Multiscale temporal tuning of the force generation complex governs cortical microtubule interactions during the first...

Dynein is an essential microtubule motor whose many roles in mitosis complicate efforts to resolve its spatiotemporally dynamic regulation. We previously established a method to classify single particles of the conserved cortical force generation complex (dynein-LIN-5^NuMA-GPR-1/2^LGN-Gα^Gαi), as free or interacting with microtubules. Here, we report the results of depleting force generation complex components and regulators. Depleting LIS-1 reversed force asymmetry, while depletion of the conserved protein phosphatase regulatory subunit SUR-6^PP2A-B55 significantly increased incidence of dynein trajectories with microtubule-interacting behavior during prophase. We next applied our classification scheme to the dynein anchor LIN-5. Microtubule-interacting LIN-5 trajectories were posteriorly enriched during anaphase, consistent with our dynein data and prior fluorescence studies. SUR-6^PP2A-B55 depletion did not alter LIN-5 kinetics, suggesting regulation of a dynein specific function rather than regulation via LIN-5. By comparing and analyzing differences in LIN-5 trajectories, we found evidence that two distinct patterns of force generation complex behavior, microtubule interactions and cortical flows, emerge at the millisecond and second time scales, respectively. Our observations provide novel insight into the regulation of cortical dynein and the coupling among the cell membrane, actomyosin cortex, force generation complex, and microtubules that positions the mitotic spindle. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text].