Role of cytoskeletal proteins phosphorylation on their cellular functions.

细胞骨架蛋白磷酸化对其细胞功能的作用。

• 批准号: 04454177
• 负责人: INAGAKI Masaki
• 金额: $ 4.22万
• 依托单位: Tokyo Metropolitan Institute of Gerontology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454177/
• 关键词: Intermediate filament; Phosphorylation; Mitosis; Phosphorylation site-specific antibodies; suc1蛋白質; ポリホスホイノシタイド; ポリホスホイノシタイド結合蛋白質

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.

蛋白质的磷酸化和去磷酸化动态地改变蛋白质的结构和功能，并且被认为是调节各种生命现象的开/关开关的形式。如果蛋白质的磷酸化和去磷酸化过程能够被可视化，特别是在每个反应的分布、位点和转变方面，那么目前可以通过间接手段来研究的相关的和各种各样的生命现象可能会更有启发性。基于这一概念，我们制备了一组可以识别细胞骨架蛋白磷酸化或去磷酸化的抗体，我们开发了四种抗体，可以区分胶质细胞酸性蛋白（GFAP）中Thr 7，Ser 8，Ser 13和Ser 34的磷酸化状态，GFAP是由星形胶质细胞的胶质纤维构成的蛋白质。免疫荧光显微镜显示，Ser 8残基在所有的细胞质胶质纤维最初磷酸化时，星形胶质细胞进入有丝分裂。在胞质分裂中，磷酸-Ser 8残基被去磷酸化，而在分裂沟处的丝中的Thr 7、Ser 13和Ser 34成为优选的磷酸化位点。磷酸酶在中期-后期转换时作为有丝分裂胶质丝磷酸酶发挥作用。

项目成果

Matsuoka,Y.,Inagaki,M.et al.: "Two different protein kinases act on a different time schedule as glial filament kinase during mitosis" EMBO Journal. 11. 2895-2902 (1992)

Matsuoka,Y.,Inagaki,M.等人：“两种不同的蛋白激酶在有丝分裂过程中以不同的时间安排发挥作用，就像神经胶质丝激酶一样”EMBO 杂志。

Krebs, E.G.and Beavo, J.A.: "Phosphorylation-dephosphorylation of enzymes." Annu. Rev. Biochem.48. 831-959 (1979)

Krebs, E.G. 和 Beavo, J.A.：“酶的磷酸化-去磷酸化。”

Furuhashi,K.,Inagaki,M.et al.: "Inositol phospholipidsーinduced suppression of Fーactin gelating activity of smooth muscle filamin." Biochem.Biophys.Res.Commun.184. 1261-1265 (1992)

Furuhashi, K., Inagaki, M. 等人：“肌醇磷脂诱导的平滑肌丝蛋白胶凝活性的抑制。”Biochem.Biophys.Res.Commun.184 (1992)。

Greengard, P.: "Phosphorylated proteins as physiological effectors : protein phosphorylation may be a final common pathway for many biological regulatory agents." Science. 199. 146-152 (1978)

Greengard, P.：“磷酸化蛋白质作为生理效应器：蛋白质磷酸化可能是许多生物调节剂的最终共同途径。”

Furuhashi,K.,Inagaki,M.et al.: "Phosphorylation by actin kinase of the pointed endーdomain on the actin molecule." The Journal of Biological Chemistry. 267. 9326-9330 (1992)

Furuhashi, K., Inagaki, M. 等人：“肌动蛋白分子上尖端结构域的磷酸化”，《生物化学杂志》267. 9326-9330 (1992)。