Podosome Force Generation Machinery: A Local Balance between Protrusion at the Core and Traction at the Ring
Podosome Force Generation Machinery: A Local Balance between Protrusion at the Core and Traction at the Ring
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DOI：
10.1021/acsnano.7b00622
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发表时间：
2017-04-01
期刊：
影响因子：
17.1
通讯作者：
Poincloux, Renaud
中科院分区：
文献类型：
--
作者：
Bouissou, Angs;Proag, Amsha;Poincloux, Renaud
在纳米尺度上确定细胞如何产生和传递机械力是理解众多生理和病理过程的主要技术挑战。足体是一种亚微米的细胞结构，其核心是柱状的F-肌动蛋白核心，周围环绕着一环黏附蛋白，具有突出和探测细胞外基质的独特能力。使用突出力显微镜，我们以前已经证明了单个足体在细胞外环境中产生局部纳米级的突起。然而，细胞力是如何分布的，以实现这种突起机制，目前还不清楚。为了研究突出力产生的分子机制，我们进行了力学模拟，并开发了纳米尺度的结构和力学测量的定量图像分析。首先，在计算机模拟中表明，由足体造成的衬底的变形需要粘附环所在的紧邻核心外围的局部牵引力来平衡突出力。其次，我们证明了肌动蛋白聚合和突出力的产生都需要三环蛋白。第三，使用Donald，一种提供20 nm各向同性定位精度的3D纳米技术，我们将力的产生与足体环内talin的分子延伸相关联，这需要纽菌素和帕西林，表明该环保持机械张力。我们的工作表明，环是一个张力的位置，平衡核心的突出。这种反作用力的局部耦合形成了突起的基础，并揭示了足体作为纳米级自主力发生器的作用。
Determining how cells generate and trans duce mechanical forces at the nanoscale is a major technical challenge for the understanding of numerous physiological and pathological processes. Podosomes are submicrometer cell structures with a columnar F-actin core surrounded by a ring of adhesion proteins, which possess the singular ability to protrude into and probe the extracellular matrix. Using protrusion force microscopy, we have previously shown that single podosomes produce local nanoscale protrusions on the extracellular environment. However, how cellular forces are distributed to allow this protruding mechanism is still unknown. To investigate the molecular machinery of protrusion force generation, we performed mechanical simulations and developed quantitative image analyses of nanoscale architectural and mechanical measurements. First, in silico modeling showed that the deformations of the substrate made by podosomes require protrusion forces to be balanced by local traction forces at the immediate core periphery where the adhesion ring is located. Second, we showed that three-ring proteins are required for actin polymerization and protrusion force generation. Third, using DONALD, a 3D nanoscopy technique that provides 20 nm isotropic localization precision, we related force generation to the molecular extension of talin within the podosome ring, which requires vinculin and paxillin, indicating that the ring sustains mechanical tension. Our work demonstrates that the ring is a site of tension, balancing protrusion at the core. This local coupling of opposing forces forms the basis of protrusion and reveals the podosome as a nanoscale autonomous force generator.