When is cytokinesis

Originally, discoidins were suggested to play a role in mediating cell—cell interaction owing to their lectin properties. However, Discoidin I was recently found to interact genetically and biochemically with a key protein involved in cell contractility and cytokinesis, namely cortexillin I Robinson and Spudich, ; Kothari et al. Myosin II and the actin crosslinker Cortexillin I are essential components of the CN and their mechanosensory ability and accumulation at the cleavage furrow were proven to be vital for the progression and completion of cytokinesis Faix et al.

Interestingly, discoidin also acts as a genetic suppressor of cortexillin I null mutants through a selection for genetic suppressors where discoidin overexpression could rescue the developmental phenotypes of cortexillin I null cells Robinson and Spudich, Collectively, discoidin proteins might provide a scaffolding function and help recruit essential components of the CN system to the division site during cytokinesis.

Another possibility is that the discoidins may help to directly connect the CN to the plasma membrane. It is not fully understood in any system how the CN is attached to the membrane, particularly in the context of cytokinesis. Dividing Dictyostelium cells show an enrichment of glycosylated proteins at the cleavage furrow membrane as the lectin concanavalin A preferentially labels the cleavage furrow membrane Faix et al. Thus, the combination of observations suggests a provocative hypothesis that extracellular discoidin might also interact with the cortical cytoskeleton through a transmembrane domain containing protein.

The human discoidin domain receptor DDR1 protein does exactly this, except it contains a transmembrane domain and interacts with non-muscle myosin IIA Huang et al. Invasive tumors and associated metastases show elevated expression of DDR1, suggesting its role as a promoter of tumor cell invasion Yang et al.

Discoidin was not the first sugar-related protein that was speculated to be crucial in cytokinesis-related processes. Mucin is also a glycosylated protein and is a component of ring canals and intercellular bridges, structures formed by incomplete cytokinesis, in various cell types and tissues in Drosophila melanogaster Robinson and Cooley, ; Kramerova and Kramerov, Even though the exact functions of mucinoprotein in ring canal networks are not well-understood, it is possible that it might be an important constituent of a complex or backbone that facilitates the assembly of these structures.

Further studies are needed to elaborate on the exact roles and functions of discoidin and other sugar-binding proteins that take part in cytokinesis. Another unexpected class of membrane proteins that were found to play a role in cytokinesis are the highly-conserved chloride intracellular channels CLICs. Comprised of six members that exist primarily as membrane-bound proteins, CLICs are involved in diverse cellular functions, including cell-cell interactions, angiogenesis, and membrane potential regulation Rual et al.

Multiple studies have also implicated CLIC4 in cancer progression. The expression level of CLIC4 was found to be gradually decreased in squamous cancer cells as they transformed from benign to malignant, a prominent indicator of a role in mitosis regulation Suh et al.

Additionally, CLIC4 is a direct molecular interactor of profilin-1 and a component in the RhoA-mDia2 signaling pathway that induces cortical actin polymerization Argenzio et al. A previous proteomics study investigating the biochemical changes at the cell surface during cell division revealed a significant enrichment of both CLIC4 and CLIC1 on the surface of rounded up mitotic cells compared with flat interphase cells Ozlu et al.

It was also previously found that cortical actin controls cell cycle progression, and tumor cells lose their dependence on cortical branched actin Molinie et al. Through their role in maintaining cortical and CN stability, CLICs might facilitate normal mitotic progression and prevent the transformation from benignity to malignancy in cancer cells.

Overall, CLIC4 and CLIC1 are two additional membrane-associated proteins that are recruited to the cortex, where they potentially help anchor the CN, allowing for proper contraction and cell division. While the exact functions of many RNPs are still being defined, studies in some systems indicate that RNPs contribute to cytokinesis. In Dictyostelium , overexpression of RNP-1A suppresses the effect of nocodazole, a microtubule inhibitor, on cellular growth of Dictyostelium discoideum Zhou et al.

This protein has protective functions on the microtubule ends, and also localizes to the polar cortex in dividing cells and to the leading edges of migrating cells. These strongly suggest a role of RNP-1A in regulating microtubule dynamics, possibly through stabilizing the linkage between microtubules and the cortex. In the same study, using fluorescence cross-correlation spectroscopy, RNP-1A was found to interact strongly with cortexillin I.

