SUMMARY Background Cytoplasmic dynein motility along microtubules is critical for diverse cellular processes ranging from vesicular transport to nuclear envelope breakdown to mitotic spindle alignment. In yeast, we have proposed a regulated-offloading model to explain how dynein motility drives microtubule sliding along the cortex, powering transport of the nucleus into the mother-bud neck [1, 2]: the dynein regulator She1 limits dynein offloading by gating the recruitment of dynactin to the astral microtubule plus end, a prerequisite for offloading to the cortex. However, whether She1 subsequently affects cortically anchored dynein activity during microtubule sliding is unclear. Results Using single-molecule motility assays, we show that She1 strongly inhibits dynein movement along microtubules, acting directly on the motor domain in a manner independent of dynactin. She1 has no effect on the motility of either Kip2, a kinesin that utilizes the same microtubule track as dynein, or human kinesin-1, demonstrating the specificity of She1 for the dynein motor. At single-molecule resolution, She1 binds tightly to and exhibits diffusional behavior along microtubules. Diffusive She1 collides with and pauses motile dynein motors, prolonging their attachment to the microtubule. Furthermore, Aurora B/Ipl1 directly phosphorylates She1 and this modification appears to enhance the diffusive behavior of She1 along microtubules and its potency against dynein. In cells, She1 dampens productive microtubule-cortex interactions specifically in the mother compartment, polarizing spindle movements toward the bud cell. Conclusions Our data reveal how inhibitory microtubule-associated proteins selectively regulate motor activity to achieve unidirectional nuclear transport, and demonstrate a direct link between cell cycle machinery and dynein pathway activity.

Cortical force generators play a central role in the orientation and positioning of the mitotic spindle. In higher eukaryotes, asymmetrically localized cortical polarity determinants recruit or activate such force generators, which, through interactions with astral microtubules, position the mitotic spindle at the future site of cytokinesis. Recent studies in budding yeast have shown that, rather than the cell cortex, the astral microtubules themselves may provide polarity cues that are needed for asymmetric pulling on the mitotic spindle. Such asymmetry has been shown to be required for proper spindle positioning, and consequently faithful and accurate chromosome segregation. In this review, we highlight results that have shed light on spindle orientation in this classical model of asymmetric cell division, and review findings that may shed light on similar processes in higher eukaryotes.