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Sample records for cell division protein

  1. Bacterial cell division proteins as antibiotic targets

    NARCIS (Netherlands)

    T. den Blaauwen; J.M. Andreu; O. Monasterio

    2014-01-01

    Proteins involved in bacterial cell division often do not have a counterpart in eukaryotic cells and they are essential for the survival of the bacteria. The genetic accessibility of many bacterial species in combination with the Green Fluorescence Protein revolution to study localization of protein

  2. Escherichia coli cell division protein FtsZ is a guanine nucleotide binding protein.

    OpenAIRE

    Mukherjee, A; Dai, K; Lutkenhaus, J

    1993-01-01

    FtsZ is an essential cell division protein in Escherichia coli that forms a ring structure at the division site under cell cycle control. The dynamic nature of the FtsZ ring suggests possible similarities to eukaryotic filament forming proteins such as tubulin. In this study we have determined that FtsZ is a GTP/GDP binding protein with GTPase activity. A short segment of FtsZ is homologous to a segment in tubulin believed to be involved in the interaction between tubulin and guanine nucleoti...

  3. CyDiv, a conserved and novel filamentous Cyanobacteria cell division protein involved in septum localization.

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    Dinka eMandakovic

    2016-02-01

    Full Text Available Cell division in bacteria has been studied mostly in Escherichia coli and Bacillus subtilis, model organisms for Gram-negative and Gram-positive bacteria, respectively. However, cell division in filamentous cyanobacteria is poorly understood. Here, we identified a novel protein, named CyDiv (Cyanobacterial Division, encoded by the all2320 gene in Anabaena sp. PCC 7120. We show that CyDiv plays a key role during cell division. CyDiv has been previously described only as an exclusive and conserved hypothetical protein in filamentous cyanobacteria. Using polyclonal antibodies against CyDiv, we showed that it localizes at different positions depending on cell division timing: poles, septum, in both daughter cells, but also in only one of the daughter cells. The partial deletion of CyDiv gene generates partial defects in cell division, including severe membrane instability and anomalous septum localization during late division. The inability to complete knock out CyDiv strains suggests that it is an essential gene. In silico structural protein analyses and our experimental results suggest that CyDiv is an FtsB/DivIC-like protein, and could therefore, be part of an essential late divisome complex in Anabaena sp. PCC 7120.

  4. CyDiv, a Conserved and Novel Filamentous Cyanobacterial Cell Division Protein Involved in Septum Localization

    Science.gov (United States)

    Mandakovic, Dinka; Trigo, Carla; Andrade, Derly; Riquelme, Brenda; Gómez-Lillo, Gabriela; Soto-Liebe, Katia; Díez, Beatriz; Vásquez, Mónica

    2016-01-01

    Cell division in bacteria has been studied mostly in Escherichia coli and Bacillus subtilis, model organisms for Gram-negative and Gram-positive bacteria, respectively. However, cell division in filamentous cyanobacteria is poorly understood. Here, we identified a novel protein, named CyDiv (Cyanobacterial Division), encoded by the all2320 gene in Anabaena sp. PCC 7120. We show that CyDiv plays a key role during cell division. CyDiv has been previously described only as an exclusive and conserved hypothetical protein in filamentous cyanobacteria. Using polyclonal antibodies against CyDiv, we showed that it localizes at different positions depending on cell division timing: poles, septum, in both daughter cells, but also in only one of the daughter cells. The partial deletion of CyDiv gene generates partial defects in cell division, including severe membrane instability and anomalous septum localization during late division. The inability to complete knock out CyDiv strains suggests that it is an essential gene. In silico structural protein analyses and our experimental results suggest that CyDiv is an FtsB/DivIC-like protein, and could therefore, be part of an essential late divisome complex in Anabaena sp. PCC 7120. PMID:26903973

  5. Localization of Cell Division Protein FtsQ by Immunofluorescence Microscopy in Dividing and Nondividing Cells of Escherichia coli

    Science.gov (United States)

    Buddelmeijer, Nienke; Aarsman, Mirjam E. G.; Kolk, Arend H. J.; Vicente, Miguel; Nanninga, Nanne

    1998-01-01

    The localization of cell division protein FtsQ in Escherichia coli wild-type cells was studied by immunofluorescence microscopy with specific monoclonal antibodies. FtsQ could be localized to the division site in constricting cells. FtsQ could also localize to the division site in ftsQ1(Ts) cells grown at the permissive temperature. A hybrid protein in which the cytoplasmic domain and the transmembrane domain were derived from the γ form of penicillin-binding protein 1B and the periplasmic domain was derived from FtsQ was also able to localize to the division site. This result indicates that the periplasmic domain of FtsQ determines the localization of FtsQ, as has also been concluded by others for the periplasmic domain of FtsN. Noncentral FtsQ foci were found in the area of the cell where the nucleoid resides and were therefore assumed to represent sites where the FtsQ protein is synthesized and simultaneously inserted into the cytoplasmic membrane. PMID:9829918

  6. Interaction of Mouse Pem Protein and Cell Division Cycle 37 Homolog

    Institute of Scientific and Technical Information of China (English)

    Fen GUO; Yue-Qin LI; Shi-Qian LI; Zhi-Wen LUO; Xin ZHANG; Dong-Sheng TANG; Tian-Hong ZHOU

    2005-01-01

    Mouse Pem, a homeobox gene, encodes a protein consisting of 210 amino acid residues. To study the function of mouse Pem protein, we used the yeast two-hybrid system to screen the library of 7-day mouse embryo with full-length mouse Pem eDNA. Fifty-two colonies were obtained after 1.57×108 colonies were screened by nutrition limitation and β-galactosidase assay. Seven individual insert fragments were obtained from the library, and three of them were identified, one of which was confirmed to be the cell division cycle 37 (Cdc37) homolog gene by sequencing. The interaction between mouse Pem and Cdc37homolog was then confirmed by glutathione S-transferase pull-down assay, and the possible interaction model was suggested.

  7. Targeting the Wolbachia cell division protein FtsZ as a new approach for antifilarial therapy.

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    Zhiru Li

    2011-11-01

    Full Text Available The use of antibiotics targeting the obligate bacterial endosymbiont Wolbachia of filarial parasites has been validated as an approach for controlling filarial infection in animals and humans. Availability of genomic sequences for the Wolbachia (wBm present in the human filarial parasite Brugia malayi has enabled genome-wide searching for new potential drug targets. In the present study, we investigated the cell division machinery of wBm and determined that it possesses the essential cell division gene ftsZ which was expressed in all developmental stages of B. malayi examined. FtsZ is a GTPase thereby making the protein an attractive Wolbachia drug target. We described the molecular characterization and catalytic properties of Wolbachia FtsZ. We also demonstrated that the GTPase activity was inhibited by the natural product, berberine, and small molecule inhibitors identified from a high-throughput screen. Furthermore, berberine was also effective in reducing motility and reproduction in B. malayi parasites in vitro. Our results should facilitate the discovery of selective inhibitors of FtsZ as a novel anti-symbiotic approach for controlling filarial infection. NOTE: The nucleotide sequences reported in this paper are available in GenBank™ Data Bank under the accession number wAlB-FtsZ (JN616286.

  8. The RNA binding protein Larp1 regulates cell division, apoptosis and cell migration.

    Science.gov (United States)

    Burrows, Carla; Abd Latip, Normala; Lam, Sarah-Jane; Carpenter, Lee; Sawicka, Kirsty; Tzolovsky, George; Gabra, Hani; Bushell, Martin; Glover, David M; Willis, Anne E; Blagden, Sarah P

    2010-09-01

    The RNA binding protein Larp1 was originally shown to be involved in spermatogenesis, embryogenesis and cell-cycle progression in Drosophila. Our data show that mammalian Larp1 is found in a complex with poly A binding protein and eukaryote initiation factor 4E and is associated with 60S and 80S ribosomal subunits. A reduction in Larp1 expression by siRNA inhibits global protein synthesis rates and results in mitotic arrest and delayed cell migration. Consistent with these data we show that Larp1 protein is present at the leading edge of migrating cells and interacts directly with cytoskeletal components. Taken together, these data suggest a role for Larp1 in facilitating the synthesis of proteins required for cellular remodelling and migration. PMID:20430826

  9. The Snail protein family regulates neuroblast expression of inscuteable and string, genes involved in asymmetry and cell division in Drosophila.

    Science.gov (United States)

    Ashraf, S I; Ip, Y T

    2001-12-01

    Delaminated neuroblasts in Drosophila function as stem cells during embryonic central nervous system development. They go through repeated asymmetric divisions to generate multiple ganglion mother cells, which divide only once more to produce postmitotic neurons. Snail, a zinc-finger transcriptional repressor, is a pan-neural protein, based on its extensive expression in neuroblasts. Previous results have demonstrated that Snail and related proteins, Worniu and Escargot, have redundant and essential functions in the nervous system. We show that the Snail family of proteins control central nervous system development by regulating genes involved in asymmetry and cell division of neuroblasts. In mutant embryos that have the three genes deleted, the expression of inscuteable is significantly lowered, while the expression of other genes that participate in asymmetric division, including miranda, staufen and prospero, appears normal. The deletion mutants also have much reduced expression of string, suggesting that a key component that drives neuroblast cell division is abnormal. Consistent with the gene expression defects, the mutant embryos lose the asymmetric localization of prospero RNA in neuroblasts and lose the staining of Prospero protein that is normally present in ganglion mother cells. Simultaneous expression of inscuteable and string in the snail family deletion mutant efficiently restores Prospero expression in ganglion mother cells, demonstrating that the two genes are key targets of Snail in neuroblasts. Mutation of the dCtBP co-repressor interaction motifs in the Snail protein leads to reduction of the Snail function in central nervous system. These results suggest that the Snail family of proteins control both asymmetry and cell division of neuroblasts by activating, probably indirectly, the expression of inscuteable and string. PMID:11731456

  10. Universal Protein Distributions in a Model of Cell Growth and Division

    CERN Document Server

    Brenner, Naama; Osmanovic, Dino; Rabin, Yitzhak; Salman, Hanna; Stein, D L

    2015-01-01

    Protein distributions measured under a broad set of conditions in bacteria and yeast exhibit a universal skewed shape, with variances depending quadratically on means. For bacteria these properties are reproduced by protein accumulation and division dynamics across generations. We present a stochastic growth-and-division model with feedback which captures these observed properties. The limiting copy number distribution is calculated exactly, and a single parameter is found to determine the distribution shape and the variance-to-mean relation. Estimating this parameter from bacterial temporal data reproduces the measured universal distribution shape with high accuracy, and leads to predictions for future experiments.

  11. Radmis, a novel mitotic spindle protein that functions in cell division of neural progenitors.

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    Takahito Yumoto

    Full Text Available Developmental dynamics of neural stem/progenitor cells (NSPCs are crucial for embryonic and adult neurogenesis, but its regulatory factors are not fully understood. By differential subtractive screening with NSPCs versus their differentiated progenies, we identified the radmis (radial fiber and mitotic spindle/ckap2l gene, a novel microtubule-associated protein (MAP enriched in NSPCs. Radmis is a putative substrate for the E3-ubiquitin ligase, anaphase promoting complex/cyclosome (APC/C, and is degraded via the KEN box. Radmis was highly expressed in regions of active neurogenesis throughout life, and its distribution was dynamically regulated during NSPC division. In embryonic and perinatal brains, radmis localized to bipolar mitotic spindles and radial fibers (basal processes of dividing NSPCs. As central nervous system development proceeded, radmis expression was lost in most brain regions, except for several neurogenic regions. In adult brain, radmis expression persisted in the mitotic spindles of both slowly-dividing stem cells and rapid amplifying progenitors. Overexpression of radmis in vitro induced hyper-stabilization of microtubules, severe defects in mitotic spindle formation, and mitotic arrest. In vivo gain-of-function using in utero electroporation revealed that radmis directed a reduction in NSPC proliferation and a concomitant increase in cell cycle exit, causing a reduction in the Tbr2-positive basal progenitor population and shrinkage of the embryonic subventricular zone. Besides, radmis loss-of-function by shRNAs induced the multipolar mitotic spindle structure, accompanied with the catastrophe of chromosome segregation including the long chromosome bridge between two separating daughter nuclei. These findings uncover the indispensable role of radmis in mitotic spindle formation and cell-cycle progression of NSPCs.

  12. An archaebacterial homologue of the essential eubacterial cell division protein FtsZ.

    OpenAIRE

    Baumann, P; Jackson, S P

    1996-01-01

    Life falls into three fundamental domains--Archaea, Bacteria, and Eucarya (formerly archaebacteria, eubacteria, and eukaryotes,. respectively). Though Archaea lack nuclei and share many morphological features with Bacteria, molecular analyses, principally of the transcription and translation machineries, have suggested that Archaea are more related to Eucarya than to Bacteria. Currently, little is known about the archaeal cell division apparatus. In Bacteria, a crucial component of the cell d...

  13. SecA is required for membrane targeting of the cell division protein DivIVA in vivo

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    SvenHalbedel

    2014-02-01

    Full Text Available The conserved protein DivIVA is involved in different morphogenetic processes in Gram-positive bacteria. In Bacillus subtilis, the protein localises to the cell division site and cell poles, and functions as a scaffold for proteins that regulate division site selection, and for proteins that are required for sporulation. To identify other proteins that bind to DivIVA, we performed an in vivo cross-linking experiment. A possible candidate that emerged was the secretion motor ATPase SecA. SecA mutants have been described that inhibit sporulation, and since DivIVA is necessary for sporulation, we examined the localisation of DivIVA in these mutants. Surprisingly, DivIVA was delocalised, suggesting that SecA is required for DivIVA targeting. To further corroborate this, we performed SecA depletion and inhibition experiments, which provided further indications that DivIVA localisation depends on SecA. Cell fractionation experiments showed that SecA is important for binding of DivIVA to the cell membrane. This was unexpected since DivIVA does not contain a signal sequence, and is able to bind to artificial lipid membranes in vitro without support of other proteins. SecA is required for protein secretion and membrane insertion, and therefore its role in DivIVA localisation is likely indirect. Possible alternative roles of SecA in DivIVA folding and/or targeting are discussed.

