Step Of Cell Division Research Articles

The contractile ring

What is the contractile ring? During cell division, the contractile ring generates the constricting force to separate one cell into two cells. Formed during cytokinesis, the last step of cell division, the contractile ring is composed of filamentous actin (F-actin) and the motor protein myosin-2, along with additional structural and regulatory proteins. The contractile ring forms under the surface of the plasma membrane and is linked to the plasma membrane such that, when it constricts, it creates a cleavage furrow that partitions the cell in two.

Cytokinesis, ploidy and aneuploidy

Cytokinesis is the last step of cell division that physically separates the daughter cells. As such, it ensures the proper inheritance of both nuclear and cytoplasmic contents. Accomplishment of cytokinesis in eukaryotes is dictated by several key events: establishment of the division plane, furrow ingression through contraction of an actomyosin ring and abscission via membrane fusion. Most mammalian somatic cells are diploid. Polyploidy can result from cytokinesis failure and may contribute to the development of pathologies such as cancer. However, polyploidy is essential for cellular differentiation and function in some contexts (eg hepatocytes, megakaryocytes and others). Consequently, the degree of ploidy and the achievement of cytokinesis must be tightly regulated throughout an organism and among different cell types. In this review we will highlight several examples of normal and pathological polyploidy, focusing on those caused by a controlled failure or dysregulation of cytokinesis, respectively. Last, we propose therapeutic routes to control cytokinesis to restore or block cell division.

A new job for ancient extracellular matrix proteins: Hemicentins stabilize cleavage furrows.

Interactions between extracellular matrix (ECM) proteins and transmembrane receptors mediate changes in cell shape during cell migration, adhesion, differentiation and polarization. Cytokinesis is the final step in cell division as cells employ a contractile ring composed of actin and myosin to partition one cell into two. During the partition process, an invagination in nascent membrane forms a new extracellular space called the cleavage furrow. Despite the dramatic changes in cell shape during cytokinesis, existing models include no role for the ECM. In a recent paper, we show that hemicentins assemble in the cleavage furrow of C. elegans germ cells and mouse embryo blastomeres. Hemicentin depletion results in membrane destabilization, cleavage furrow retraction and cytokinesis failure. The data suggest that hemicentins and other ECM proteins stabilize the cleavage furrow during cytokinesis of multiple cell types.

A new job for ancient extracellular matrix proteins

Interactions between extracellular matrix (ECM) proteins and transmembrane receptors mediate changes in cell shape during cell migration, adhesion, differentiation and polarization. Cytokinesis is the final step in cell division as cells employ a contractile ring composed of actin and myosin to partition one cell into two. During the partition process, an invagination in nascent membrane forms a new extracellular space called the cleavage furrow. Despite the dramatic changes in cell shape during cytokinesis, existing models include no role for the ECM. In a recent paper, we show that hemicentins assemble in the cleavage furrow of C. elegans germ cells and mouse embryo blastomeres. Hemicentin depletion results in membrane destabilization, cleavage furrow retraction and cytokinesis failure. The data suggest that hemicentins and other ECM proteins stabilize the cleavage furrow during cytokinesis of multiple cell types.

A model for the Escherichia coli FtsB/FtsL/FtsQ cell division complex

BackgroundBacterial division is produced by the formation of a macromolecular complex in the middle of the cell, called the divisome, formed by more than 10 proteins. This process can be divided into two steps, in which the first is the polymerization of FtsZ to form the Z ring in the cytoplasm, and then the sequential addition of FtsA/ZipA to anchor the ring at the cytoplasmic membrane, a stage completed by FtsEX and FtsK. In the second step, the formation of the peptidoglycan synthesis machinery in the periplasm takes place, followed by cell division. The proteins involved in connecting both steps in cell division are FtsQ, FtsB and FtsL, and their interaction is a crucial and conserved event in the division of different bacteria. These components are small bitopic membrane proteins, and their specific function seems to be mainly structural. The purpose of this study was to obtain a structural model of the periplasmic part of the FtsB/FtsL/FtsQ complex, using bioinformatics tools and experimental data reported in the literature.ResultsTwo oligomeric models for the periplasmic region of the FtsB/FtsL/FtsQ E. coli complex were obtained from bioinformatics analysis. The FtsB/FtsL subcomplex was modelled as a coiled-coil based on sequence information and several stoichiometric possibilities. The crystallographic structure of FtsQ was added to this complex, through protein-protein docking. Two final structurally-stable models, one trimeric and one hexameric, were obtained. The nature of the protein-protein contacts was energetically favourable in both models and the overall structures were in agreement with the experimental evidence reported.ConclusionsThe two models obtained for the FtsB/FtsL/FtsQ complex were stable and thus compatible with the in vivo periplasmic complex structure. Although the hexameric model 2:2:2 has features that indicate that this is the most plausible structure, the ternary complex 1:1:1 cannot be discarded. Both models could be further stabilized by the binding of the other proteins of the divisome. The bioinformatics modelling of this kind of protein complex, whose function is mainly structural, provide useful information. Experimental results should confirm or reject these models and provide new data for future bioinformatics studies to refine the models.

