Talin determines the nanoscale architecture of focal adhesions
Talin determines the nanoscale architecture of focal adhesions
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DOI：
10.1073/pnas.1512025112
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发表时间：
2015-09-01
影响因子：
11.1
通讯作者：
Kanchanawong, Pakorn
中科院分区：
文献类型：
--
作者：
Liu, Jaron;Wang, Yilin;Kanchanawong, Pakorn
深入了解分子机器如何执行其生物功能取决于对组件的空间组织、连接性、几何形状和组织层次结构的了解。然而，这些参数是难以确定的多组分组件，如整合素为基础的局灶性粘连（FA）。我们以前应用三维超分辨率荧光显微镜探测主要FA组分的空间组织，观察整合素和肌动蛋白细胞骨架之间的蛋白质纳米级分层。在这里，我们结合联合收割机超分辨率成像技术与蛋白质工程的方法来研究如何产生这样的纳米结构。我们证明，塔林发挥了关键的结构作用，在调节脂肪酸的纳米结构，类似于一个分子的统治者。Talin对角地跨越FA核心，其N末端在膜上，C末端划分FA/应力纤维界面。相比之下，黏着斑蛋白被发现是用于FA纳米结构的规格。重组类似物的塔林与修改的长度重演其极化方向，但改变了FA/应力纤维界面的线性方式，符合其模块化结构，并牵连的整合素-塔林-肌动蛋白复合物作为主要的机械联动FA。塔林被发现是类似于97 nm的长度和取向在类似于15度相对于质膜。我们的研究结果确定塔林作为FA纳米级组织的主要决定因素，并建议如何多个细胞的力量可能会整合在粘附位点。
Insight into howmolecularmachines perform their biological functions depends on knowledge of the spatial organization of the components, their connectivity, geometry, and organizational hierarchy. However, these parameters are difficult to determine in multicomponent assemblies such as integrin-based focal adhesions (FAs). We have previously applied 3D superresolution fluorescence microscopy to probe the spatial organization of major FA components, observing a nanoscale stratification of proteins between integrins and the actin cytoskeleton. Here we combine superresolution imaging techniques with a protein engineering approach to investigate how such nanoscale architecture arises. We demonstrate that talin plays a key structural role in regulating the nanoscale architecture of FAs, akin to a molecular ruler. Talin diagonally spans the FA core, with its N terminus at the membrane and C terminus demarcating the FA/stress fiber interface. In contrast, vinculin is found to be dispensable for specification of FA nanoscale architecture. Recombinant analogs of talin with modified lengths recapitulated its polarized orientation but altered the FA/stress fiber interface in a linear manner, consistent with its modular structure, and implicating the integrin-talin-actin complex as the primary mechanical linkage in FAs. Talin was found to be similar to 97 nm in length and oriented at similar to 15 degrees relative to the plasma membrane. Our results identify talin as the primary determinant of FA nanoscale organization and suggest how multiple cellular forces may be integrated at adhesion sites.