Heme allocation and disruptions in asthma and the failing heart

哮喘和心脏衰竭中的血红素分配和干扰

• 批准号: 10516435
• 负责人: Elizabeth A. Sweeny
• 金额: $ 24.9万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10516435
• 关键词: Heme; allocation; disruptions; asthma; failing

The heme protein NADPH oxidase 5 (NOX5) is a transmembrane signaling enzyme which produces superoxide in response to elevated intracellular calcium levels and is emerging as an exciting player in immunity and the cardiovascular system. NOX5 is crucial for proper vascular contraction and appears to be a nexus between cellular redox and calcium signaling, As the most recently discovered member of the NOX family of enzymes, details of NOX5 regulation and its role in the cell remain poorly resolved. It has, however, been implicated in numerous human diseases including cancers, diabetes and cardiovascular disorders. Elucidating details of NOX5 regulation and its role in cardiovascular health and disease is crucial to our understanding of normal cellular functions and how these become disrupted in disease. Based on preliminary data from the K99 phase, the R00 phase will focus on investigating the role of NOX5 in cardiomyocyte function and its contribution to the initiation and progression of atrial fibrillation (AF) (Aim 1) and to probe the significance of novel protein:protein interactions identified in the K99 phase which link NOX5 and the actin cytoskeleton, mitochondria, RNA regulation and stress response systems (Aim 2). Aim 1 will focus on understanding how NOX5 knockdown and overexpression affect gene and protein expression, calcium flux, and cellular metabolism in induced pluripotent stem cell (iPSC) derived cardiomyocytes, and then using patient tissue from hearts in AF or sinus rhythm to test hypothesizes generated from the iPSC derived cardiomyocyte system. Aim 2 will use a model cell system (HEK293 cells and HEK293 cells overexpressing NOX5) as well as iPSC derived cardiomyocytes to probe the interactome of NOX5 in response to stimuli and to understand how these interactions affect NOX5 activity and localization, the actin cytoskeleton, calcium flux, cellular metabolism, gene expression and the stress response system. This project will uncover crucial details about the role of NOX5 in the heart and in the broader context of cellular homeostasis. It will also lay important groundwork for identifying molecular factors responsible for the switch between physiological and pathological responses and identify interactions and interaction networks ideal for further study using purified components for use in reconstitution assays, structural biology projects, mechanistic studies using biochemical approaches and drug discovery projects.

血红素蛋白 NADPH 氧化酶 5 (NOX5) 是一种跨膜信号酶，可产生 超氧化物对细胞内钙水平升高做出反应，并正在成为一个令人兴奋的参与者 免疫力和心血管系统。 NOX5 对于适当的血管收缩至关重要，并且似乎 细胞氧化还原和钙信号传导之间的联系，作为最近发现的成员 NOX 酶家族、NOX5 调节细节及其在细胞中的作用仍不清楚。它有， 然而，它与许多人类疾病有关，包括癌症、糖尿病和心血管疾病 失调。阐明 NOX5 调节的细节及其在心血管健康和疾病中的作用至关重要 帮助我们了解正常细胞功能以及这些功能如何在疾病中受到破坏。基于 从 K99 阶段的初步数据来看，R00 阶段将重点研究 NOX5 在 心肌细胞功能及其对心房颤动 (AF) 发生和进展的贡献（目标 1） 并探讨新蛋白质的重要​​性：在 K99 相中确定的蛋白质相互作用将 NOX5 和肌动蛋白细胞骨架、线粒体、RNA 调节和应激反应系统（目标 2）。目的 1 将重点了解 NOX5 敲低和过度表达如何影响基因和蛋白质 诱导多能干细胞 (iPSC) 衍生的表达、钙通量和细胞代谢 心肌细胞，然后使用 AF 或窦性心律患者心脏组织来检验假设 由 iPSC 衍生的心肌细胞系统产生。目标 2 将使用模型细胞系统（HEK293 细胞 和过表达 NOX5 的 HEK293 细胞以及 iPSC 衍生的心肌细胞来探测相互作用组 NOX5 对刺激的反应并了解这些相互作用如何影响 NOX5 活性 定位、肌动蛋白细胞骨架、钙通量、细胞代谢、基因表达和应激 响应系统。该项目将揭示 NOX5 在心脏和身体中的作用的重要细节 细胞稳态的更广泛背景。它还将为识别分子奠定重要基础。 负责生理和病理反应之间转换的因素并确定 相互作用和相互作用网络非常适合使用纯化的成分进行进一步研究 重构测定、结构生物学项目、使用生化方法的机制研究和 药物发现项目。