Phosphorylation controls spatial and temporal activities of motor-PRC1 complexes to complete mitosis.

During mitosis, spindle architecture alters as chromosomes segregate into daughter cells. The microtubule crosslinker protein regulator of cytokinesis 1 (PRC1) is essential for spindle stability, chromosome segregation and completion of cytokinesis, but how it recruits motors to the central spindle to coordinate the segregation of chromosomes is unknown. Here, we combine structural and cell biology approaches to show that the human CENP-E motor, which is essential for chromosome capture and alignment by microtubules, binds to PRC1 through a conserved hydrophobic motif. This binding mechanism is also used by Kinesin-4 Kif4A:PRC1. Using in vitro reconstitution, we demonstrate that CENP-E slides antiparallel PRC1-crosslinked microtubules. We find that the regulation of CENP-E -PRC1 interaction is spatially and temporally coupled with relocalization to overlapping microtubules in anaphase. Finally, we demonstrate that the PRC1-microtubule motor interaction is essential in anaphase to control chromosome partitioning, retain central spindle integrity and ensure cytokinesis. Taken together our findings reveal the molecular basis for the cell cycle regulation of motor-PRC1 complexes to couple chromosome segregation and cytokinesis.

Reconstitution of an active human CENP-E motor.

CENP-E is a large kinesin motor protein which plays pivotal roles in mitosis by facilitating chromosome capture and alignment, and promoting microtubule flux in the spindle. So far, it has not been possible to obtain active human CENP-E to study its molecular properties. Xenopus CENP-E motor has been characterized in vitro and is used as a model motor; however, its protein sequence differs significantly from human CENP-E. Here, we characterize human CENP-E motility in vitro. Full-length CENP-E exhibits an increase in run length and longer residency times on microtubules when compared to CENP-E motor truncations, indicating that the C-terminal microtubule-binding site enhances the processivity when the full-length motor is active. In contrast with constitutively active human CENP-E truncations, full-length human CENP-E has a reduced microtubule landing rate in vitro, suggesting that the non-motor coiled-coil regions self-regulate motor activity. Together, we demonstrate that human CENP-E is a processive motor, providing a useful tool to study the mechanistic basis for how human CENP-E drives chromosome congression and spindle organization during human cell division.

Leaving no-one behind: how CENP-E facilitates chromosome alignment.

Chromosome alignment and biorientation is essential for mitotic progression and genomic stability. Most chromosomes align at the spindle equator in a motor-independent manner. However, a subset of polar kinetochores fail to bi-orient and require a microtubule motor-based transport mechanism to move to the cell equator. Centromere Protein E (CENP-E/KIF10) is a kinesin motor from the Kinesin-7 family, which localizes to unattached kinetochores during mitosis and utilizes plus-end directed microtubule motility to slide mono-oriented chromosomes to the spindle equator. Recent work has revealed how CENP-E cooperates with chromokinesins and dynein to mediate chromosome congression and highlighted its role at aligned chromosomes. Additionally, we have gained new mechanistic insights into the targeting and regulation of CENP-E motor activity at the kinetochore. Here, we will review the function of CENP-E in chromosome congression, the pathways that contribute to CENP-E loading at the kinetochore, and how CENP-E activity is regulated during mitosis.