TRANSGENIC MICE WITH ALTERED CALCIUM HANDLING

钙处理能力改变的转基因小鼠

• 批准号: 6188439
• 负责人: Evangelia G Kranias
• 金额: $ 35.6万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6188439
• 关键词: animal breeding; animal colony; calcium metabolism; calcium transporting ATPase; disease /disorder model; gene expression; gene targeting; genetic promoter element; genetically modified animals; laboratory mouse; model design /development; mutant; phospholamban; phosphorylation; polymerase chain reaction; sarcoplasmic reticulum; second messengers; smooth muscle

The sarcoplasmic recticulum (SR) is an internal membrane system in muscle, which functions as a Ca2+-sink during relaxation as as a Ca2+-source during contraction. Relaxation is mediated by the transport of Ca2+into the SR lumen by the Ca2+-ATPase (SERCA2), which is under regulation by phospholamban (PLB) in cardiac, slow-twitch skeletal and smooth muscles. Dephosphorylated PLB is an inhibitor of the affinity of the SR Ca2+-pump for Ca2+and phosphorylation relieves this inhibition. Alterations is in the expression levels of PLB or the SR Ca2+-ATPasehave been linked to altered Ca2+ homeostasis and deterioration of cellular function in several diseases. While transgenic mice have been rccently generated, which elucidated the functional role of altered PLB expression in vivo, focused on cardiac muscle and the physiological significance of PLB in other muscle and non-muscle tissues is not well understood. Thus, the objectives of the present proposal are to generate mouse models, with altered expressionof PLB or the SR Ca2+-ATPase to better define the function of each of these two key Ca2+-handling protein in vivo. Specifically, we will generate mice: a) overexpressing PLB and its phosphorylation mutants in either smooth or soleus muscle. Studies in these models coupled with studies in the PLB knockout mouse will elucidate the functional role of PLB in smooth and soleus muscles and define the second messanger pathways regulating these muscles through phosphorylation of PLB; b) overexpressing PLB in multiple tissues and under the control of an inducible promoter to achieve tight temporal and quantitative control of PLB expression in a reversible manner. These models will permit evaluation of the role of temporal alterations in PLB expression levels on cellular function; and c) overexpressing each of the SERCA2 isoforms (SERCA2a or SERCA2b) or conditionally ablating SERCA2 expression in a tissue specific manner. The models with altered SERCA2 expression levels will elucidate the role of this protein in the intact animal. Overall, the proposed animal models will provide valuable and unique systems for the biomedical community at large to carry out further studies on elucidating the functional role of PLB and SERCA2 in intracellular calcium handling in health and disease.

肌质晶状体（SR）是一种内部膜系统 肌肉，在放松期间用作Ca2+-sink的肌肉 收缩期间CA2+-SOURCE。 放松是由运输介导的 Ca2+通过Ca2+-ATPase（SERCA2）的Ca2+of of ca2+ 在心脏，慢速骨骼和 平滑的肌肉。 去磷酸化的PLB是亲和力的抑制剂 Ca2+和磷酸化的SR Ca2+泵可以缓解这种抑制作用。 变化在PLB或SR Ca2+-atpaseHave的表达水平中 与CA2+稳态改变和细胞恶化有关 在多种疾病中起作用。 虽然转基因小鼠已经被摩擦 生成的，阐明了PLB改变的功能作用 体内表达，专注于心肌和生理 PLB在其他肌肉和非肌肉组织中的重要性不好 理解。 因此，本提议的目标是 生成鼠标模型，具有PLB或SR的表达式变化 Ca2+-ATPase更好地定义了这两个键的功能 Ca2+ - 在体内进行蛋白质。 具体来说，我们将生成小鼠：a） 过表达PLB及其磷酸化突变体的光滑或 比目鱼肌。 这些模型的研究与PLB中的研究结合 敲除小鼠将阐明PLB在平滑和 比目鱼肌肉并定义了第二个Messanger途径来调节这些 通过PLB的磷酸化肌肉； b）过表达PLB 多个组织并在诱导型启动子的控制下 在A中实现PLB表达的紧密时间和定量控制 可逆的方式。 这些模型将允许评估 PLB表达水平的时间改变在细胞功能上；和 c）过表达每个SERCA2同工型（SERCA2A或SERCA2B）或 有条件地以特定于组织的方式烧毁SERCA2表达。 SERCA2表达水平改变的模型将阐明角色 完整动物中的蛋白质。 总体而言，提议的动物 模型将为生物医学提供宝贵而独特的系统 整个社区，以进一步阐明 PLB和SERCA2在细胞内钙处理中的功能作用 健康与疾病。