Graduate Program in Biophysical Sciences, The University of Chicago, Chicago, Illinois; Institute of Biophysical Dynamics, The University of Chicago, Chicago, Illinois.
Institute of Biophysical Dynamics, The University of Chicago, Chicago, Illinois; Pritzker School of Molecular Engineering, The University of Chicago, Chicago, Illinois.
Biophys J. 2024 Jan 16;123(2):157-171. doi: 10.1016/j.bpj.2023.12.004. Epub 2023 Dec 6.
The actomyosin cytoskeleton generates mechanical forces that power important cellular processes, such as cell migration, cell division, and mechanosensing. Actomyosin self-assembles into contractile networks and bundles that underlie force generation and transmission in cells. A central step is the assembly of the myosin II filament from myosin monomers, regulation of which has been extensively studied. However, myosin filaments are almost always found as clusters within the cell cortex. While recent studies characterized cluster nucleation dynamics at the cell periphery, how myosin clusters grow on stress fibers remains poorly characterized. Here, we utilize a U2OS osteosarcoma cell line with endogenously tagged myosin II to measure the myosin cluster size distribution in the lamella of adherent cells. We find that myosin clusters can grow with Rho-kinase (ROCK) activity alone in the absence of myosin motor activity. Time-lapse imaging reveals that myosin clusters grow via increased myosin association to existing clusters, which is potentiated by ROCK-dependent myosin filament assembly. Enabling myosin motor activity allows further myosin cluster growth through myosin association that is dependent on F-actin architecture. Using a toy model, we show that myosin self-affinity is sufficient to recapitulate the experimentally observed myosin cluster size distribution, and that myosin cluster sizes are determined by the pool of myosin available for cluster growth. Together, our findings provide new insights into the regulation of myosin cluster sizes within the lamellar actomyosin cytoskeleton.
肌动球蛋白细胞骨架产生机械力,为细胞迁移、细胞分裂和机械感知等重要细胞过程提供动力。肌动球蛋白自组装成收缩网络和束,为细胞内的力产生和传递提供基础。一个关键步骤是肌球蛋白单体组装成肌球蛋白丝,其调控已得到广泛研究。然而,肌球蛋白丝几乎总是在细胞皮层内以簇的形式存在。虽然最近的研究描述了细胞边缘处簇核形成的动力学,但肌球蛋白簇在应力纤维上的生长仍然知之甚少。在这里,我们利用内源性标记肌球蛋白 II 的 U2OS 骨肉瘤细胞系来测量贴壁细胞板层中肌球蛋白簇的大小分布。我们发现,在没有肌球蛋白运动活性的情况下,肌球蛋白簇可以仅在 Rho 激酶(ROCK)活性的作用下生长。延时成像揭示了肌球蛋白簇通过增加与现有簇的肌球蛋白结合而生长,这是由 ROCK 依赖性肌球蛋白丝组装增强的。使肌球蛋白运动活性能够通过依赖 F-肌动蛋白结构的肌球蛋白结合进一步促进肌球蛋白簇的生长。使用一个玩具模型,我们表明肌球蛋白的自我亲和力足以再现实验观察到的肌球蛋白簇大小分布,并且肌球蛋白簇的大小由可用于簇生长的肌球蛋白池决定。总之,我们的研究结果为板层肌动球蛋白细胞骨架中肌球蛋白簇大小的调控提供了新的见解。