Molecular subdivision of the cortex of dividing Tetrahymena is coupled with the formation of the fission zone.

In contrast to a mitotic-spindle-associated bipolar cytokinesis, the cytokinesis of polarized ciliates is preceded by a reorganization of the cortex into dual metameric patterns for prospective daughter cells and then separated by a transverse fission line. This study concerns relations between the generation of cortical metamery and the formation of the fission line in an amicronuclear (i.e., without mitotic spindle) ciliate, Tetrahymena pyriformis. The fission line appears in the division of T. pyriformis as a transverse line formed by equatorial gaps in the meridional ciliary rows, with the second oral structure (OA2) formed posterior to it. It was found that the metamery of cortical morphogenesis is expressed by the appearance of increased MPM2 antibody binding in dividing cells in an apical area and posterior to the fission line gaps, including patterned changes of this binding in both oral apparatuses (OA1 and OA2), and by a reciprocal decrease of binding of an anti-epiplasm antibody. These tested antigens are localized to different cortical structures, but in predividing cells both uniformly show formation of the fission line contrast of labeling. A serine/threonine kinase inhibitor, 6-dimethylaminopurine (6-DMAP), was applied to dividing T. pyriformis at specific stages: (1) if 6-DMAP was added to early dividing cells, it prevented cells from initiating cytokinesis. (2) If 6-DMAP was added to cells at stages close to the physiological transition point of cell division, it yielded either (i) a partial formation of the fission line on the ventral side, combined with modified growth of undivided cortex adjacent to the fission line, with abnormal cytokinesis, or (ii) variable anterior displacement of the complete fission line, which contracted slowly but uniformly. (3) If 6-DMAP was applied during cytokinesis, it did not delay cell division, but daughter cells become abnormal and underwent an incomplete oral reorganization. These results suggest that the generation of metamerism in the cortex of T. pyriformis involves differentiation of the asymmetric fission zone. At least four stage-dependent 6-DMAP-sensitive effects jointly control the progress of cell division and the mutual spatial relations between the generation of metamery and the appearance, completeness, and position of the fission zone in the cortex of polarized T. pyriformis.     

Structural changes in the cell wall of Schizosaccharomyces pombe during cell division

Streiblová, Eva (Czechoslovak Academy of Sciences, Prague, Czechoslovakia), I. Málek, and K. Beran. Structural changes in the cell wall of Schizosaccharomyces pombe during cell division. J. Bacteriol. 91:428-435. 1966.-Individual stages of growing and dividing cells of Schizosaccharomyces pombe were studied by means of fluorescence and electron microscopy with the use of metal-shadowed isolated walls, replicas, and ultrathin sections. Vegetative cells were found to contain division scars (six at the most); their formation and structure are described. More data on the growth of arthrospores were obtained. New structural observations were made on the architecture of the cell wall (original wall ring, polar cell wall, plug wall band, additional wall ring). Structural changes of cell surfaces and lateral walls during fission are represented schematically to the fourth generation. The question of origin of the septum is discussed, and on this basis the entire structure of the cell wall is interpreted.     

Sid2p, a spindle pole body kinase that regulates the onset of cytokinesis.

The fission yeast Schizosaccharomyces pombe divides by medial fission through the use of an actomyosin contractile ring. Precisely at the end of anaphase, the ring begins to constrict and the septum forms. Proper coordination of cell division with mitosis is crucial to ensure proper segregation of chromosomes to daughter cells. The Sid2p kinase is one of several proteins that function as part of a novel signaling pathway required for initiation of medial ring constriction and septation. Here, we show that Sid2p is a component of the spindle pole body at all stages of the cell cycle and localizes transiently to the cell division site during medial ring constriction and septation. A medial ring and an intact microtubule cytoskeleton are required for the localization of Sid2p to the division site. We have established an in vitro assay for measuring Sid2p kinase activity, and found that Sid2p kinase activity peaks during medial ring constriction and septation. Both Sid2p localization to the division site and activity depend on the function of all of the other septation initiation genes: cdc7, cdc11, cdc14, sid1, spg1, and sid4. Thus, Sid2p, a component of the spindle pole body, by virtue of its transient localization to the division site, appears to determine the timing of ring constriction and septum delivery in response to activating signals from other Sid gene products.     

