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: 10542447
• 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/10542447
• 关键词: Actins; Affect; Asthma; Atrial Fibrillation; Biochemical; Biological Assay; Blood Vessels; Calcium; Calcium Signaling; Cardiac Myocytes; Cardiovascular Diseases; Cardiovascular system; Cause of Death; Cell model; Cell physiology; Cells; Cessation of life; Cytoskeleton; Data; Diabetes Mellitus; Disease; Disease Progression; Enzymes; Family; Gene Expression; Heart; Heart failure; Heme; Hemeproteins; Homeostasis; Immunity; Link; Malignant Neoplasms; Membrane; Metabolism; Mitochondria; Molecular; NADPH Oxidase; Oxidation-Reduction; Pathologic; Patients; Phase; Physiological; RNA; Regulation; Role; Signal Transduction; Sinus; Stimulus; Stroke; Superoxides; System; Testing; Tissues; United States; Work; biological adaptation to stress; cardiovascular health; drug discovery; human disease; induced pluripotent stem cell derived cardiomyocytes; knock-down; member; novel; overexpression; prevent; protein expression; protein protein interaction; reconstitution; release of sequestered calcium ion into cytoplasm; response; structural biology

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）是一种跨膜信号传导酶， 超氧化物响应细胞内钙水平升高，并正在成为一个令人兴奋的球员， 免疫力和心血管系统。NOX5对血管的正常收缩至关重要， 细胞氧化还原和钙信号之间的联系，作为最新发现的成员， NOX家族的酶，NOX 5调节的细节及其在细胞中的作用仍然没有得到很好的解决。是的， 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。 紊乱阐明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 我们对正常细胞功能的理解以及这些功能在疾病中是如何被破坏的。基于 根据K99阶段的初步数据，R00阶段将重点研究NOX 5在以下方面的作用： 心肌细胞功能及其对房颤（AF）发生和发展的作用（目的1） 并探索在K99期鉴定的新蛋白质：蛋白质相互作用的意义， NOX5和肌动蛋白细胞骨架，线粒体，RNA调节和应激反应系统（目的2）。目的 1将专注于了解NOX5敲低和过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或窦性心律的心脏患者组织来测试假设 由iPSC衍生的心肌细胞系统产生。目的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，以探测相互作用组。 NOX5对刺激的反应，并了解这些相互作用如何影响NOX5的活性， 定位、肌动蛋白细胞骨架、钙流、细胞代谢、基因表达和应激 响应系统该项目将揭示有关NOX5在心脏中的作用的关键细节， 更广泛的细胞内稳态的背景下。这也将为分子鉴定奠定重要基础 负责生理和病理反应之间的转换的因素，并确定 相互作用和相互作用网络是进一步研究的理想选择， 重组分析，结构生物学项目，使用生物化学方法的机制研究， 药物发现项目。