Kinesin motors and their associated microtubule tracks are essential for long-distance transport of cellular cargos. Intracellular activity and proper recruitment of kinesins is regulated by biochemical signaling, cargo adaptors, microtubule-associated proteins, and mechanical forces. In this study, we found that the effect of opposing forces on the kinesin-microtubule attachment duration depends strongly on experimental assay geometry. Using optical tweezers and the conventional single-bead assay, we show that detachment of kinesin from the microtubule is likely accelerated by forces vertical to the long axis of the microtubule due to contact of the single bead with the underlying microtubule. We used the three-bead assay to minimize the vertical force component and found that when the opposing forces are mainly parallel to the microtubule, the median value of attachment durations between kinesin and microtubules can be up to 10-fold longer than observed using the single-bead assay. Using the three-bead assay, we also found that not all microtubule protofilaments are equivalent interacting substrates for kinesin and that the median value of attachment durations of kinesin varies by more than 10-fold, depending on the relative angular position of the forces along the circumference of the microtubule. Thus, depending on the geometry of forces across the microtubule, kinesin can switch from a fast detaching motor (median attachment duration <0.2 s) to a persistent motor that sustains attachment (median attachment duration >3 s) at high forces (5 pN). Our data show that the load-bearing capacity of the kinesin motor is highly variable and can be dramatically affected by off-axis forces and forces across the microtubule lattice, which has implications for a range of cellular activities, including cell division and organelle transport.

中文翻译：

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通过力几何和微管轨道调节运动蛋白的承载能力

驱动蛋白马达及其相关的微管轨道对于细胞货物的长途运输至关重要。细胞内活动和驱动蛋白的适当募集受生化信号、货物适配器、微管相关蛋白和机械力的调节。在这项研究中，我们发现相反的力对驱动蛋白-微管附着持续时间的影响在很大程度上取决于实验测定几何学。我们使用光学镊子和传统的单珠测定法表明，由于单珠与下面的微管接触，垂直于微管长轴的力可能会加速驱动蛋白与微管的分离。我们使用三珠试验来最小化垂直力分量，发现当相反的力主要平行于微管时，驱动蛋白和微管之间的附着持续时间的中值可以比使用单珠测定法观察到的时间长 10 倍。使用三珠测定法，我们还发现并非所有微管原丝都是驱动蛋白的等效相互作用底物，并且驱动蛋白附着持续时间的中值变化超过 10 倍，具体取决于力沿微管的相对角位置微管的周长。因此，根据穿过微管的力的几何形状，驱动蛋白可以从快速分离马达（中位附着持续时间 <0.2 秒）切换到持续附着的持久马达（中位附着持续时间 >3 秒）在高力 (5 pN) .