During the normal mitotic phase of the mammalian cell cycle, division of the somatic mother cell into two genetically identical daughter cells occurs through two processes: 1) mitosis—a process in which chromosomes in a cell nucleus are separated into two identical sets and 2) cytokinesis—a process following mitosis where the cytoplasm, organelles, and cell membrane are divided into two new cells. Execution of cytokinesis requires precise spatial and temporal activation of proteins that coordinate cell cycle control and cytoskeletal reorganization for the final separation of two daughter cells. It is of fundamental importance to the study of cell biology and of significant biological relevance to understand this highly regulated and complex process because cytokinesis failure leads to both centrosome amplification and production of tetraploid cells, which may set the stage for tumor initiation.
Both coordinated temporal regulation and physical-mechanical regulation of the cell is required for proper cytokinesis. The major mitotic initiator, Cyclin-dependent kinase 1 (CDK1), is an important player in the temporal regulation of cytokinesis, as CDK1 inactivation is required for proper mitotic exit and initiation of cytokinesis. The mechanical regulation of cytokinesis is split into four different stages which include: 1) specification of the cleavage plane—where the central regulator of cytokinesis, the small GTPase RHOA, is recruited to the cleavage site, 2) ingression of the cleavage furrow—where formation of an actomyosin ring and myosin-dependent motor activity causes cleavage furrow ingression, 3) formation of the midbody (a.k.a. Flemming body) and stabilization of the cytokinetic furrow—where regulatory cytokinesis proteins coordinately function to stabilize interaction between the central spindle, a microtubule based structure which forms in between segregating chromosomes during anaphase, and the actomyosin ring, and 4) abscission—when cytoplasmic contents are finally separated through narrowing of the furrow and vesicle formation and trafficking. These four stages of cytokinesis are a series of linked processes that require the proper execution of the previous stage, therefore defects at any step of the cascade can result in cytokinesis failure. My thesis work revealed that YAP depletion causes a cytokinetic defect due to misregulation of the first and second stages of cytokinesis as evidenced by disrupted midbody protein localization.
Fig. 1 Mitosis and Cytokinesis (Scholey et al. Cell Division, Nature, 2003).
In order for a cell to execute proper cell division, mitosis and cytokinesis must be coordinated. It is important that initiation of cytokinesis and furrow formation does not occur until the chromosomes have begun to segregate at the onset of anaphase. This temporal control is achieved through a number of overlapping mechanisms, which either prevent or promote the initiation of furrow and anaphase central spindle formation. The major cell cycle regulator, CDK1, is a serine/threonine kinase that phosphorylates a variety of target substrates to regulate cell cycle progression. CDK1 forms a complex with mitotic cyclin B1 and upon activation, promotes spindle assembly and chromatid alignment, thus initiating mitosis. Phosphorylation of CDK1 at Tyr15 in humans inactivates the protein and leads to the activation of the ubiquitin-protein ligase APC/CDC20, which regulates chromatid segregation and degradation cyclin B1. Thus, coordinated CDK1 inactivation leads to the final events of mitosis and initiation of cytokinesis.
Mitotic microtubules send positive signals to initiate furrowing at the correct place in the cell by regulating the RHOA pathway. Microtubules coordinate the localized activation of RHOA in a narrow zone at the future cleavage furrow site. Perturbations that broaden the zone of RHOA activation can lead to furrow ingression or even failure to furrow. In order to form a narrow zone of RHOA activation, RHOA activators such as the centralspindlin complex, which initiates the formation of the central spindle, and ECT2, a RHOA guanine nucleotide exchange factor, need to be recruited to the central spindle by microtubules to establish a strong, spatially restricted signal region for promoting proper temporal initiation of cytokinesis.
Upon further constriction of the furrow at later stages of cytokinesis, the central spindle region develops into a structure known as the mammalian midbody. The midbody is a morphologically dense structure that gradually narrows into a tubular intercellular bridge that transiently connects the two daughter cells. The midbody becomes the site of daughter cell partitioning in a process called abscission. Several cytokinesis-coupled events converge at the midbody to regulate the separation of the two daughter cells, including degradation of cell cycle regulators, cytoskeletal rearrangements, membrane trafficking, and plasma membrane remodeling. Although its precise composition is not known, the midbody consists of tightly bundled interpolar microtubules with a midbody ring, a central protein ring containing numerous microtubule interacting proteins critical for cytokinesis, such as vesicle transport machinery (ie. ESCRT), actomyosin, and, importantly, RHOA regulatory components.