Mitosis is a type of cell division in which one cell (the mother) divides to produce two new cells (the daughters) that are genetically identical to itself. In the context of the cell cycle, mitosis is the part of the division process in which the DNA of the cell's nucleus is split into two equal sets of chromosomes. Comparing mitosis and meiosis. Chromosomal crossover in meiosis I. Phases of meiosis I. Phases of meiosis II. This is the currently selected item. Sexual life cycles. Practice: Meiosis. Cell cycle regulation, cancer, and stem cells.
Sullivan, in, 2001Mitosis, the process of somatic cell division, has been one of the most closely studied cellular processes since microscopists first witnessed dividing cells. Cell proliferation through mitosis is fundamental to development, growth, and tissue maintenance and so influences human biology and medicine at fundamental levels. Mitosis is primarily a large-scale mechanical reorganization of the cell in which chromosome segregation and cytoplasmic fission are precisely choreographed to provide error-free cell replication. In parallel, an intricate network of regulatory enzymes and interactions guides the cell through mitosis.
Defects in mitotic regulation are central in the establishment, growth, and genomic instability of human tumors. The geometry of cell division is critical for properly partitioning cells with different cytoplasmic contents in early development and contributes to tissue architecture through directed asymmetry. This evolving view of mitosis has resolved some of the earliest puzzles of mitosis and defined new questions at the molecular and cellular levels that will profoundly impact human health. Franziska Teusel.
Mayer, in, 2018 AbstractMitosis belongs to the most appealing cellular processes. Yet, the highly dynamic and complex nature of mitosis represents a major challenge when it comes to the functional dissection of mitotic proteins. Due to their fast and often reversible mode of action, small molecules have proven themselves as invaluable tools to dissect mitotic processes. In this chapter, we provide a broad overview of available compounds affecting mitosis. We discuss the different application fields of small molecules and important aspects that have to be considered when using them.
Finally, we provide two detailed protocols for the application of small molecules to study mitosis in tissue culture cells. Mitosis and migration of stromal cells are noted approximately 8–12 hours after the initial corneal injury.
13 Initially, most cells undergoing mitosis appear to be keratocytes, but corneal fibroblasts and other cells may make subsequent contributions to this response. This cellular mitosis response provides corneal fibroblasts and other cells that participate in corneal wound healing and replenish the stroma. Once again, localization of the stromal mitosis response is related to the type of injury.
Thus, in PRK stromal mitosis tends to occur in the anterior stroma, as well as in the peripheral and posterior stroma outside the zone of apoptosis ( Figure 3.2). In LASIK, stromal mitosis occurs at the periphery of the flap where the epithelium was injured, and anterior and posterior to the lamellar cut. Stromal cell mitosis at 24 hours after photorefractive keratectomy. Arrows indicate cells in the stroma that stain for Ki-67, a marker for mitosis.
Blue is the 4’,6-diamidino-2-phenylindole (DAPI) stain for the nucleus that stains all cells. 500× magnification.Mitosis and migration of stromal cells are regulated by cytokines released from the epithelium and its basement membrane. For example, PDGF is produced by corneal epithelium and bound to basement membrane due to heparin-binding properties of the cytokine. It is released from the epithelial basement membrane after injury and stimulates mitosis of corneal fibroblasts. It is also highly chemotactic to corneal fibroblasts, tending to attract them to the source of the cytokine. Thus, in PRK, for example, PDGF released from the injured epithelium and basement membrane stimulates surviving keratocytes in the peripheral and posterior stroma to undergo mitosis and the daughter cells are attracted to the ongoing PDGF release and repopulate the anterior stroma. Other cytokines such as TGF-β also likely contribute to this keratocyte/corneal fibroblast mitosis and migration.
