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A cell cycle timer for asymmetric cell
division
Asymmetric cell division is important for generating cell diversity during development. While much recent progress has been made on discovering the molecular mechanisms of mitotic spindle displacement during asymmetric cell divisions, far less is known about how spindle displacement is timed precisely. A conserved timing mechanism is known to govern progression through mitosis, although this has never been linked to timely spindle displacement. This mechanism involves the anaphase-promoting complex (APC), its activator CDC20/fizzy, and its degradation target cyclin-dependent kinase (CDK). We have found that these components can function as a timer for asymmetric spindle positioning. In the C. elegans zygote, the mitotic spindle begins to move to an asymmetric position at a precise time, just after chromosome congression to the metaphase plate. We found that reducing the function of CDC20/fizzy, the APC or the proteasome caused delays in spindle displacement. Conversely, inactivating CDK caused premature spindle displacement. Inactivating CDK could rescue most of the delay produced by reduction of APC function, suggesting that the APC functions in spindle positioning through its role in CDK inactivation, as it does in mitotic progression. Premature CDK inactivation resulted in premature displacement of incompletely assembled mitotic spindles. We conclude that in normal C. elegans embryos, asymmetric positioning of the mitotic spindle is delayed for a short time until the APC inactivates CDK, and that this delay prevents premature displacement of incompletely assembled mitotic spindles. This work reveals that a link exists between mitotic cell cycle progression and asymmetric cell division, a new link in cell and developmental biology. We speculate that this link might be evolutionarily conserved, because the mitotic spindle is displaced at a similar stage of mitosis during asymmetric cell divisions in diverse systems. Automating measurements of microtubule stability at the cell cortex The one-cell C. elegans embryo divides asymmetrically along an anterior-posterior axis, which causes the anterior cell to be larger than the posterior cell. Prior to anaphase onset, the mitotic spindle shifts from a central location in the embryo towards the posterior cortex. Astral microtubules that reach the embryo’s cortex are regulated by several players, including the PAR polarity proteins, heterotrimeric G proteins, and LET-99. By imaging the plus ends of individual astral microtubules reaching the cortex, Labbé et al. (2003) determined that the residence times of microtubules at the cortex are not uniform across the embryo. Microtubules at the cortex in anterior regions are about 15% more stable than microtubules at the cortex in posterior regions of the embryo, and this difference is dependent on the PAR proteins and heterotrimeric G proteins. Our collaborators at UNC and we have created a computer program that automates the measurements of astral microtubule residence times at the embryo’s cortex. This program locates microtubule tips at the cell cortex in a tubulin:GFP film and quantifies where and how long each microtubule resides at the cortex. We expect that automating these measurements will facilitate quantifying the microtubule stabilities in embryos of several previously untested genetic backgrounds, from which we hope to further our understanding of the molecular mechanisms that regulate microtubule dynamics at the cell cortex. In addition, our program allows us to visualize individual microtubules in novel ways that generate new, testable hypotheses regarding the regulation of microtubule dynamics.
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Erin McCarthy