Force measurements on cargoes in living cells reveal collective dynamics of microtubule motors
Force measurements on cargoes in living cells reveal collective dynamics of microtubule motors
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DOI：
10.1073/pnas.1215462109
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发表时间：
2012-11-06
影响因子：
11.1
通讯作者：
Goldman, Yale E.
中科院分区：
文献类型：
--
作者：
Hendricks, Adam G.;Holzbaur, Erika L. F.;Goldman, Yale E.
许多细胞内的货物在正负端定向的运动蛋白的驱动下，沿沿着微管双向运动。为了探测在细胞内运输过程中施加在货物上的力，我们检查了吞噬到活的哺乳动物巨噬细胞中的乳胶珠。这些乳胶珠隔室（LBC）被包裹在膜中，并通过内源性驱动蛋白-1、驱动蛋白-2和动力蛋白马达的补充沿着细胞骨架运输。LBC的大小和折射率使它们非常适合用光学陷阱进行操作。我们开发的方法，提供在复杂的细胞环境中的光阱的原位校准，考虑到货物和局部粘弹性的细胞质之间的任何变化。我们发现，施加在LBC上的中心和外围方向的力具有相似的幅度，最大力类似于20 pN。在大于10 pN的力事件期间，我们经常在两个方向上观察到8 nm的步进，这表明多个电机的步进是相关的。这些观察结果表明，LBC的双向运输是由在力平衡附近操作的稳定绑定电机的相对团队驱动的。
Many cellular cargoes move bidirectionally along microtubules, driven by teams of plus-and minus-end-directed motor proteins. To probe the forces exerted on cargoes during intracellular transport, we examined latex beads phagocytosed into living mammalian macrophages. These latex bead compartments (LBCs) are encased in membrane and transported along the cytoskeleton by a complement of endogenous kinesin-1, kinesin-2, and dynein motors. The size and refractive index of LBCs makes them well-suited for manipulation with an optical trap. We developed methods that provide in situ calibration of the optical trap in the complex cellular environment, taking into account any variations among cargoes and local viscoelastic properties of the cytoplasm. We found that centrally and peripherally directed forces exerted on LBCs are of similar magnitude, with maximum forces of similar to 20 pN. During force events greater than 10 pN, we often observe 8-nm steps in both directions, indicating that the stepping of multiple motors is correlated. These observations suggest bidirectional transport of LBCs is driven by opposing teams of stably bound motors that operate near force balance.