Working strokes produced by curling protofilaments at disassembling microtubule tips: Their role in kinetochore motility and microtubule dynamics

The disassembly of microtubules can generate force and drive intracellular motility. During mitosis, for example, chromosomes remain persistently attached via kinetochores to the tips of disassembling microtubules, which pull the sister chromatids apart. According to the popular conformational wave hypothesis, such force generation requires that protofilaments curl outward from the disassembling tips to exert pulling force directly on kinetochores. However, there are other possible mechanisms and rigorously testing the conformational wave idea will require modifying the mechanical and energetic properties of curling protofilaments. By direct measurement of working strokes generated in vitro by curling protofilaments, we previously showed that their mechanical energy output can be increased by adding magnesium, and that yeast microtubules generate larger and more energetic working strokes than bovine microtubules. These increases in work output occurred independently of any change in the bending stiffness or intrinsic curvature of the protofilament curls, and were largely attributable to increases in contour length alone. Here we investigate the role of protofilament curls in the motility of the yeast kinetochore and explore the relationship between tubulin bending stiffness and microtubule dynamics.