Load-Sharing Mechanism of Cytoplasmic Dynein

Abstract

Cytoplasmic dynein is a homodimeric AAA+ motor protein responsible for minus end-directed transport along microtubules. The two heads of dynein are not strictly coordinated during processive motility, unlike those of dimeric kinesins and myosins. Therefore, tail-tethered optical trapping experiments yield insufficient information about the force production of individual heads. Here, we connected a trapped bead to a dynein dimer through a rigid DNA tether and directly measured the stepping motion of a single head under load. We observed that even though the step size of a head decreases under load, the head only stalls at half the stall force of the dimer and must rely on its partner head to walk against near-stall loads. The bulk of the force is produced when a head is bound to the microtubule, and stepping direction in the unbound state can be reversed by as little as 0.5 pN of backward load. Force-velocity curves indicate that the forward stepping rate of a dynein head is strongly inhibited by external force, while the backward rate is slow and nearly force-independent. Our results provide evidence for tension gating in dynein motility and lead us to propose a load-sharing mechanism between the two heads of a dynein dimer.