Binding of 14-3-3β regulates the kinase activity and subcellular localization of testicular protein kinase 1
Binding of 14-3-3β regulates the kinase activity and subcellular localization of testicular protein kinase 1
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DOI：
10.1074/jbc.m104620200
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发表时间：
2001-11-16
影响因子：
4.8
通讯作者：
Mizuno, K
中科院分区：
文献类型：
--
作者：
Toshima, JY;Toshima, J;Mizuno, K
睾丸蛋白激酶1 （TESK1）是一种丝氨酸/苏氨酸激酶，可磷酸化cofilin并诱导肌动蛋白细胞骨架重组。整合素介导的信号通路可刺激TESK1的激酶活性，但其调控机制尚不清楚。利用酵母双杂交系统鉴定出14-3-3 β是TESK1的结合蛋白。在体外和体内共沉淀试验中，TESK1和14-3-3 β之间的特异性相互作用变得明显。14-3-3 β通过TESK1的c端区域与TESK1相互作用，其作用方式依赖于RXXSXP基序中Ser-439的磷酸化。14-3-3 β结合抑制TESK1的激酶活性。在细胞在纤维连接蛋白上扩散的过程中，TESK1/14-3-3 β相互作用显著降低，其时间过程与TESK1激酶活性的增加呈负相关。因此，14-3-3 β与TESK1/14-3-3 β复合物的分离可能参与了整合素介导的TESK1激活。在HeLa细胞中，当细胞涂有纤维连接蛋白时，TESK1和14-3-3 β聚集在细胞周围，而在未受刺激的细胞中，它们弥散分布在细胞质中。我们认为14-3-3 β在整合素刺激下调节TESK1激酶活性和将TESK1定位到细胞粘附位点中发挥重要作用。
Testicular protein kinase 1 (TESK1) is a serine/threonine kinase that phosphorylates cofilin and induces actin cytoskeletal reorganization. The kinase activity of TESK1 is stimulated by integrin-mediated signaling pathways, but the mechanism of regulation has remained unknown. By using the yeast two-hybrid system, we identified 14-3-3 beta to be the binding protein of TESK1. Specific interaction between TESK1 and 14-3-3 beta became evident in in vitro and in vivo co-precipitation assays. 14-3-3 beta interacts with TESK1 through the C-terminal region of TESK1 and in a manner dependent on the phosphorylation of Ser-439 within an RXXSXP motif. Binding of 14-3-3 beta inhibited the kinase activity of TESK1. During cell spreading on fibronectin, the TESK1/14-3-3 beta interaction significantly decreased, in a time course that inversely correlated with increase in TESK1 kinase activity. Thus, the dissociation of 14-3-3 beta from a TESK1/14-3-3 beta complex is likely to be involved in the integrin-mediated TESK1 activation. In HeLa cells, TESK1, together with 14-3-3 beta, accumulated at the cell periphery when cells were plated on fibronectin, whereas they were diffusely distributed in the cytoplasm in the case of non-stimulated cells. We propose that 14-3-3 beta plays important roles in regulating the kinase activity of TESK1 and localizing TESK1 to cell adhesion sites following integrin stimulation.