Related Experiment Video
Updated: Apr 17, 2026

Identification of Cyclin-dependent Kinase 1 Specific Phosphorylation Sites by an In Vitro Kinase Assay
Published on: May 3, 2018
Sequential multisite phospho-regulation of KNL1-BUB3 interfaces at mitotic kinetochores
Mathijs Vleugel1, Manja Omerzu2, Vincent Groenewold2
1Molecular Cancer Research, University Medical Center Utrecht, 3584 CG Utrecht, The Netherlands.
Abstract:
Regulated recruitment of the kinase-adaptor complex BUB1/BUB3 to kinetochores is crucial for correcting faulty chromosome-spindle attachments and for spindle assembly checkpoint (SAC) signaling. BUB1/BUB3 localizes to kinetochores by binding phosphorylated MELT motifs (MELpT) in the kinetochore scaffold KNL1. Human KNL1 has 19 repeats that contain a MELT-like sequence. The repeats are, however, larger than MELT, and repeat sequences can vary significantly. Using systematic screening, we show that only a limited number of repeats is "active." Repeat activity correlates with the presence of a vertebrate-specific SHT motif C-terminal to the MELT sequence. SHT motifs are phosphorylated by MPS1 in a manner that requires prior phosphorylation of MELT. Phospho-SHT (SHpT) synergizes with MELpT in BUB3/BUB1 binding in vitro and in cells, and human BUB3 mutated in a predicted SHpT-binding surface cannot localize to kinetochores. Our data show sequential multisite regulation of the KNL1-BUB1/BUB3 interaction and provide mechanistic insight into evolution of the KNL1-BUB3 interface.
More Related Videos
08:33Combining Mitotic Cell Synchronization and High Resolution Confocal Microscopy to Study the Role of Multifunctional Cell Cycle Proteins During Mitosis
Published on: December 5, 2017
07:20Simultaneous Visualization of the Dynamics of Crosslinked and Single Microtubules In Vitro by TIRF Microscopy
Published on: February 18, 2022
Related Concept Videos
Attachment of Sister Chromatids
Disassembly of Intermediate Filaments
Keratin proteins, found at the cell periphery near cell junctions, undergo a cycle of assembly and disassembly. In Type...
Separation of Sister Chromatids
At the onset of anaphase, separase, a proteolytic enzyme, is...
Anaphase A and B
Plus-end depolymerization releases tubulin heterodimers from the terminal region of the microtubule. As tubulin subunits are lost, the Ndc80 complexes detach...
The Spindle Assembly Checkpoint
Many proteins function together to control the spindle assembly checkpoint. Mutations affecting these proteins may allow cells to proceed into anaphase prematurely, resulting in the...
M-Cdk Drives Transition Into Mitosis
Cyclin-dependent kinases, or Cdks, work in concert with cyclins to control cell cycle transitions. M-Cdk, a complex of Cdk1 bound to M cyclin, is a well-known example of this coordinated control that drives the transition from the G2 to the M phase.
M cyclin...