Generation of Directed Motion: How Random Steps in Cytoskeletal Systems can lead to Processive Movement

细胞内转运有很多种：从浸入胞质溶胶中的组分的纯粹扩散性随机运动到分子马达的高度定向性确定性易位。后者甚至可以连续执行多个步骤，这种能力被称为持续合成能力。我们将在这里研究的是随机运动和高度进行性运动之间的广泛范围。虽然大多数马达蛋白与其货物严格结合，但其中一些只是松散地连接。例如，在细胞分裂过程中，微管彼此相对滑动是由扩散锚定的驱动蛋白马达促进的。在这个过程中产生了什么力量，这种扩散锚定的好处是什么，类似于“降低”的持续性？另一方面，细胞骨架驱动的持续性也可以增加：蛋白质的构象增强了聚合驱动的肌动蛋白丝的热棘轮样向前运动。聚合过程中的步长和力与没有聚合的情况相比如何？而且，在一个小的肌动蛋白束中，肌动蛋白丝的随机和真正的非进行性解聚能产生足够的熵变化，使组装体沿着束轴收缩吗？使用单分子技术应用于重建在体外测定，我们将表征（i）微管微管滑动驱动的扩散锚定驱动蛋白-14马达，（ii）肌动蛋白丝的聚合存在下的肌动蛋白，和（iii）收缩的肌动蛋白束诱导的丝解聚。我们认为这些过程是随机的，扩散性元素和定向的，确定性成分之间的细胞内相互作用的原型系统。该项目的结果预计将增加我们对生物系统如何利用不同运动模式以及这些模式如何有利地组合和相互转换的（生物）物理理解。

Intracellular transport comes in a wide variety: from the purely diffusive, random motion of components immersed in the cytosol to the highly directed, deterministic translocation of molecular motors. The latter may even perform multiple steps in a row, an ability referred to as processivity. It is the wide spectrum between random and highly processive motion that we will investigate here. While most motor proteins are rigidly bound to their cargo, some of them are only loosely attached. For example, during cell division the relative sliding of microtubules against each other is facilitated by diffusively anchored kinesin motors. What forces are generated during this process and what are the benefits of such diffusive anchorage, similar to a 'lowered' processivity? On the other hand, the processivity of cytoskeletal actuation can also be increased: the protein formin enhances the polymerization-driven, thermal-ratchet-like forward motion of actin filaments. How do step size and force during polymerization compare to the case without formin? And, can the rather random and truly non-processive depolymerization of actin filaments in a small actin bundle generate a sufficient entropic change to contract the assembly along the bundle axis? Using single-molecule techniques applied to reconstituted in vitro assays, we will characterize (i) the microtubule-microtubule sliding actuated by diffusively anchored kinesin-14 motors, (ii) the polymerization of actin filaments in the presence of formin, and (iii) the contraction of actin bundles induced by filament depolymerization. We regard these processes as prototypical systems for the intracellular interplay between random, diffusive elements and directed, deterministic constituents. The results of the project are expected to add to our (bio)physical understanding of how biological systems utilize different modes of movement as well as how these modes can be beneficially combined and interconverted.