Role of calmodulin kinases on the regulation of Ca^<2+> signaling of central nervous systems

钙调蛋白激酶对中枢神经系统Ca^2信号传导调节的作用

基本信息

批准号：
10470484
负责人：
YAMAUCHI Takashi
金额：
$ 8.51万
依托单位：
The University of Tokushima
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2001
项目状态：
已结题

项目摘要

Neuronal Ca^<2+> /calmodulin-dependent protein kinase II (CaM kinase II) regulates important neuronal functions, including neurotransmitter synthesis and neurotransmitter release, modulation of ion channel activity, synaptic plasticity, and gene expression. The following results were obtained in this study; 1. when CaM kinase II was introduced in neuronal cells, neurite outgrowth was stimulated, indicating that CaM kinase II played a role stimulation of neurite outgrowth; 2. Ca^<2+> independent activity of the kinase autophosphorylated at Thr-286 involved in neurite outgrowth; 3. more than 28 protein substrates of CaM kinase II were found in the postsynaptic density (PSD), and almost all of these proteins were identified by protein sequencing and mass spectrometry; 4. when the developmental change of the kinase was investigated during the neural differentiation of cultured cells, it was found that CaM kinase II was upregulated during the neuronal differentiation; 5. cell type distinctive changes of splicing pattern of δ isoform were found not only during development of rat brain tissue, but also during differentiation of cultured neuronal cells, indicating that alternatively spliced variants of δ isoform of CaM kinase II were expressed during neural differentiation; 6. when we used the deletion mutants for β-specific insertions to explore the difference between α and β CaM kinase II in enzymatic properties, it was found that β specific insertion of β CaM kinase II played an important role in the cellular distribution of the kinase; 7 The gene encoding the β isoform of rat Ca^<2+>/calmodulin-dependent protein kinase II was cloned, and its structure and exon-intron organization were determined; 8. When the promoter activity of the gene was analyzed using nueronal and non-neuronal cells, neuronal cell type-specific promoter activity was found in the 5'-upstream region of the gene.

神经元Ca^2+/钙调蛋白依赖性蛋白激酶II(CaM激酶II)调节重要的神经元功能，包括神经递质合成和神经递质释放、离子通道活性的调节、突触可塑性和基因表达。本研究；1.当CaM激酶II被引入神经元细胞时，神经突生长受到刺激，表明CaM激酶II起到刺激神经突生长的作用；参与神经突生长的Thr-286处的激酶自磷酸化的Ca^2+独立活性；3.在突触后密度(PSD)中发现了超过28种CaM激酶II的蛋白质底物，并且几乎所有这些蛋白质均被鉴定。 4.通过蛋白测序和质谱分析培养细胞神经分化过程中激酶的发育变化，发现CaM激酶II在神经元分化过程中表达上调；不仅在大鼠脑组织的发育过程中，而且在培养的神经元细胞的分化过程中都发现了 δ 同工型剪接模式的独特变化，表明 CaM 激酶 II 的 δ 同工型的选择性剪接变体在神经分化过程中表达 6；当我们使用β特异性插入的缺失突变体来探索α和β CaM激酶II在酶学特性上的差异时，发现β CaM激酶II的β特异性插入在细胞分布中发挥着重要作用。 7 克隆编码大鼠Ca^2+/钙调蛋白依赖性蛋白激酶IIβ亚型的基因，并确定其结构和外显子-内含子组织； 8.利用神经元和内切酶分析该基因的启动子活性；在非神经元细胞中，在该基因的 5'-上游区域发现了神经元细胞类型特异性启动子活性。