In cells lacking centrosomes, such as those found in female meiosis, chromosomes must nucleate and stabilize microtubules in order to form a bipolar spindle. Here we report the identification of Dasra A and Dasra B, two new components of the vertebrate chromosomal passenger complex containing Incenp, Survivin, and the kinase Aurora B, and demonstrate that this complex is required for chromatin-induced microtubule stabilization and spindle formation. The failure of microtubule stabilization caused by depletion of the chromosomal passenger complex was rescued by codepletion of the microtubule-depolymerizing kinesin MCAK, whose activity is negatively regulated by Aurora B. By contrast, we present evidence that the Ran-GTP pathway of chromatin-induced microtubule nucleation does not require the chromosomal passenger complex, indicating that the mechanisms of microtubule assembly by these two pathways are distinct. We propose that the chromosomal passenger complex regulates local MCAK activity to permit spindle formation via stabilization of chromatin-associated microtubules.

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Motility is a fundamentally important property of most members of the kinesin superfamily, but a rare subset of kinesins are also able to alter microtubule dynamics. At kinetochore-microtubule plus ends, the kinesin-8 family member Kif18A is essential to align mitotic chromosomes at the spindle equator during cell division, but how it accomplishes this function is unclear. We report here that Kif18A is a plus-end-directed motor that inhibits the polymerization dynamics of microtubule plus ends without destabilizing them, distinguishing Kif18A from the budding yeast ortholog Kip3. In interphase cells, Kif18A uses this activity to reduce the overall dynamicity of microtubule plus ends and effectively constrains the distance over which plus ends grow and shrink. Our findings suggest that kinesin-8 family members have developed biochemically distinct activities throughout evolution and have implications for how Kif18A affects kinetochore-microtubule plus-end dynamics during mitosis in animal cells.

The KinI kinesin MCAK is a microtubule depolymerase important for governing spindle microtubule dynamics during chromosome segregation. The dynamic nature of spindle assembly and chromosome-microtubule interactions suggest that mechanisms must exist that modulate the activity of MCAK, both spatially and temporally. In Xenopus extracts, MCAK associates with and is stimulated by the inner centromere protein ICIS. The inner centromere kinase Aurora B also interacts with ICIS and MCAK raising the possibility that Aurora B may regulate MCAK activity as well. Herein, we demonstrate that recombinant Aurora B-INCENP inhibits Xenopus MCAK activity in vitro in a phosphorylationdependent manner. Substituting endogenous MCAK in Xenopus extracts with the alanine mutant XMCAK-4A, which is resistant to inhibition by Aurora B-INCENP, led to assembly of mono-astral and monopolar structures instead of bipolar spindles. The size of these structures and extent of tubulin polymerization in XMCAK-4A extracts indicate that XM-CAK-4A is not defective for microtubule dynamics regulation throughout the cytoplasm. We further demonstrate that the ability of XMCAK-4A to localize to inner centromeres is abolished. Our results show that MCAK regulation of cytoplasmic and spindle-associated microtubules can be differentiated by Aurora B-dependent phosphorylation, and they further demonstrate that this regulation is required for bipolar meiotic spindle assembly. INTRODUCTIONAssembly of the microtubule-based bipolar spindle, the apparatus that powers chromosome segregation during mitosis and meiosis (M phase), is driven by the inherent dynamic instability of microtubules, as well as by proteins that alter microtubule dynamics (Hyman and Karsenti, 1996;Wittmann et al., 2001). Accordingly, M-phase onset triggers global, cytoplasmic changes in microtubule dynamics that expedite spindle assembly, most notably an increase in the frequency of catastrophes, the transition from microtubule growth to shrinkage (Belmont et al., 1990;Verde et al., 1992;Rusan et al., 2001;Kinoshita et al., 2002). In Xenopus egg extracts, the KinI kinesin MCAK, classified by its internally located motor domain (Wordeman and Mitchison, 1995;Walczak et al., 1996), is essential for governing this change in catastrophe frequency (Walczak et al., 1996;Tournebize et al., 2000). Biochemical studies indicate that MCAK likely accomplishes catastrophe induction directly by catalytically depolymerizing microtubules from their protofilament ends (Desai et al., 1999b;Hunter et al., 2003).Although spindle assembly demands global regulation of microtubule dynamics, proper spindle function also requires precise control of microtubule dynamics regulation at defined locations within the spindle. Because accurate chromosome segregation requires that each kinetochore of a pair of sister chromatids be attached to microtubules derived from opposing spindle poles (biorientation), it is particularly important to control the interaction of spindle microtubules with kinetochores (Rieder and Salmon, 1...