Sequential Multisite Phospho-Regulation of KNL1-BUB3 Interfaces at Mitotic Kinetochores
Copyright policy )Sequential Multisite Phospho-Regulation of KNL1-BUB3 Interfaces at Mitotic Kinetochores
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.
- Utrecht University Netherlands
- University Medical Center Utrecht Netherlands
Repetitive Sequences, Amino Acid, Models, Molecular, Amino Acid Motifs, Immunoblotting, Molecular Sequence Data, Mitosis, Cell Cycle Proteins, Protein Serine-Threonine Kinases, Time-Lapse Imaging, Humans, Amino Acid Sequence, Phosphorylation, Kinetochores, Poly-ADP-Ribose Binding Proteins, Molecular Biology, Sequence Homology, Amino Acid, Nocodazole, Cell Biology, Protein-Tyrosine Kinases, Protein-Serine-Threonine Kinases, Tubulin Modulators, Protein Structure, Tertiary, Mutation, M Phase Cell Cycle Checkpoints, RNA Interference, Microtubule-Associated Proteins, HeLa Cells, Protein Binding
Repetitive Sequences, Amino Acid, Models, Molecular, Amino Acid Motifs, Immunoblotting, Molecular Sequence Data, Mitosis, Cell Cycle Proteins, Protein Serine-Threonine Kinases, Time-Lapse Imaging, Humans, Amino Acid Sequence, Phosphorylation, Kinetochores, Poly-ADP-Ribose Binding Proteins, Molecular Biology, Sequence Homology, Amino Acid, Nocodazole, Cell Biology, Protein-Tyrosine Kinases, Protein-Serine-Threonine Kinases, Tubulin Modulators, Protein Structure, Tertiary, Mutation, M Phase Cell Cycle Checkpoints, RNA Interference, Microtubule-Associated Proteins, HeLa Cells, Protein Binding
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