A force-sensitive mutation reveals a spindle assembly checkpoint-independent role for dynein in anaphase progression

Abstract

ABSTRACTThe cytoplasmic dynein-1 (dynein) motor organizes cells by shaping microtubule networks and moving a large variety of cargoes along them. However, dynein’s diverse roles complicatein vivostudies of its functions significantly. To address this issue, we have used gene editing to generate a series of missense mutations inDrosophilaDynein heavy chain (Dhc). We find that mutations associated with human neurological disease cause a range of defects in larval and adult flies, including impaired cargo trafficking in neurons. We also describe a novel mutation in the microtubule-binding domain (MTBD) of Dhc that, remarkably, causes metaphase arrest of mitotic spindles in the embryo but does not impair other dynein-dependent processes. We demonstrate that the mitotic arrest is independent of dynein’s well-established roles in silencing the spindle assembly checkpoint.In vitroreconstitution and optical trapping assays reveal that the mutation only impairs the performance of dynein under load.In silicoall-atom molecular dynamics simulations show that this effect correlates with increased flexibility of the MTBD, as well as an altered orientation of the stalk domain, with respect to the microtubule. Collectively, our data point to a novel role of dynein in anaphase progression that depends on the motor operating in a specific load regime. More broadly, our work illustrates how cytoskeletal transport processes can be dissectedin vivoby manipulating mechanical properties of motors.