During oocyte meiotic cell divisions, bipolar spindles assemble without centrosomes, and the mechanisms that mediate pole assembly and limit it to a bipolar state are unknown. Using live cell imaging and the oocyte meiotic spindle pole marker GFP::ASPM-1, we have found that multiple small pole foci initially are present early in meiotic spindle assembly and then coalesce to form two foci and a bipolar spindle. Thus-as has been observed by live imaging of microtubules in mouse oocytes1-in the absence of centrosomal organizing centers, multiple small poles initially form and then coalesce to produce a bipolar spindle. We have gained insight into this coalescence from a screen for meiotic spindle assembly defective mutants, in which we identified two temperature-sensitive alleles of
klp-7, the lone MCAK/kinesin-13 gene in C. elegans. MCAKs are known to function as microtubule depolymerases, and we found that extra microtubules accumulated during oocyte meiosis I in
klp-7(-) mutants, and the multiple GFP::ASPM-1 foci that formed early in meiosis I consistently coalesced to form three and sometimes four foci that persisted throughout most of meiosis I. Moreover, paired chromosome homologues frequently aligned along more than one pole-to-pole axis, and the chromosomes frequently segregated as three distinct masses during anaphase. We conclude that KLP-7 is required for the coalescence of multiple early pole foci to a bipolar state, and that in the absence of KLP-7 extra functional poles persist. Because MCAKs are thought to function at kinetochores to detach inappropriate (syntelic) kinetochore-microtubule (k-MT) attachments2, we hypothesized that in the absence of KLP-7/MCAK, persistent syntelic k-MT attachments result in abnormal spindle tension that interferes with the coalescence of early pole foci and results in the persistence of additional poles. To test this hypothesis, we knocked down components of the kinetochore complex Ndc-80, which bind microtubules to mediate kinetochore attachment3, and found that Ndc-80 depletion restored bipolarity in most
klp-7(-) oocytes. We therefore propose a model in which KLP-7/MCAK promotes early pole foci coalescence and spindle bipolarity by regulating k-MT attachment to relieve imbalances in spindle tension, and we are further investigating the influences of kinetochore function on the acentrosomal process of oocyte meiotic spindle assembly.1. Schuh and Ellenberg, 2007 Cell 130: 484-98.2. Wordeman et al., 2007 The Journal of Cell Biology 179: 869-793. Cheeseman et al., 2006 Cell 127: 983-97.