The full significance of these interactions is still under investigation. The recognition of RNA-binding proteins as potential effectors of cytokinesis was not entirely surprising, as CAR-1, another RNA-binding protein found at P granules and the mitotic spindle during cell division, was identified using an RNA-interference screen for genes essential for cytokinesis in C.

In car-1 -depleted embryos, cleavage furrow ingression was significantly disrupted, and the anaphase spindle structure was greatly defective Audhya et al. As the study was conducted primarily in early C. This effect was similar to a previously characterized RNA-binding protein, CPEB, whose inhibition also results in spindle structural defects Groisman et al. Overall, these roles of RNA-binding proteins in cytokinesis regulation elucidates an important crosstalk between local protein translation and steps in cell division.

It is tempting to speculate that the unique localization and interactions of these RNP-1 proteins facilitates localized mRNA translation during cytokinesis when cells are undergoing dynamic and robust cell shape changes. The importance of RNA helicases in regulating cell cycle progression and mitosis is not entirely surprising. Indeed, since the progression of the cell cycle requires the coordination of enormous cohorts of differentially expressed proteins through each step, cells need mechanisms to tightly regulate mRNA transcription, translation, and protein degradation to facilitate this dynamic process.

For example, RNA helicases have been implicated to play critical roles in mRNA export pathways that are tightly linked to gene products involved in mitosis regulation Strasser and Hurt, ; Herold et al.

Depletion of these RNA helicases, as a result, leads to mitotic progression defects. However, unexpected functions of several RNA helicases in cytokinesis independent from its traditional roles in regulating mitotically expressed proteins have been discovered. Embryos depleted of cgh-1 had penetrant sterility while those partially depleted exhibited perturbed localization of CAR-1 and phenocopied car-1 -depleted cells with defects in microtubule structures.

Interestingly, CGH-1 is the C. The exact functions of DDXs in regulating cell cycle and tumorigenesis remain controversial. Knockdown of DDX3 in mice embryonic cells led to reduced growth and proliferation Li et al. However, a recent study elucidated a possible mechanism through which DDX3, another member of the DDX family, positively controls cytokinesis and subsequently acts as a tumor suppressor. DDX3 localizes to centrosomes throughout the cell cycle and prevents chromosome misalignment by inactivating and aggregating supernumerary centrosomes Chen et al.

DDX silencing subsequently led to chromosome misalignment, segregation defects, and eventually cell death. Moreover, DDX3 localizes to the midbody during late cell division Chen et al. These studies support the possibility of novel roles of RNA helicases in maintaining structural integrity of essential mitotic and cytokinesis components, therefore ensuring successful completion of cell division.

Thus, free importins may function as a molecular ruler, or buffer, for which an optimal level is required to maintain the appropriate cortical recruitment of anillin.

Ras-related nuclear protein Ran is a small G-protein involved in transporting various proteins and cellular components in and out of the nucleus through the nuclear pore complex. The function of Ran is tightly regulated by the GTP gradient across the nuclear membrane. Ran has been implicated in numerous cellular processes such as DNA synthesis, nuclear envelope structure, and cell cycle progression Sazer and Dasso, Mounting evidence now suggests that Ran also participates in regulating cytokinesis.

Recent studies revealed that chromatin-associated signals can regulate the cortical dynamics during cytokinesis Norden et al. Due to its concentration gradient with the peak occurring around the chromatin, chromatin-associated Ran-GTP is potentially also a regulator of the cortex. During meiosis of mouse oocytes, chromatin positioned near the cortex induces the formation of an actin cap via Ran-GTP, and the placement of DNA-coated beads near the cortex induces cortical polarity independent of microtubules Deng et al.

Active Ran regulates human anillin during anaphase and decreasing Ran-GTP leads to the ectopic localization of anillin and myosin to the cell poles and blocks proper furrowing Beaudet et al. These studies suggested that a chromatin-associated Ran-GTP gradient may function as a molecular ruler that helps recruit and organize essential components at the cortex. In another study, Ran-GTP was found to positively regulate the actomyosin cortex for pseudocleavage furrowing in the early Drosophila embryo development Silverman-Gavrila et al.

