Developmental changes in phosphorylation state of neurofilament proteins in the chick embryonic optic nerve.
Developmental changes in phosphorylation state of neurofilament proteins in the chick embryonic optic nerve.
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DOI：
10.1016/0012-1606(89)90080-8
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发表时间：
1989-07
影响因子：
2.7
通讯作者：
M. Go;H. Tanaka;K. Obata;S. Fujita
中科院分区：
文献类型：
--
作者：
M. Go;H. Tanaka;K. Obata;S. Fujita
应用鸡神经细丝高(180K)和中(160K)高相对分子质量亚基高度磷酸化成分的单抗(MAb)82E10，对鸡胚视神经中神经丝蛋白(NFP)磷酸化状态的发育变化进行了组织化学和生化研究。用该单抗对发育中的胚胎视神经切片进行酶免疫组织化学研究。随着发育阶段的不同，染色模式也有明显的变化。在6天的胚胎(E6)中，整个横截面被染色，而在E10中，只有大约一半的横截面被染色。在E14年，几乎整个横截面区域都没有染色。此后，免疫反应性重新出现并逐渐增强，以至于在E20整个横截面再次呈免疫阳性。对不同发育阶段胚胎的视神经进行了电泳法和免疫印迹分析。在胚胎14天，随着组织化学改变的时间进程，核因子-M部位的82E10免疫反应出现一过性丢失。蛋白质染色的二维凝胶进一步表明，在胚胎14岁时，高度磷酸化的NF-M形式从胚胎视神经中暂时消失，而较低磷酸化的形式持续整个胚胎发育阶段。为了了解胚胎视神经中82E10免疫反应性与视网膜组织和时序组织的关系，采用视网膜注射荧光染料DiI作为所选纤维的顺行追踪标记。胚胎视神经内纤维排列有序，提示胚胎视神经中82E10免疫反应活性的有序丧失反映了视神经轴突的时间顺序。胚胎视神经中NFP磷酸化状态的这些变化可能反映了发育过程中特定阶段神经元细胞骨架的动态变化。
Developmental changes in the phosphorylation state of neurofilament proteins (NFPs) in the chick embryonic optic nerve were histochemically and biochemically studied using monoclonal antibody (MAb) 82E10 specific to the highly phosphorylated components of high (180K)- and middle (160K)-molecular-weight subunits of neurofilament (NF) in the chicken. Cross sections of developing embryonic optic nerve were studied by enzyme immunohistochemistry using this MAb. The staining pattern showed marked changes with the developmental stage. In 6-day embryos (E6) the entire cross section was stained, whereas in E10 only about a ventroposterior half of the cross section was stained. In E14 nearly the entire area of the cross section became unstained. Thereafter, the immunoreactivity reappeared and gradually increased, such that in E20 the entire cross section became immunopositive again. Electrophoretic and immunoblot analyses were made on optic nerves dissected out of embryos of various stages. The 82E10 immunoreactivity at the position of NF-M underwent a transient loss in E14 in parallel with the time course of histochemical change. Two-dimensional gels stained for protein further showed that the highly phosphorylated form of NF-M is transiently lost from embryonic optic nerve in E14, while the less phosphorylated form persists throughout the embryonic developmental stages. In order to understand the orderly loss of the 82E10 immunoreactivity in relation to retinotopic and chronotopic organizations of the fibers in the embryonic optic nerve, retinal injection of a fluorescent dye DiI as an anterograde tracing marker for selected fibers was utilized. An ordered arrangement of the fibers was present within the embryonic optic pathway, suggesting that the orderly loss of the 82E10 immunoreactivity in the embryonic optic nerve reflects the chronological order of the optic axons. These changes in the phosphorylation state of NFPs in the embryonic optic nerve presumably reflect dynamic changes of the neuronal cytoskeleton at certain stages during development.