Regulation of Erythrocyte Volume Homeostasis

红细胞容量稳态的调节

• 批准号: 9382408
• 负责人: PATRICK G GALLAGHER
• 金额: $ 42.96万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-15 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9382408
• 关键词: Address; Adverse effects; Affect; Anemia; Animal Model; Biological; Biological Assay; Calcium; Cations; Cell Count; Cell model; Cell physiology; Cells; Clinical; Critical Pathways; Data; Dehydration; Disease; Electrophysiology (science); Erythrocyte Aging; Erythrocyte Survival; Erythrocyte volume; Erythrocytes; Erythroid Cells; Esthesia; Exhibits; Genes; Genetic; Genetic study; Genomics; Goals; Hemolytic Anemia; Hereditary Spherocytosis; Homeostasis; Hydration status; Inherited; Knock-in; Knock-in Mouse; Knowledge; Laboratories; Lead; Link; Maintenance; Malaria; Mediating; Membrane Proteins; Molecular; Mus; Mutation; Names; Pathway interactions; Patients; Permeability; Phenotype; Physiological; Play; Process; Proteins; Proteomics; Regulation; Role; Severity of illness; Sickle Cell; Sickle Cell Anemia; Sodium Chloride; Stress; Stretching; Structure; Syndrome; Technology; Thalassemia; Variant; Water; Work; base; beta Thalassemia; cell type; clinical phenotype; cohort; gain of function; gain of function mutation; genetic variant; human disease; improved; in vivo; in vivo Model; indexing; innovation; loss of function; mouse model; multidisciplinary; mutant; novel; protein expression; rare variant; solute; trafficking

PROJECT SUMMARY/ABSTRACT PIEZO1 has recently been identified as the long sought after protein involved in mammalian mechano- sensation and stretch-activated cation channel activation. We have discovered mutations in PIEZO1 lead to hereditary xerocytosis, a hemolytic anemia characterized by primary erythrocyte dehydration, indicating PIEZO1 plays a critical role in cellular volume homeostasis. PIEZO1 is a candidate for the unidentified stretch- induced calcium-activated cation pathways in the red blood cell that play critical roles in erythrocyte aging, malaria invasion, and circulatory sheer stress. Preliminary data also indicate that PIEZO1 is an excellent candidate for Psickle, an unidentified cation permeability pathway induced by deoxygenation in sickle erythrocytes, at the initiation of the dehydration cascade. Therefore, Psickle is of fundamental importance to sickle cell pathobiology. Despite its importance, we have no knowledge of the mechanisms controlling PIEZO1 expression, structure, or function in erythroid cells. The overall goal of this proposal is to begin to elucidate the molecular mechanisms involved in PIEZO1 regulation and function in erythroid cells. Our preliminary studies indicate that variants of the PIEZO1 gene occur in hereditary xerocytosis and sickle cell disease patients and that these variants are associated with alterations in erythrocyte hydration. The goal of aim one is identification of genetic variants influencing erythrocyte hydration in patients with sickle cell disease and increased numbers of dense cells and characterization of the effect of these mutations on PIEZO1 expression, structure and function. The goal of aim two is to create a murine model of hereditary xerocytosis and analyze the influence of HX-associated Piezo1 gain of function mutations in vivo on wild type and sickle cell backgrounds. The goal of aim 3 is the characterization of the influence of Piezo1 loss of function on erythrocyte function in wild type and SCD erythrocytes to better understand molecular mechanisms regulating erythrocyte volume homeostasis, to address the hypothesis that PIEZO1 mediates the cation currents of Psickle, the unidentified, incipient transport pathway critical for dehydration of sickle erythrocytes, and to assess the influence of Piezo1 deficiency on parameters of sickle cell disease. To address the influence of mutations on PIEZO1 structure and function, functional cell-based assays of PIEZO1 membrane protein expression, trafficking, and electrophysiology in a novel, in vivo stably-transfected, single-copy, inducible cell model of PIEZO1 expression will be performed. Physiologic studies of mature erythrocytes from genetically modified mice and affected patients will be performed under a variety of cellular conditions. PIEZO1 is expressed in many cell types, indicating PIEZO1 likely mediates important functions in a wide variety of cells. Thus studies in erythroid cells may yield mechanistic or biological principles generalizable to many critical cellular processes or human diseases.

项目总结/摘要 PIEZO 1最近已被鉴定为长期寻求的蛋白质，其参与哺乳动物的机械- 感觉和牵张激活阳离子通道激活。我们已经发现PIEZO 1的突变导致 遗传性干细胞症，一种以原发性红细胞脱水为特征的溶血性贫血，表明 PIEZO 1在细胞体积稳态中起关键作用。PIEZO 1是一个候选人的身份不明的延伸- 在红细胞中诱导钙激活阳离子途径，在红细胞老化中起关键作用， 疟疾侵袭和循环系统的绝对压力初步数据还表明，PIEZO 1是一种优秀的 Psickle的候选者，一种由Sickle中脱氧诱导的未鉴定阳离子渗透途径 红细胞，在脱水级联反应的开始。因此，Psickle对于 镰状细胞病理学尽管它的重要性，我们没有知识的机制控制PIEZO 1 在红系细胞中的表达、结构或功能。本提案的总体目标是开始阐明 参与PIEZO 1在红系细胞中的调节和功能的分子机制。我们的初步研究 表明PIEZO 1基因变体出现在遗传性干细胞症和镰状细胞病患者中， 这些变异与红细胞水合作用的改变有关。目标一的目标是认同 影响镰状细胞病患者红细胞水合作用的遗传变异， 和表征这些突变对PIEZO 1表达、结构和功能的影响。 功能目的二是建立遗传性干细胞症的小鼠模型，分析 在野生型和镰状细胞背景下，HX相关的Piezo 1在体内获得功能突变。的目标 目的3是表征野生型中Piezo 1功能丧失对红细胞功能的影响， SCD红细胞，以更好地了解调节红细胞体积稳态的分子机制， 解决PIEZO 1介导Psickle阳离子电流的假设， Piezo 1缺乏对镰状红细胞脱水至关重要的途径，并评估Piezo 1缺乏对 镰状细胞病的参数。为了解决突变对PIEZO 1结构和功能的影响， PIEZO 1膜蛋白表达，运输和电生理学的功能性细胞为基础的测定， 将进行PIEZO 1表达的新型体内稳定转染的单拷贝诱导型细胞模型。 将对来自转基因小鼠和受影响患者的成熟红细胞进行生理学研究。 在各种细胞条件下进行。PIEZO 1在许多细胞类型中表达，表明PIEZO 1 可能在多种细胞中调节重要功能。因此，在红系细胞中的研究可能会产生 可推广到许多关键细胞过程或人类疾病的机械或生物学原理。