Microtubule stability and the cell cortex: the role of MCAK in microtubule-cortex interactions

Dynamic microtubules are important for cell shape and maintaining genomic integrity during mitosis. The kinesin-13 family consists of middle-motor kinesins that depolymerize microtubules. The first part of this work characterizes the three mammalian kinesin-13s, Kif2A, Kif2B and Kif2C in human tissue culture cells, comparing their interphase and mitotic localizations and depolymerization activities. Kif2C, hereafter referred to as mitotic centromere-associated kinesin (MCAK) is the only mammalian kinesin-13 that tracks on the tips of polymerizing microtubules. This activity is seen in both interphase and mitosis but the function of this localization has not been investigated.;The second part of this work examines the function of MCAK localized to the tips of astral microtubules during mitosis. We depleted MCAK from mammalian tissue culture cells, examining their progression through mitosis and general mitotic function in the absence of MCAK. Depletion of MCAK results in longer metaphase spindles with strikingly long astral microtubules relative to control cells. At the onset of anaphase, live imaging reveals an extraordinary spindle rocking phenotype during late anaphase, wherein the entire mitotic spindle oscillates along the spindle axis from one proto-daughter cell to the other. The rocking of the spindle triggers a concurrent oscillation in the cortical non-muscle myosin II that continues to drive the oscillation of the spindle. Despite this dramatic contractile activity the cleavage furrow persists in closing, often on one side of the spindle, resulting in incorrect segregation of DNA and binucleation in 30% of cells exhibiting the phenotype. This spindle rocking phenotype can be mimicked by the application of the microtubule stabilizing drug taxol, which also causes long, stable astral microtubules. Additionally, both MCAK depletion and taxol treatment cause a delocalization of dynein from astral microtubules, indicating that these treatments may interfere with the cell's spindle positioning mechanism. These results demonstrate the importance of MCAK in maintaining astral microtubule length during mitosis, to allow for proper spindle positioning.