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Zwint-1 is a novel Aurora B substrate required for the assembly of a dynein-binding platform on kinetochores.
- Source :
-
Molecular biology of the cell [Mol Biol Cell] 2011 Sep; Vol. 22 (18), pp. 3318-30. Date of Electronic Publication: 2011 Jul 20. - Publication Year :
- 2011
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Abstract
- Aurora B (AurB) is a mitotic kinase responsible for multiple aspects of mitotic progression, including assembly of the outer kinetochore. Cytoplasmic dynein is an abundant kinetochore protein whose recruitment to kinetochores requires phosphorylation. To assess whether AurB regulates recruitment of dynein to kinetochores, we inhibited AurB using ZM447439 or a kinase-dead AurB construct. Inhibition of AurB reduced accumulation of dynein at kinetochores substantially; however, this reflected a loss of dynein-associated proteins rather than a defect in dynein phosphorylation. We determined that AurB inhibition affected recruitment of the ROD, ZW10, zwilch (RZZ) complex to kinetochores but not zwint-1 or more-proximal kinetochore proteins. AurB phosphorylated zwint-1 but not ZW10 in vitro, and three novel phosphorylation sites were identified by tandem mass spectrometry analysis. Expression of a triple-Ala zwint-1 mutant blocked kinetochore assembly of RZZ-dependent proteins and induced defects in chromosome movement during prometaphase. Expression of a triple-Glu zwint-1 mutant rendered cells resistant to AurB inhibition during prometaphase. However, cells expressing the triple-Glu mutant failed to satisfy the spindle assembly checkpoint (SAC) at metaphase because poleward streaming of dynein/dynactin/RZZ was inhibited. These studies identify zwint-1 as a novel AurB substrate required for kinetochore assembly and for proper SAC silencing at metaphase.
- Subjects :
- Amino Acid Substitution
Animals
Aurora Kinase B
Aurora Kinases
Benzamides pharmacology
Dynactin Complex
HeLa Cells
Humans
Intracellular Signaling Peptides and Proteins genetics
M Phase Cell Cycle Checkpoints
Metaphase
Microscopy, Fluorescence
Microtubule-Associated Proteins metabolism
Mutagenesis, Site-Directed
Nuclear Proteins genetics
Phosphorylation
Protein Serine-Threonine Kinases antagonists & inhibitors
Quinazolines pharmacology
Rats
Single-Cell Analysis
Time-Lapse Imaging
Cytoplasmic Dyneins metabolism
Intracellular Signaling Peptides and Proteins metabolism
Kinetochores metabolism
Nuclear Proteins metabolism
Protein Serine-Threonine Kinases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1939-4586
- Volume :
- 22
- Issue :
- 18
- Database :
- MEDLINE
- Journal :
- Molecular biology of the cell
- Publication Type :
- Academic Journal
- Accession number :
- 21775627
- Full Text :
- https://doi.org/10.1091/mbc.E11-03-0213