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Molecular mechanism of the initial process of neuronal aging.-perturbation in transport and polymerization-depolymerization dynamics of axonal cytoskeleton.

神经元衰老初始过程的分子机制。轴突细胞骨架运输和聚合-解聚动力学的扰动。

基本信息

批准号：
07458204
负责人：
KOMIYA Yoshiaki
金额：
$ 4.8万
依托单位：
Gunma University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

Studies on the regulatory mechanism of polymerization-depolymerization and transport of axonal neurofilament was carried out and the following new findings were obtained.1).Within 24 hours after intra-peritoneal injection of beta, beta'-iminodipropionitrile (IDPN), phosphorylation of NF-H subunit (apparent molecular weight 200,000) was transiently decreased in the rat sciatic motor axons. This is earlier than the first appearance of neurological symptoms caused by IDPN (about 3 days after injection), and than the disturbance of neurofilament transport within the motor axons of IDPN-treated rat (about 1-2 days after injection).2).When preparation was performed in the presence of phosphatase inhibitors, the solubility of NF-H after treatment with the buffer containing 1% Triton-100 was increased to about 20% of the total, compared to that in the absence of those inhibitors. In contrast NF-L subunit (apparent molecular weight 68,000) remained insoluble irrespective of the presence or absence of phosphatase inhibitors.When N-terminal domain of NF-H was phosphorylated by c-AMP-dependent protein kinase in vitro, it was solubilized together with NF-L subunit, suggesting depolymerization of whole neurofilament.When axonal neurofilaments were labeled and their transport was analyzed, solubility of NF-H was minimum at the peak of transport of labeled neurofilament protein, and maximum at the leading edge and the tail of the transport, suggesting the close association between transported form of neurofilament and solubility of NF-H.

对轴突神经丝聚合-解聚和转运的调控机制进行了研究，获得以下新发现：1）腹腔注射β，β ′-亚氨基二丙腈（IDPN）后24小时内，大鼠坐骨神经运动轴突NF-H亚单位（表观分子量200，000）的磷酸化水平短暂降低。这比IDPN引起的神经系统症状的首次出现要早（注射后约3天），然后IDPN治疗的大鼠运动轴突内的神经丝转运障碍（注射后约1-2天）。2）当在磷酸酶抑制剂存在下进行制备时，与不存在这些抑制剂的情况相比，在用含有1%Triton-100的缓冲液处理后，NF-H的溶解度增加到总溶解度的约20%。相反，NF-L亚基当NF-H的N端结构域在体外被cAMP依赖性蛋白激酶磷酸化时，它与NF-L亚基一起溶解，表明整个神经丝解聚。NF-H的溶解度在标记的神经丝蛋白转运峰处最小，在转运的前沿和尾部最大，表明神经丝蛋白的转运形式与NF-H的溶解度密切相关。