Conformational Dynamics of the Focal Adhesion Targeting Domain Control Specific Functions of Focal Adhesion Kinase in Cells
Conformational Dynamics of the Focal Adhesion Targeting Domain Control Specific Functions of Focal Adhesion Kinase in Cells
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DOI：
10.1074/jbc.m114.593632
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发表时间：
2015-01-02
影响因子：
4.8
通讯作者：
Girault, Jean-Antoine
中科院分区：
文献类型：
--
作者：
Kadare, Gress;Gervasi, Nicolas;Girault, Jean-Antoine
背景：Focal adhesion kinase （FAK）通过其Focal adhesion targeting （FAT）结构域（一个四螺旋束）在Focal adhesion中富集。结果：促进或阻止第一螺旋打开的突变对FAK的生化特性及其在细胞中的功能有深远的影响。结论：FAT的开闭能力对FAK的功能至关重要。意义：这为FAT构象动力学的功能重要性提供了证据。局灶黏附激酶（FA）通过膜受体的信号传导调节细胞存活和运动。FAK的c端FA靶向（FAT）结构域具有多种功能，包括通过paxillin结合募集FA。Tyr上的FAT磷酸化（925）促进了FA的分解，并通过Grb2结合连接到MAPK途径，但需要解离FAT四螺旋束的第一螺旋（H1）。我们通过比较含有野生型或突变FAT的FAK分子与受损或促进H1开口的特性，研究了细胞中H1开口的重要性。这些突变不会改变FAK的激活，但会选择性地影响其细胞功能，包括自我结合、Tyr（925）磷酸化、paxillin结合、FA靶向和周转。位于激酶和FAT结构域之间的Tyr（861）的磷酸化也因打开FAT束的突变而增强。同样，ERK对bombesin的磷酸化也会随着FAT的打开而增加。虽然FAK分子的突变倾向于FAT打开在FAs募集不足，他们有效地恢复FA周转和FAK缺陷细胞的细胞形状。相反，阻止H1打开的突变明显损害了FAK的功能。我们的数据支持FAT结构域的构象动力学及其与分子其他部分的功能相互作用的生物学重要性。
Background: Focal adhesion kinase (FAK) is enriched at focal adhesions through its focal adhesion targeting (FAT) domain, a four-helix bundle. Results: Mutations that facilitate or prevent opening of the first helix have profound consequences on the biochemical properties of FAK and its function in cells. Conclusion: The ability of FAT to open and close is essential for FAK function. Significance: This provides evidence for the functional importance of the conformational dynamics of FAT.Focal adhesion (FA) kinase (FAK) regulates cell survival and motility by transducing signals from membrane receptors. The C-terminal FA targeting (FAT) domain of FAK fulfils multiple functions, including recruitment to FAs through paxillin binding. Phosphorylation of FAT on Tyr(925) facilitates FA disassembly and connects to the MAPK pathway through Grb2 association, but requires dissociation of the first helix (H1) of the four-helix bundle of FAT. We investigated the importance of H1 opening in cells by comparing the properties of FAK molecules containing wild-type or mutated FAT with impaired or facilitated H1 openings. These mutations did not alter the activation of FAK, but selectively affected its cellular functions, including self-association, Tyr(925) phosphorylation, paxillin binding, and FA targeting and turnover. Phosphorylation of Tyr(861), located between the kinase and FAT domains, was also enhanced by the mutation that opened the FAT bundle. Similarly phosphorylation of Ser(910) by ERK in response to bombesin was increased by FAT opening. Although FAK molecules with the mutation favoring FAT opening were poorly recruited at FAs, they efficiently restored FA turnover and cell shape in FAK-deficient cells. In contrast, the mutation preventing H1 opening markedly impaired FAK function. Our data support the biological importance of conformational dynamics of the FAT domain and its functional interactions with other parts of the molecule.