Center for Molecular Spectroscopy and Dynamics, Institute for Basic Science, Seoul 02841, Korea.
Center for Breast Cancer, National Cancer Center, Goyang 10408, Korea.
J Phys Chem Lett. 2020 Dec 3;11(23):10233-10241. doi: 10.1021/acs.jpclett.0c02103. Epub 2020 Nov 18.
Focal adhesions (FAs) are dynamic protein nanostructures that form mechanical links between cytoskeletal actin fibers and the extracellular matrix. Here, we demonstrate that interferometric scattering (iSCAT) microscopy, a high-speed and time-unlimited imaging technique, can uncover the real-time dynamics of nanoscopic nascent adhesions (NAs). The high sensitivity and stability of the iSCAT signal enabled us to trace the whole life span of each NA spontaneously nucleated under a lamellipodium. Such high-throughput and long-term image data provide a unique opportunity for statistical analysis of adhesion dynamics. Moreover, we directly revealed that FAs play critical roles in both the extrusion of filopodia as nucleation sites on the leading edge and the one-dimensional transport of cargos along cytoskeletal fibers as fiber docking sites. These experimental results show that iSCAT is a sensitive tool for tracking real-time dynamics of nanoscopic objects involved in endogenous and exogenous biological processes in living cells.
粘着斑（FAs）是一种动态的蛋白质纳米结构，它在细胞骨架肌动蛋白纤维和细胞外基质之间形成机械连接。在这里，我们证明了干涉散射（iSCAT）显微镜是一种高速且无时间限制的成像技术，可以揭示纳米新生粘着斑（NAs）的实时动力学。iSCAT 信号的高灵敏度和稳定性使我们能够追踪在片状伪足下自发形成的每个 NA 的整个生命周期。这种高通量和长期的图像数据为粘附动力学的统计分析提供了独特的机会。此外，我们直接揭示了 FAs 在丝状伪足的挤出过程中作为核位点以及沿着细胞骨架纤维的货物的一维运输过程中作为纤维对接位点都发挥着关键作用。这些实验结果表明，iSCAT 是一种灵敏的工具，可用于跟踪活细胞内参与内源性和外源性生物过程的纳米级物体的实时动力学。