2Corneal fibroblasts derived from keratocytes produce collagen, glycosaminoglycans, collagenases, gelatinases, and metalloproteinases 18 used to restore corneal stromal integrity and function. These cells also produce cytokines such as EGF, HGF, and KGF that direct mitosis, migration, and differentiation of the overlying healing epithelium. 1,2,19 After total epithelialization, the fibronectin clot disappears and the nonkeratinized stratified epithelium is re-established. Anutosh Ganguly. Fernando Cabral, in, 2013 4.2 Microtubule Detachment in SpindlesMitotic cells present an especially difficult challenge because they are frequently round, lack a nucleus, and have two spindle poles rather than a single centrosome nucleating microtubules. Mitotic cells are also thicker than interphase cells thus causing poor contrast and difficulty tracking individual microtubules in the three-dimensional space. Other cells such as PtK2 remain well attached during mitosis but are still difficult to image because of the high density of microtubules in the region near the spindle poles.
We have been able to partially overcome problems associated with the rounded morphology of mitotic CHO cells by examining those cells during prophase, a stage at which they remain well attached. Despite being early in mitosis, detachment events are already elevated during prophase compared to interphase ( Yang et al., 2010).
We have not been able to reliably measure detachment at later stages of mitosis, and so we cannot say with any confidence that detachment remains elevated following prophase. However, others have reported a high detachment frequency during anaphase in LLCPK1 cells, and they proposed that detachment is required for microtubule rearrangements that may be occurring during that stage of mitosis ( Rusan & Wadsworth, 2005). It is possible that detachment plays a similar role during prophase to hasten the conversion of the cytoplasmic microtubules into mitotic spindle fibers. Alternatively, detachment may be taking place throughout mitosis to provide microtubule fragments for the construction and dynamics of the mitotic spindle apparatus ( Ganguly et al., 2010; Yang et al., 2007).Given the problems associated with the measurement of microtubule detachment from spindle poles, we have used interphase cells as a substitute to follow changes that are likely to also take place during mitosis.
The rationale for this approach comes from the observation that perturbations such as drug treatment, mutant tubulin expression, overexpression of class V β-tubulin, and overexpression of MCAK that interfere with spindle function and cell division alter the detachment frequency not only during mitosis but also during interphase ( Bhattacharya et al., 2011; Ganguly, Yang, & Cabral, 2011b; Ganguly, Yang, Pedroza, et al., 2011; Ganguly et al., 2010; Yang et al., 2010). Jie Qiao, in, 2019 Mitotic DivisionMitotic division takes place when PGCs migrate to the female gonads and differentiate into oogonia 6. Mitosis involves a nuclear division that produces two genetically identical daughter cells through the stages of prophase, prometaphase, metaphase, anaphase, and telophase. Mitosis is a reasonably short stage of the cell cycle, preceded by the long interphase. In preparation for the division, the cell synthesizes mRNA and proteins during gap 1 (G1-pase) of the interphase. Then the cell progresses to the synthesis (S-phase) of the interphase, during which the whole genome encoded within double DNA strands is replicated.
Subsequently, rapid growth and protein synthesis occur during gap 2 (G2-phase) of the interphase, which ends with the onset of prophase, a first phase of mitosis (M-phase).Prophase involves condensation and separation of the duplicated DNA into sister chromatids, each of which corresponds to one set of the duplicated genome. Nuclear chromatin becomes visible in the light microscope as chromosomes. During prometaphase of mitosis, the nuclear membrane dissolves and spindle fibers attach to the chromosomal centromeres. This is followed by alignment of sister chromatids along the middle of the cell in metaphase (metaphase plate). Chromosomes then segregate during anaphase to generate two genetically identical nuclei, receiving one copy of each chromosome. Afterward, chromosomes disperse in telophase and spindle fibers gradually disappear.
The process is accompanied by cytokinesis, which results in the division of cytoplasm around each nucleus and the production of two identical daughter cells. All the cell cycle phases round sequentially under a strict surveillance of the checkpoint proteins to ensure an adequate cycle progression.Mitosis in the vertebrate is activated by cyclin-dependent kinase Cdk1 (also known as Cdc2), which is dependent on cyclin B. Other proteins and protein kinases involved in the regulation of mitosis include Cdc25C, Wee1, and Myt1 18.
An important mitotic checkpoint (also known as the spindle assembly checkpoint) involves a complex signaling cascade that ensures uninterrupted chromosome segregation, and arrests aberrant mitotic division via inhibition of the anaphase-promoting complex/cyclosome (APC/C), an E3 ubiquitin ligase. Defects in this signaling complex have been linked with tumor formation 19. Additional, a less common mitotic derangement is tripolar mitosis, which results in the production of three daughter cells with uneven distribution of genetic material 20.
