JCB_201611105 2047..2058

During eukaryotic cell division, coordination between the mitotic spindle, which partitions duplicated chromosomes, and the cytokinetic contractile network, which partitions the cytoplasm, ensures that each daughter cell inherits a genome (Rappaport and Ebstein, 1965). In animal cells, the position of the spindle determines the site of cytokinesis, which is critical during asymmetric cell divisions that underlie development and tissue homeostasis for guiding the inheritance of cytoplasmic fate determinants and the positions of daughter cells. In unicellular organisms such as budding yeast, the spindle must be brought to a predetermined site of cytokinesis between the mother and bud. In all of these scenarios, the spindle is positioned through mechanisms that integrate polarity signals into force generation by the microtubule cytoskeleton (McNally, 2013). Spindle movement in many species and cellular contexts is driven by cytoplasmic dynein, a large multisubunit protein complex that uses ATP hydrolysis to power motility toward the minus ends of microtubules. Dynein moves the spindle by generating pulling forces on astral microtubules, which are nucleated at microtubule-organizing centers and project into the cytoplasm, toward the cell cortex (Carminati and Stearns, 1997; Skop and White, 1998; Adames and Cooper, 2000; Grill et al., 2001; Kiyomitsu and Cheeseman, 2013). Observations in different organisms suggest that dynein can generate pulling forces either through end-on interactions with microtubules at the cortex or by sliding microtubules laterally along the cortex. In budding yeast, dynein is delivered on the plus ends of astral microtubules to the cortex, where it attaches to its cortical receptor Num1 (Lee et al., 2005). Then, dynein is activated, and its minus end–directed motility slides the astral microtubule along the cortex and past the anchored motor, drawing the spindle toward the cortex (Adames and Cooper, 2000; Lammers and Markus, 2015). In the Caenorhabditis elegans zygote, dynein accumulates at the anterior cell cortex and captures astral microtubules in an end-on manner (Labbé et al., 2003; Redemann et al., 2010). These microtubule ends quickly undergo catastrophe but remain at the cortex, and their depolymerization is thought to generate force to pull the spindle toward the anterior (Kozlowski et al., 2007). Whereas end-on interactions predominate during anaphase in C. elegans zygotes, lateral sliding interactions are observed during telophase and are highly abundant in embryos that generate free microtubule fragments by ectopic expression of the microtubule-severing protein katanin (Kozlowski et al., 2007; Gusnowski and Srayko, 2011). This suggests dynein can alternate between different modes of force generation. Consistent with this, experiments by Laan et al. (2012) demonstrate that dynein purified from budding yeast can generate force by capturing plus ends at a fabricated barrier, promoting catastrophe and maintaining attachment as the microtubules depolymerize. In this scenario, dynein is thought to promote catastrophe by holding the microtubule end close to the barrier. Microtubule ends can be induced to catastrophe by colliding with barriers, which is thought to impede the arrival of new tubulin subunits (Janson et al., 2003). Thus, dynein How dynein motors accurately move cargoes is an important question. In budding yeast, dynein moves the mitotic spindle to the predetermined site of cytokinesis by pulling on astral microtubules. In this study, using high-resolution imaging in living cells, we discover that spindle movement is regulated by changes in microtubule plus-end dynamics that occur when dynein generates force. Mutants that increase plus-end stability increase the frequency and duration of spindle movements, causing positioning errors. We find that dynein plays a primary role in regulating microtubule dynamics by destabilizing microtubules. In contrast, the dynactin complex counteracts dynein and stabilizes microtubules through a mechanism involving the shoulder subcomplex and the cytoskeletal-associated protein glycine-rich domain of Nip100/ p150glued. Our results support a model in which dynein destabilizes its microtubule substrate by using its motility to deplete dynactin from the plus end. We propose that interplay among dynein, dynactin, and the stability of the microtubule substrate creates a mechanism that regulates accurate spindle positioning. Dynein is regulated by the stability of its microtubule track