  14. Structure of the bacterial cell division determinant GpsB and its interaction with penicillin-binding proteins.

    Science.gov (United States)

    Rismondo, Jeanine; Cleverley, Robert M; Lane, Harriet V; Großhennig, Stephanie; Steglich, Anne; Möller, Lars; Mannala, Gopala Krishna; Hain, Torsten; Lewis, Richard J; Halbedel, Sven

    2016-03-01

    Each bacterium has to co-ordinate its growth with division to ensure genetic stability of the population. Consequently, cell division and growth are tightly regulated phenomena, albeit different bacteria utilise one of several alternative regulatory mechanisms to maintain control. Here we consider GpsB, which is linked to cell growth and division in Gram-positive bacteria. ΔgpsB mutants of the human pathogen Listeria monocytogenes show severe lysis, division and growth defects due to distortions of cell wall biosynthesis. Consistent with this premise, GpsB interacts both in vitro and in vivo with the major bi-functional penicillin-binding protein. We solved the crystal structure of GpsB and the interaction interfaces in both proteins are identified and validated. The inactivation of gpsB results in strongly attenuated virulence in animal experiments, comparable in degree to classical listerial virulence factor mutants. Therefore, GpsB is essential for in vitro and in vivo growth of a highly virulent food-borne pathogen, suggesting that GpsB could be a target for the future design of novel antibacterials. PMID:26575090

  15. Nanoengineering: Super symmetry in cell division

    Science.gov (United States)

    Huang, Kerwyn Casey

    2015-08-01

    Bacterial cells can be sculpted into different shapes using nanofabricated chambers and then used to explore the spatial adaptation of protein oscillations that play an important role in cell division.

  16. The Effect of Olive Oil Mill Effluent on the Mitotic Cell Division and Total Protein Amount of the Root Tips of Triticum aestivumL.

    OpenAIRE

    Aybeke, Mehmet; OLGUN, Göksel

    2000-01-01

    In this work sitotoxic and mutagenic effects Olive Oil Mill Effluent (OOME) on the root tips of Triticum aestivumL. were investigated. In this purpose, germination rate of seeds, mitotic division abnormalities and total protein amounts were evaluated. The seeds kept in various OOME concentrastions, it was determinated that germination rate decreased, whilst mitotic abnormalities and mitotic cell division frequency increased. Especially, the increased cell division frequency was of signif...

  17. Actin related protein complex subunit 1b controls sperm release, barrier integrity and cell division during adult rat spermatogenesis.

    Science.gov (United States)

    Kumar, Anita; Dumasia, Kushaan; Deshpande, Sharvari; Gaonkar, Reshma; Balasinor, N H

    2016-08-01

    Actin remodeling is a vital process for signaling, movement and survival in all cells. In the testes, extensive actin reorganization occurs at spermatid-Sertoli cell junctions during sperm release (spermiation) and at inter Sertoli cell junctions during restructuring of the blood testis barrier (BTB). During spermiation, tubulobulbar complexes (TBCs), rich in branched actin networks, ensure recycling of spermatid-Sertoli cell junctional molecules. Similar recycling occurs during BTB restructuring around the same time as spermiation occurs. Actin related protein 2/3 complex is an essential actin nucleation and branching protein. One of its subunits, Arpc1b, was earlier found to be down-regulated in an estrogen-induced rat model of spermiation failure. Also, Arpc1b was found to be estrogen responsive through estrogen receptor beta in seminiferous tubule culture. Here, knockdown of Arpc1b by siRNA in adult rat testis led to defects in spermiation caused by failure in TBC formation. Knockdown also compromised BTB integrity and caused polarity defects of mature spermatids. Apart from these effects pertaining to Sertoli cells, Arpc1b reduction perturbed ability of germ cells to enter G2/M phase thus hindering cell division. In summary, Arpc1b, an estrogen responsive gene, is a regulator of spermiation, mature spermatid polarity, BTB integrity and cell division during adult spermatogenesis. PMID:27113856

  18. Direct interaction between the cell division protein FtsZ and the cell differentiation protein SpoIIE

    OpenAIRE

    Lucet, Isabelle; Feucht, Andrea; Yudkin, Michael D.; Errington, Jeffery

    2000-01-01

    SpoIIE is a bifunctional protein with two critical roles in the establishment of cell fate in Bacillus subtilis. First, SpoIIE is needed for the normal formation of the asymmetrically positioned septum that forms early in sporulation and separates the mother cell from the prespore compartment. Secondly, SpoIIE is essential for the activation of the first compartment-specific transcription factor σF in the prespore. After initiation of sporulation, SpoIIE localizes to the potential asymmetric ...

  19. Oriented Cell Division in the C. elegans Embryo Is Coordinated by G-Protein Signaling Dependent on the Adhesion GPCR LAT-1.

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    Antje Müller

    2015-10-01

    Full Text Available Orientation of spindles and cell division planes during development of many species ensures that correct cell-cell contacts are established, which is vital for proper tissue formation. This is a tightly regulated process involving a complex interplay of various signals. The molecular mechanisms underlying several of these pathways are still incompletely understood. Here, we identify the signaling cascade of the C. elegans latrophilin homolog LAT-1, an essential player in the coordination of anterior-posterior spindle orientation during the fourth round of embryonic cell division. We show that the receptor mediates a G protein-signaling pathway revealing that G-protein signaling in oriented cell division is not solely GPCR-independent. Genetic analyses showed that through the interaction with a Gs protein LAT-1 elevates intracellular cyclic AMP (cAMP levels in the C. elegans embryo. Stimulation of this G-protein cascade in lat-1 null mutant nematodes is sufficient to orient spindles and cell division planes in the embryo in the correct direction. Finally, we demonstrate that LAT-1 is activated by an intramolecular agonist to trigger this cascade. Our data support a model in which a novel, GPCR-dependent G protein-signaling cascade mediated by LAT-1 controls alignment of cell division planes in an anterior-posterior direction via a metabotropic Gs-protein/adenylyl cyclase pathway by regulating intracellular cAMP levels.

  20. Oriented Cell Division in the C. elegans Embryo Is Coordinated by G-Protein Signaling Dependent on the Adhesion GPCR LAT-1

    Science.gov (United States)

    Fiedler, Franziska; Sastradihardja, Tania; Binder, Claudia; Schnabel, Ralf; Kungel, Jana; Rothemund, Sven; Hennig, Christian; Schöneberg, Torsten; Prömel, Simone

    2015-01-01

    Orientation of spindles and cell division planes during development of many species ensures that correct cell-cell contacts are established, which is vital for proper tissue formation. This is a tightly regulated process involving a complex interplay of various signals. The molecular mechanisms underlying several of these pathways are still incompletely understood. Here, we identify the signaling cascade of the C. elegans latrophilin homolog LAT-1, an essential player in the coordination of anterior-posterior spindle orientation during the fourth round of embryonic cell division. We show that the receptor mediates a G protein-signaling pathway revealing that G-protein signaling in oriented cell division is not solely GPCR-independent. Genetic analyses showed that through the interaction with a Gs protein LAT-1 elevates intracellular cyclic AMP (cAMP) levels in the C. elegans embryo. Stimulation of this G-protein cascade in lat-1 null mutant nematodes is sufficient to orient spindles and cell division planes in the embryo in the correct direction. Finally, we demonstrate that LAT-1 is activated by an intramolecular agonist to trigger this cascade. Our data support a model in which a novel, GPCR-dependent G protein-signaling cascade mediated by LAT-1 controls alignment of cell division planes in an anterior-posterior direction via a metabotropic Gs-protein/adenylyl cyclase pathway by regulating intracellular cAMP levels. PMID:26505631

  1. A mechanism for ParB-dependent waves of ParA, a protein related to DNA segregation during cell division in prokaryotes

    DEFF Research Database (Denmark)

    Hunding, Axel; Gerdes, Kenn; Charbon, Gitte Ebersbach

    2003-01-01

    Prokaryotic plasmids encode partitioning (par) loci involved in segregation of DNA to daughter cells at cell division. A functional fusion protein consisting of Walker-type ParA ATPase and green fluorescent protein (Gfp) oscillates back and forth within nucleoid regions with a wave period of about...

  2. Polarized Cell Division of Chlamydia trachomatis.

    Science.gov (United States)

    Abdelrahman, Yasser; Ouellette, Scot P; Belland, Robert J; Cox, John V

    2016-08-01

    Bacterial cell division predominantly occurs by a highly conserved process, termed binary fission, that requires the bacterial homologue of tubulin, FtsZ. Other mechanisms of bacterial cell division that are independent of FtsZ are rare. Although the obligate intracellular human pathogen Chlamydia trachomatis, the leading bacterial cause of sexually transmitted infections and trachoma, lacks FtsZ, it has been assumed to divide by binary fission. We show here that Chlamydia divides by a polarized cell division process similar to the budding process of a subset of the Planctomycetes that also lack FtsZ. Prior to cell division, the major outer-membrane protein of Chlamydia is restricted to one pole of the cell, and the nascent daughter cell emerges from this pole by an asymmetric expansion of the membrane. Components of the chlamydial cell division machinery accumulate at the site of polar growth prior to the initiation of asymmetric membrane expansion and inhibitors that disrupt the polarity of C. trachomatis prevent cell division. The polarized cell division of C. trachomatis is the result of the unipolar growth and FtsZ-independent fission of this coccoid organism. This mechanism of cell division has not been documented in other human bacterial pathogens suggesting the potential for developing Chlamydia-specific therapeutic treatments. PMID:27505160

  3. The WD40 repeat protein NEDD1 functions in microtubule organization during cell division in Arabidopsis thaliana.

    Science.gov (United States)

    Zeng, C J Tracy; Lee, Y-R Julie; Liu, Bo

    2009-04-01

    Although cells of flowering plants lack a structurally defined microtubule-organizing center like the centrosome, organization of the spindles and phragmoplasts in mitosis is known to involve the evolutionarily conserved gamma-tubulin complex. We have investigated the function of Arabidopsis thaliana NEDD1, a WD40 repeat protein related to the animal NEDD1/GCP-WD protein, which interacts with the gamma-tubulin complex. The NEDD1 protein decorates spindle microtubules (MTs) preferentially toward spindle poles and phragmoplast MTs toward their minus ends. A T-DNA insertional allele of the single NEDD1 gene was isolated and maintained in heterozygous sporophytes, and NEDD1's function in cell division was analyzed in haploid microspores produced by the heterozygote. In approximately half of the dividing microspores exhibiting aberrant MT organization, spindles were no longer restricted to the cell periphery and became abnormally elongated. After mitosis, MTs aggregated between reforming nuclei but failed to appear in a bipolar configuration. Consequently, defective microspores did not form a continuous cell plate, and two identical nuclei were produced with no differentiation into generative and vegetative cells. Our results support the notion that the plant NEDD1 homolog plays a critical role in MT organization during mitosis, and its function is likely linked to that of the gamma-tubulin complex. PMID:19383896

  4. Genes involved in cell division in mycoplasmas

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    Frank Alarcón

    2007-01-01

    Full Text Available Bacterial cell division has been studied mainly in model systems such as Escherichia coli and Bacillus subtilis, where it is described as a complex process with the participation of a group of proteins which assemble into a multiprotein complex called the septal ring. Mycoplasmas are cell wall-less bacteria presenting a reduced genome. Thus, it was important to compare their genomes to analyze putative genes involved in cell division processes. The division and cell wall (dcw cluster, which in E. coli and B. subtilis is composed of 16 and 17 genes, respectively, is represented by only three to four genes in mycoplasmas. Even the most conserved protein, FtsZ, is not present in all mycoplasma genomes analyzed so far. A model for the FtsZ protein from Mycoplasma hyopneumoniae and Mycoplasma synoviae has been constructed. The conserved residues, essential for GTP/GDP binding, are present in FtsZ from both species. A strong conservation of hydrophobic amino acid patterns is observed, and is probably necessary for the structural stability of the protein when active. M. synoviae FtsZ presents an extended amino acid sequence at the C-terminal portion of the protein, which may participate in interactions with other still unknown proteins crucial for the cell division process.

  5. Evolution and tinkering: what do a protein kinase, a transcriptional regulator and chromosome segregation/cell division proteins have in common?

    Science.gov (United States)

    Derouiche, Abderahmane; Shi, Lei; Kalantari, Aida; Mijakovic, Ivan

    2016-02-01

    In this study, we focus on functional interactions among multi-domain proteins which share a common evolutionary origin. The examples we develop are four Bacillus subtilis proteins, which all possess an ATP-binding Walker motif: the bacterial tyrosine kinase (BY-kinase) PtkA, the chromosome segregation protein Soj (ParA), the cell division protein MinD and a transcription regulator SalA. These proteins have arisen via duplication of the ancestral ATP-binding domain, which has undergone fusions with other functional domains in the process of divergent evolution. We point out that these four proteins, despite having very different physiological roles, engage in an unusually high number of binary functional interactions. Namely, MinD attracts Soj and PtkA to the cell pole, and in addition, activates the kinase function of PtkA. SalA also activates the kinase function of PtkA, and it gets phosphorylated by PtkA as well. The consequence of this phosphorylation is the activation of SalA as a transcriptional repressor. We hypothesize that these functional interactions remain preserved during divergent evolution and represent a constraint on the process of evolutionary "tinkering", brought about by fusions of different functional domains. PMID:26286503

  6. Control of apoptosis by asymmetric cell division.

    Science.gov (United States)

    Hatzold, Julia; Conradt, Barbara

    2008-04-01

    Asymmetric cell division and apoptosis (programmed cell death) are two fundamental processes that are important for the development and function of multicellular organisms. We have found that the processes of asymmetric cell division and apoptosis can be functionally linked. Specifically, we show that asymmetric cell division in the nematode Caenorhabditis elegans is mediated by a pathway involving three genes, dnj-11 MIDA1, ces-2 HLF, and ces-1 Snail, that directly control the enzymatic machinery responsible for apoptosis. Interestingly, the MIDA1-like protein GlsA of the alga Volvox carteri, as well as the Snail-related proteins Snail, Escargot, and Worniu of Drosophila melanogaster, have previously been implicated in asymmetric cell division. Therefore, C. elegans dnj-11 MIDA1, ces-2 HLF, and ces-1 Snail may be components of a pathway involved in asymmetric cell division that is conserved throughout the plant and animal kingdoms. Furthermore, based on our results, we propose that this pathway directly controls the apoptotic fate in C. elegans, and possibly other animals as well. PMID:18399720

  7. Expression, purification and crystallization of the cell-division protein YgfE from Escherichia coli

    International Nuclear Information System (INIS)

    An open reading frame from E. coli MG1655 has been cloned, expressed and purified. Crystals obtained from the purified recombinant protein have been obtained in a variety of different forms diffracting to 1.8 Å resolution

  8. Ser/Thr protein kinase StkP regulates cell division in Streptococcus pneumoniae

    Czech Academy of Sciences Publication Activity Database

    Nováková, Linda; Bezoušková, Silvia; Pompach, Petr; Branny, Pavel

    Bratislava : Československá spoločnosť mikrobiologická, 2010. s. 196-196. ISBN 970-80-970477-8-8 Institutional research plan: CEZ:AV0Z50200510 Keywords : protein kinase Subject RIV: EE - Microbiology, Virology

  9. Dynamics of pre-replication complex proteins during the cell division cycle.

    OpenAIRE

    Prasanth, Supriya G.; Méndez, Juan; Prasanth, Kannanganattu V.; Stillman, Bruce

    2004-01-01

    Replication of the human genome every time a cell divides is a highly coordinated process that ensures accurate and efficient inheritance of the genetic information. The molecular mechanism that guarantees that many origins of replication fire only once per cell-cycle has been the area of intense research. The origin recognition complex (ORC) marks the position of replication origins in the genome and serves as the landing pad for the assembly of a multiprotein, pre-replicative complex (pre-R...

