线粒体钙信号中的作用尚未确定。我们建议 利用结合了基因操作、高分辨率活细胞的多方面方法 显微镜下分析内质网和线粒体动力学，线粒体 活性分析和行为分析，以确定这些基因产物以及选择的作用。 12例具有线粒体健康和神经元健康。

PROJECT SUMMARY/ABSTRACT Calcium is a critical mediator of many cellular processes, including muscle contraction, mitochondrial activity, transcription, cell division, and synaptic vesicle release. Paradoxically, calcium can also trigger cell death and cellular necrosis. Therefore, dysregulation in calcium signaling can disrupt cellular homeostasis. Consequently, calcium levels need to be tightly controlled. Indeed, defective calcium signaling has been implicated in many neurodegenerative diseases, muscular dystrophies and heart disease. To understand the mechanisms regulating calcium signaling and how defects in this process can lead to cellular dysfunction, we are exploiting the model system Caenorhabditis elegans to identify critical processes that are involved in the regulation of calcium handling. We have recently demonstrated that SEL-12, the C. elegans presenilin ortholog, has a role in mediating endoplasmic reticulum-mitochondrial calcium homeostasis. Disruption of presenilin function results in an increase in mitochondrial calcium levels and alters mitochondrial metabolism promoting protein homeostasis collapse and neurodegeneration. Presenilin is a highly conserved protein found from plants to humans that is extensively found on endomembrane structures (e.g., endoplasmic reticulum and lysosome) of most cell types. However, the role presenilin has in the endomembrane system is not clear. Importantly, mutations in human presenilin are the most common cause of early onset familial Alzheimer's disease. Despite the identification of the involvement of presenilin in Alzheimer's disease over 20 years ago, the functional consequences of mutations in presenilin causing Alzheimer's disease are not understood. To gain further insight into the role presenilin has in mitochondrial calcium homeostasis and neuronal fitness, we have developed a novel and highly selective RNA interference screen to identify gene products that meditate the elevated endoplasmic reticulum to mitochondrial calcium signaling observed in sel-12 mutants. From this screen, we have identified several proteins known to mediate endoplasmic reticulum calcium release and mitochondrial calcium uptake but we also identified several gene products with an uncharacterized role in endoplasmic reticulum and mitochondrial calcium signaling. We propose to utilize a multifaceted approach that combines genetic manipulation, high resolution live cell microscopy to analyze endoplasmic reticulum and mitochondrial dynamics, mitochondrial activity assays, and behavioral assays to determine the role these gene products as well as sel- 12 have in mitochondrial health and neuronal fitness.