PLK1 Phosphorylates Mitotic Centromere-associated Kinesin and Promotes Its Depolymerase Activity

Abstract

During cell division, interaction between kinetochores and dynamic spindle microtubules governs chromosome movements. The microtubule depolymerase mitotic centromere-associated kinesin (MCAK) is a key regulator of mitotic spindle assembly and dynamics. However, the regulatory mechanisms underlying its depolymerase activity during the cell cycle remain elusive. Here, we showed that PLK1 is a novel regulator of MCAK in mammalian cells. MCAK interacts with PLK1 in vitro and in vivo. The neck and motor domain of MCAK associates with the kinase domain of PLK1. MCAK is a novel substrate of PLK1, and the phosphorylation stimulates its microtubule depolymerization activity of MCAK in vivo. Overexpression of a polo-like kinase 1 phosphomimetic mutant MCAK causes a dramatic increase in misaligned chromosomes and in multipolar spindles in mitotic cells, whereas overexpression of a nonphosphorylatable MCAK mutant results in aberrant anaphase with sister chromatid bridges, suggesting that precise regulation of the MCAK activity by PLK1 phosphorylation is critical for proper microtubule dynamics and essential for the faithful chromosome segregation. We reasoned that dynamic regulation of MCAK phosphorylation by PLK1 is required to orchestrate faithful cell division, whereas the high levels of PLK1 and MCAK activities seen in cancer cells may account for a mechanism underlying the pathogenesis of genomic instability.

Functional analysis of phosphorylation of the mitotic centromere-associated kinesin by Aurora B kinase in human tumor cells

“…3C), in line with previous reports. 16,32 Comparable results were also obtained in HCT116 cells (Fig. S3B-E).…”

“…Additionally, a part of mutant transfected was not able to establish a bipolar spindle, leading to multipolar spindles, possibly resulted from deregulated microtubule nucleation or defective cytokinesis due to severely altered MCAK's activity. 16,32 We show further that this phosphorylation of MCAK regulates its localization on the kinetochore/centromere region in cells: the hyperactive MCAK S192A favored the kinetochores, whereas the inactive form MCAK S192D preferred the centromere region. These data suggest that mitotic failures could be ascribed to altered catalytic activity as well as to deregulated localization of MCAK at the centromere/kinetochore region, where a precise balance between active and inactive MCAK is required to correct defective kinetochore-MT attachments.…”

“…8,9 To fulfill these functions, MCAK has to be tempospatially regulated via phosphorylation events by various kinases like Aurora A, Aurora B, cyclin-dependent kinase 1 (Cdk1) and Polo-like kinase 1 (Plk1). [10][11][12][13][14] Deregulation of MCAK induces defects in spindle assembly, chromosome congression and segregation in mammalian cells, [12][13][14][15][16] leading further to chromosome instability, a hallmark of tumors. In support of this notion, overexpression of MCAK associates with a more invasive and metastatic phenotype and poor prognosis for breast, gastric and colorectal cancer patients.…”

EB1 acetylation by P300/CBP-associated factor (PCAF) ensures accurate kinetochore–microtubule interactions in mitosis

“…A central characteristic of the kinetochore-spindle interface is its capacity to orchestrate stable and dynamic associations, whereas bound microtubules are polymerizing or depolymerizing. Such properties would be best coordinated by distinct but cooperative kinetochore-microtubule binding sites regulated by signaling cascades (23)(24)(25)(26). In the mitotic spindle, EB1 interacts with its cargo proteins, APC, CLASP, MCAK, and TIP150, to achieve chromosome segregation.…”

Signaling-dependent Phosphorylation of Mitotic Centromere-associated Kinesin Regulates Microtubule Depolymerization and Its Centrosomal Localization

“…The ability of MCAK to attenuate or accelerate MT depolymerization ensures proper attachment of microtubules to kinetochores. The MCAK activity is tightly regulated by mitotic kinases such as Aurora-A, Aurora-B, Plk1, and Cdk1 (5)(6)(7)(8)(9)(10)(11)(12).…”