  10. Modeling of the dynamic pole-to-pole oscillations of the min proteins in bacterial cell division: The effect of an external field

    CERN Document Server

    Modchang, C; Triampo, W; Ngamsaad, W; Nuttawut, N; Tang, I M; Lenbury, Y; Modchang, Charin; Kanthang, Paisan; Triampo, Wannapong; Ngamsaad, Waipot; Nuttawut, Narin; Lenbury, Yongwimol

    2004-01-01

    One of the most important steps in the developmental process of the bacteria cell at the cellular level is the determination of the middle of the cell and the proper placement of the septum, these being essential to the division of the cell. In E. coli, this step depends on the proteins MinC, MinD, and MinE. Exposure to a constant electric field may cause the bacteria cell division mechanism to change, resulting in an abnormal cytokinesis. To see the effects of an external field e.g., an electric or magnetic field on this process, we have solved a set of deterministic reaction diffusion equations, which incorporate the influence of an electric field. We have found some changes in the dynamics of the oscillations of the min proteins from pole to pole. The numerical results show some interesting effects, which are qualitatively in good agreement with some experimental results.

  11. LocZ is a new cell division protein involved in proper septum placement in Streptococcus pneumoniae

    Czech Academy of Sciences Publication Activity Database

    Holečková, Nela; Doubravová, Linda; Massidda, Orietta; Molle, Virginie; Buriánková, Karolína; Benada, Oldřich; Kofroňová, Olga; Ulrych, Aleš; Branny, Pavel

    2015-01-01

    Roč. 6, č. 1 (2015), s. 1-13. ISSN 2150-7511 R&D Projects: GA ČR GAP207/12/1568; GA ČR GAP302/12/0256 Institutional support: RVO:61388971 Keywords : cell division * septum placement * Streptococcus pneumoniae Subject RIV: EE - Microbiology, Virology Impact factor: 6.786, year: 2014

  12. The cytological changes of tobacco zygote and proembryo cells induced by beta-glucosyl Yariv reagent suggest the involvement of arabinogalactan proteins in cell division and cell plate formation

    Directory of Open Access Journals (Sweden)

    Yu Miao

    2012-08-01

    Full Text Available Abstract Background In dicotyledonous plant, the first asymmetric zygotic division and subsequent several cell divisions are crucial for proembryo pattern formation and later embryo development. Arabinogalactan proteins (AGPs are a family of extensively glycosylated cell surface proteins that are thought to have important roles in various aspects of plant growth and development, including embryogenesis. Previous results from our laboratory show that AGPs are concerned with tobacco egg cell fertilization and zygotic division. However, how AGPs interact with other factors involved in zygotic division and proembryo development remains unknown. Results In this study, we used the tobacco in vitro zygote culture system and series of meticulous cell biology techniques to investigate the roles of AGPs in zygote and proembryo cell division. For the first time, we examined tobacco proembryo division patterns detailed to every cell division. The bright-field images and statistical results both revealed that with the addition of an exogenous AGPs inhibitor, beta-glucosyl Yariv (beta-GlcY reagent, the frequency of aberrant division increased remarkably in cultured tobacco zygotes and proembryos, and the cell plate specific locations of AGPs were greatly reduced after beta-GlcY treatment. In addition, the accumulations of new cell wall materials were also significantly affected by treating with beta-GlcY. Detection of cellulose components by Calcofluor white stain showed that strong fluorescence was located in the newly formed wall of daughter cells after the zygotic division of in vivo samples and the control samples from in vitro culture without beta-GlcY treatment; while there was only weak fluorescence in the newly formed cell walls with beta-GlcY treatment. Immunocytochemistry examination with JIM5 and JIM7 respectively against the low- and high-esterified pectins displayed that these two pectins located in opposite positions of zygotes and proembryos in

  13. Disruption of an M. tuberculosis Membrane Protein Causes a Magnesium-dependent Cell Division Defect and Failure to Persist in Mice

    Science.gov (United States)

    Goodsmith, Nichole; Guo, Xinzheng V.; Vandal, Omar H.; Vaubourgeix, Julien; Wang, Ruojun; Botella, Hélène; Song, Shuang; Bhatt, Kamlesh; Liba, Amir; Salgame, Padmini; Schnappinger, Dirk; Ehrt, Sabine

    2015-01-01

    The identification of Mycobacterium tuberculosis genes necessary for persistence in vivo provides insight into bacterial biology as well as host defense strategies. We show that disruption of M. tuberculosis membrane protein PerM (Rv0955) resulted in an IFN-γ-dependent persistence defect in chronic mouse infection despite the mutant’s near normal growth during acute infection. The perM mutant required increased magnesium for replication and survival; incubation in low magnesium media resulted in cell elongation and lysis. Transcriptome analysis of the perM mutant grown in reduced magnesium revealed upregulation of cell division and cell wall biosynthesis genes, and live cell imaging showed PerM accumulation at the division septa in M. smegmatis. The mutant was acutely sensitive to β-lactam antibiotics, including specific inhibitors of cell division-associated peptidoglycan transpeptidase FtsI. Together, these data implicate PerM as a novel player in mycobacterial cell division and pathogenesis, and are consistent with the hypothesis that immune activation deprives M. tuberculosis of magnesium. PMID:25658098

  14. Regulation of cell division in higher plants

    Energy Technology Data Exchange (ETDEWEB)

    Jacobs, T.W.

    1992-01-01

    Cell division is arguably the most fundamental of all developmental processes. In higher plants, mitotic activity is largely confined to foci of patterned cell divisions called meristems. From these perpetually embryonic tissues arise the plant's essential organs of light capture, support, protection and reproduction. Once an adequate understanding of plant cell mitotic regulation is attained, unprecedented opportunities will ensue for analyzing and genetically controlling diverse aspects of development, including plant architecture, leaf shape, plant height, and root depth. The mitotic cycle in a variety of model eukaryotic systems in under the control of a regulatory network of striking evolutionary conservation. Homologues of the yeast cdc2 gene, its catalytic product, p34, and the cyclin regulatory subunits of the MPF complex have emerged as ubiquitous mitotic regulators. We have cloned cdc2-like and cyclin genes from pea. As in other eukaryotic model systems, p34 of Pisum sativum is a subunit of a high molecular weight complex which binds the fission yeast p13 protein and displays histone H1 kinase activity in vitro. Our primary objective in this study is to gain baseline information about the regulation of this higher plant cell division control complex in non-dividing, differentiated cells as well as in synchronous and asynchronous mitotic cells. We are investigating cdc2 and cyclin expression at the levels of protein abundance, protein phosphorylation and quaternary associations.

  15. The Monofunctional Glycosyltransferase of Escherichia coli Localizes to the Cell Division Site and Interacts with Penicillin-Binding Protein 3, FtsW, and FtsN▿ ‡

    OpenAIRE

    Derouaux, Adeline; Wolf, Benoît; Fraipont, Claudine; Breukink, Eefjan; Nguyen-Distèche, Martine; Terrak, Mohammed

    2007-01-01

    The monofunctional peptidoglycan glycosyltransferase (MtgA) catalyzes glycan chain elongation of the bacterial cell wall. Here we show that MtgA localizes at the division site of Escherichia coli cells that are deficient in PBP1b and produce a thermosensitive PBP1a and is able to interact with three constituents of the divisome, PBP3, FtsW, and FtsN, suggesting that MtgA may play a role in peptidoglycan assembly during the cell cycle in collaboration with other proteins.

  16. Impact of the cell division cycle on gene circuits

    Science.gov (United States)

    Bierbaum, Veronika; Klumpp, Stefan

    2015-12-01

    In growing cells, protein synthesis and cell growth are typically not synchronous, and, thus, protein concentrations vary over the cell division cycle. We have developed a theoretical description of genetic regulatory systems in bacteria that explicitly considers the cell division cycle to investigate its impact on gene expression. We calculate the cell-to-cell variations arising from cells being at different stages in the division cycle for unregulated genes and for basic regulatory mechanisms. These variations contribute to the extrinsic noise observed in single-cell experiments, and are most significant for proteins with short lifetimes. Negative autoregulation buffers against variation of protein concentration over the division cycle, but the effect is found to be relatively weak. Stronger buffering is achieved by an increased protein lifetime. Positive autoregulation can strongly amplify such variation if the parameters are set to values that lead to resonance-like behaviour. For cooperative positive autoregulation, the concentration variation over the division cycle diminishes the parameter region of bistability and modulates the switching times between the two stable states. The same effects are seen for a two-gene mutual-repression toggle switch. By contrast, an oscillatory circuit, the repressilator, is only weakly affected by the division cycle.

  17. Abnormal number cell division of human thyroid anaplastic carcinoma cell line, SW 1736

    Directory of Open Access Journals (Sweden)

    Keiichi Ikeda

    2015-12-01

    Full Text Available Cell division, during which a mother cell usually divides into two daughter cells during one cell cycle, is the most important physiological event of cell biology. We observed one-to-four cell division during imaging of live SW1736 human thyroid anaplastic carcinoma cells transfected with a plasmid expressing the hybrid protein of green fluorescent protein and histone 2B (plasmid eGFP-H2B. Analysis of the images revealed a mother cell divided into four daughter cells. And one of the abnormally divided daughter cells subsequently formed a dinucleate cell.

  18. Asymmetric cell division: a persistent issue?

    OpenAIRE

    Aakre, Christopher D.; Laub, Michael T.

    2012-01-01

    Heterogeneity within a clonal population of cells can increase survival in the face of environmental stress. In a recent issue of Science, Aldridge et al. (2012) demonstrate that cell division in mycobacteria is asymmetric, producing daughter cells that differ in size, growth rate, and susceptibility to antibiotics.

  19. Cell division activity during apical hook development

    NARCIS (Netherlands)

    Raz, V.; Koornneef, M.

    2001-01-01

    Growth during plant development is predominantly governed by the combined activities of cell division and cell elongation. The relative contribution of both activities controls the growth of a tissue. A fast change in growth is exhibited at the apical hypocotyl of etiolated seedlings where cells gro

  20. The Escherichia coli Cell Division Protein and Model Tat Substrate SufI (FtsP) Localizes to the Septal Ring and Has a Multicopper Oxidase-Like Structure

    OpenAIRE

    Tarry, Michael; Arends, S. J. Ryan; Roversi, Pietro; Piette, Evan; Sargent, Frank; Berks, Ben C.; Weiss, David S.; Lea, Susan M.

    2009-01-01

    The Escherichia coli protein SufI (FtsP) has recently been proposed to be a component of the cell division apparatus. The SufI protein is also in widespread experimental use as a model substrate in studies of the Tat (twin arginine translocation) protein transport system. We have used SufI-GFP (green fluorescent protein) fusions to show that SufI localizes to the septal ring in the dividing cell. We have also determined the structure of SufI by X-ray crystallography to a resolution of 1.9 Å. ...

  1. Control of cell division in Streptococcus pneumoniae by the conserved Ser/Thr protein kinase StkP

    Czech Academy of Sciences Publication Activity Database

    Beilharz, K.; Nováková, Linda; Fadda, D.; Branny, Pavel; Massida, O.; Veening, J.-W.

    2012-01-01

    Roč. 109, č. 15 (2012), s. 905-913. ISSN 0027-8424 R&D Projects: GA AV ČR IAA600200801; GA MŠk LH12055 Institutional research plan: CEZ:AV0Z50200510 Keywords : PENICILLIN-BINDING PROTEINS * SERINE/THREONINE KINASE * MYCOBACTERIUM- TUBERCULOSIS Subject RIV: EE - Microbiology, Virology Impact factor: 9.737, year: 2012

  2. Oriented Cell Division in the C. elegans Embryo Is Coordinated by G-Protein Signaling Dependent on the Adhesion GPCR LAT-1

    OpenAIRE

    Antje Müller; Jana Winkler; Franziska Fiedler; Tania Sastradihardja; Claudia Binder; Ralf Schnabel; Jana Kungel; Sven Rothemund; Christian Hennig; Torsten Schöneberg; Simone Prömel

    2015-01-01

    Orientation of spindles and cell division planes during development of many species ensures that correct cell-cell contacts are established, which is vital for proper tissue formation. This is a tightly regulated process involving a complex interplay of various signals. The molecular mechanisms underlying several of these pathways are still incompletely understood. Here, we identify the signaling cascade of the C. elegans latrophilin homolog LAT-1, an essential player in the coordination of a...

  3. An electrostatic model for biological cell division

    CERN Document Server

    Faraggi, Eshel

    2010-01-01

    Probably the most fundamental processes for biological systems is their ability to create themselves through the use of cell division and cell differentiation. In this work a simple physical model is proposed for biological cell division. The model consists of a positive ionic gradient across the cell membrane, and concentration of charge at the nodes of the spindle and on the chromosomes. A simple calculation, based on Coulomb's Law, shows that under such circumstances a chromosome will tend to break up to its constituent chromatids and that the chromatids will be separated by a distance that is an order of thirty percent of the distance between the spindle nodes. Further repulsion between the nodes will tend to stretch the cell and eventually break the cell membrane between the separated chromatids, leading to cell division. The importance of this work is in continuing the understanding of the electromagnetic basis of cell division and providing it with an analytical model. A central implication of this and...

  4. LocZ Is a New Cell Division Protein Involved in Proper Septum Placement in Streptococcus pneumoniae

    Czech Academy of Sciences Publication Activity Database

    Holečková, Nela; Doubravová, Linda; Massida, O.; Molle, V.; Buriánková, Karolína; Benada, Oldřich; Kofroňová, Olga; Ulrych, Aleš; Branny, Pavel

    2015-01-01

    Roč. 6, č. 1 (2015). ISSN 2150-7511 R&D Projects: GA ČR GAP302/12/0256; GA ČR GAP207/12/1568 Institutional support: RVO:61388971 Keywords : Z-RING FORMATION * ESCHERICHIA-COLI-CELLS * CHROMOSOME SEGREGATION Subject RIV: EE - Microbiology, Virology Impact factor: 6.786, year: 2014

  5. Involvement of YODA and mitogen activated protein kinase 6 in Arabidopsis post-embryogenic root development through auxin up-regulation and cell division plane orientation

    Czech Academy of Sciences Publication Activity Database

    Smékalová, V.; Luptovčiak, I.; Komis, G.; Šamajová, O.; Ovečka, M.; Doskočilová, A.; Takáč, T.; Vadovič, P.; Novák, Ondřej; Pechan, T.; Ziemann, A.; Košútová, P.; Šamaj, J.

    2014-01-01

    Roč. 203, č. 4 (2014), s. 1175-1193. ISSN 0028-646X R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : Arabidopsis * cell division plane * MAP65-1 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.672, year: 2014

  6. The Escherichia coli cell division protein and model Tat substrate SufI (FtsP) localizes to the septal ring and has a multicopper oxidase-like structure.

