Minimal features of paxillin that are required for the tyrosine phosphorylation of focal adhesion kinase.
Minimal features of paxillin that are required for the tyrosine phosphorylation of focal adhesion kinase.
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DOI：
10.1042/bj20051241
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发表时间：
2006-01
期刊：
影响因子：
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通讯作者：
R. Wade;S. V. Vande Pol
中科院分区：
文献类型：
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作者：
R. Wade;S. V. Vande Pol
FAK（局灶黏附激酶）的酪氨酸磷酸化调节整合素与细胞外基质和生长因子受体相互作用产生的信号。这个过程的一个关键步骤是FAK的Tyr397的磷酸化，它为Src家族激酶、PI3K（磷酸肌肽3-激酶）和Shc （Src同源性和胶原同源性）创造了一个结合位点。完整的Tyr397位点对于fak介导的细胞迁移、生存信号和对可溶性生长因子的完全响应的调节是必需的。我们之前的研究表明，胚胎干细胞中FAK的酪氨酸磷酸化需要连接蛋白paxillin [Wade， Bohl and Vande Pol (2002) Oncogene 21,96 -107]。在本文中，我们确定了支持FAK酪氨酸磷酸化和Tyr397磷酸化所需的paxillin的最小结构特征。Paxillin含有n端富含亮氨酸的LD基元，直接结合FAK和c端四个LIM结构域（Lin-11， is -1和Mec-3）。我们发现paxillin LIM结构域1、2和3都是FAK酪氨酸磷酸化所必需的，而LIM4是可缺性的。除了paxillin LIM结构域1、2和3外，在胚胎干细胞中，paxillin上还需要一个LD基序来支持FAK酪氨酸磷酸化。支持FAK酪氨酸磷酸化的LD基序存在序列和空间要求。有趣的是，在体外不能结合FAK的合成LD基序能够完全支持FAK酪氨酸磷酸化，这表明LD基序与FAK的最小相互作用就足够了。我们的研究结果表明，paxillin至少有四个不同的结构域支持至少三个不同的功能，每个功能都是FAK酪氨酸磷酸化所必需的。
Tyrosine phosphorylation of FAK (focal adhesion kinase) regulates signalling that results from the interaction of integrins with extracellular matrix and growth factor receptors. A critical step in this process is the phosphorylation of Tyr397 of FAK, which creates a binding site for Src family kinases, PI3K (phosphoinositide 3-kinase) and Shc (Src homology and collagen homology). An intact Tyr397 site is required for FAK-mediated regulation of cell migration, survival signals and full responsiveness to soluble growth factors. We showed previously that the adaptor protein paxillin is required for the overall tyrosine phosphorylation of FAK in embryonic stem cells [Wade, Bohl and Vande Pol (2002) Oncogene 21, 96-107]. In the present paper, we identify the minimal structural features of paxillin that are required to support overall FAK tyrosine phosphorylation and Tyr397 phosphorylation. Paxillin contains N-terminal leucine-rich LD motifs that bind directly to FAK and four LIM (Lin-11, Isl-1 and Mec-3) domains in the C-terminus. We show that paxillin LIM domains 1, 2 and 3 are each required for FAK tyrosine phosphorylation, while LIM4 is dispensable. In addition to paxillin LIM domains 1, 2 and 3, a single LD motif on paxillin is required to support FAK tyrosine phosphorylation in embryonic stem cells. Both sequence and spatial requirements exist for LD motifs to support FAK tyrosine phosphorylation. Interestingly, synthetic LD motifs that fail to bind FAK in vitro are able to fully support FAK tyrosine phosphorylation, indicating that minimal interactions of LD motifs with FAK suffice. Our results demonstrate at least four distinct structural domains of paxillin support at least three distinct functions that are each required for FAK tyrosine phosphorylation.