Mitosis involves substantial rearrangements of all major structural elements leading to a reversible rounding of the cell. Given the role of phosphorylation in modifying the arrangement of keratin filaments, an elevated level of keratin phosphorylation is to be expected during mitosis. This notion was fully supported by experimental observations that are summarized in Table 3 and exemplified in Figs. Specifically, of the type II keratins K4, K5, K6, and K8 are phosphorylated by mitogen-regulated kinases in proliferating epithelial cells ( Table 3) at the highly conserved LLS/TPL motif in the H1 head domain suggesting an evolutionary conserved function of keratin phosphorylation in mitosis ( Liao et al., 1997; Toivola et al., 2002). The higher level of phosphorylated keratins in the soluble fraction as compared to the insoluble fraction during mitosis further indicated a role of phosphorylation in shifting the solubility equilibrium of the keratin pool ( Baribault et al., 1989; Celis et al., 1983; Chou and Omary, 1993; Fey et al., 1983; Lane et al., 1982). Additionally, keratin filament reorganization into granules is often observed during mitosis ( Figs. 2 and 3; Aubin et al., 1980; Baribault et al., 1989; Fey et al., 1983; Horwitz et al., 1981; Jones et al., 1985; Lane et al., 1982; Schwarz et al., 2015; Windoffer and Leube, 2001).
Mitotic human vulva carcinoma A431 and cervix carcinoma HeLa present both a diffuse and granular cytoplasmic keratin pattern, whereas the keratin network of rat kangaroo kidney-derived PtK2 cells does not disassemble into granules but is only redistributed ( Horwitz et al., 1981). Another report showed that the percentage of transformed human epithelial amnion cells containing mitotic keratin granules increased from prophase onwards, reaching a peak during late anaphase/early telophase and plummeting during late telophase. In contrast, normal mitotic amnion cells did not show this characteristic keratin reorganization during mitosis, in spite of similar keratin phosphorylation levels as the transformed cells ( Celis et al., 1983). This suggests that additional factors such as mitosis duration or overall kinase activity levels influence keratin reorganization during mitosis. Moreover, preferential phosphorylation of K8 over K18 was observed during mitosis, in rat hepatocytes.
However, in this case, phosphorylation of both K8 and K18 upon triton X-100 permeabilization suggested that keratin phosphorylation during mitosis may also depend on the accessibility of their phosphorylation sites ( Baribault et al., 1989). Mitotic keratin restructuring may influence the kinetics and efficiency of cell division as has been suggested for vimentin phosphorylation ( Ikawa et al., 2014). Moreover, in vivo monitoring of K8 restructuring during the trophoectodermal cell divisions in blastocysts of K8-YFP knockin mice provided direct proof for the significance of mitotic keratin reorganization in the native tissue context ( Schwarz et al., 2015).
Keratin/Keratin ResidueSourceReferencesK55/K49Rat hepatocytesBaribault et al. (1989)K8S431HT29 cells aLiao et al. (1997)BHK cells bKu and Omary (1997)K18S52HT29 cells aLiao et al. (1995a)K18S33HT29 cells aKu et al. (1998b) and Ku et al. (2002b)K4T133 cHuman esophageal epitheliumToivola et al.
(2002)K5T150 cKC cells dK6T145 cHuman epidermisK8S73 cHT29 cells aLiao et al. (1997)Murine mitotic basal crypt cells of the intestineToivola et al.
(2002)Murine regenerating hepatocytesA431 cells eWoll et al. (2007)a Human colorectal adenocarcinoma-derived cell line. B Syrian golden hamster kidney fibroblast-derived cell line. C These residues are located in the conserved LLS/TPL motif of the type II keratins. D Human foreskin primary keratinocytes.
E Human vulva squamous cell carcinoma-derived cell line. Keratin filament network disassembly and reassembly during mitosis. The images in (A–F) show Hoechst stains and corresponding fluorescence of human keratin 5-enhanced yellow fluorescent protein chimeras in methanol-acetone fixed keratinocyte-derived cell line HaCaT B10 ( Moch et al., 2013) during different phases of mitosis (A′–F′). Note the reversible keratin granule formation that is induced at late prophase. Accumulation of keratin granules is typically seen in the cleavage furrow (arrows in D′ and E′).