    Science.gov (United States)

    Tarry, Michael; Arends, S J Ryan; Roversi, Pietro; Piette, Evan; Sargent, Frank; Berks, Ben C; Weiss, David S; Lea, Susan M

    2009-02-20

    The Escherichia coli protein SufI (FtsP) has recently been proposed to be a component of the cell division apparatus. The SufI protein is also in widespread experimental use as a model substrate in studies of the Tat (twin arginine translocation) protein transport system. We have used SufI-GFP (green fluorescent protein) fusions to show that SufI localizes to the septal ring in the dividing cell. We have also determined the structure of SufI by X-ray crystallography to a resolution of 1.9 A. SufI is structurally related to the multicopper oxidase superfamily but lacks metal cofactors. The structure of SufI suggests it serves a scaffolding rather than an enzymatic role in the septal ring and reveals regions of the protein likely to be involved in the protein-protein interactions required to assemble SufI at the septal ring. PMID:19135451

  7. Molecular evolution in bacteria: cell division

    OpenAIRE

    Trevors J.T.

    1998-01-01

    Molecular evolution in bacteria is examined with an emphasis on the self-assembly of cells capable of primitive division and growth during early molecular evolution. Also, the possibility that some type of encapsulation structure preceeded biochemical pathways and the assembly of genetic material is examined. These aspects will be considered from an evolutionary perspective.

  8. Regulation of cell division in higher plants. Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Jacobs, T.W.

    1992-07-01

    Cell division is arguably the most fundamental of all developmental processes. In higher plants, mitotic activity is largely confined to foci of patterned cell divisions called meristems. From these perpetually embryonic tissues arise the plant`s essential organs of light capture, support, protection and reproduction. Once an adequate understanding of plant cell mitotic regulation is attained, unprecedented opportunities will ensue for analyzing and genetically controlling diverse aspects of development, including plant architecture, leaf shape, plant height, and root depth. The mitotic cycle in a variety of model eukaryotic systems in under the control of a regulatory network of striking evolutionary conservation. Homologues of the yeast cdc2 gene, its catalytic product, p34, and the cyclin regulatory subunits of the MPF complex have emerged as ubiquitous mitotic regulators. We have cloned cdc2-like and cyclin genes from pea. As in other eukaryotic model systems, p34 of Pisum sativum is a subunit of a high molecular weight complex which binds the fission yeast p13 protein and displays histone H1 kinase activity in vitro. Our primary objective in this study is to gain baseline information about the regulation of this higher plant cell division control complex in non-dividing, differentiated cells as well as in synchronous and asynchronous mitotic cells. We are investigating cdc2 and cyclin expression at the levels of protein abundance, protein phosphorylation and quaternary associations.

  9. Polarity in Stem Cell Division: Asymmetric Stem Cell Division in Tissue Homeostasis

    OpenAIRE

    Yamashita, Yukiko M; Yuan, Hebao; Cheng, Jun; Hunt, Alan J.

    2010-01-01

    Many adult stem cells divide asymmetrically to balance self-renewal and differentiation, thereby maintaining tissue homeostasis. Asymmetric stem cell divisions depend on asymmetric cell architecture (i.e., cell polarity) within the cell and/or the cellular environment. In particular, as residents of the tissues they sustain, stem cells are inevitably placed in the context of the tissue architecture. Indeed, many stem cells are polarized within their microenvironment, or the stem cell niche, a...

  10. Cell division in Corynebacterineae

    Directory of Open Access Journals (Sweden)

    CatrionaDonovan

    2014-04-01

    Full Text Available Bacterial cells must coordinate a number of events during the cell cycle. Spatio-temporal regulation of bacterial cytokinesis is indispensable for the production of viable, genetically identical offspring. In many rod-shaped bacteria, precise midcell assembly of the division machinery relies on inhibitory systems such as Min and Noc. In rod-shaped Actinobacteria, for example Corynebacterium glutamicum and Mycobacterium tuberculosis, the divisome assembles in the proximity of the midcell region, however more spatial flexibility is observed compared to Escherichia coli and Bacillus subtilis. Actinobacteria represent a group of bacteria that spatially regulate cytokinesis in the absence of recognizable Min and Noc homologs. The key cell division steps in E. coli and B. subtilis have been subject to intensive study and are well understood. In comparison, only a minimal set of positive and negative regulators of cytokinesis are known in Actinobacteria. Nonetheless, the timing of cytokinesis and the placement of the division septum is coordinated with growth as well as initiation of chromosome replication and segregation. We summarize here the current knowledge on cytokinesis and division site selection in the Actinobacteria suborder Corynebacterineae.

  11. Alignment of cell division axes in directed epithelial cell migration

    International Nuclear Information System (INIS)

    Cell division is an essential dynamic event in tissue remodeling during wound healing, cancer and embryogenesis. In collective migration, tensile stresses affect cell shape and polarity, hence, the orientation of the cell division axis is expected to depend on cellular flow patterns. Here, we study the degree of orientation of cell division axes in migrating and resting epithelial cell sheets. We use microstructured channels to create a defined scenario of directed cell invasion and compare this situation to resting but proliferating cell monolayers. In experiments, we find a strong alignment of the axis due to directed flow while resting sheets show very weak global order, but local flow gradients still correlate strongly with the cell division axis. We compare experimental results with a previously published mesoscopic particle based simulation model. Most of the observed effects are reproduced by the simulations. (paper)

  12. Cell division pattern influences gene expression in the shoot apical meristem

    OpenAIRE

    Wyrzykowska, Joanna; Fleming, Andrew

    2003-01-01

    The shoot apical meristem of angiosperms shows a highly conserved cellular architecture in which a change of cell division orientation correlates with early events of leaf initiation. However, the causal role of this altered cellular parameter in leaf formation is debatable. We have used the dynamin-like protein phragmoplastin as a tool to modify the pattern of cell division within the apical meristem. Taking a microinduction approach, we show that local alteration in cell division orientatio...

  13. Kinetics of cell division in epidermal maintenance

    CERN Document Server

    Klein, Allon M; Jones, Philip H; Simons, Benjamin D

    2007-01-01

    The rules governing cell division and differentiation are central to understanding the mechanisms of development, aging and cancer. By utilising inducible genetic labelling, recent studies have shown that the clonal population in transgenic mouse epidermis can be tracked in vivo. Drawing on these results, we explain how clonal fate data may be used to infer the rules of cell division and differentiation underlying the maintenance of adult murine tail-skin. We show that the rates of cell division and differentiation may be evaluated by considering the long-time and short-time clone fate data, and that the data is consistent with cells dividing independently rather than synchronously. Motivated by these findings, we consider a mechanism for cancer onset based closely on the model for normal adult skin. By analysing the expected changes to clonal fate in cancer emerging from a simple two-stage mutation, we propose that clonal fate data may provide a novel method for studying the earliest stages of the disease.

  14. Formation of a cylindrical bridge in cell division

    Science.gov (United States)

    Citron, Daniel; Schmidt, Laura E.; Reichl, Elizabeth; Ren, Yixin; Robinson, Douglas; Zhang, Wendy W.

    2007-11-01

    In nature, the shape transition associated with the division of a mother cell into two daughter cells proceeds via a variety of routes. In the cylinder-thinning route, which has been observed in Dictyostelium and most animal cells, the mother cell first forms a broad bridge-like region, also known as a furrow, between two daughter cells. The furrow then rapidly evolves into a cylindrical bridge, which thins and eventually severs the mother cell into two. The fundamental mechanism underlying this division route is not understood. Recent experiments on Dictyostelium found that, while the cylinder-thinning route persists even when key actin cross-linking proteins are missing, it is disrupted by the removal of force-generating myosin-II proteins. Other measurements revealed that mutant cells lacking myosin-II have a much more uniform tension over the cell surface than wild-type cells. This suggests that tension variation may be important. Here we use a fluid model, previously shown to reproduce the thinning dynamics [Zhang & Robinson, PNAS 102, 7186 (2005)], to test this idea. Consistent with the experiments, the model shows that the cylinder formation process occurs regardless of the exact viscoelastic properties of the cell. In contrast to the experiments, a tension variation in the model hinders, rather then expedites, the cylinder formation.

  15. Cell adhesion in regulation of asymmetric stem cell division

    OpenAIRE

    Yamashita, Yukiko M

    2010-01-01

    Adult stem cells inevitably communicate with their cellular neighbors within the tissues they sustain. Indeed, such communication, particularly with components of the stem cell niche, is essential for many aspects of stem cell behavior, including the maintenance of stem cell identity and asymmetric cell division. Cell adhesion mediates this communication by placing stem cells in close proximity to the signaling source and by providing a polarity cue that orients stem cells. Here, I review the...

  16. Cell Division, Differentiation and Dynamic Clustering

    CERN Document Server

    Kaneko, K; Kaneko, Kunihiko; Yomo, Tetsuya

    1993-01-01

    A novel mechanism for cell differentiation is proposed, based on the dynamic clustering in a globally coupled chaotic system. A simple model with metabolic reaction, active transport of chemicals from media, and cell division is found to show three successive stages with the growth of the number of cells; coherent growth, dynamic clustering, and fixed cell differentiation. At the last stage, disparity in activities, germ line segregation, somatic cell differentiation, and homeochaotic stability against external perturbation are found. Our results, in consistency with the experiments of the preceding paper, imply that cell differentiation can occur without a spatial pattern. From dynamical systems viewpoint, the new concept of ``open chaos" is proposed, as a novel and general scenario for systems with growing numbers of elements, also seen in economics and sociology.A

  17. Monitoring structural changes in intrinsically disordered proteins using QCM-D: application to the bacterial cell division protein ZipA.

    Science.gov (United States)

    Mateos-Gil, Pablo; Tsortos, Achilleas; Vélez, Marisela; Gizeli, Electra

    2016-05-01

    The sensitivity of QCM-D to molecular hydrodynamic properties is applied in this work to study conformational changes of the intrinsically disordered protein ZipA. Acoustic measurements can clearly follow ZipA's unstructured domain expansion and contraction with salt content and be correlated with changes in the hydrodynamic radius of 1.8 nm or less. PMID:27109863

  18. Chromosome replication, cell growth, division and shape: a personal perspective.

    Science.gov (United States)

    Zaritsky, Arieh; Woldringh, Conrad L

    2015-01-01

    The origins of Molecular Biology and Bacterial Physiology are reviewed, from our personal standpoints, emphasizing the coupling between bacterial growth, chromosome replication and cell division, dimensions and shape. Current knowledge is discussed with historical perspective, summarizing past and present achievements and enlightening ideas for future studies. An interactive simulation program of the bacterial cell division cycle (BCD), described as "The Central Dogma in Bacteriology," is briefly represented. The coupled process of transcription/translation of genes encoding membrane proteins and insertion into the membrane (so-called transertion) is invoked as the functional relationship between the only two unique macromolecules in the cell, DNA and peptidoglycan embodying the nucleoid and the sacculus respectively. We envision that the total amount of DNA associated with the replication terminus, so called "nucleoid complexity," is directly related to cell size and shape through the transertion process. Accordingly, the primary signal for cell division transmitted by DNA dynamics (replication, transcription and segregation) to the peptidoglycan biosynthetic machinery is of a physico-chemical nature, e.g., stress in the plasma membrane, relieving nucleoid occlusion in the cell's center hence enabling the divisome to assemble and function between segregated daughter nucleoids. PMID:26284044

  19. ARC3 is a stromal Z-ring accessory protein essential for plastid division

    Science.gov (United States)

    Maple, Jodi; Vojta, Lea; Soll, Jurgen; Møller, Simon G

    2007-01-01

    In plants, chloroplast division is an integral part of development, and these vital organelles arise by binary fission from pre-existing cytosolic plastids. Chloroplasts arose by endosymbiosis and although they have retained elements of the bacterial cell division machinery to execute plastid division, they have evolved to require two functionally distinct forms of the FtsZ protein and have lost elements of the Min machinery required for Z-ring placement. Here, we analyse the plastid division component accumulation and replication of chloroplasts 3 (ARC3) and show that ARC3 forms part of the stromal plastid division machinery. ARC3 interacts specifically with AtFtsZ1, acting as a Z-ring accessory protein and defining a unique function for this family of FtsZ proteins. ARC3 is involved in division site placement, suggesting that it might functionally replace MinC, representing an important advance in our understanding of the mechanism of chloroplast division and the evolution of the chloroplast division machinery. PMID:17304239

  20. Onset of cell division in maize germination: action of auxins

    International Nuclear Information System (INIS)

    Seed germination implies metabolic reactivation, synthesis of macromolecules and onset of cell division. During maize germination, meristematic tissues of embryos re-initiate cell division asynchronically. Since auxins are known to stimulate cell division, they asked how auxins might regulate cell cycle re-initiation. Embryonic tissues were incubated with and without auxins. A pulse of either 3H-thymidine or 32P-ortophosphate was given to the tissues. Mitotic indexes were determined and % of labeled mitotic cells recorded. Results indicated that meristematic cells re-initiate cell division either from G1 or G2 phases. Auxin stimulated differentially the cell division process of these cells. 32P incorporation into cytoplasmic or nucleic histones was measured. Auxins stimulated this incorporation. Active turnover of histone phosphorylation occurred simultaneously to the cell division process. It is suggested that auxins might regulate the cell cycle by phosphorylation-dephosphorylation of histones

  1. Cell Shape and Cell Division in Fission Yeast Minireview

    OpenAIRE

    Piel, Matthieu; Tran, Phong T.

    2009-01-01

    The fission yeast Schizosaccharomyces pombe has served as an important model organism for investigating cellular morphogenesis. This unicellular rod-shaped fission yeast grows by tip extension and divides by medial fission. In particular, microtubules appear to define sites of polarized cell growth by delivering cell polarity factors to the cell tips. Microtubules also position the cell nucleus at the cell middle, marking sites of cell division. Here, we review the microtubule-dependent mecha...

  2. A DNA damage-induced, SOS-independent checkpoint regulates cell division in Caulobacter crescentus.

    Directory of Open Access Journals (Sweden)

    Joshua W Modell

    2014-10-01

    Full Text Available Cells must coordinate DNA replication with cell division, especially during episodes of DNA damage. The paradigm for cell division control following DNA damage in bacteria involves the SOS response where cleavage of the transcriptional repressor LexA induces a division inhibitor. However, in Caulobacter crescentus, cells lacking the primary SOS-regulated inhibitor, sidA, can often still delay division post-damage. Here we identify didA, a second cell division inhibitor that is induced by DNA damage, but in an SOS-independent manner. Together, DidA and SidA inhibit division, such that cells lacking both inhibitors divide prematurely following DNA damage, with lethal consequences. We show that DidA does not disrupt assembly of the division machinery and instead binds the essential division protein FtsN to block cytokinesis. Intriguingly, mutations in FtsW and FtsI, which drive the synthesis of septal cell wall material, can suppress the activity of both SidA and DidA, likely by causing the FtsW/I/N complex to hyperactively initiate cell division. Finally, we identify a transcription factor, DriD, that drives the SOS-independent transcription of didA following DNA damage.

