Intracellular Electrophysiology: An electrochemical atlas of organelles

项目总结 这项提议的长期目标是建立一个细胞器的电化学图谱，以指导 神经退行性疾病背景下细胞器间接触的合理操作。一种新的模式 在细胞器的水平上，细胞内的通信正在出现，由此两个并列的膜 细胞器通过细胞质表面的蛋白质-蛋白质相互作用而物理连接，称为 细胞器间的接触。离子和小分子活跃地从一个细胞器转移到另一个细胞器。 这些接触，穿过两个密封的膜。细胞器间的接触对细胞功能、组织至关重要 动态平衡和生理学，因为它们调节从脂肪代谢到细胞凋亡的各种过程。 然而，我们仍然不知道是什么信号启动了接触形成，也不知道是什么开启了化学传输 跨接触，我们也不能区分功能接触和功能障碍接触。因此，虽然我们 我们知道蛋白质中的特定突变会破坏接触，导致各种神经退行性疾病 仍然不知道如何恢复这些接触和治疗那些疾病。 我假设细胞器的电化学状态，单独的和接触的，将通知哪些途径和 分子应该有针对性地纠正疾病状态下的异常接触。我的理由是，如果我们抽象 在分子细节上，细胞器间的接触类似于神经元突触。即使在突触中，离子也会继续流动 穿过两个密封的、毗邻的膜。就像跨神经元膜的离子传输机制一样 由Hodgkin和Huxley的电化学模型揭示，这是细胞器膜的一个类似模型 将揭示跨细胞器间接触的离子流动机制以及哪些特定流动在疾病中受到影响。 我建议建立一个细胞器的电化学图谱，作为研究健康和健康中接触的普遍参考 疾病。本图集将是包括主要细胞器的电化学模型的方程式的概要， 独自一人和有联系的人。通过使我们能够区分正常和异常接触者，我预计地图集将揭示 跨越疾病的共同途径，可以针对这些途径恢复与受影响离子流的接触。 无法测定细胞器中的离子或电压，阻碍了电化学的发展。 它们的膜的模型。在过去的十年里，我的实验室开发了一个化学平台来量化离子和 细胞器中的电压。通过将电生理学整合到这个平台上，我建议现在制定出 细胞膜在隔离和接触状态下的电化学性质，并使电化学反应 细胞器图集。我们将应用图谱来阐明钙离子是如何通过异常接触的 内质网和溶酶体可被纠正以恢复帕金森病患者的溶酶体钙离子。 在许多神经退行性疾病中，调节失调的溶酶体钙离子是一个共同的因素，而 电化学图谱是它的开创性能力，它揭示了可以作为治疗靶点的常见途径 疾病交叉的方式。

PROJECT SUMMARY The long-term objective of this proposal is to build an electrochemical atlas of organelles to guide the rational manipulation of inter-organelle contacts in the context of neurodegenerative diseases. A new mode of intracellular communication is emerging at the level of organelles, whereby the membranes of two juxtaposed organelles are physically connected, via protein-protein interactions on their cytoplasmic faces, referred to as inter-organelle contacts. Ions and small molecules are actively transferred from one organelle to the other across these contacts, traversing two sealed membranes. Inter-organelle contacts are vital to cell function, tissue homeostasis and physiology because they regulate processes ranging from lipid metabolism to apoptosis. However, we still do not know what signals initiate contact formation or what switches on chemical transport across contacts, nor can we discriminate between functional and dysfunctional contacts. Hence, although we know of specific mutations in proteins that disrupt contact, leading to diverse neurodegenerative diseases, we still do not know how to restore these contacts and treat those diseases. I posit that the electrochemical states of organelles, alone and in contact, will inform which pathways and molecules should be targeted to rectify aberrant contacts in disease states. My rationale is that, if we abstract out the molecular details, inter-organelle contacts resemble neuronal synapses. Even in synapses, ions flow on cue across two sealed, abutting membranes. Just as ion-transport mechanisms across neuronal membranes were revealed by Hodgkin and Huxley’s electrochemical model, an analogous model of organelle membranes will reveal ion flow mechanisms across inter-organelle contacts and which specific flows are impacted in disease. I propose to build an electrochemical atlas of organelles as a universal reference to study contacts in health and disease. This atlas will be a compendium of equations comprising electrochemical models of major organelles, alone and in contact. By enabling us to discriminate normal and aberrant contacts, I envisage the atlas will reveal common pathways across diseases that can be targeted to restore contacts with impacted ion flows. The inability to assay ions or voltage in organelles has prevented the development of electrochemical models of their membranes. Over the last decade, my lab developed a chemical platform to quantify ions and voltage in organelles. By integrating electrophysiology to this platform, I propose to now map out the electrochemical characteristics of organelle membranes in isolation and in contact, and make an electrochemical atlas of organelles. We will apply the atlas to elucidate how Ca2+ flow across aberrant contacts between the endoplasmic reticulum and the lysosome can be rectified to restore lysosomal Ca2+ in parkinsonism. Dysregulated lysosomal Ca2+ is a common factor across many neurodegenerative diseases and the value of the electrochemical atlas is its pioneering ability to reveal common pathways that can be targeted for treatment in a disease cross-cutting manner.