Role of cytoskeletal proteins phosphorylation on their cellular functions.

蛋白质的磷酸化和去磷酸化动态地改变了蛋白质的结构和功能，被认为是调节各种生命现象的一种开/关开关。如果蛋白质的磷酸化和去磷酸化能够可视化，特别是从每个反应的分布、位置和转变方面来看，那么目前只能通过间接手段研究的相关和各种重要现象可能会更有启示意义。基于这一概念，我们制备了一组抗体，可以识别细胞骨架蛋白是磷酸化还是去磷酸化。我们制备了四种抗体，分别区分星形胶质细胞胶质丝构成的胶质纤维酸性蛋白（GFAP）中Thr7、Ser8、Ser13和Ser34的磷酸化状态。免疫荧光显微镜显示，当星形胶质细胞进入有丝分裂时，所有细胞质胶质丝中的Ser8残基最初被磷酸化。在细胞分裂过程中，磷酸丝氨酸8残基被去磷酸化，而丝氨酸裂解沟处的Thr7、Ser13和Ser34成为磷酸化的首选位点。在这种情况下，至少有两种不同类型的蛋白激酶在有丝分裂过程中以不同的时间表作为有丝分裂胶质丝激酶。一种磷酸酶(s)作为有丝分裂胶质丝磷酸酶，在过渡后期。

Phosphorylation and dephosphorylation of proteins dynamically alter the structure and function of proteins and are regarded as a form of an on/off switch regulating various vital phenomena. If the phosphorylation and dephosphorylation of proteins could be visualized, especially in terms of the distribution, site and transition of each reaction, the relevant and various vital phenomena, that currently can be investigated merily by indirect means, might be more revealing. Based on this concept, we prepared a group of antibodies that can identify whether a cytoskeletal protein is phosphorylated or dephosphorylated.We developed four antibodies which distinguish phosphorylated states of Thr7, Ser8, Ser13 and Ser34 in glial fibrillary acidic protein (GFAP), a protein constituted of glial filaments of astroglial cells. Immunofluorescence microscopy shows that Ser8 residues in all cytoplasmic glial filaments are initially phosphorylated when the astroglial cells enter mitosis. In cytokinesis, the phospho-Ser8 residues become dephosphorylated, whereas Thr7, Ser13 and Ser34 in the filaments at the cleavage furrow become the preferred sites of phosphorylation.Such being the case, at least two different types of protein kinases act as mitotic glial filament kinases, at a different time schedule during mitosis. A phosphatase (s) acts as a mitotic glial filament phosphatase, at the time of meta-anaphase transition.