  3. A DNA damage-induced, SOS-independent checkpoint regulates cell division in Caulobacter crescentus.

    Science.gov (United States)

    Modell, Joshua W; Kambara, Tracy K; Perchuk, Barrett S; Laub, Michael T

    2014-10-01

    Cells must coordinate DNA replication with cell division, especially during episodes of DNA damage. The paradigm for cell division control following DNA damage in bacteria involves the SOS response where cleavage of the transcriptional repressor LexA induces a division inhibitor. However, in Caulobacter crescentus, cells lacking the primary SOS-regulated inhibitor, sidA, can often still delay division post-damage. Here we identify didA, a second cell division inhibitor that is induced by DNA damage, but in an SOS-independent manner. Together, DidA and SidA inhibit division, such that cells lacking both inhibitors divide prematurely following DNA damage, with lethal consequences. We show that DidA does not disrupt assembly of the division machinery and instead binds the essential division protein FtsN to block cytokinesis. Intriguingly, mutations in FtsW and FtsI, which drive the synthesis of septal cell wall material, can suppress the activity of both SidA and DidA, likely by causing the FtsW/I/N complex to hyperactively initiate cell division. Finally, we identify a transcription factor, DriD, that drives the SOS-independent transcription of didA following DNA damage. PMID:25350732

  4. Activation of cell divisions in legume nodulation

    DEFF Research Database (Denmark)

    Nadzieja, Marcin

    2016-01-01

    organogenesis. Coordination of these two interdependent processes results in formation of nodules - bacterial accommodating structures where fixation of atmospheric nitrogen takes place. Plant hormones such as auxin and cytokinin play important roles in nodulation. In some legumes the infection process...... was shown to require auxin signalling. Cytokinin, in contrast, exert a negative regulation of bacterial entry into the root. During organogenesis, auxin and cytokinin maxima are known to accompany nodule primordia development and together regulate progression through the cell cycle. Moreover, application...... of auxin transport inhibitors or cytokinin alone was shown to induce cortical cell divisions in the absence of rhizobia in certain legume species. While the roles of auxin and cytokinin in nodulation have been studied extensively, the precise timing, location and means of molecular crosstalk between...

  5. Mammalian aPKC/Par polarity complex mediated regulation of epithelial division orientation and cell fate

    International Nuclear Information System (INIS)

    Oriented cell division is a key regulator of tissue architecture and crucial for morphogenesis and homeostasis. Balanced regulation of proliferation and differentiation is an essential property of tissues not only to drive morphogenesis but also to maintain and restore homeostasis. In many tissues orientation of cell division is coupled to the regulation of differentiation producing daughters with similar (symmetric cell division, SCD) or differential fate (asymmetric cell division, ACD). This allows the organism to generate cell lineage diversity from a small pool of stem and progenitor cells. Division orientation and/or the ratio of ACD/SCD need to be tightly controlled. Loss of orientation or an altered ratio can promote overgrowth, alter tissue architecture and induce aberrant differentiation, and have been linked to morphogenetic diseases, cancer and aging. A key requirement for oriented division is the presence of a polarity axis, which can be established through cell intrinsic and/or extrinsic signals. Polarity proteins translate such internal and external cues to drive polarization. In this review we will focus on the role of the polarity complex aPKC/Par3/Par6 in the regulation of division orientation and cell fate in different mammalian epithelia. We will compare the conserved function of this complex in mitotic spindle orientation and distribution of cell fate determinants and highlight common and differential mechanisms in which this complex is used by tissues to adapt division orientation and cell fate to the specific properties of the epithelium

  6. Mammalian aPKC/Par polarity complex mediated regulation of epithelial division orientation and cell fate

    Energy Technology Data Exchange (ETDEWEB)

    Vorhagen, Susanne; Niessen, Carien M., E-mail: carien.niessen@uni-koeln.de

    2014-11-01

    Oriented cell division is a key regulator of tissue architecture and crucial for morphogenesis and homeostasis. Balanced regulation of proliferation and differentiation is an essential property of tissues not only to drive morphogenesis but also to maintain and restore homeostasis. In many tissues orientation of cell division is coupled to the regulation of differentiation producing daughters with similar (symmetric cell division, SCD) or differential fate (asymmetric cell division, ACD). This allows the organism to generate cell lineage diversity from a small pool of stem and progenitor cells. Division orientation and/or the ratio of ACD/SCD need to be tightly controlled. Loss of orientation or an altered ratio can promote overgrowth, alter tissue architecture and induce aberrant differentiation, and have been linked to morphogenetic diseases, cancer and aging. A key requirement for oriented division is the presence of a polarity axis, which can be established through cell intrinsic and/or extrinsic signals. Polarity proteins translate such internal and external cues to drive polarization. In this review we will focus on the role of the polarity complex aPKC/Par3/Par6 in the regulation of division orientation and cell fate in different mammalian epithelia. We will compare the conserved function of this complex in mitotic spindle orientation and distribution of cell fate determinants and highlight common and differential mechanisms in which this complex is used by tissues to adapt division orientation and cell fate to the specific properties of the epithelium.

  7. Chromokinesin: Kinesin superfamily regulating cell division through chromosome and spindle.

    Science.gov (United States)

    Zhong, Ai; Tan, Fu-Qing; Yang, Wan-Xi

    2016-09-01

    Material transportation is essential for appropriate cellular morphology and functions, especially during cell division. As a motor protein moving along microtubules, kinesin has several intracellular functions. Many kinesins play important roles in chromosome condensation and separation and spindle organization during the cell cycle. Some of them even can directly bind to chromosomes, as a result, these proteins are called chromokinesins. Kinesin-4 and kinesin-10 family are two major families of chromokinesin and many members can regulate some processes, both in mitosis and meiosis. Their functions have been widely studied. Here, we summarize current knowledge about known chromokinesins and introduce their intracellular features in accordance with different families. Furthermore, we have also introduced some new-found but unconfirmed kinesins which may have a relationship with chromosomes or the cell cycle. PMID:27196062

  8. Plastid chaperonin proteins Cpn60α and Cpn60β are required for plastid division in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Osteryoung Katherine W

    2009-04-01

    Full Text Available Abstract Background Plastids arose from a free-living cyanobacterial endosymbiont and multiply by binary division as do cyanobacteria. Plastid division involves nucleus-encoded homologs of cyanobacterial division proteins such as FtsZ, MinD, MinE, and ARC6. However, homologs of many other cyanobacterial division genes are missing in plant genomes and proteins of host eukaryotic origin, such as a dynamin-related protein, PDV1 and PDV2 are involved in the division process. Recent identification of plastid division proteins has started to elucidate the similarities and differences between plastid division and cyanobacterial cell division. To further identify new proteins that are required for plastid division, we characterized previously and newly isolated plastid division mutants of Arabidopsis thaliana. Results Leaf cells of two mutants, br04 and arc2, contain fewer, larger chloroplasts than those of wild type. We found that ARC2 and BR04 are identical to nuclear genes encoding the plastid chaperonin 60α (ptCpn60α and chaperonin 60β (ptCpn60β proteins, respectively. In both mutants, plastid division FtsZ ring formation was partially perturbed though the level of FtsZ2-1 protein in plastids of ptcpn60β mutants was similar to that in wild type. Phylogenetic analyses showed that both ptCpn60 proteins are derived from ancestral cyanobacterial proteins. The A. thaliana genome encodes two members of ptCpn60α family and four members of ptCpn60β family respectively. We found that a null mutation in ptCpn60α abolished greening of plastids and resulted in an albino phenotype while a weaker mutation impairs plastid division and reduced chlorophyll levels. The functions of at least two ptCpn60β proteins are redundant and the appearance of chloroplast division defects is dependent on the number of mutant alleles. Conclusion Our results suggest that both ptCpn60α and ptCpn60β are required for the formation of a normal plastid division apparatus, as

  9. Microtubule networks for plant cell division.

    Science.gov (United States)

    de Keijzer, Jeroen; Mulder, Bela M; Janson, Marcel E

    2014-09-01

    During cytokinesis the cytoplasm of a cell is divided to form two daughter cells. In animal cells, the existing plasma membrane is first constricted and then abscised to generate two individual plasma membranes. Plant cells on the other hand divide by forming an interior dividing wall, the so-called cell plate, which is constructed by localized deposition of membrane and cell wall material. Construction starts in the centre of the cell at the locus of the mitotic spindle and continues radially towards the existing plasma membrane. Finally the membrane of the cell plate and plasma membrane fuse to form two individual plasma membranes. Two microtubule-based cytoskeletal networks, the phragmoplast and the pre-prophase band (PPB), jointly control cytokinesis in plants. The bipolar microtubule array of the phragmoplast regulates cell plate deposition towards a cortical position that is templated by the ring-shaped microtubule array of the PPB. In contrast to most animal cells, plants do not use centrosomes as foci of microtubule growth initiation. Instead, plant microtubule networks are striking examples of self-organizing systems that emerge from physically constrained interactions of dispersed microtubules. Here we will discuss how microtubule-based activities including growth, shrinkage, severing, sliding, nucleation and bundling interrelate to jointly generate the required ordered structures. Evidence mounts that adapter proteins sense the local geometry of microtubules to locally modulate the activity of proteins involved in microtubule growth regulation and severing. Many of the proteins and mechanisms involved have roles in other microtubule assemblies as well, bestowing broader relevance to insights gained from plants. PMID:25136380

  10. Concise Review: Asymmetric Cell Divisions in Stem Cell Biology

    Directory of Open Access Journals (Sweden)

    Florian Murke

    2015-11-01

    Full Text Available Somatic stem cells are rare cells with unique properties residing in many organs and tissues. They are undifferentiated cells responsible for tissue regeneration and homeostasis, and contain both the capacity to self-renew in order to maintain their stem cell potential and to differentiate towards tissue-specific, specialized cells. However, the knowledge about the mechanisms controlling somatic stem cell fate decisions remains sparse. One mechanism which has been described to control daughter cell fates in selected somatic stem cell systems is the process of asymmetric cell division (ACD. ACD is a tightly regulated and evolutionary conserved process allowing a single stem or progenitor cell to produce two differently specified daughter cells. In this concise review, we will summarize and discuss current concepts about the process of ACD as well as different ACD modes. Finally, we will recapitulate the current knowledge and our recent findings about ACD in human hematopoiesis.

  11. Huntingtin Regulates Mammary Stem Cell Division and Differentiation

    Directory of Open Access Journals (Sweden)

    Salah Elias

    2014-04-01

    Full Text Available Little is known about the mechanisms of mitotic spindle orientation during mammary gland morphogenesis. Here, we report the presence of huntingtin, the protein mutated in Huntington’s disease, in mouse mammary basal and luminal cells throughout mammogenesis. Keratin 5-driven depletion of huntingtin results in a decreased pool and specification of basal and luminal progenitors, and altered mammary morphogenesis. Analysis of mitosis in huntingtin-depleted basal progenitors reveals mitotic spindle misorientation. In mammary cell culture, huntingtin regulates spindle orientation in a dynein-dependent manner. Huntingtin is targeted to spindle poles through its interaction with dynein and promotes the accumulation of NUMA and LGN. Huntingtin is also essential for the cortical localization of dynein, dynactin, NUMA, and LGN by regulating their kinesin 1-dependent trafficking along astral microtubules. We thus suggest that huntingtin is a component of the pathway regulating the orientation of mammary stem cell division, with potential implications for their self-renewal and differentiation properties.

  12. Huntingtin regulates mammary stem cell division and differentiation.

    Science.gov (United States)

    Elias, Salah; Thion, Morgane S; Yu, Hua; Sousa, Cristovao Marques; Lasgi, Charlène; Morin, Xavier; Humbert, Sandrine

    2014-04-01

    Little is known about the mechanisms of mitotic spindle orientation during mammary gland morphogenesis. Here, we report the presence of huntingtin, the protein mutated in Huntington's disease, in mouse mammary basal and luminal cells throughout mammogenesis. Keratin 5-driven depletion of huntingtin results in a decreased pool and specification of basal and luminal progenitors, and altered mammary morphogenesis. Analysis of mitosis in huntingtin-depleted basal progenitors reveals mitotic spindle misorientation. In mammary cell culture, huntingtin regulates spindle orientation in a dynein-dependent manner. Huntingtin is targeted to spindle poles through its interaction with dynein and promotes the accumulation of NUMA and LGN. Huntingtin is also essential for the cortical localization of dynein, dynactin, NUMA, and LGN by regulating their kinesin 1-dependent trafficking along astral microtubules. We thus suggest that huntingtin is a component of the pathway regulating the orientation of mammary stem cell division, with potential implications for their self-renewal and differentiation properties. PMID:24749073

  13. Study of genes induced by ionizing radiations at Arabidopsis thaliana: identification and molecular characterization of the ATGR1 gene, a new gene encoding a protein involved in plant cell division

    International Nuclear Information System (INIS)

    DNA damage, that can be experimentally introduced by ionizing radiation (IR), induces complex signal transduction pathways leading to cell recovery or, alternatively to programmed cell death if damages are too severe. To identify the inducible components of the response to genotoxic stress in plants, we have screened by Differential Display for mRNAs that rapidly and strongly accumulate after IR treatment in A. thaliana cells. We have characterized ATGR1, a new single copy Arabidopsis gene encoding a PEST-box protein of unknown function. In unstressed plant organs the ATGR1 mRNA is hardly detectable, whereas the protein is present in extracts prepared from roots, shoot meristems and inflorescences, that all contain large amounts of actively dividing cells. This pattern is confirmed by immuno localisation on tissue sections that shows constitutive ATGR1 protein expression covering the root elongation zone, the shoot meristem, leaf primordial and the ovules of developing flowers. Histochemical analysis of transgenic plants expressing the GUS reporter gene under the control of the ATGR1 promoter, demonstrate that the developmental and tissue-specific profile of ATGR1 protein expression is conferred by the gene promoter. The massive, transient and dose-dependent accumulation of ATGR1 transcripts after IR treatment observed in all plant organs does not lead to significant changes in ATGR1 protein pattern. Stable ATGR1 protein overexpression, as exemplified by transgenic A. thaliana plants that contain a 35S promoter-ATGR1 gene fusion, does not induce notable changes of the overall ATGR1 protein level, but leads to male and female sterility. The cause of sterility is a lack of correct chromosome assembly and distribution at the stage metaphase II of meiosis. Taken together our results show that i) ATGR1 gene expression is associated to cell division during plant development ii) the ATGR1 protein level is regulated at the transcriptional and post-transcriptional level iii

  14. Mechanisms of daughter cell-size control during cell division.

    Science.gov (United States)

    Kiyomitsu, Tomomi

    2015-05-01

    Daughter cell size is tightly regulated during cell division. In animal cells, the position of the anaphase spindle specifies the cell cleavage site to dictate the relative size of the daughter cells. Although spindle orientation is regulated by dynein-dependent cortical pulling forces exerted on astral microtubules in many cell types, it was unclear how these forces are precisely regulated to center or displace the spindle. Recently, intrinsic signals derived from chromosomes or spindle poles have been demonstrated to regulate dynein-dependent pulling forces in symmetrically dividing cells. Unexpectedly, myosin-dependent contractile forces have also been shown to control spindle position by altering the cellular boundaries during anaphase. In this review, I discuss how dynein- and myosin-dependent forces are coordinately regulated to control daughter cell size. PMID:25548067

  15. Smurfs have "fused" into the asymmetric division of stem cells

    Institute of Scientific and Technical Information of China (English)

    Steven Y. Cheng; Ying E. Zhang

    2011-01-01

    @@ The asymmetric cell division is the way in which a stem cell divides into one daughter stem cell and one differentiated daughter cell.This process is one of the key principles of developmental biology that ensures the perpetual supply of stem cells while allowing a particular cell lineage to be populated.During Drosophila oogenesis, the fate of the daughter stem cell produced from the asymmetric division of germline stem cells (GSCs) is specified by Decapentaplegic (Dpp), but the other daughter cell has almost equal access to the Dpp signal.

