Serine and threonine phosphorylation of the paxillin LIM domains regulates paxillin focal adhesion localization and cell adhesion to fibronectin
Serine and threonine phosphorylation of the paxillin LIM domains regulates paxillin focal adhesion localization and cell adhesion to fibronectin
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DOI：
10.1091/mbc.9.7.1803
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发表时间：
1998-07-01
影响因子：
3.3
通讯作者：
Turner, CE
中科院分区：
文献类型：
--
作者：
Brown, MC;Perrotta, JA;Turner, CE
我们之前已经证明，巴西林的LIM结构域作为该蛋白的焦点黏附(FA)靶向基序发挥作用。在目前的研究中，我们已经确定了Paxlin LIM2和LIM3作为丝氨酸/苏氨酸激酶的结合位点和底物的能力。在CHO.K1细胞与纤维连接蛋白黏附后，LIM2和LIM3相关的激酶的活性被刺激，因此，研究了LIM结构域的磷酸化在调节与纤维连接蛋白黏附后的帕西林的亚细胞定位中的作用。利用禽巴西林-CHO.K1模型体系，探讨了巴西林磷酸化在巴西林定位FAs中的作用。我们发现，模拟LIM结构域磷酸化的帕西林突变加速了纤维连接蛋白诱导的帕西林定位到局灶性接触。此外，阻断LIM结构域的磷酸化降低了细胞与纤维连接蛋白的黏附，而结构性LIM结构域的磷酸化显著增加了细胞黏附纤维连接蛋白的能力。FA对纤维连接蛋白的靶向性和细胞黏附的增强是LIM结构域磷酸化所特有的，因为因纤维连接蛋白黏附而被磷酸化的巴西林的氨基末端酪氨酸和丝氨酸残基的突变对FA的定位或细胞黏附没有影响。这首次证明了通过LIM结构域的磷酸化来调节蛋白质定位，并提出了一种新的调节LIM结构域功能的机制。此外，这些结果还首次证明了巴西林参与了整合素介导的信号转导。
We have previously shown that the LIM domains of paxillin operate as the focal adhesion (FA)-targeting motif of this protein. In the current study, we have identified the capacity of paxillin LIM2 and LIM3 to serve as binding sites for, and substrates of serine/threonine kinases. The activities of the LIM2- and LIM3-associated kinases were stimulated after adhesion of CHO.K1 cells to fibronectin; consequently, a role for LIM domain phosphorylation in regulating the subcellular localization of paxillin after adhesion to fibronectin was investigated. An avian paxillin-CHO.K1 model system was used to explore the role of paxillin phosphorylation in paxillin localization to FAs. We found that mutations of paxillin that mimicked LIM domain phosphorylation accelerated fibronectin-induced localization of paxillin to focal contacts. Further, blocking phosphorylation of the LIM domains reduced cell adhesion to fibronectin, whereas constitutive LIM domain phosphorylation significantly increased the capacity of cells to adhere to fibronectin. The potentiation of FA targeting and cell adhesion to fibronectin was specific to LIM domain phosphorylation as mutation of the amino-terminal tyrosine and serine residues of paxillin that are phosphorylated in response to fibronectin adhesion had no effect on the rate of FA localization or cell adhesion. This represents the first demonstration of the regulation of protein localization through LIM domain phosphorylation and suggests a novel mechanism of regulating LIM domain function. Additionally, these results provide the first evidence that paxillin contributes to "inside-out" integrin-mediated signal transduction.