Ran controls pseudocleavage furrow organization independently of its role in regulating the microtubule cytoskeleton. Further, disruption of the Ran pathway prevented pseudocleavage furrow formation and restricted the depth and duration of furrow ingression of those pseudocleavage furrows that did form. To further elucidate the mechanism through which Ran-GTP acts to regulate cytokinesis, the authors found that Ran is required for the pseudocleavage furrow localization of the septin, peanut, a protein whose association with anillin contributes to stabilization of the CN.

Therefore, Ran-GTP plays an important role in pseudocleavage furrow ingression in syncytial embryos. Overall, Ran-GTP acts as a microtubule-independent pathway that regulates polarization of contractile protein anillin a process traditionally thought to require microtubules to couple signals from the chromatin to the cortex and to ensure robustness of cytokinesis. Lamins are important constituents of the nuclear lamina in eukaryotes. Apart from their functions in maintaining the structural integrity of the nuclear pores and membranes, lamins are crucial for the assembly and disassembly of the nuclear envelope during mitosis Lopez-Soler et al.

In addition to its canonical roles within the nucleus, lamin B, a ubiquitously expressed type of lamin, helps regulate mitosis. In M-phase-arrested Xenopus egg extracts, lamin B associates with the mitotic spindles as well as the surrounding regions of the spindle. When expression of lamin B was reduced using siRNA in HeLa cells, typical spindle defects such as poor spindle morphology and lack of chromosome segregation were detected, suggesting a role for lamin B in facilitating spindle assembly Tsai et al.

In the same study, the authors also reported that lamin B is also an essential component in the formation of the mitotic matrix, allowing for tethering for other spindle assembly factors. Dominant negative forms of lamin B significantly disrupted this matrix formation, leading to severing of the mitotic spindles and cytokinesis arrest. These results indicate the significant role of lamin B in maintaining the integrity of mitotic spindles during cytokinesis.

The prevailing idea to explain these observations is that the disassembled lamin B is dispersed throughout the cytoplasm during mitosis. This pattern of localization of lamin B elucidates its novel role in regulating cytokinesis.

Interestingly, association of lamin B with the mitotic spindle is regulated by Ran-GTP, eliciting the possibility of an entirely novel pathway of mitotic spindle regulation involving Ran and its downstream targets Ma et al. Strikingly, ESCRT-III-mediated membrane fusion facilitates reintegration of lagging chromosome fragments into micronuclei through lamin-based channels during cytokinesis in Drosophila , emphasizing the intricate integration between the abscission and chromosome segregation machineries during cytokinesis Warecki et al.

Methylmalonate-semialdehyde dehydrogenase mmsdh is an enzyme typically thought of being located in the mitochondria and that helps catalyze the production of propionyl- and acetyl-CoA Kedishvili et al.

Recent studies suggests a possible novel role of mmsdh in regulating cytokinesis. Previously identified in a genetic suppression screen in Dictyostelium , over-expression of mmsdh suppressed the dominant-negative phenotype of a myosin II phosphomimetic Ren et al. For context, to generate force, the non-muscle myosin II must polymerize into bipolar filaments, and this assembly is regulated by heavy chain phosphorylation.

The myosin II phosphomimetic mimics this phosphorylated state and impairs bipolar filament assembly Egelhoff et al. Mmsdh promoted myosin II phosphomimetic accumulation at the cortex and cleavage furrow in myoII null cells, thus potentially acting as a modulator of myosin II function Ren et al. Importantly, mmsdh showed up again as a molecular interactor of cortexillin I and IQGAP2, core mechanosensory proteins in the contractility system Kothari et al. It is possible that mmsdh functions in cytokinesis through a metabolic pathway that leads to post-translational modification of several contractile enzymes, including myosin II.

Mmsdh catalyzes the degradation of valine, leading to the production of propionyl-CoA. Indeed, post-transtional modifications are important in modulating proteins involved in cytokinesis.

In mammalian cells, anillin and myosin IIA are acetylated during cytokinesis, and histone deacetylase HDAC inhibitors lead to cytokinetic defects, consistent with this concept Chuang et al. AMPK is present in the nucleus and cytoplasm, and its intracellular distribution is regulated by stress and cell growth Kodiha et al.