  16. Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

    Directory of Open Access Journals (Sweden)

    Piotr Koprowski

    Full Text Available Bacterial mechano-sensitive (MS channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

  17. Streptomyces: A Screening Tool for Bacterial Cell Division Inhibitors

    Science.gov (United States)

    Jani, Charul; Tocheva, Elitza I.; McAuley, Scott; Craney, Arryn; Jensen, Grant J.; Nodwell, Justin

    2016-01-01

    Cell division is essential for spore formation but not for viability in the filamentous streptomycetes bacteria. Failure to complete cell division instead blocks spore formation, a phenotype that can be visualized by the absence of gray (in Streptomyces coelicolor) and green (in Streptomyces venezuelae) spore-associated pigmentation. Despite the lack of essentiality, the streptomycetes divisome is similar to that of other prokaryotes. Therefore, the chemical inhibitors of sporulation in model streptomycetes may interfere with the cell division in rod-shaped bacteria as well. To test this, we investigated 196 compounds that inhibit sporulation in S. coelicolor. We show that 19 of these compounds cause filamentous growth in Bacillus subtilis, consistent with impaired cell division. One of the compounds is a DNA-damaging agent and inhibits cell division by activating the SOS response. The remaining 18 act independently of known stress responses and may therefore act on the divisome or on divisome positioning and stability. Three of the compounds (Fil-1, Fil-2, and Fil-3) confer distinct cell division defects on B. subtilis. They also block B. subtilis sporulation, which is mechanistically unrelated to the sporulation pathway of streptomycetes but is also dependent on the divisome. We discuss ways in which these differing phenotypes can be used in screens for cell division inhibitors. PMID:25256667

  18. Ploidy-Dependent Unreductional Meiotic Cell Division in Polyploid Wheat

    Science.gov (United States)

    Meiosis includes one round of DNA replication and two successive nuclear divisions, i.e. meiosis I (reductional) and meiosis II (equational). This specialized cell division reduces chromosomes in half and generates haploid gametes in sexual reproduction of eukaryotes. It ensures faithful transmiss...

  19. Phosphorus deficiency inhibits cell division but not growth in the dinoflagellate Amphidinium carterae

    Directory of Open Access Journals (Sweden)

    Meizhen eLi

    2016-06-01

    Full Text Available Phosphorus (P is an essential nutrient element for the growth of phytoplankton. How P deficiency affects population growth and the cell division cycle in dinoflagellates has only been studied in some species, and how it affects photosynthesis and cell growth remains poorly understood. In the present study, we investigated the impact of P deficiency on the cell division cycle, the abundance of the carbon-fixing enzyme Rubisco, and other cellular characteristics in the Gymnodiniales peridinin-plastid species Amphidinium carterae. We found that under P-replete condition, the cell cycle actively progressed in the culture in a 24-hour diel cycle with daily growth rates markedly higher than the P-deficient cultures, in which cells were arrested in the G1 phase and cell size significantly enlarged. The results suggest that, as in previously studied dinoflagellates, P deficiency likely disenables A. carterae to complete DNA duplication or check-point protein phosphorylation. We further found that under P-deficient condition, overall photosystem II quantum efficiency (Fv/Fm ratio and Rubisco abundance decreased but not significantly, while cellular contents of carbon, nitrogen, and proteins increased significantly. These observations indicated that under P-deficiency, this dinoflagellate was able to continue photosynthesis and carbon fixation, such that proteins and photosynthetically fixed carbon could accumulate resulting in continued cell growth in the absence of division. This is likely an adaptive strategy thereby P-limited cells can be ready to resume the cell division cycle upon resupply of phosphorus.

  20. Stochastic modeling of cell growth with symmetric or asymmetric division.

    Science.gov (United States)

    Marantan, Andrew; Amir, Ariel

    2016-07-01

    We consider a class of biologically motivated stochastic processes in which a unicellular organism divides its resources (volume or damaged proteins, in particular) symmetrically or asymmetrically between its progeny. Assuming the final amount of the resource is controlled by a growth policy and subject to additive and multiplicative noise, we derive the recursive integral equation describing the evolution of the resource distribution over subsequent generations and use it to study the properties of stable resource distributions. We find conditions under which a unique stable resource distribution exists and calculate its moments for the class of affine linear growth policies. Moreover, we apply an asymptotic analysis to elucidate the conditions under which the stable distribution (when it exists) has a power-law tail. Finally, we use the results of this asymptotic analysis along with the moment equations to draw a stability phase diagram for the system that reveals the counterintuitive result that asymmetry serves to increase stability while at the same time widening the stable distribution. We also briefly discuss how cells can divide damaged proteins asymmetrically between their progeny as a form of damage control. In the appendixes, motivated by the asymmetric division of cell volume in Saccharomyces cerevisiae, we extend our results to the case wherein mother and daughter cells follow different growth policies. PMID:27575162

  1. Stochastic modeling of cell growth with symmetric or asymmetric division

    Science.gov (United States)

    Marantan, Andrew; Amir, Ariel

    2016-07-01

    We consider a class of biologically motivated stochastic processes in which a unicellular organism divides its resources (volume or damaged proteins, in particular) symmetrically or asymmetrically between its progeny. Assuming the final amount of the resource is controlled by a growth policy and subject to additive and multiplicative noise, we derive the recursive integral equation describing the evolution of the resource distribution over subsequent generations and use it to study the properties of stable resource distributions. We find conditions under which a unique stable resource distribution exists and calculate its moments for the class of affine linear growth policies. Moreover, we apply an asymptotic analysis to elucidate the conditions under which the stable distribution (when it exists) has a power-law tail. Finally, we use the results of this asymptotic analysis along with the moment equations to draw a stability phase diagram for the system that reveals the counterintuitive result that asymmetry serves to increase stability while at the same time widening the stable distribution. We also briefly discuss how cells can divide damaged proteins asymmetrically between their progeny as a form of damage control. In the appendixes, motivated by the asymmetric division of cell volume in Saccharomyces cerevisiae, we extend our results to the case wherein mother and daughter cells follow different growth policies.

  2. Regulation of cell division in higher plants. Final technical report

    Energy Technology Data Exchange (ETDEWEB)

    Jacobs, Thomas W.

    2000-02-29

    Research in the latter part of the grant period was divided into two parts: (1) expansion of the macromolecular tool kit for studying plant cell division; (2) experiments in which the roles played by plant cell cycle regulators were to be cast in the light of the emerging yeast and animal cell paradigm for molecular control of the mitotic cycle. The first objectives were accomplished to a very satisfactory degree. With regard to the second part of the project, we were driven to change our objectives for two reasons. First, the families of cell cycle control genes that we cloned encoded such closely related members that the prospects for success at raising distinguishing antisera against each were sufficiently dubious as to be impractical. Epitope tagging is not feasible in Pisum sativum, our experimental system, as this species is not realistically transformable. Therefore, differentiating the roles of diverse cyclins and cyclin-dependent kinases was problematic. Secondly, our procedure for generating mitotically synchronized pea root meristems for biochemical studies was far too labor intensive for the proposed experiments. We therefore shifted our objectives to identifying connections between the conserved proteins of the cell cycle engine and factors that interface it with plant physiology and development. In this, we have obtained some very exciting results.

  3. Dynamics of Tetrahymena macronuclear lamina during cell division

    Institute of Scientific and Technical Information of China (English)

    CHENBIN; ZHONGHEZHAI

    1994-01-01

    During mitosis,the nuclear lamina in higher eukaryotic cells undergoes a distinctly morphological change.It breaks down into lamin polymers or monomers at prophase.At telophase,the lamins reassemble around the condensed chromatin to form the layer of lamina.Using antiserum to mammalian lamins,we studied the dynamics of lamina during cell division in the macronuleus of Tetrahymena shanghaiensis,which divided in the way of amitosis.In contrast to those in higher animal cells,the typical perinuclear lamin distribution in the macronucleus persisted throughout the whole cell cycle.It was further found that in some synchronized cells,the lamin distribution bisplayed an unusual pattern consisting of a series of spots within the macronucleus.Using South-western hybridization,we found that the purified 66 KD lamin in Tetrahymena showed specific affinity with the telomere DNA sequence in the same species.Therefore,we propose that pattern of immunofluorescence may be due to the interaction of lamin protein with the nucleoli and the condensed chromatins in the macronucleus.

  4. The Saccharomyces cerevisiae centromere protein Slk19p is required for two successive divisions during meiosis.

    OpenAIRE

    Zeng, X.; Saunders, W S

    2000-01-01

    Meiotic cell division includes two separate and distinct types of chromosome segregation. In the first segregational event the sister chromatids remain attached at the centromere; in the second the chromatids are separated. The factors that control the order of chromosome segregation during meiosis have not yet been identified but are thought to be confined to the centromere region. We showed that the centromere protein Slk19p is required for the proper execution of meiosis in Saccharomyces c...

  5. Stationary Size Distributions of Growing Cells with Binary and Multiple Cell Division

    Science.gov (United States)

    Rading, M. M.; Engel, T. A.; Lipowsky, R.; Valleriani, A.

    2011-10-01

    Populations of unicellular organisms that grow under constant environmental conditions are considered theoretically. The size distribution of these cells is calculated analytically, both for the usual process of binary division, in which one mother cell produces always two daughter cells, and for the more complex process of multiple division, in which one mother cell can produce 2 n daughter cells with n=1,2,3,… . The latter mode of division is inspired by the unicellular algae Chlamydomonas reinhardtii. The uniform response of the whole population to different environmental conditions is encoded in the individual rates of growth and division of the cells. The analytical treatment of the problem is based on size-dependent rules for cell growth and stochastic transition processes for cell division. The comparison between binary and multiple division shows that these different division processes lead to qualitatively different results for the size distribution and the population growth rates.

  6. Biased DNA Segregation during Stem Cell Division

    OpenAIRE

    Anversa, Piero; Leri, Annarosa; Kajstura, Jan

    2012-01-01

    Adult skeletal muscle stem cells are a heterogeneous cell population characterized by a small subset of undifferentiated cells that express at high level the paired/homeodomain gene Pax7. This category of satellite cells divides predominantly by asymmetric chromatid segregation generating a daughter cell that carries the mother DNA and retains stem cell property, and a daughter cell that inherits the newly-synthesized DNA and acquires the myocyte lineage.1

  7. Periplasmic Acid Stress Increases Cell Division Asymmetry (Polar Aging of Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Michelle W Clark

    Full Text Available Under certain kinds of cytoplasmic stress, Escherichia coli selectively reproduce by distributing the newer cytoplasmic components to new-pole cells while sequestering older, damaged components in cells inheriting the old pole. This phenomenon is termed polar aging or cell division asymmetry. It is unknown whether cell division asymmetry can arise from a periplasmic stress, such as the stress of extracellular acid, which is mediated by the periplasm. We tested the effect of periplasmic acid stress on growth and division of adherent single cells. We tracked individual cell lineages over five or more generations, using fluorescence microscopy with ratiometric pHluorin to measure cytoplasmic pH. Adherent colonies were perfused continually with LBK medium buffered at pH 6.00 or at pH 7.50; the external pH determines periplasmic pH. In each experiment, cell lineages were mapped to correlate division time, pole age and cell generation number. In colonies perfused at pH 6.0, the cells inheriting the oldest pole divided significantly more slowly than the cells inheriting the newest pole. In colonies perfused at pH 7.50 (near or above cytoplasmic pH, no significant cell division asymmetry was observed. Under both conditions (periplasmic pH 6.0 or pH 7.5 the cells maintained cytoplasmic pH values at 7.2-7.3. No evidence of cytoplasmic protein aggregation was seen. Thus, periplasmic acid stress leads to cell division asymmetry with minimal cytoplasmic stress.

  8. Microtubule networks for plant cell division

    NARCIS (Netherlands)

    Keijzer, de Jeroen; Mulder, B.M.; Janson, M.E.

    2014-01-01

    During cytokinesis the cytoplasm of a cell is divided to form two daughter cells. In animal cells, the existing plasma membrane is first constricted and then abscised to generate two individual plasma membranes. Plant cells on the other hand divide by forming an interior dividing wall, the so-called

  9. Size-independent symmetric division in extraordinarily long cells

    NARCIS (Netherlands)

    N. Pende; N. Leisch; H.R. Gruber-Vodicka; N.R. Heindl; J. Ott; T. den Blaauwen; S. Bulgheresi

    2014-01-01

    Two long-standing paradigms in biology are that cells belonging to the same population exhibit little deviation from their average size and that symmetric cell division is size limited. Here, ultrastructural, morphometric and immunocytochemical analyses reveal that two Gammaproteobacteria attached t

  10. Relevant parameters in models of cell division control

    CERN Document Server

    Grilli, Jacopo; Kennard, Andrew S; Lagomarsino, Marco Cosentino

    2016-01-01

    A recent burst of dynamic single-cell growth-division data makes it possible to characterize the stochastic dynamics of cell division control in bacteria. Different modeling frameworks were used to infer specific mechanisms from such data, but the links between frameworks are poorly explored, with relevant consequences for how well any particular mechanism can be supported by the data. Here, we describe a simple and generic framework in which two common formalisms can be used interchangeably: (i) a continuous-time division process described by a hazard function and (ii) a discrete-time equation describing cell size across generations (where the unit of time is a cell cycle). In our framework, this second process is a discrete-time Langevin equation with a simple physical analogue. By perturbative expansion around the mean initial size (or inter-division time), we show explicitly how this framework describes a wide range of division control mechanisms, including combinations of time and size control, as well a...

  11. Primitive human hematopoietic cells give rise to differentially specified daughter cells upon their initial cell division.

    NARCIS (Netherlands)

    Giebel, B.; Zhang, T.; Beckmann, J.; Spanholtz, J.; Wernet, P.; Ho, A.; Punzel, M.

