Imaging and functional regulation of microenvironment in subcellular organelle

亚细胞器微环境的成像和功能调控

基本信息

批准号：
15390064
负责人：
MATSUI Hideki
金额：
$ 9.86万
依托单位：
OKAYAMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15390064/
关键词：
Calcineurin Calpain Calpastatin Neuronal cell death Glutamate Kainate FK506 Protein therapy

项目摘要

Excessive glutamate as an excitotoxic inducer plays an important role in acute and chronic neuronal cell death including ischemia, trauma, Huntington's disease, Alzheimer's disease, and Amyotrophic Lateral Sclerosis. Despite years of investigation, the precise mechanism involved in glutamate-induced excitotoxic neuronal cell death has not been well characterized.Calcineurin(CaN) and calpain, a Ca^<2+>/calmodulin (CaM)-dependent protein phosphatase and a Ca^<2+>-dependent cysteine protease, respevtively, are thought to be two of the most important Ca^<2+>-dependent effectors involved in neuronal cell death. Although the mechanism of CaN-mediated neuronal cell death remains controversial, one proposed mechanism that calcineurin triggers dysfunction of the mitochondria in degenerative neurons has been strongly supportedOn the other hand, calpain regulates physiological signal transduction by cleaving cytoskeletal proteins, membrane proteins and enzymes and is also thought to play a critic … More al role in a form of neuronal cell death involved in the pathogenesis of several diseases such as brain injury, Alzheimer's disease and focal or global cerebral ischemia. Some substrates of calpain such as p35, a specific activator of cyclin-dependent kinase 5(Cdk5) have been suggested as key molecules in the induction of neuronal cell death.In this research, we showed that during glutamate neuroexcitotoxicity, calcineurin A(CnA) is directly cleaved by calpain in vitro and in vivo, resulting in the enzyme being converted to an active form. Mass spectrometry identified the three cleavage sites in CnA and two of them were constitutively active forms. Overexpression of the cleaved CnA induced caspase activity and neuronal cell death. Calpain inhibitors not only blocked the cleavage of calcineurin, but also protected against neuronal cell death in vitro and in vivo. These results indicate that calcineurin is a crucial target for calpain and the calpain-mediated activation of calcineurin triggers excitotoxic neurodegeneration. This further shows the direct cross talk between phosphatase and protease in pathopysiology. Less

过量的谷氨酸作为兴奋性毒性诱导剂在急性和慢性神经元细胞死亡中发挥重要作用，包括缺血、创伤、亨廷顿舞蹈病、阿尔茨海默病和肌萎缩性侧索硬化症，尽管经过多年的研究，谷氨酸诱导的兴奋性毒性神经元细胞死亡的确切机制仍然存在。尚未得到很好的表征。钙调神经磷酸酶 (CaN) 和钙蛋白酶，一种 Ca^2+/钙调蛋白 (CaM) 依赖性蛋白磷酸酶和 Ca^<2+> 依赖性半胱氨酸蛋白酶分别被认为是参与神经元细胞死亡的两个最重要的 Ca^<2+> 依赖性效应物，尽管 CaN 介导的神经元死亡机制。细胞死亡仍然存在争议，钙调神经磷酸酶触发退行性神经元线粒体功能障碍的一种机制得到了强烈支持。另一方面，钙蛋白酶通过裂解细胞骨架蛋白、膜蛋白和细胞膜蛋白来调节生理信号转导。酶，也被认为在涉及多种疾病发病机制的神经元细胞死亡中发挥重要作用，例如脑损伤、阿尔茨海默病和局灶性或整体性脑缺血。钙蛋白酶的一些底物，例如 p35，是一种酶。细胞周期蛋白依赖性激酶 5 (Cdk5) 的特异性激活剂已被认为是诱导神经元细胞死亡的关键分子。在这项研究中，我们表明，在谷氨酸神经兴奋毒性过程中，钙调神经磷酸酶 A(CnA) 在体外和体内均被钙蛋白酶直接裂解，导致该酶转化为活性形式。质谱鉴定了 CnA 中的三个裂解位点，其中两个是裂解的 CnA 的组成型活性形式。诱导的半胱天冬酶活性和神经元细胞死亡不仅可以阻断钙调神经磷酸酶的裂解，而且可以在体外和体内防止神经元细胞死亡。这些结果表明钙调磷酸酶是钙蛋白酶的重要靶标，钙调磷酸酶介导的激活会引发兴奋性神经变性，这进一步表明磷酸酶和蛋白酶之间的直接相互作用在病理生理学中较少。