The structural reorganization is coupled to keratin phosphorylation (see Fig. Scale bars = 10 μm.Interestingly, A431 and HeLa cells show a concentration of keratin granules in the cleavage furrow during mitosis.
3D–E′ highlights the enrichment of keratin granules in this region of dividing immortalized human HaCaT keratinocytes. The cleavage furrow harbors several kinases such as Rho-kinase and Aurora B which phosphorylate and thus regulate intermediate filament reorganization, which is essential for efficient cytokinesis ( Horwitz et al., 1981; Kawajiri et al., 2003; Kosako et al., 1999; Yasui et al., 1998). Furthermore, p38 mitogen-activated protein kinase, which plays an important role in cell proliferation, colocalizes with keratin granules during mitosis in A431 cells ( Woll et al., 2007). This indicates the necessity of a kinase-enriched environment for keratin reorganization during mitosis.
It is likely also relevant for other cell types that do not form keratin granules during mitosis. In these instances, the locally increased kinase activities could sever the keratin network at the cleavage furrow for distribution of filamentous keratin into both daughter cells. Moreover, keratins are substrates of mitotic kinases as shown by in vitro phosphorylation of K8-S431 through mitogen-activated protein kinase and cdc2 kinase ( Ku and Omary, 1997).The hyperphosphorylation of keratins during mitosis also modulates their binding to associated proteins. This is well demonstrated by the requirement of K18-S33 phosphorylation for an interaction between K18 and 14-3-3 family of proteins, facilitating normal mitotic progression ( Liao et al., 1996).
Although K18-S33A mice did not have defects in liver regeneration, they displayed abnormal mitotic arrest-related figures such as tripolar and angular mitotic bodies and anomalous proportions of mitotic stages. Furthermore, retention of 14-3-3-ζ in the nucleus of K18-S33A hepatocytes suggested a role of keratin phosphorylation in modulating 14-3-3 distribution, which is an important sequestering and compartmentalizing factor for multiple proteins ( Ku et al., 1998a, 2002b; Liao et al., 1996). Thus, keratin phosphorylation during mitosis may not only contribute to cytoskeletal reordering but may also impact the function of keratin-associated proteins. Iskra Yanakieva.
Caren Norden, in, 2018 1 IntroductionMitosis is the process during which the duplicated DNA of a cell is distributed equally between two daughter cells in the form of condensed chromosomes. Most of the basic mechanisms of mitosis are conserved between organisms, from yeast, to Tetrahymena, to animal and plant cells. Mitosis has been studied in detail in diverse systems with some focus on cell lines in culture ( Yanagida, 2014) and unicellular organisms like yeast. While in these systems mitosis is mainly a means of reproduction, in multicellular organisms it is crucial to drive growth during development and later serves to maintain adult tissues during homeostasis. The life of both animals and plants starts with a single zygotic cell that, through coordinated mitotic divisions, gives rise to a wide variety of cell and tissue types functioning in concert in the adult organism. Thus, to understand how embryonic tissues grow and mature correctly, it is important to explore in detail the mechanisms that control mitosis in the context of development.To generate such an understanding, we here concentrate on means to investigate three basic questions that arise when studying the role of mitosis in embryonic development: (1) How do mitotic patterns give rise to the observed tissue growth and how do tissues take shape? (2) Are the basic mechanisms of mitosis conserved when mitosis takes place in different tissues during development?
(3) How do developmental lineages arise over multiple mitotic cycles from progenitor cells?To be able to address these questions, different imaging approaches can be employed. Depending on the question posed, quantifications based on fixed tissue imaging or live imaging with durations ranging from minutes to multiple days would be necessary to find an answer. For example, to understand the contribution of mitosis to shape and growth patterns in different tissues, it is important to determine the fraction of cells that are in mitosis in a given time, a parameter known as mitotic index. This is usually done by imaging multiple samples of fixed tissue. Conversely, to understand the basic mechanisms of mitosis, it is necessary to study the dynamics of the mitotic apparatus live. This can be achieved by specific labelling of intracellular components and direct observation of these components with lower or higher temporal but always high spatial resolution.