    2006-01-01

    It is often predicted that stem cells divide asymmetrically, creating a daughter cell that maintains the stem-cell capacity, and 1 daughter cell committed to differentiation. While asymmetric stem-cell divisions have been proven to occur in model organisms (eg, in Drosophila), it remains illusive wh

  12. The TCP4 transcription factor of Arabidopsis blocks cell division in yeast at G1 → S transition

    International Nuclear Information System (INIS)

    Highlights: → TCP4 is a class II TCP transcription factor, that represses cell division in Arabidopsis. → TCP4 expression in yeast retards cell division by blocking G1 → S transition. → Genome-wide expression studies and Western analysis reveals stabilization of cell cycle inhibitor Sic1, as possible mechanism. -- Abstract: The TCP transcription factors control important aspects of plant development. Members of class I TCP proteins promote cell cycle by regulating genes directly involved in cell proliferation. In contrast, members of class II TCP proteins repress cell division. While it has been postulated that class II proteins induce differentiation signal, their exact role on cell cycle has not been studied. Here, we report that TCP4, a class II TCP protein from Arabidopsis that repress cell proliferation in developing leaves, inhibits cell division by blocking G1 → S transition in budding yeast. Cells expressing TCP4 protein with increased transcriptional activity fail to progress beyond G1 phase. By analyzing global transcriptional status of these cells, we show that expression of a number of cell cycle genes is altered. The possible mechanism of G1 → S arrest is discussed.

  13. Structural and functional studies of MinD ATPase: implications for the molecular recognition of the bacterial cell division apparatus

    OpenAIRE

    Hayashi, Ikuko; Oyama, Takuji; Morikawa, Kosuke

    2001-01-01

    Proper placement of the bacterial cell division site requires the site-specific inactivation of other potential division sites. In Escherichia coli, selection of the correct mid-cell site is mediated by the MinC, MinD and MinE proteins. To clarify the functional role of the bacterial cell division inhibitor MinD, which is a membrane-associated ATPase that works as an activator of MinC, we determined the crystal structure of a Pyrococcus furiosus MinD homologue complexed with a substrate analo...

  14. AtPPR2, an Arabidopsis pentatricopeptide repeat protein, binds to plastid 23S rRNA and plays an important role in the first mitotic division during gametogenesis and in cell proliferation during embryogenesis

    OpenAIRE

    Lu, Yuqing; Li, Cong; Wang, Hai; Chen, Hao; Berg, Howard; Xia, Yiji

    2011-01-01

    Pentatricopeptide repeat (PPR) proteins are mainly involved in regulating post-transcriptional processes in mitochondria and plastids, including chloroplasts. Mutations in the Arabidopsis PPR2 gene have previously been found to cause defects in seed development and reduced transmission through male and female gametophytes. However, the exact function of AtPPR2 has not been defined. We found that a loss-of-function mutation of AtPPR2 leads to arrest of the first mitotic division during both ma...

  15. Polyalkoxyflavonoids as inhibitors of cell division

    Science.gov (United States)

    Semenov, V. V.; Semenova, M. N.

    2015-02-01

    Being structural analogues of natural microtubule-destabilizing cytostatics, polyalkoxyflavonoids represent a promising class of compounds for anticancer drug design. The review covers synthetic routes to various polyalkoxyflavonoids and the results of biological assays in vitro on human cancer cells and in vivo using sea urchin embryos as a model. Mechanisms of action and structure-relationship activity for polyalkoxyflavonoids are discussed. The bibliography includes 151 references.

  16. A polymerization–depolymerization model for generation of contractile force during bacterial cell division

    Indian Academy of Sciences (India)

    Biplab Ghosh; Anirban Sain

    2008-08-01

    During the last phase of cell division in bacteria, a polymeric ring forms at the division site. The ring, made of intracellular proteins, anchors to the cell wall and starts to contract. That initiates a dividing septum to close in, like the shutter of a camera, eventually guillotining the cell into two daughters. All through, the ring remains at the leading edge of the septum and seems to power its closure. It is not understood why does the ring contract. We propose a theoretical model to explain this. It is worth mentioning that a similar contraction phenomenon occurs for the actin ring in eukaryotes, but there it is due to motor proteins, which however, are absent in bacteria.

  17. Dielectric modelling of cell division for budding and fission yeast

    International Nuclear Information System (INIS)

    The frequency dependence of complex permittivity or the dielectric spectrum of a system including a cell in cell division has been simulated by a numerical technique based on the three-dimensional finite difference method. Two different types of cell division characteristic of budding and fission yeast were examined. The yeast cells are both regarded as a body of rotation, and thus have anisotropic polarization, i.e. the effective permittivity of the cell depends on the orientation of the cell to the direction of an applied electric field. In the perpendicular orientation, where the rotational axis of the cell is perpendicular to the electric field direction, the dielectric spectra for both yeast cells included one dielectric relaxation and its intensity depended on the cell volume. In the parallel orientation, on the other hand, two dielectric relaxations appeared with bud growth for budding yeast and with septum formation for fission yeast. The low-frequency relaxation was shifted to a lower frequency region by narrowing the neck between the bud and the mother cell for budding yeast and by increasing the degree of septum formation for fission yeast. After cell separation, the low-frequency relaxation disappeared. The simulations well interpreted the oscillation of the relative permittivity of culture broth found for synchronous cell growth of budding yeast

  18. Dielectric modelling of cell division for budding and fission yeast

    Science.gov (United States)

    Asami, Koji; Sekine, Katsuhisa

    2007-02-01

    The frequency dependence of complex permittivity or the dielectric spectrum of a system including a cell in cell division has been simulated by a numerical technique based on the three-dimensional finite difference method. Two different types of cell division characteristic of budding and fission yeast were examined. The yeast cells are both regarded as a body of rotation, and thus have anisotropic polarization, i.e. the effective permittivity of the cell depends on the orientation of the cell to the direction of an applied electric field. In the perpendicular orientation, where the rotational axis of the cell is perpendicular to the electric field direction, the dielectric spectra for both yeast cells included one dielectric relaxation and its intensity depended on the cell volume. In the parallel orientation, on the other hand, two dielectric relaxations appeared with bud growth for budding yeast and with septum formation for fission yeast. The low-frequency relaxation was shifted to a lower frequency region by narrowing the neck between the bud and the mother cell for budding yeast and by increasing the degree of septum formation for fission yeast. After cell separation, the low-frequency relaxation disappeared. The simulations well interpreted the oscillation of the relative permittivity of culture broth found for synchronous cell growth of budding yeast.

  19. Genetic Dissection of the Sporulation Protein SpoIIE and Its Role in Asymmetric Division in Bacillus subtilis†

    OpenAIRE

    Carniol, Karen; Ben-Yehuda, Sigal; King, Nicole; Losick, Richard

    2005-01-01

    SpoIIE is a dual-function protein in Bacillus subtilis that contributes to the switch from medial to polar cell division during sporulation and is responsible for activating the cell-specific transcription factor σF. SpoIIE consists of an N-terminal domain with 10 membrane-spanning segments (region I), a C-terminal phosphatase domain (region III), and a central domain (region II) of uncertain function. To investigate the role of SpoIIE in polar division, we took advantage of a system for effi...

  20. Asymmetric cell division of stem cells in the lung and other systems

    Directory of Open Access Journals (Sweden)

    AhmedH. K.El-Hashash

    2014-07-01

    Full Text Available New insights have been added to identification, behavior and cellular properties of embryonic and tissue-specific stem cells over the last few years. The modes of stem cell division, asymmetric versus symmetric, are tightly regulated during development and regeneration. The proper choice of a stem cell to divide asymmetrically or symmetrically has great consequences for development and disease because inappropriate asymmetric division disrupts organ morphogenesis, whereas uncontrolled symmetric division induces tumorigenesis. Therefore, understanding the behavior of lung stem cells could identify innovative solutions for restoring normal morphogenesis and/or regeneration of different organs. In this concise review, we describe recent studies in our laboratory about the mode of division of lung epithelial stem cells. We also compare asymmetric cell division in the lung stem cells with other tissues in different organisms.

  1. Stem cell regulation: Implications when differentiated cells regulate symmetric stem cell division.

    Science.gov (United States)

    Høyem, Marte Rørvik; Måløy, Frode; Jakobsen, Per; Brandsdal, Bjørn Olav

    2015-09-01

    We use a mathematical model to show that if symmetric stem cell division is regulated by differentiated cells, then changes in the population dynamics of the differentiated cells can lead to changes in the population dynamics of the stem cells. More precisely, the relative fitness of the stem cells can be affected by modifying the death rate of the differentiated cells. This result is interesting because stem cells are less sensitive than differentiated cells to environmental factors, such as medical therapy. Our result implies that stem cells can be manipulated indirectly by medical treatments that target the differentiated cells. PMID:25997796

  2. Polar flagellar biosynthesis and a regulator of flagellar number influence spatial parameters of cell division in Campylobacter jejuni.

    Directory of Open Access Journals (Sweden)

    Murat Balaban

    2011-12-01

    Full Text Available Spatial and numerical regulation of flagellar biosynthesis results in different flagellation patterns specific for each bacterial species. Campylobacter jejuni produces amphitrichous (bipolar flagella to result in a single flagellum at both poles. These flagella confer swimming motility and a distinctive darting motility necessary for infection of humans to cause diarrheal disease and animals to promote commensalism. In addition to flagellation, symmetrical cell division is spatially regulated so that the divisome forms near the cellular midpoint. We have identified an unprecedented system for spatially regulating cell division in C. jejuni composed by FlhG, a regulator of flagellar number in polar flagellates, and components of amphitrichous flagella. Similar to its role in other polarly-flagellated bacteria, we found that FlhG regulates flagellar biosynthesis to limit poles of C. jejuni to one flagellum. Furthermore, we discovered that FlhG negatively influences the ability of FtsZ to initiate cell division. Through analysis of specific flagellar mutants, we discovered that components of the motor and switch complex of amphitrichous flagella are required with FlhG to specifically inhibit division at poles. Without FlhG or specific motor and switch complex proteins, cell division occurs more often at polar regions to form minicells. Our findings suggest a new understanding for the biological requirement of the amphitrichous flagellation pattern in bacteria that extend beyond motility, virulence, and colonization. We propose that amphitrichous bacteria such as Campylobacter species advantageously exploit placement of flagella at both poles to spatially regulate an FlhG-dependent mechanism to inhibit polar cell division, thereby encouraging symmetrical cell division to generate the greatest number of viable offspring. Furthermore, we found that other polarly-flagellated bacteria produce FlhG proteins that influence cell division, suggesting that

  3. Functional Genomic Analysis of Systemic Cell Division Regulation in Legumes

    International Nuclear Information System (INIS)

    Legumes develop root nodules from pluripotent stem cells in the root pericycle in response to mitogenic activation by a decorated chitin-like nodulation factor synthesized in Rhizobium bacteria. The soybean genes encoding the receptor for such signals were cloned using map-based cloning approaches. Pluripotent cells in the root pericycle and the outer or inner cortex undergo repeated cell divisions to initiate a composite nodule primordium that develops to a functional nitrogen-fixing nodule. The process itself is autoregulated, leading to the characteristic nodulation of the upper root system. Autoregulation of nodulation (AON) in all legumes is controlled in part by a leucine-rich repeat receptor kinase gene (GmNARK). Mutations of GmNARK, and its other legume orthologues, result in abundant nodulation caused by the loss of a yet-undefined negative nodulation repressor system. AON receptor kinases are involved in perception of a long distance, root-derived signal, to negatively control nodule proliferation. GmNARK and LjHAR1 are expressed in phloem parenchyma. GmNARK kinase domain interacts with Kinase Associated Protein Phosphatase (KAPP). NARK gene expression did not mirror biological NARK activity in nodulation control, as q-RT-PCR in soybean revealed high NARK expression in roots, root tips, leaves, petioles, stems and hypocotyls, while shoot and root apical meristems were devoid of NARK RNA. High through-put transcript analysis in soybean leaf and root indicated that major genes involved in JA synthesis or response are preferentially down-regulated in leaf but not root of wild type, but not NARK mutants, suggesting that AON signaling may in part be controlled by events relating to hormone metabolism. Ethylene and abscisic acid insensitive mutants of L. japonicus are described. Nodulation in legumes has significance to global economies and ecologies, as the nitrogen input into the biosphere allows food, feed and biofuel production without the inherent costs

  4. SepG coordinates sporulation-specific cell division and nucleoid organization in Streptomyces coelicolor.

    Science.gov (United States)

    Zhang, Le; Willemse, Joost; Claessen, Dennis; van Wezel, Gilles P

    2016-04-01

    Bacterial cell division is a highly complex process that requires tight coordination between septum formation and chromosome replication and segregation. In bacteria that divide by binary fission a single septum is formed at mid-cell, a process that is coordinated by the conserved cell division scaffold protein FtsZ. In contrast, during sporulation-specific cell division in streptomycetes, up to a hundred rings of FtsZ (Z rings) are produced almost simultaneously, dividing the multinucleoid aerial hyphae into long chains of unigenomic spores. This involves the active recruitment of FtsZ by the SsgB protein, and at the same time requires sophisticated systems to regulate chromosome dynamics. Here, we show that SepG is required for the onset of sporulation and acts by ensuring that SsgB is localized to future septum sites. Förster resonance energy transfer imaging suggests direct interaction between SepG and SsgB. The beta-lactamase reporter system showed that SepG is a transmembrane protein with its central domain oriented towards the cytoplasm. Without SepG, SsgB fails to localize properly, consistent with a crucial role for SepG in the membrane localization of the SsgB-FtsZ complex. While SsgB remains associated with FtsZ, SepG re-localizes to the (pre)spore periphery. Expanded doughnut-shaped nucleoids are formed in sepG null mutants, suggesting that SepG is required for nucleoid compaction. Taken together, our work shows that SepG, encoded by one of the last genes in the conserved dcw cluster of cell division and cell-wall-related genes in Gram-positive bacteria whose function was still largely unresolved,coordinates septum synthesis and chromosome organization in Streptomyces. PMID:27053678

  5. Rab24 is required for normal cell division.

    Science.gov (United States)

    Militello, Rodrigo D; Munafó, Daniela B; Berón, Walter; López, Luis A; Monier, Solange; Goud, Bruno; Colombo, María I

    2013-05-01

    Rab24 is an atypical member of the Rab GTPase family whose distribution in interphase cells has been characterized; however, its function remains largely unknown. In this study, we have analyzed the distribution of Rab24 throughout cell division. We have observed that Rab24 was located at the mitotic spindle in metaphase, at the midbody during telophase and in the furrow during cytokinesis. We have also observed partial co-localization of Rab24 and tubulin and demonstrated its association to microtubules. Interestingly, more than 90% of transiently transfected HeLa cells with Rab24 presented abnormal nuclear connections (i.e., chromatin bridges). Furthermore, in CHO cells stably transfected with GFP-Rab24wt, we observed a large percentage of binucleated and multinucleated cells. In addition, these cells presented an extremely large size and multiple failures in mitosis, as aberrant spindle formation (metaphase), delayed chromosomes (telophase) and multiple cytokinesis. A marked increase in binucleated, multinucleated and multilobulated nucleus formation was observed in HeLa cells depleted of Rab24. We also present evidence that a fraction of Rab24 associates with microtubules. In addition, Rab24 knock down resulted in misalignment of chromosomes and abnormal spindle formation in metaphase leading to the appearance of delayed chromosomes during late telophase and failures in cytokinesis. Our findings suggest that an adequate level of Rab24 is necessary for normal cell division. In summary, Rab24 modulates several mitotic events, including chromosome segregation and cytokinesis, perhaps through the interaction with microtubules. PMID:23387408

  6. Cell division control by the Chromosomal Passenger Complex

    Energy Technology Data Exchange (ETDEWEB)

    Waal, Maike S. van der; Hengeveld, Rutger C.C.; Horst, Armando van der; Lens, Susanne M.A., E-mail: s.m.a.lens@umcutrecht.nl

    2012-07-15

    The Chromosomal Passenger Complex (CPC) consisting of Aurora B kinase, INCENP, Survivin and Borealin, is essential for genomic stability by controlling multiple processes during both nuclear and cytoplasmic division. In mitosis it ensures accurate segregation of the duplicated chromosomes by regulating the mitotic checkpoint, destabilizing incorrectly attached spindle microtubules and by promoting the axial shortening of chromosomal arms in anaphase. During cytokinesis the CPC most likely prevents chromosome damage by imposing an abscission delay when a chromosome bridge connects the two daughter cells. Moreover, by controlling proper cytoplasmic division, the CPC averts tetraploidization. This review describes recent insights on how the CPC is capable of conducting its various functions in the dividing cell to ensure chromosomal stability.