In addition to its role as an energy sensor, AMPK was found to act as a suppressor of cell proliferation. Enhanced activation of AMPK inhibits tumor growth and oncogenic transformation of several cancer cell lines Zakikhani et al. Many studies followed to elucidate the mechanisms of AMPK regulating cell division. Substantial evidence now suggests that AMPK plays obligatory role in chromosome segregation and cytokinesis completion.

Furthermore, kinase activity of Plk1, a major regulator of mitotic progression, regulates localization and activation of AMPK at the mitotic apparatus. Studies in budding yeast reveal similar observations. Loss of Snf1, the budding yeast ortholog of AMPK, caused cells to display aberrant spindle alignment, further highlighting its role in regulating the mitotic spindle Tripodi et al. The exact mechanisms and functions of AMPK in the context of cytokinesis regulation remain to be elucidated, but AMPK could be central to physically and temporally tethering the energy state of a cell to cell cycle progression.

While substantial progress has been made with regard to the intricacies of the CN and mitotic spindle formation and function, we are now in position to appreciate that cytokinesis results from the function of a truly integrated and collaborative cellular system Figure 2. Further, these examples of unusual suspects in cytokinesis can help motivate us to think beyond the scope of just a few sub-cellular systems and pathways that drive a cellular process. We also should take into consideration that under physiological contexts, a cellular system is under constant environmental pressures that can be chemical and mechanical in nature.

Cells are exquisitely exceptional at coping with these challenges. Recent studies have drawn upon this principle to identify new roles of known factors as well as new unknown factors. For instance, a study recently published used mechanically loaded cellular systems to study cytokinesis in the context of mechanics and identified new functions of several factors Singh et al. The authors found that factors such as MEL and LIN-5, canonically involved in regulating myosin II function and spindle positioning, respectively, are also involved in regulating cortical rotation, further elucidating how cells simultaneously respond to and create forces during cytokinesis.

Figure 2. Piecing together the unusual suspects of the cytokinesis system. A An overview of cytokinesis with both the usual and unusual suspects, represented by the jigsaw pieces. Expanding our understanding of cytokinesis by studying these unusual suspects allows us to fully appreciate an intricate cellular system complete with cross talks, redundancy, and network integration. B Membrane-associated proteins. Upon activation of RhoA, CLICs localize to the cortex where they bind various cortical components, such as profilin-1 and ezrin.

Discoidin, through its interaction with the actin crosslinker cortexillin I and its regulator IQGAP2, might facilitate the assembly of the contractility network. C RNA-associated proteins. RNP-1A through its protection of microtubules during cytokinesis may stabilize the mitotic spindle, ensuring successful cytokinesis. On the other hand, RNP-1B interacts with cortexillin I and might play a role in regulating the contractility network.

D Nuclear proteins. In addition, lamin B, an interactor of Ran, is crucial for mitotic spindle assembly. E Metabolic enzymes. Along with its original known biochemical activity in valine degradation, leading to the production of propionyl- and acetyl-CoA, mmsdh may promote the post-translational modification of contractile proteins, such as myosin II.

AMPK is also important for mitotic apparatus assembly and regulation. Activation of AMPK by Plk1 may be important to couple cellular bioenergetic state to cell cycle progression. Expanding our understanding of cytokinesis by studying these unusual suspects allows us to fully appreciate an intricate cellular system loaded with cross talks, redundancy, and network integration.

By doing so, we will begin to better understand how other systems operate. For example, RNPs localize to and help organize P-granules, leading to the concept of phase transitions. In most species , cohesin is largely removed from the arms of the sister chromatids during prophase, allowing the individual sister chromatids to be resolved. Cohesin is retained, however, at the most constricted part of the chromosome, the centromere Figure 9. During prophase, the spindle also begins to form as the two pairs of centrioles move to opposite poles and microtubules begin to polymerize from the duplicated centrosomes.

Prometaphase begins with the abrupt fragmentation of the nuclear envelope into many small vesicles that will eventually be divided between the future daughter cells.

