CLASP2 binding to curved microtubule tips promotes flux and stabilizes kinetochore attachments.

CLASPs are conserved microtubule plus-end-tracking proteins that suppress microtubule catastrophes and independently localize to kinetochores during mitosis. Thus, CLASPs are ideally positioned to regulate kinetochore-microtubule dynamics required for chromosome segregation fidelity, but the underlying mechanism remains unknown. Here, we found that human CLASP2 exists predominantly as a monomer in solution, but it can self-associate through its C-terminal kinetochore-binding domain. Kinetochore localization was independent of self-association, and driving monomeric CLASP2 to kinetochores fully rescued normal kinetochore-microtubule dynamics, while partially sustaining mitosis. CLASP2 kinetochore localization, recognition of growing microtubule plus-ends through EB-protein interaction, and the ability to associate with curved microtubule protofilaments through TOG2 and TOG3 domains independently sustained normal spindle length, timely spindle assembly checkpoint satisfaction, chromosome congression, and faithful segregation. Measurements of kinetochore-microtubule half-life and poleward flux revealed that CLASP2 regulates kinetochore-microtubule dynamics by integrating distinctive microtubule-binding properties at the kinetochore-microtubule interface. We propose that kinetochore CLASP2 suppresses microtubule depolymerization and detachment by binding to curved protofilaments at microtubule plus-ends.

Plk1 puts a (Has)pin on the mitotic histone code.

Haspin is an atypical mitotic kinase that phosphorylates histone H3 on threonine 3 (H3T3), which is required to target Aurora B to centromeres. However, how Haspin is activated upon mitotic entry remained unknown. Two independent studies, published in Molecular Cell and in this issue of EMBO reports by Ghenoiu et al and Zhou et al, respectively, now show that Plk1 is responsible for Haspin activation as a H3T3 kinase. These results shed light on the spatiotemporal regulation of Aurora B to ensure mitotic fidelity.

Drosophila Polo regulates the spindle assembly checkpoint through Mps1-dependent BubR1 phosphorylation.

Maintenance of genomic stability during eukaryotic cell division relies on the spindle assembly checkpoint (SAC) that prevents mitotic exit until all chromosomes are properly attached to the spindle. Polo is a mitotic kinase proposed to be involved in SAC function, but its role has remained elusive. We demonstrate that Polo and Aurora B functional interdependency comprises a positive feedback loop that promotes Mps1 kinetochore localization and activity. Expression of constitutively active Polo restores normal Mps1 kinetochore levels even after Aurora B inhibition, highlighting a role for Polo in Mps1 recruitment to unattached kinetochores downstream of Aurora B. We also show that Mps1 kinetochore localization is required for BubR1 hyperphosphorylation and formation of the 3F3/2 phosphoepitope. This is essential to allow recruitment of Cdc20 to unattached kinetochores and the assembly of anaphase-promoting complex/cyclosome-inhibitory complexes to levels that ensure long-term SAC activity. We propose a model in which Polo controls Mps1-dependent BubR1 phosphorylation to promote Cdc20 kinetochore recruitment and sustained SAC function.

A tumor suppressor role of the Bub3 spindle checkpoint protein after apoptosis inhibition.

Most solid tumors contain aneuploid cells, indicating that the mitotic checkpoint is permissive to the proliferation of chromosomally aberrant cells. However, mutated or altered expression of mitotic checkpoint genes accounts for a minor proportion of human tumors. We describe a Drosophila melanogaster tumorigenesis model derived from knocking down spindle assembly checkpoint (SAC) genes and preventing apoptosis in wing imaginal discs. Bub3-deficient tumors that were also deficient in apoptosis displayed neoplastic growth, chromosomal aneuploidy, and high proliferative potential after transplantation into adult flies. Inducing aneuploidy by knocking down CENP-E and preventing apoptosis does not induce tumorigenesis, indicating that aneuploidy is not sufficient for hyperplasia. In this system, the aneuploidy caused by a deficient SAC is not driving tumorigenesis because preventing Bub3 from binding to the kinetochore does not cause hyperproliferation. Our data suggest that Bub3 has a nonkinetochore-dependent function that is consistent with its role as a tumor suppressor.

POLO ensures chromosome bi-orientation by preventing and correcting erroneous chromosome-spindle attachments.

Correct chromosome segregation during cell division requires bi-orientation at the mitotic spindle. Cells possess mechanisms to prevent and correct inappropriate chromosome attachment. Sister kinetochores assume a 'back-to-back' geometry on chromosomes that favors amphitelic orientation but the regulation of this process and molecular components are unknown. Abnormal chromosome-spindle interactions do occur but are corrected through the activity of Aurora B, which destabilizes erroneous attachments. Here, we address the role of Drosophila POLO in chromosome-spindle interactions and show that, unlike inhibition of its activity, depletion of the protein results in bipolar spindles with most chromosomes forming stable attachments with both sister kinetochores bound to microtubules from the same pole in a syntelic orientation. This is partly the result of impaired localization and activity of Aurora B but also of an altered centromere organization with abnormal distribution of centromeric proteins and shorter interkinetochore distances. Our results suggests that POLO is required to promote amphitelic attachment and chromosome bi-orientation by regulating both the activity of the correction mechanism and the architecture of the centromere.