The fission yeast Schizosaccharomyces pombe in continuous culture

The fission yeast Schizosaccharomyces pombe was cultivated in a chemostat at dilution rates of D = 0.03, 0.05, 0.10, and 0.20 h(-1). After steady state had been reached, the amount of dry matter, number of cells, concentration of residual sugar, yield coefficient (Y), and some morphological properties of the cells were estimated. Curves reflecting the dry mass, number of cells, and cell mean volume show a changing coordination between the growth rate and the rate of cell division, with respect to D. In addition, it could be concluded that in dividing cells the cell septum is localized asymmetrically; Two nonidentical cells differing both in length and volume result. The degree of asymmetry is a function of the dilution rate.     

A stochastic, molecular model of the fission yeast cell cycle: role of the nucleocytoplasmic ratio in cycle time regulation

We propose a stochastic version of a recently published, deterministic model of the molecular mechanism regulating the mitotic cell cycle of fission yeast, Schizosaccharomyces pombe. Stochasticity is introduced in two ways: (i) by considering the known asymmetry of cell division, which produces daughter cells of slightly different sizes; and (ii) by assuming that the nuclear volumes of the two newborn cells may also differ. In this model, the accumulation of cyclins in the nucleus is proportional to the ratio of cytoplasmic to nuclear volumes. We have simulated the cell-cycle statistics of populations of wild-type cells and of wee1(-) mutant cells. Our results are consistent with well known experimental observations.     

The multiprotein exocyst complex is essential for cell separation in Schizosaccharomyces pombe

Schizosaccharomyces pombe cells divide by medial fission through the use of an actomyosin-based contractile ring. A mulitlayered division septum is assembled in concert with ring constriction. Finally, cleavage of the inner layer of the division septum results in the liberation of daughter cells. Although numerous studies have focused on actomyosin ring and division septum assembly, little information is available on the mechanism of cell separation. Here we describe a mutant, sec8-1, that is defective in cell separation but not in other aspects of cytokinesis. sec8-1 mutants accumulate about 100-nm vesicles and have reduced secretion of acid phosphatase, suggesting that they are defective in exocytosis. Sec8p is a component of the exocyst complex. Using biochemical methods, we show that Sec8p physically interacts with other members of the exocyst complex, including Sec6p, Sec10p, and Exo70p. These exocyst proteins localize to regions of active exocytosis-at the growing ends of interphase cells and in the medial region of cells undergoing cytokinesis-in an F-actin-dependent and exocytosis-independent manner. Analysis of a number of mutations in various exocyst components has established that these components are essential for cell viability. Interestingly, all exocyst mutants analyzed appear to be able to elongate and to assemble division septa but are defective for cell separation. We therefore propose that the fission yeast exocyst is involved in targeting of enzymes responsible for septum cleavage. We further propose that cell elongation and division septum assembly can continue with minimal levels of exocyst function.     

The price of independence: cell separation in fission yeast

The ultimate goal of cell division is to give rise to two viable independent daughter cells. A tight spatial and temporal regulation between chromosome segregation and cytokinesis ensures the viability of the daughter cells. Schizosaccharomyces pombe, commonly known as fission yeast, has become a leading model organism for studying essential and conserved mechanisms of the eukaryotic cell division process. Like many other eukaryotic cells it divides by binary fission and the cleavage furrow undergoes ingression due to the contraction of an actomyosin ring. In contrast to mammalian cells, yeasts as cell-walled organisms, also need to form a division septum made of cell wall material to complete the process of cytokinesis. The division septum is deposited behind the constricting ring and it will constitute the new ends of the daughter cells. Cell separation also involves cell wall degradation and this process should be precisely regulated to avoid cell lysis. In this review, we will give a brief overview of the whole cytokinesis process in fission yeast, from the positioning and assembly of the contractile ring to the final step of cell separation, and the problems generated when these processes are not precise.     