In case the aim is to trace the lineage of a single cell, cells and their progeny need to be tracked over long periods of time in live specimens. Distinct microscopy approaches can be used to achieve these different goals. While for fixed samples phototoxicity is not an issue, for live observation of mitotic events, it is important to note that fluorescence microscopy is not completely noninvasive due to the illumination of the sample with high-intensity light ( Magidson & Khodjakov, 2013). Phototoxic effects can perturb the process of interest and need to be kept to a minimum. The amount of inflicted photodamage depends on the method of illumination of the focal plane (reviewed in Icha, Weber, Waters, & Norden, 2017), as well as the temporal resolution and signal-to-noise ratio (SNR) during acquisition ( Laissue, Alghamdi, Tomancak, Reynaud, & Shroff, 2017).
The desired SNR, temporal and spatial resolution for each experiment depend on the scientific question asked and need to be carefully balanced to reduce phototoxicity. To provide the balance required by each experimental setup different microcopy systems can be used. Laser-scanning confocal microscopy (LSCM) provides excellent SNR and spatial resolution but during multiple scans the sample can accumulate photodamage. As noted, phototoxicity is not a concern when imaging fixed tissues, and one can benefit from the LSCM's ability to acquire images with high spatial resolution in a single time point. However, this is not the case when living samples need to be imaged at short time intervals. When high temporal resolution and low phototoxicity need to be balanced with satisfactory spatial resolution, a spinning disk confocal microscope (SDCM) can be a better solution due to lower photodamage.
When the aim is to image the developing organism over long periods of time to trace cell linages, for example, high temporal resolution can be sacrificed to keep phototoxicity to a minimum. The state of the art microscope to accommodate such imaging experiment is the light-sheet fluorescence microscope (LSFM).Regardless of the choice of microscope, imaging of living embryos can be hampered if the studied model organism is developing slowly, or develops in utero. Zebrafish emerged as an excellent model system for all the listed imaging approaches due its rapid ex utero development and the transparency of embryos. Thus, mitotic events in diverse tissues and developmental dynamics can be observed directly in situ.One tissue type in which mitotic events have been investigated in the developing zebrafish are neuroepithelia ( Baye & Link, 2007; Del Bene, Wehman, Link, & Baier, 2008; Geldmacher-Voss, Reugels, Pauls, & Campos-Ortega, 2003; Strzyz et al., 2015; Weber et al., 2014). These are highly proliferative tissues consisting of progenitor cells that gives rise to parts of the central nervous system, including the retina.
It was shown that proliferation and mitotic events in these tissues are sensitive to photodamage ( Icha et al., 2017) and thus they serve as a good example of why it is critical to balance spatial and temporal resolution with phototoxicity in studies of mitosis during development.Here, we summarize three protocols that illustrate how different microcopy systems can be used to study mitosis in developing zebrafish neuroepithelia. First, to understand how growth depends on the rates and patterns of mitosis in a given tissue, we show how to determine its mitotic index using a LSCM. Second, we explain how to observe the dynamics of intracellular components during mitosis in the embryo and to assess what fails during perturbation by using imaging with high temporal resolution in SDCM. Third, we show how to follow cell lineages during consecutive cell cycles in the course of development using a LSFM.
Before going into these details, however, in the next paragraphs we briefly outline the imaging principles of the three microscopes mentioned. In, 2008 What Is Mitosis?Mitosis is the tightly regulated process of cell division that includes both nuclear division (karyokinesis) and the division of cytoplasm to two daughter cells (cytokinesis). This process can be divided into distinct phases including prophase, prometaphase, metaphase, anaphase, telophase, and finally, cytokinesis. The products of mitosis are two daughter cells that have identical DNA content that is also identical to the DNA content of the original parental cell. Depending on the nature of the parental cell, the daughters may be essentially identical in phenotype, or they may differ. In the case of undifferentiated adult stem cells or progenitor cells, one daughter may remain undifferentiated (self-renewing), whereas the other becomes committed to a differentiated lineage.
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