Phenylpropanoids inhibit protofilament formation of Escherichia coli cell division protein FtsZ

The earliest step in cell division in bacteria is the assembly of FtsZ, an essential cell division protein, into a ring at the division site. FtsZ has GTPase activity and can assemble in vitro to form protein filaments. The present work involved the study of eight phenylpropanoids (cinnamic, p-coumaric, caffeic, chlorogenic, ferulic, 3,4-dimethoxycinnamic and 2,4,5-trimethoxycinnamic acids and eugenol) as inhibitors of Escherichia coli FtsZ. Phenylpropanoids make up the majority of our diet and act as antibacterial agents. Polymerization and GTPase inhibition assays showed that chlorogenic and caffeic acids were the most active amongst these (IC(50) of 70 and 106 µM, respectively). Circular dichroism studies indicated that chlorogenic acid perturbed the protein conformation and electron microscopy showed distorted filaments. Bacillus subtilis 168 cells treated with the phenylpropanoids were longer when compared to the control. The highest binding energy was observed between chlorogenic acid and the homology modelled E. coli FtsZ, which was consistent with the experimental results. A strong negative correlation was observed between binding energy and inhibition of the polymerization ability. 3D-Quantitative structure-activity relationship studies using GTPase activity indicated that the presence of more hydrophilic groups around the 3'- and 4'-carbon increased the activity. The effect of stress-induced formation of cell filamentation has to be understood before confirming the role of phenylpropanoids as FtsZ inhibitors.

Nessun Dorma, a novel centralspindlin partner, is required for cytokinesis in Drosophila spermatocytes

Cytokinesis, the final step of cell division, usually ends with the abscission of the two daughter cells. In some tissues, however, daughter cells never completely separate and remain interconnected by intercellular bridges or ring canals. In this paper, we report the identification and analysis of a novel ring canal component, Nessun Dorma (Nesd), isolated as an evolutionarily conserved partner of the centralspindlin complex, a key regulator of cytokinesis. Nesd contains a pectin lyase-like domain found in proteins that bind to polysaccharides, and we present evidence that it has high affinity for β-galactosides in vitro. Moreover, nesd is an essential gene in Drosophila melanogaster, in which it is required for completion of cytokinesis during male meiosis and possibly in female germline cells. Our findings indicate that Nesd is a novel carbohydrate-binding protein that functions together with centralspindlin in late cytokinesis, thus highlighting the importance of glycosylation in this process.

MTOR, la cible fonctionnelle de la rapamycine

The discovery of rapamycin from a soil sample on Easter Island in the mid 60's marked the beginning of an exciting field of research in cell biology and medicine. While it was first used as an antifungal and as an immunosuppressive drug, more recent studies confirmed rapamycin's antiproliferative properties over a variety of solid tumors. Research aimed at identifying its mechanism of action uncovered mTOR (mammalian target of rapamycin), a protein kinase that regulates mRNA translation and protein synthesis, an essential step in cell division and proliferation. Recent evidence suggests a more complex role for mTOR in the regulation of several growth factor-stimulated protein kinases, including the proto-oncogene Akt. This article reviews mTOR function and regulation, and briefly details the future challenges for anti-cancer therapies based on mTOR inhibition.

