Division of Cardiovascular Medicine, Department of Medicine, Jichi Medical University, Shimotsuke 329-0498, Tochigi, Japan.
J Cell Sci. 2010 Mar 1;123(Pt 5):747-55. doi: 10.1242/jcs.052498.
Visualization and quantification of the dynamics of protein-protein interactions in living cells can be used to explore the macromolecular events involved in signal transduction processes. In this study, functional molecular imaging using a luciferase-based complementation method demonstrated how the integrin-linked kinase (ILK)-mediated protein complex controls downstream signals. The luciferase complementation assay showed that Akt1 preferentially binds to beta-parvin rather than to ILK within the complex. Moreover, photon flux from the interaction between beta-parvin and Akt1 increased following serum stimulation, and the beta-parvin-Akt1 interaction was dependent on phosphoinositide 3-kinase. Intriguingly, small interfering (si)RNA-mediated beta-parvin knockdown increased photon flux from the interaction between ILK and Akt1, leading to stabilization of hypoxia-inducible factor-1alpha and increased expression of vascular endothelial growth factor-A. These data from functional molecular imaging demonstrated that beta-parvin plays a regulatory role in the ILK-mediated Akt (also called protein kinase B) signaling cascades, suggesting that beta-parvin might be a crucial modulator of cell survival.
蛋白质-蛋白质相互作用在活细胞中的动态可视化和定量可用于探索信号转导过程中涉及的大分子事件。在这项研究中，使用基于荧光素酶的互补方法进行功能分子成像，展示了整合素连接激酶 (ILK) 介导的蛋白质复合物如何控制下游信号。荧光素酶互补测定表明，Akt1 优先与复合物中的β-辅肌动蛋白而不是与 ILK 结合。此外，血清刺激后，来自β-辅肌动蛋白和 Akt1 之间相互作用的光通量增加，并且β-辅肌动蛋白-Akt1 相互作用依赖于磷酸肌醇 3-激酶。有趣的是，小干扰 (si)RNA 介导的β-辅肌动蛋白敲低增加了来自 ILK 和 Akt1 之间相互作用的光通量，导致缺氧诱导因子-1α的稳定和血管内皮生长因子-A 的表达增加。来自功能分子成像的数据表明，β-辅肌动蛋白在 ILK 介导的 Akt（也称为蛋白激酶 B）信号级联中发挥调节作用，表明β-辅肌动蛋白可能是细胞存活的关键调节剂。
