Regulation and localization of CAS substrate domain tyrosine phosphorylation
Regulation and localization of CAS substrate domain tyrosine phosphorylation
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DOI：
10.1016/j.cellsig.2003.10.004
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发表时间：
2004-05-01
影响因子：
4.8
通讯作者：
Hanks, SK
中科院分区：
文献类型：
--
作者：
Fonseca, PM;Shin, NY;Hanks, SK
Crk 相关底物 (CAS) 是一种酪氨酸激酶底物，参与整合素对细胞行为的控制。 Src 家族激酶对 CAS 底物结构域 (SD) 中多个酪氨酸残基的磷酸化是促进细胞运动的主要整合素信号转导事件。在本研究中，对针对 CAS SD 磷酸酪氨酸位点的新型磷酸特异性抗体（“pCAS”抗体）进行了表征，并用于研究 CAS SD 酪氨酸磷酸化的细胞调节和定位。对分别在 CAS 和 FAK 缺陷细胞系中表达的 CAS 和粘着斑激酶 (FAK) 变体的分析表明，CAS SD 酪氨酸磷酸化基本上是由 Src 家族激酶通过两种不同的机制与 CAS 结合来实现的：直接结合到 CAS Src 结合域和通过 FAK 桥间接结合。使用 pCAS 抗体进行的细胞免疫染色显示，CAS SD 酪氨酸磷酸化仅发生在整联蛋白粘附位点，包括在延伸片状伪足边缘形成的新生焦点复合物以及细胞体下方的成熟焦点粘附。这些发现进一步证明了 FAK 作为 CAS SD 酪氨酸磷酸化的重要上游调节因子的作用，并暗示 CAS 介导的信号传导事件在细胞运动过程中促进膜突出/片状足延伸。 (C) 2003 Elsevier Inc. 保留所有权利。
Crk-associated substrate (CAS) is a tyrosine kinase substrate implicated in integrin control of cell behavior. Phosphorylation, by Src family kinases, of multiple tyrosine residues in the CAS substrate domain (SD) is a major integrin signaling event that promotes cell motility. In this study, novel phosphospecific antibodies directed against CAS SD phosphotyrosine sites ("pCAS" antibodies) were characterized and employed to investigate the cellular regulation and localization of CAS SD tyrosine phosphorylation. An analysis of CAS and focal adhesion kinase (FAK) variants expressed in CAS- and FAK-deficient cell lines, respectively, indicated that CAS SD tyrosine phosphorylation is substantially achieved by Src family kinases brought into association with CAS through two distinct mechanisms: direct binding to the CAS Src-binding domain and indirect association through a FAK bridge. Cell immumostaining with pCAS antibodies revealed that CAS SD tyrosine phosphorylation occurs exclusively at sites of integrin adhesion including both nascent focal complexes formed at the edges of extending lamellipodia as well as mature focal adhesions underlying the cell body. These findings further document a role for FAK as an important upstream regulator of CAS SD tyrosine phosphorylation and implicate CAS-mediated signaling events in promoting membrane protrusion/lamellipodium extension during cell motility. (C) 2003 Elsevier Inc. All rights reserved.