FUNCTIONAL STUDIES OF THE NA,K-ATPASE

NA,K-ATP酶的功能研究

• 批准号: 6476756
• 负责人: JERRY B LINGREL
• 金额: $ 38.25万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-01-01 至 2004-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6476756
• 关键词: animal breeding; animal infant mortality; binding sites; cardiac glycosides; chemotherapy; chimeric proteins; drug adverse effect; embryonic stem cell; enzyme activity; enzyme mechanism; gene targeting; genetically modified animals; heart function; histology; immunocytochemistry; isozymes; laboratory mouse; muscle function; protein structure function; sodium potassium exchanging ATPase; tissue /cell culture; tissue /cell preparation

DESCRIPTION (Verbatim from the Applicant's Abstract): Our laboratory is interested in various aspects of the Na,K-ATPase, including structure-function relationships and understanding the regulated expression of the isoforms of the alpha and beta subunits. We are especially interested in the physiological role this enzyme plays and have recently developed animals lacking the alpha1 and alpha2 isoforms. Of particular significance from these studies is the finding that each isoform plays a different role in heart and skeletal muscle contraction. We wish to follow up on these studies as well as pursue others that are related to the in vivo role of this enzyme. The specific aims of our grant are to use a replacement gene targeting approach to determine whether the cardiac glycoside binding site of the Na,K-ATPase plays a biological role. Such studies will help define whether natural ligands exist for this enzyme and the role that they might play. In our second aim, we will use these animals to define the role of individual alpha isoforms in mediating the positive and negative effects of cardiac glycoside therapy. A third aim relates to understanding the physiological role of each of the alpha isoforms using chimeric analysis and tissue-specific knockouts. Animals lacking the alpha2 isoform fail to survive past birth and these animals provide an opportunity for understanding the specific biological niche that this isoform plays. In order to more fully define the tissue responsile for the lethal phenotype, we plan to carry out chimeric studies with embryonic stem cells which lack both copies of the alpha2 isoform and to carry out tissue-specific knockouts. These approaches will help define which organ system(s) is responsible for the failure of the animals to develop past birth. These studies will also likely provide animals with tissues lacking the alpha2 isoform and make it possible to define the role of this isoform in that tissue.

描述（逐字摘自申请人摘要）：我们的实验室是 对 Na,K-ATP 酶的各个方面感兴趣，包括结构功能 关系并理解异构体的调节表达 α和β亚基。我们对生理作用特别感兴趣 这种酶发挥作用，最近已开发出缺乏 alpha1 和 α2亚型。这些研究特别重要的是发现 每种异构体在心脏和骨骼肌中发挥不同的作用 收缩。我们希望跟进这些研究并进行其他研究 与该酶的体内作用有关。我们的具体目标 赠款将使用替代基因靶向方法来确定是否 Na,K-ATP酶的强心苷结合位点发挥着生物学作用。这样的 研究将有助于确定这种酶是否存在天然配体以及 他们可能扮演的角色。在我们的第二个目标中，我们将利用这些动物 定义个体α同种型在介导积极和消极方面的作用 强心苷治疗的负面影响。第三个目标涉及 使用以下方法了解每种 α 异构体的生理作用 嵌合分析和组织特异性敲除。缺乏α2的动物 同种型无法在出生后存活，这些动物提供了机会 了解该亚型所发挥的特定生物位。为了 为了更全面地定义对致死表型有反应的组织，我们计划 用缺乏两个拷贝的胚胎干细胞进行嵌合研究 α2亚型并进行组织特异性敲除。这些方法 将有助于确定哪个器官系统对失败负责 动物在出生后发育。这些研究也可能为动物提供 与缺乏 α2 同种型的组织一起使用，并有可能定义其作用 该同工型在该组织中的存在。