A critical role for the type V myosin, Myo52, in septum deposition and cell fission during cytokinesis in Schizosaccharomyces pombe

Cytokinesis in fission yeast involves the coordination of septum deposition with the contraction of a cytokinetic actomyosin ring. We have examined the role of the type V myosin Myo52 in the coupling of these two events by the construction of a series of deletion mutants of the Myo52 tail and a further mutant within the ATP binding domain of the head. Each mutant protein was ectopically expressed in fission yeast cells. Each truncation was assayed for the ability to localize to the cell poles and septum (the normal cellular locations of Myo52) and to rescue the morphology defects and temperature sensitivity of a myo52Delta strain. A region within the Myo52 tail (amino acids 1320-1503), with a high degree of similarity to the vesicle-binding domain of the budding yeast type V myosin Myo2p, was essential for Myo52's role in the maintenance of growth polarity and cell division. A separate region (amino acids 1180-1320) was required for Myo52 foci to move throughout the cytoplasm; however, constructs lacking this region, but which retained the ability to dimerize still associated with actin at sites of cell growth. Not all of the Myo52 truncations which localized rescued the morphological defects of myo52Delta, demonstrating that loss of function was not simply brought about by an inability of mutant proteins to target the correct cellular location. By contrast, Myo52 motor activity was required for both localization and cellular function. myo52Delta cells were unable to efficiently localize the beta-1,3-glucan synthase, Bgs1, either at the cell poles or at the division septum, regions of cell wall deposition. Bgs1 and Myo52 localized to vesicle-like dots at the poles in interphase and these moved together to the septum at division. These data have led to the formulation of a model in which Myo52 is responsible for the delivery of Bgs1 and associated molecules to polar cell growth regions during interphase. On the commencement of septum formation, Myo52 transports Bgs1 to the cell equator, thus ensuring the accurate deposition of beta-1,3-glucan at the leading edge of the primary septum.     

Ultrastructural Explanation for Snapping Postfission Movements in Arthrobacter crystallopoietes

The ultrastructure of dividing rod-stage cells of Arthrobacter crystallopoietes was examined by electron microscopy. The cell walls consist of two layers. During cell division, the inner layer invaginates to form the septum. The outer layer does not participate in septum formation. After septum formation is completed, the two daughter cells remain attached by the outer layer of the cell wall. It appears that localized rupture of the outer layer during further wall growth is responsible for the phenomenon known as "snapping division" or "snapping postfission movement."     

Control of cell division in Paramecium tetraurelia. Effects of abrupt changes in nutrient level on accumulation of macronuclear DNA and cell mass

In the cell cycle of Paramecium there are three points of interaction between cell growth-related processes and the processes of macronuclear DNA replication and cell division: initiation of DNA synthesis, regulation of the rates of growth and DNA accumulation, and initiation of cell division. This study examines the regulation of the latter two processes by analysis of the response of each to abrupt changes in nutrient level brought about either by transferring dividing cells from a steady-state chemostat culture to medium with unlimited food, or by transferring well-fed dividing cells to exhausted medium. The rates of DNA accumulation and cell growth respond quickly to changes in nutrient level. The amounts of these cell components accumulated during the cell cycle following a shift in nutrient level are typical of those occurring during equilibrium growth under post-shift conditions. Commitment to division occurs at a fixed interval prior to fission that is similar in well-fed and nutrient-limited cells. Initiation of cell division in Paramecium is associated with accumulation of a threshold DNA increment, whose level is largely independent of nutritive conditions. The amount of DNA accumulated during the cell cycle varies with nutritional conditions because the rates of growth and DNA accumulation are affected by nutrient level; slowly growing cells accumulated relatively little DNA during the fixed interval between commitment to cell division and fission.     

Drc1p/Cps1p, a 1,3-beta-glucan synthase subunit, is essential for division septum assembly in Schizosaccharomyces pombe.

