A prelude to cleavage: Orchestrating cytokinesis onset in mammalian cells

Abstract

The molecular mechanisms underlying cytokinesis appear complex and involve redundancy at various levels. Signals from the anaphase spindle define where and when cytokinesis is initiated but the biochemical nature of these signals is still unclear (reviewed in chapter 1). Whilst the spindle midzone was considered the main determinant of cytokinesis initiation in small mammalian cells, we disagreed with this notion based on the observation that small human cells lacking an organized spindle midzone could still initiate cytokinesis (Mollinari et al., 2005; Zhu et al., 2006). This suggested alternative routes to furrow ingression in human cells. In chapter 2, we studied the regulation of spindle midzone-independent cytokinesis initiation by knockdown of PRC1. We found that cytokinesis initiation in these cells could occur in the absence of PLK1 activity. Instead, we found it required a pool of Aurora B kinase localized at the equatorial cortex. Uncovering a role for cortical-associated Aurora B in cytokinesis initiation led us to question how Aurora B reaches the equatorial cortex in anaphase. Since this localization required the presence of the kinesin-6 motor MKLP2 and astral microtubules, we tested the hypothesis that Aurora B, as part of the CPC, is transported to the cortex by MKLP2 in chapter 3. Using microscopy-based in vitro reconstitution assays with purified proteins and live cell imaging, we provide evidence that supports this hypothesis. Mass Spectrometric analysis of MKLP2 purified from human cells identified MAP7 (Microtubule Associated Protein 7, Ensconsin) and MAP7D2 as interactors of MKLP2. In chapter 4, we demonstrate that MAP7 is required for the proper localization of MKLP2 and the CPC in anaphase. Without MAP7, MKLP2 and the CPC appear as foci resembling liquid condensates between the segregating chromosomes in anaphase. Finally, in chapter 5 we discuss these new findings and how they fit in the previously discussed mechanistic models of cytokinesis initiation.