title: Distinct roles for antiparallel microtubule pairing and overlap during early spindle assembly.
abstract: During spindle assembly, microtubules may attach to kinetochores or pair to form antiparallel pairs or interpolar microtubules, which span the two spindle poles and contribute to mitotic pole separation and chromosome segregation. Events in the specification of the interpolar microtubules are poorly understood. Using three-dimensional electron tomography and analysis of spindle dynamical behavior in living cells, we investigated the process of spindle assembly. Unexpectedly, we found that the phosphorylation state of an evolutionarily conserved Cdk1 site (S360) in γ-tubulin is correlated with the number and organization of interpolar microtubules. Mimicking S360 phosphorylation (S360D) results in bipolar spindles with a normal number of microtubules but lacking interpolar microtubules. Inhibiting S360 phosphorylation (S360A) results in spindles with interpolar microtubules and high-angle, antiparallel microtubule pairs. The latter are also detected in wild-type spindles <1 μm in length, suggesting that high-angle microtubule pairing represents an intermediate step in interpolar microtubule formation. Correlation of spindle architecture with dynamical behavior suggests that microtubule pairing is sufficient to separate the spindle poles, whereas interpolar microtubules maintain the velocity of pole displacement during early spindle assembly. Our findings suggest that the number of interpolar microtubules formed during spindle assembly is controlled in part through activities at the spindle poles.
authors: Department of Biology, McGill University, Montreal, QC H3G 0B1, Canada Department of Physics, McGill University, Montreal, QC H3G 0B1, Canada Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder CO 80309.
authors: Elena
authors: E
authors: Nazarova
authors: Eileen
authors: E
authors: O'Toole
authors: Susi
authors: S
authors: Kaitna
authors: Paul
authors: P
authors: Francois
authors: Mark
authors: M
authors: Winey
authors: Jackie
authors: J
authors: Vogel
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D016203:CDC2 Protein Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D055032:Electron Microscope Tomography; D018386:Kinetochores / Q000378:metabolism* / Q000648:ultrastructure; D008870:Microtubules / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D010766:Phosphorylation; D012441:Saccharomyces cerevisiae; D008941:Spindle Apparatus / Q000235:genetics* / Q000648:ultrastructure; D013329:Structure-Activity Relationship
chemical_list: D016203:CDC2 Protein Kinase
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