Schizosaccharomyces pombe divides by medial fission through the use of an actomyosin-based contractile ring. A division septum is formed centripetally, concomitant with ring constriction. Although several genes essential for cytokinesis have been described previously, enzymes that participate in the assembly of the division septum have not been identified. Here we describe a temperature-sensitive mutation, drc1-191, that prevents division septum assembly and causes mutant cells to arrest with a stable actomyosin ring. Unlike the previously characterized cytokinesis mutants, which undergo multiple mitotic cycles, drc1-191 is the first cytokinesis mutant that arrests with two interphase nuclei. Interestingly, unlike drc1-191, drc1-null mutants proceed through multiple mitotic cycles, leading to the formation of large cells with many nuclei. drc1 is allelic to cps1, which encodes a 1,3-beta-glucan synthase subunit. We conclude that Drc1p/Cps1p is not required for cell elongation and cell growth, but plays an essential role in assembly of the division septum. Furthermore, it appears that constriction of the actomyosin ring might depend on assembly of the division septum. We discuss possible mechanisms that account for the differences in the phenotypes of the drc1-191 and the drc1-null mutants and also reflect the potential links between Drc1p and other cytokinesis regulators.     

Cell division site placement and asymmetric growth in mycobacteria

Mycobacteria are members of the actinomycetes that grow by tip extension and lack apparent homologues of the known cell division regulators found in other rod-shaped bacteria. Previous work using static microscopy on dividing mycobacteria led to the hypothesis that these cells can grow and divide asymmetrically, and at a wide range of sizes, in contrast to the cell growth and division patterns observed in the model rod-shaped organisms. In this study, we test this hypothesis using live-cell time-lapse imaging of dividing Mycobacterium smegmatis labelled with fluorescent PBP1a, to probe peptidoglycan synthesis and label the cell septum. We demonstrate that the new septum is placed accurately at mid-cell, and that the asymmetric division observed is a result of differential growth from the cell tips, with a more than 2-fold difference in growth rate between fast and slow growing poles. We also show that the division site is not selected at a characteristic cell length, suggesting this is not an important cue during the mycobacterial cell cycle.     

Cell division in baeocyte producing cyanobacteria

Summary An ultrastructural examination of cell division in two baeocyte producing cyanobacteria,Pleurocapsa minor andDermocarpa violaceae, reveals two distinct patterns of binary (transverse) fission. Septate binary fission, inPleurocapsa minor, involves centripetal synthesis and deposition of the mucopolymer cell wall layer (L 2). The ingrowth of the cytoplasmic membrane and L 1 cell wall layer, along with the synthesis of the L 2 cell wall layer, results in the formation of a prominent septum. Partitioning of the cell occurs by the constriction of the outer cell wall layers (L 3 and L 4) through the septum. InDermocarpa violaceae, constrictive binary fission occurs by the simultaneous ingrowth or constriction of the cytoplasmic membrane and all cell wall layers (L1, L2, L3, L4). Septate and constrictive binary fission may proceed symmetrically (medially) or asymmetrically (nonmedially). Multiple fission occurs regularly inDermocarpa violaceae and provides for a rapid means of reproduction when compared to binary fission. Successive radial and tangential divisions of the protoplast result in formation of many small daughter cells (baeocytes). The process of multiple fission is similar to septate binary fission with reduced septa being formed. However, constriction of the outer cell wall layers, through the septa, proceeds concurrently with septum formation.     

Fine Structure of Bacillus megaterium During Synchronous Growth

A fine-structure study of synchronously dividing Bacillus megaterium revealed the sequence of events involved in the division of the cell. First, a mesosome develops as a concentric fold of the plasma membrane at the site of septum formation. The mesosome contains membrane-bound vesicular structures, 300 to 500 A in diameter, plus a large membrane-bound structure, 2,000 A in diameter. These larger vesicles are peculiar to mesosomes in this stage of division and are not observed in the mesosomes involved in spore septum formation. The transverse septum originates within the mesosome and remains enclosed during its subsequent growth across the cell. An intimate association is observed between mesosome vesicles, mesosome membrane, and the growing edge of the transverse septum. Prior to completion of the septum, the membranes bounding the mesosome fuse, and further wall thickening occurs within the structure formed by this fusion. At this time, the septum only equals the parent cell wall in thickness. The doubling in thickness of the septum, which is required for the production of two normal daughter cell walls, occurs during a second phase of wall thickening, which is characterized by the appearance of a constriction at the base of the septum. As the constriction widens, the wall in this region thickens, forming the typical rounded poles of the daughter cells. Capsular synthesis at the poles occurs during this second phase of wall thickening. Throughout the division process, the nuclear material appears to be associated at one end with a mesosome at or near the pole of the cell and at the other end to the mesosome involved in septum formation. This association frequently takes the form of a stalklike extension of the mesosome penetrating into the chromatin fibrils.     

