Deciphering Molecular Mechanisms of Calcium Homeostasis

破译钙稳态的分子机制

• 批准号: 10406433
• 负责人: Kenneth R Norman
• 金额: $ 40.75万
• 依托单位: ALBANY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10406433
• 关键词: Affect; Alzheimer' s Disease; Behavioral Assay; Biological Assay; Biological Models; Caenorhabditis elegans; Calcium; Calcium Signaling; Cell Death; Cell division; Cell physiology; Cells; Defect; Disease; Early Onset Familial Alzheimer' s Disease; Endoplasmic Reticulum; Functional disorder; Genetic Transcription; Goals; Health; Heart Diseases; Homeostasis; Human; Lead; Lysosomes; Mediating; Mediator of activation protein; Mitochondria; Molecular; Muscle Contraction; Muscular Dystrophies; Mutation; Necrosis; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Orthologous Gene; Plants; Process; Proteins; RNA interference screen; Regulation; Research Project Grants; Resolution; Role; Structure; Synaptic Vesicles; System; age related; cell type; fitness; gene product; genetic manipulation; insight; live cell microscopy; mitochondrial metabolism; mutant; novel; presenilin; proteostasis; uptake; vesicular release

PROJECT SUMMARY 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 affect cells in different ways and to varying degrees. 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.

项目摘要 钙是许多细胞过程的关键介质，包括肌肉收缩， 线粒体活性、转录、细胞分裂和突触囊泡释放。巧合的是， 钙还可引发细胞死亡和细胞坏死。因此，钙的失调 信号传导可以以不同的方式和不同的程度影响细胞。因此，钙水平 必须严格控制。事实上，有缺陷的钙信号已经牵连在许多 神经变性疾病、肌肉萎缩症和心脏病。了解 调节钙信号传导的机制，以及这一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，以确定关键的 涉及钙处理调节的工艺。我们最近 表明SEL-12、C. elegans早老素直系同源物，在介导 内质网-线粒体钙稳态。早老素功能破坏 导致线粒体钙水平增加并改变线粒体代谢 促进蛋白质稳态崩溃和神经变性。早老素是一种 从植物到人类广泛存在于内膜上的保守蛋白质 结构（例如，内质网和溶酶体）。然而，作用 早老素在内膜系统中的作用尚不清楚。重要的是，人类中的突变 早老素是早发性家族性阿尔茨海默病的最常见原因。尽管 早老素参与阿尔茨海默病的鉴定在20多年前， 导致阿尔茨海默病的早老素突变的功能后果并不 明白为了进一步了解早老素在线粒体钙离子通道中的作用， 自稳态和神经适应，我们已经开发出一种新的和高选择性的RNA 干扰筛选以鉴定调节内质网升高的基因产物 与在SEL-12突变体中观察到的线粒体钙信号传导有关。从这个屏幕上，我们有 鉴定了几种已知介导内质网钙释放的蛋白质， 线粒体钙摄取，但我们也确定了几个基因产物， 在内质网和线粒体钙信号传导中的未表征的作用。我们提出 利用多方面的方法，结合基因操作，高分辨率活细胞， 显微镜下分析内质网和线粒体动力学，线粒体 活性测定和行为测定，以确定这些基因产物以及自噬的作用， 12在线粒体健康和神经元健康。