TheCaulobacterTol-Pal Complex Is Essential for Outer Membrane Integrity and the Positioning of a Polar Localization Factor

Cell division in Caulobacter crescentus involves constriction and fission of the inner membrane (IM) followed about 20 min later by fission of the outer membrane (OM) and daughter cell separation. In contrast to Escherichia coli, the Caulobacter Tol-Pal complex is essential. Cryo-electron microscopy images of the Caulobacter cell envelope exhibited outer membrane disruption, and cells failed to complete cell division in TolA, TolB, or Pal mutant strains. In wild-type cells, components of the Tol-Pal complex localize to the division plane in early predivisional cells and remain predominantly at the new pole of swarmer and stalked progeny upon completion of division. The Tol-Pal complex is required to maintain the position of the transmembrane TipN polar marker, and indirectly the PleC histidine kinase, at the cell pole, but it is not required for the polar maintenance of other transmembrane and membrane-associated polar proteins tested. Coimmunoprecipitation experiments show that both TolA and Pal interact directly or indirectly with TipN. We propose that disruption of the trans-envelope Tol-Pal complex releases TipN from its subcellular position. The Caulobacter Tol-Pal complex is thus a key component of cell envelope structure and function, mediating OM constriction at the final step of cell division as well as the positioning of a protein localization factor.

A MAP for Bundling Microtubules

Microtubules assemble into arrays of bundled filaments that are critical for multiple steps in cell division, including anaphase and cytokinesis. Recent structural and functional studies, including two papers in this issue of Cell (Bieling et al., 2010; Subramanian et al., 2010), demonstrate how the MAP65 protein PRC1 crosslinks microtubules and cooperates with kinesin motors to control the dynamics and size of bundled regions.

Small molecules discovered in a pathway screen target the Rho pathway in cytokinesis.

We report the discovery of small molecules that target the Rho pathway, a central regulator of cytokinesis, the final step in cell division. We have developed a method to target a small molecule screen towards a specific pathway, which should be widely applicable to study any signaling pathway. In a chemical genetic variant of a classical modifier screen, we used RNA interference (RNAi) to sensitize cells and identified small molecules that suppressed or enhanced the RNAi phenotype. We have discovered promising candidate molecules, which we named Rhodblock 1–8, and we identified the target of Rhodblock 6 as Rho kinase. Several Rhodblocks inhibit a function of the Rho pathway in cells: the correct localization of phosphorylated myosin light chain during cytokinesis. Rhodblocks differentially perturb Rho pathway proteins in cells and can be used to dissect the mechanism of the Rho pathway during cytokinesis.

Cindr Interacts with Anillin to Control Cytokinesis in Drosophila melanogaster

Cytokinesis, the final step of cell division, conventionally proceeds to cell separation by abscission, or complete cytokinesis, but may in certain tissues be incomplete, yielding daughter cells that are interconnected in syncytia by stable intercellular bridges. The mechanisms that determine complete versus incomplete cytokinesis are not known. Here we report a novel in vivo role of the Drosophila CD2AP/CIN85 ortholog Cindr in both complete and incomplete cytokinesis. We also show evidence for the presence of persistent intercellular bridges in the major larval imaginal disc epithelia. During conventional division of both cultured and embryonic cells, Cindr localizes to cleavage furrows, intercellular bridges, and midbodies. Moreover, in cells undergoing incomplete cytokinesis in the female germline and the somatic ovarian follicle cell and larval imaginal disc epithelia, Cindr localizes to arrested cleavage furrows and stable intercellular bridges, respectively. In these structures, Cindr colocalizes with the essential cytokinesis regulator Anillin. We show that Cindr interacts with Anillin and that depletion of either Cindr or Anillin gives rise to binucleate cells and fewer intercellular bridges in vivo. We propose that Cindr and Anillin cooperate to promote intercellular bridge stability during incomplete cytokinesis in Drosophila melanogaster.

BRCA1 and BRCA2: breast/ovarian cancer susceptibility gene products and participants in DNA double-strand break repair

BRCA1 and BRCA2 are tumor suppressor genes, familial mutations in which account for approximately 5% of breast cancer cases in the USA annually. Germ line mutations in BRCA1 that truncate or inactivate the protein lead to a cumulative risk of breast cancer, by age 70, of up to 80%, whereas the risk of ovarian cancer is 30-40%. For germ line BRCA2 mutations, the breast cancer cumulative risk approaches 50%, whereas for ovarian cancers, it is between 10 and 15%. Both BRCA1 and BRCA2 are involved in maintaining genome integrity at least in part by engaging in DNA repair, cell cycle checkpoint control and even the regulation of key mitotic or cell division steps. Unsurprisingly, the complete loss of function of either protein leads to a dramatic increase in genomic instability. How they function in maintaining genome integrity after the onset of DNA damage will be the focus of this review.