The cdc7 protein kinase is a dosage dependent regulator of septum formation in fission yeast.

Mutation of the Schizosaccharomyces pombe cdc7 gene prevents formation of the division septum and cytokinesis. We have cloned the cdc7 gene and show that it encodes a protein kinase which is essential for cell division. In the absence of cdc7 function, spore germination, DNA synthesis and mitosis are unaffected, but cells are unable to initiate formation of the division septum. Overexpression of p120cdc7 causes cell cycle arrest; cells complete mitosis and then undergo multiple rounds of septum formation without cell cleavage. This phenotype, which is similar to that resulting from inactivation of cdc16 protein, requires the kinase activity of p120cdc7. Mutations inactivating the early septation gene, cdc11, suppress the formation of multiple septa and allow cells to proliferate normally. If formation of the division septum is prevented by inactivation of either cdc14 or cdc15, p120cdc7 overproduction does not interfere with other events in the mitotic cell cycle. Septation is not induced by overexpression of p120cdc7 in G2 arrested cells, indicating that it does not bypass the normal dependency of septation upon initiation of mitosis. These findings indicate that the p120cdc7 protein kinase plays a key role in initiation of septum formation and cytokinesis in fission yeast and suggest that p120cdc7 interacts with the cdc11 protein in the control of septation.     

Second thoughts on septation by the fission yeast, <Emphasis Type="Italic">Schizosaccharomyces pombe</Emphasis>: pull vs. push mechanisms with an appendix – dimensional modelling of the flat and variable septa

The correlation of contraction by an actomyosin band with the closing of the septum of dividing cells of the fission yeast, Schizosaccharomyces pombe, cannot suggest cause-and-effect because contraction would be apparent whether the membrane enveloping the centripetally closing septum were pulled or were pushed. Thus the common observation of contraction is not critical. Diagrams of published electron micrographs of dividing wild-type fission yeasts illustrate variable (tilted) septal images that are counterintuitive to a pull model. Circumference calculations based on those images suggest that some variable forms might be only 6% closed even though their two-dimensional profiles would be 50% closed, if they were not tilted. Development of multiseptate forms of cdc4-8 and cdc4-377 temperature sensitive mutants incubated at their restrictive temperature was followed. These multiseptate forms are shown to have functional (functional in terms of generating divided uninucleate cytoplasts) but grotesque septa which are formed in the absence of actomyosin bands. By contrast, the myosin of the plant phragmoplast is not properly oriented for contractility, and Dictyostelium (attached cells) and Saccharomyces (mutants) have been shown to divide in the absence of myosin II, just as S. pombe does (above). Hence contractility, the essence of a pull model for septum closure, would seem to be non-essential. Other, non-contractile mechanisms of myosin are emphasized, and a push model becomes a rational default hypothesis. The essence of push models is that their synthesis/assembly mechanisms are driving force sufficient for septum closure.     

Contributions of turgor pressure, the contractile ring, and septum assembly to forces in cytokinesis in fission yeast

A paradigm of cytokinesis in animal cells is that the actomyosin contractile ring provides the primary force to divide the cell [1]. In the fission yeast Schizosaccharomyces pombe, cytokinesis also involves a conserved cytokinetic ring, which has been generally assumed to provide the force for cleavage [2-4] (see also [5]). However, in contrast to animal cells, cytokinesis in yeast cells also requires the assembly of a cell wall septum [6], which grows centripetally inward as the ring closes. Fission yeast, like other walled cells, also possess high (MPa) turgor pressure [7-9]. Here, we show that turgor pressure is an important factor in the mechanics of cytokinesis. Decreasing effective turgor pressure leads to an increase in cleavage rate, suggesting that the inward force generated by the division apparatus opposes turgor pressure. The contractile ring, which is predicted to provide only a tiny fraction of the mechanical stress required to overcome turgor, is largely dispensable for ingression; once septation has started, cleavage can continue in the absence of the contractile ring. Scaling arguments and modeling suggest that the large forces for cytokinesis are not produced by the contractile ring but are driven by the assembly of cell wall polymers in the growing septum.