The breakdown of the nuclear membrane is an essential step for spindle assembly. Because the centrosomes are located outside the nucleus in animal cells, the microtubules of the developing spindle do not have access to the chromosomes until the nuclear membrane breaks apart. Prometaphase is an extremely dynamic part of the cell cycle. Microtubules rapidly assemble and disassemble as they grow out of the centrosomes, seeking out attachment sites at chromosome kinetochores, which are complex platelike structures that assemble during prometaphase on one face of each sister chromatid at its centromere.

As prometaphase ensues, chromosomes are pulled and tugged in opposite directions by microtubules growing out from both poles of the spindle, until the pole-directed forces are finally balanced. Sister chromatids do not break apart during this tug-of-war because they are firmly attached to each other by the cohesin remaining at their centromeres. At the end of prometaphase, chromosomes have a bi-orientation, meaning that the kinetochores on sister chromatids are connected by microtubules to opposite poles of the spindle.

Next, chromosomes assume their most compacted state during metaphase, when the centromeres of all the cell's chromosomes line up at the equator of the spindle.

Metaphase is particularly useful in cytogenetics , because chromosomes can be most easily visualized at this stage.

Furthermore, cells can be experimentally arrested at metaphase with mitotic poisons such as colchicine. Video microscopy shows that chromosomes temporarily stop moving during metaphase.

A complex checkpoint mechanism determines whether the spindle is properly assembled, and for the most part, only cells with correctly assembled spindles enter anaphase. Figure 10 Figure Detail. Figure 9. The progression of cells from metaphase into anaphase is marked by the abrupt separation of sister chromatids. A major reason for chromatid separation is the precipitous degradation of the cohesin molecules joining the sister chromatids by the protease separase Figure Two separate classes of movements occur during anaphase.

During the first part of anaphase, the kinetochore microtubules shorten, and the chromosomes move toward the spindle poles. During the second part of anaphase, the spindle poles separate as the non-kinetochore microtubules move past each other.

These latter movements are currently thought to be catalyzed by motor proteins that connect microtubules with opposite polarity and then "walk" toward the end of the microtubules. Mitosis ends with telophase, or the stage at which the chromosomes reach the poles. The nuclear membrane then reforms, and the chromosomes begin to decondense into their interphase conformations. Telophase is followed by cytokinesis, or the division of the cytoplasm into two daughter cells. The daughter cells that result from this process have identical genetic compositions.

Cheeseman, I. Molecular architecture of the kinetochore-microtubule interface. Nature Reviews Molecular Cell Biology 9 , 33—46 doi Cremer, T. Chromosome territories, nuclear architecture and gene regulation in mammalian cells. Nature Reviews Genetics 2 , — doi Hagstrom, K. Condensin and cohesin: More than chromosome compactor and glue.

Nature Reviews Genetics 4 , — doi Hirano, T. At the heart of the chromosome: SMC proteins in action. Nature Reviews Molecular Cell Biology 7 , — doi Mitchison, T. Mitosis: A history of division.

Nature Cell Biology 3 , E17—E21 doi Paweletz, N. Walther Flemming: Pioneer of mitosis research. Nature Reviews Molecular Cell Biology 2 , 72—75 doi Satzinger, H. Theodor and Marcella Boveri: Chromosomes and cytoplasm in heredity and development. Nature Reviews Genetics 9 , — doi Chromosome Mapping: Idiograms. Human Chromosome Translocations and Cancer. Karyotyping for Chromosomal Abnormalities. Prenatal Screen Detects Fetal Abnormalities. Synteny: Inferring Ancestral Genomes.

Telomeres of Human Chromosomes. Chromosomal Abnormalities: Aneuploidies. Chromosome Abnormalities and Cancer Cytogenetics. Copy Number Variation and Human Disease. Nuclear envelopes reassemble around each set of chromosomes. During cytokinesis in animal cells, actin filaments form a contractile ring in the plasma membrane to create a cleavage furrow, which eventually pinches the cell into two.

In plant cells, vesicles from the Golgi apparatus carrying glucose, enzymes and structural proteins join to form a new cell plate at the location of the former metaphase plate. The growing cell plate fuses with the plasma membranes on each side, eventually forming a new cell wall that divides the cell into two. Mitosis is now complete, generating two daughter cells that are identical to the parent cell. In most human cells, mitosis accounts for about one hour of the approximately hour cell cycle.

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