The impact of ColRS two-component system and TtgABC efflux pump on phenol tolerance of Pseudomonas putida becomes evident only in growing bacteria

BackgroundWe have recently found that Pseudomonas putida deficient in ColRS two-component system is sensitive to phenol and displays a serious defect on solid glucose medium where subpopulation of bacteria lyses. The latter phenotype is significantly enhanced by the presence of phenol in growth medium. Here, we focused on identification of factors affecting phenol tolerance of the colR-deficient P. putida.ResultsBy using transposon mutagenesis approach we identified a set of phenol-tolerant derivatives of colR-deficient strain. Surprisingly, half of independent phenol tolerant clones possessed miniTn5 insertion in the ttgABC operon. However, though inactivation of TtgABC efflux pump significantly enhanced phenol tolerance, it did not affect phenol-enhanced autolysis of the colR mutant on glucose medium indicating that phenol- and glucose-caused stresses experienced by the colR-deficient P. putida are not coupled. Inactivation of TtgABC pump significantly increased the phenol tolerance of the wild-type P. putida as well. Comparison of phenol tolerance of growing versus starving bacteria revealed that both ColRS and TtgABC systems affect phenol tolerance only under growth conditions and not under starvation. Flow cytometry analysis showed that phenol strongly inhibited cell division and to some extent also caused cell membrane permeabilization to propidium iodide. Single cell analysis of populations of the ttgC- and colRttgC-deficient strains revealed that their membrane permeabilization by phenol resembles that of the wild-type and the colR mutant, respectively. However, cell division of P. putida with inactivated TtgABC pump seemed to be less sensitive to phenol than that of the parental strain. At the same time, cell division appeared to be more inhibited in the colR-mutant strain than in the wild-type P. putida.ConclusionsColRS signal system and TtgABC efflux pump are involved in the phenol tolerance of P. putida. However, as they affect phenol tolerance of growing bacteria only, this indicates that they participate in the regulation of processes which are active during the growth and/or cell division. Single cell analysis data indicated that the cell division step of cell cycle is particularly sensitive to the toxic effect of phenol and its inhibition can be considered as an adaptive response under conditions of phenol stress.

A view from the top: new ligands controlling stomatal development in Arabidopsis

A view from the top: new ligands controlling stomatal development in Arabidopsis

Evidence that the tumor-suppressor protein BRCA2 does not regulate cytokinesis in human cells

Germline mutations in the tumor-suppressor gene BRCA2 predispose to breast and ovarian cancer. BRCA2 plays a well-established role in maintaining genome stability by regulating homologous recombination. BRCA2 has more recently been implicated in cytokinesis, the final step of cell division, but the molecular basis for this remains unknown. We have used time-lapse microscopy, recently developed cytokinesis assays and BAC recombineering (bacterial artificial chromosome recombinogenic engineering) to investigate the function and localization of BRCA2 during cell division. Our analysis suggests that BRCA2 does not regulate cytokinesis in human cells. Thus, cytokinesis defects are unlikely to contribute to chromosomal instability and tumorigenesis in BRCA2-related cancers.

Role of staphylococcal wall teichoic acid in targeting the major autolysin Atl

Staphylococcal cell separation depends largely on the bifunctional autolysin Atl that is processed to amidase-R(1,2) and R(3)-glucosaminidase. These murein hydrolases are targeted via repeat domains (R) to the septal region of the cell surface, thereby allowing localized peptidoglycan hydrolysis and separation of the dividing cells. Here we show that targeting of the amidase repeats is based on an exclusion strategy mediated by wall teichoic acid (WTA). In Staphylococcus aureus wild-type, externally applied repeats (R(1,2)) or endogenously expressed amidase were localized exclusively at the cross-wall region, while in Delta tagO mutant that lacks WTA binding was evenly distributed on the cell surface, which explains the increased fragility and autolysis susceptibility of the mutant. WTA prevented binding of Atl to the old cell wall but not to the cross-wall region suggesting a lower WTA content. In binding studies with ConcanavalinA-fluorescein (ConA-FITC) conjugate that binds preferentially to teichoic acids, ConA-FITC was bound throughout the cell surface with the exception of the cross wall. ConA binding suggest that either content or polymerization of WTA gradually increases with distance from the cross-wall. By preventing binding of Atl, WTA directs Atl to the cross-wall to perform the last step of cell division, namely separation of the daughter cells.

Expression pattern of the septin gene family in acute myeloid leukemias with and without MLL-SEPT fusion genes

Septins are proteins associated with crucial steps in cell division and cellular integrity. In humans, 14 septin genes have been identified, of which five ( SEPT2, SEPT5, SEPT6, SEPT9, and SEPT11) are known to participate in reciprocal translocations with the MLL gene in myeloid neoplasias. We have recently shown a significant down-regulation of both SEPT2 and MLL in myeloid neoplasias with the MLL-SEPT2 fusion gene. In this study, we examined the expression pattern of the other 13 known septin genes in altogether 67 cases of myeloid neoplasia, including three patients with the MLL-SEPT2 fusion gene, four with MLL-SEPT6 fusion, and three patients with the MLL-SEPT9 fusion gene. When compared with normal controls, a statistically significant down-regulation was observed for the expression of both MLL (6.4-fold; p = 0.008) and SEPT6 (1.7-fold; p = 0.002) in MLL-SEPT6 leukemia. Significant down-regulation of MLL was also found in MLL-MLLT3 leukemias. In addition, there was a trend for SEPT9 down-regulation in MLL-SEPT9 leukemias (4.6-fold; p = 0.077). Using hierarchical clustering analysis to compare acute myeloid leukemia genetic subgroups based on their similarity of septin expression changes, we found that MLL-SEPT2 and MLL-SEPT6 neoplasias cluster together apart from the remaining subgroups and that PML-RARA leukemia presents under-expression of most septin family genes.

Spindle Assembly Checkpoint Protein Dynamics Reveal Conserved and Unsuspected Roles in Plant Cell Division

BackgroundIn eukaryotes, the spindle assembly checkpoint (SAC) ensures that chromosomes undergoing mitosis do not segregate until they are properly attached to the microtubules of the spindle.Methodology/Principal FindingsWe investigated the mechanism underlying this surveillance mechanism in plants, by characterising the orthogolous SAC proteins BUBR1, BUB3 and MAD2 from Arabidopsis. We showed that the cell cycle-regulated BUBR1, BUB3.1 and MAD2 proteins interacted physically with each other. Furthermore, BUBR1 and MAD2 interacted specifically at chromocenters. Following SAC activation by global defects in spindle assembly, these three interacting partners localised to unattached kinetochores. In addition, in cases of ‘wait anaphase’, plant SAC proteins were associated with both kinetochores and kinetochore microtubules. Unexpectedly, BUB3.1 was also found in the phragmoplast midline during the final step of cell division in plants.Conclusions/SignificanceWe conclude that plant BUBR1, BUB3.1 and MAD2 proteins may have the SAC protein functions conserved from yeast to humans. The association of BUB3.1 with both unattached kinetochore and phragmoplast suggests that in plant, BUB3.1 may have other roles beyond the spindle assembly checkpoint itself. Finally, this study of the SAC dynamics pinpoints uncharacterised roles of this surveillance mechanism in plant cell division.

Phosphorylation-Dependent Protein Interactions at the Spindle Midzone Mediate Cell Cycle Regulation of Spindle Elongation

The metaphase-to-anaphase transition is one of the most dramatic and highly regulated steps in cell division. At anaphase onset the protease separase dissolves sister chromatid cohesion. Simultaneously, the mitotic spindle elongates as interpolar microtubules (iMTs) slide apart at the spindle midzone, ensuring chromosome segregation. However, it remains unclear how spindle elongation is coordinated with cell cycle progression. Here we demonstrate that phosphorylation of the midzone organizer Ase1 controls localization and function of Cin8, a kinesin-5 that slides iMTs relative to each other. Phosphorylation of Ase1 by Cdk1 (cyclin-dependent kinase) inhibits Cin8 binding to iMTs, preventing bending and collapse of the metaphase spindle. In anaphase Ase1 dephosphorylation by the separase-activated phosphatase Cdc14 is necessary and sufficient for Cin8 recruitment to the midzone, where it drives spindle elongation. Our results reveal that sliding forces at the midzone are activated by separase and explain how spindle elongation is triggered with anaphase entry.