  7. Arv1 promotes cell division by recruiting IQGAP1 and myosin to the cleavage furrow.

    Science.gov (United States)

    Sundvold, Hilde; Sundvold-Gjerstad, Vibeke; Malerød-Fjeld, Helle; Haglund, Kaisa; Stenmark, Harald; Malerød, Lene

    2016-03-01

    Cell division is strictly regulated by a diversity of proteins and lipids to ensure proper duplication and segregation of genetic material and organelles. Here we report a novel role of the putative lipid transporter ACAT-related protein required for viability 1 (Arv1) during telophase. We observed that the subcellular localization of Arv1 changes according to cell cycle progression and that Arv1 is recruited to the cleavage furrow in early telophase by epithelial protein lost in neoplasm (EPLIN). At the cleavage furrow Arv1 recruits myosin heavy chain 9 (MYH9) and myosin light chain 9 (MYL9) by interacting with IQ-motif-containing GTPase-activating protein (IQGAP1). Consequently the lack of Arv1 delayed telophase-progression, and a strongly increased incidence of furrow regression and formation of multinuclear cells was observed both in human cells in culture and in follicle epithelial cells of egg chambers of Drosophila melanogaster in vivo. Interestingly, the cholesterol-status at the cleavage furrow did not affect the recruitment of either IQGAP1, MYH9 or MYL. These results identify a novel function for Arv1 in regulation of cell division through promotion of the contractile actomyosin ring, which is independent of its lipid transporter activity. PMID:27104745

  8. Cell Division Behaviour in a Heterogeneous Swarm Environment

    OpenAIRE

    Erskine, Adam; Herrmann, J. Michael

    2013-01-01

    We present a system of virtual particles that interact using simple kinetic rules. It is known that heterogeneous mixtures of particles are producing particularly interesting behaviours. Here we present a two-species swarm in which a behaviour emerges that resembles cell division. We show that the dividing behaviour exists across a narrow but finite band of parameters and for a wide range of population sizes. In a two dimensional environment the swarm's characteristics and dynamism manifests ...

  9. EzrA: a spectrin-like scaffold in the bacterial cell division machinery

    Directory of Open Access Journals (Sweden)

    Robert M Cleverley

    2015-01-01

    Full Text Available Much progress has been made in identifying the components of the divisome, the assembly of proteins that undertakes the vital process of cell division in bacteria. However, how the highly interdependent processes on either side of the membrane are coordinated during division is a major unresolved question. How is the degradation and synthesis of the cell wall on the outside of the cell coordinated with cytokinesis and membrane fission, which are driven from the inside of the cell by the tubulin homologue FtsZ? A possible key mediator of such coordination is the membrane protein EzrA, as it interacts both with FtsZ and the penicillin binding proteins (PBPs that synthesize peptidoglycan. Cleverley et al. [Nature Communications (2014 5, 5421] have recently solved the crystal structure of the cytoplasmic domain of B. subtilis EzrA, which points to an important scaffolding role for EzrA in the divisome. The structure resembles the eukaryotic, cytoskeletal spectrin proteins, which link actin filaments in the cytoskeleton and also connect the actin cytoskeleton to membrane-bound integrin proteins.

  10. ParA encoded on chromosome II of Deinococcus radiodurans binds to nucleoid and inhibits cell division in Escherichia coli

    Indian Academy of Sciences (India)

    Vijaya Kumar Charaka; Kruti P Mehta; H S Misra

    2013-09-01

    Bacterial genome segregation and cell division has been studied mostly in bacteria harbouring single circular chromosome and low-copy plasmids. Deinococcus radiodurans, a radiation-resistant bacterium, harbours multipartite genome system. Chromosome I encodes majority of the functions required for normal growth while other replicons encode mostly the proteins involved in secondary functions. Here, we report the characterization of putative P-loop ATPase (ParA2) encoded on chromosome II of D. radiodurans. Recombinant ParA2 was found to be a DNA-binding ATPase. E. coli cells expressing ParA2 showed cell division inhibition and mislocalization of FtsZ-YFP and those expressing ParA2-CFP showed multiple CFP foci formation on the nucleoid. Although, in trans expression of ParA2 failed to complement SlmA loss per se, it could induce unequal cell division in slmAminCDE double mutant. These results suggested that ParA2 is a nucleoid-binding protein, which could inhibits cell division in E. coli by affecting the correct localization of FtsZ and thereby cytokinesis. Helping slmAminCDE mutant to produce minicells, a phenotype associated with mutations in the `Min’ proteins, further indicated the possibility of ParA2 regulating cell division by bringing nucleoid compaction at the vicinity of septum growth.

  11. Patterns of Stem Cell Divisions Contribute to Plant Longevity.

    Science.gov (United States)

    Burian, Agata; Barbier de Reuille, Pierre; Kuhlemeier, Cris

    2016-06-01

    The lifespan of plants ranges from a few weeks in annuals to thousands of years in trees. It is hard to explain such extreme longevity considering that DNA replication errors inevitably cause mutations. Without purging through meiotic recombination, the accumulation of somatic mutations will eventually result in mutational meltdown, a phenomenon known as Muller's ratchet. Nevertheless, the lifespan of trees is limited more often by incidental disease or structural damage than by genetic aging. The key determinants of tree architecture are the axillary meristems, which form in the axils of leaves and grow out to form branches. The number of branches is low in annual plants, but in perennial plants iterative branching can result in thousands of terminal branches. Here, we use stem cell ablation and quantitative cell-lineage analysis to show that axillary meristems are set aside early, analogous to the metazoan germline. While neighboring cells divide vigorously, axillary meristem precursors maintain a quiescent state, with only 7-9 cell divisions occurring between the apical and axillary meristem. During iterative branching, the number of branches increases exponentially, while the number of cell divisions increases linearly. Moreover, computational modeling shows that stem cell arrangement and positioning of axillary meristems distribute somatic mutations around the main shoot, preventing their fixation and maximizing genetic heterogeneity. These features slow down Muller's ratchet and thereby extend lifespan. PMID:27161504

  12. Differential Management of the Replication Terminus Regions of the Two Vibrio cholerae Chromosomes during Cell Division

    OpenAIRE

    Gaëlle Demarre; Elisa Galli; Leila Muresan; Evelyne Paly; Ariane David; Christophe Possoz; François-Xavier Barre

    2014-01-01

    The replication terminus region (Ter) of the unique chromosome of most bacteria locates at mid-cell at the time of cell division. In several species, this localization participates in the necessary coordination between chromosome segregation and cell division, notably for the selection of the division site, the licensing of the division machinery assembly and the correct alignment of chromosome dimer resolution sites. The genome of Vibrio cholerae, the agent of the deadly human disease choler...

  13. Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

    Science.gov (United States)

    Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

    2010-05-01

    Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

  14. Lessons from development: A role for asymmetric stem cell division in cancer

    OpenAIRE

    Powell, Anne E.; Shung, Chia-Yi; Saylor, Katherine W.; Müllendorf, Karin A.; Weiss, Joseph B.; Wong, Melissa H.

    2009-01-01

    Asymmetric stem cell division has emerged as a major regulatory mechanism for physiologic control of stem cell numbers. Reinvigoration of the cancer stem cell theory suggests that tumorigenesis may be regulated by maintaining the balance between asymmetric and symmetric cell division. Therefore, mutations affecting this balance could result in aberrant expansion of stem cells. Although a number of molecules have been implicated in regulation of asymmetric stem cell division, here, we highligh...

  15. Asymmetric Cell Division in T Lymphocyte Fate Diversification.

    Science.gov (United States)

    Arsenio, Janilyn; Metz, Patrick J; Chang, John T

    2015-11-01

    Immunological protection against microbial pathogens is dependent on robust generation of functionally diverse T lymphocyte subsets. Upon microbial infection, naïve CD4(+) or CD8(+) T lymphocytes can give rise to effector- and memory-fated progeny that together mediate a potent immune response. Recent advances in single-cell immunological and genomic profiling technologies have helped elucidate early and late diversification mechanisms that enable the generation of heterogeneity from single T lymphocytes. We discuss these findings here and argue that one such mechanism, asymmetric cell division, creates an early divergence in T lymphocyte fates by giving rise to daughter cells with a propensity towards the terminally differentiated effector or self-renewing memory lineages, with cell-intrinsic and -extrinsic cues from the microenvironment driving the final maturation steps. PMID:26474675

  16. Bacillus thuringiensis peptidoglycan hydrolase SleB171 involved in daughter cell separation during cell division.

    Science.gov (United States)

    Li, Hua; Hu, Penggao; Zhao, Xiuyun; Yu, Ziniu; Li, Lin

    2016-04-01

    Whole-genome analyses have revealed a putative cell wall hydrolase gene (sleB171) that constitutes an operon with two other genes (ypeBandyhcN) of unknown function inBacillus thuringiensisBMB171. The putative SleB171 protein consists of 259 amino acids and has a molecular weight of 28.3 kDa. Gene disruption ofsleB171in the BMB171 genome causes the formation of long cell chains during the vegetative growth phase and delays spore formation and spore release, although it has no significant effect on cell growth and the ultimate release of the spores. The inseparable vegetative cells were nearly restored through the complementation ofsleB171expression. Real-time quantitative polymerase chain reaction analysis revealed thatsleB171is mainly active in the vegetative growth phase, with a maximum activity at the early stationary growth phase. Western blot analysis also confirmed thatsleB171is preferentially expressed during the vegetative growth phase. These results demonstrated that SleB171 plays an essential role in the daughter cell separation during cell division. PMID:26922318

  17. Omics and modeling approaches approaches for understanding regulation of asymmetric cell divisions in Arabidopsis and other angiosperm plants.

    NARCIS (Netherlands)

    Kajala, K.; Ramakrishna, A.; Fisher, A.; Bergmann, D.C.; Smet, De I.; Sozzani, R.; Weijers, D.; Brady, S.M.

    2014-01-01

    Background Asymmetric cell divisions are formative divisions that generate daughter cells of distinct identity. These divisions are coordinated by either extrinsic (‘niche-controlled’) or intrinsic regulatory mechanisms and are fundamentally important in plant development. Scope This review describe

  18. Role of SCHIZORIZA in asymmetric cell division, cell fate segregation and specification in Arabidopsis root development

    NARCIS (Netherlands)

    Jansweijer, V.M.A.

    2013-01-01

    Multicellular organisms develop their large variety of cell types from just one single cell, the zygote. Both plants and animals use asymmetric cell division to establish a multicellular body plan How different cell and tissue types are determined, how patterns are created and maintained, and which

  19. Cell cycle related /sup 125/IUDR-induced-division delay

    International Nuclear Information System (INIS)

    A series of experiments were run to determine if /sup 125/I-decays, in /sup 125/IUdR labeled DNA, specifically accumulated at 1, 3, 5, 7 and 9 hours after plating labeled mitotic cells caused a change in the rate or time of cell entry into mitosis. To accomplish this, a pool of labeled mitotic cells was selected in mitosis and plated in replicate flasks. /sup 125/I decays were accumulated in groups of cells by cooling (40C) for 2 hours starting at the designated times. After rewarding, colcemid was added to arrest cells in mitosis. The rate of cell progression into mitosis for each cell cycle time of accumulation was determined by scoring the mitotic index of cells sampled as a function of time after addition of the colcemid. The results are summarized: (1) Decays from /sup 125/I in /sup 125/I(UdR) labeled DNA reduced the rate of cell progression into mitosis and delayed the time of initiation of mitosis. (2) The reduced rate of progression and the delayed time of initiation of mitosis were independent of the cell cycle time that /sup 125/I-decays were accumulated. (3) The reduced rate of progression after cell cycle accumulation of /sup 125/I decay was statistically indistinguishable from the corresponding controls. (4) The delayed initiation of mitosis after specific cell cycle accumulation of /sup 125/I- decays was greater than the corresponding control. The relationship of these data to DNA and non-DNA division delay target(s) is emphasized

  20. Patterns of cell division revealed by transcriptional regulation of genes during the cell cycle in plants.

    OpenAIRE

    Fobert, P R; Coen, E S; Murphy, G. J.; Doonan, J H

    1994-01-01

    Transcripts from five cell cycle related genes accumulate in isolated cells dispersed throughout the actively dividing regions of plant meristems. We propose that this pattern reflects gene expression during particular phases of the cell division cycle. The high proportion of isolated cells suggests that synchrony between daughter cells is rapidly lost following mitosis. This is the first time that such a cell specific expression pattern has been described in a higher organism. Counterstainin...

  1. Enzymatically Inactive Procaspase 1 stabilizes the ASC Pyroptosome and Supports Pyroptosome Spreading during Cell Division.

    Science.gov (United States)

    Stein, Robert; Kapplusch, Franz; Heymann, Michael Christian; Russ, Susanne; Staroske, Wolfgang; Hedrich, Christian Michael; Rösen-Wolff, Angela; Hofmann, Sigrun Ruth

    2016-08-26

    Caspase-1 is a key player during the initiation of pro-inflammatory innate immune responses, activating pro-IL-1β in so-called inflammasomes. A subset of patients with recurrent febrile episodes and systemic inflammation of unknown origin harbor mutations in CASP1 encoding caspase-1. CASP1 variants result in reduced enzymatic activity of caspase-1 and impaired IL-1β secretion. The apparent paradox of reduced IL-1β secretion but systemic inflammation led to the hypothesis that CASP1 mutations may result in variable protein interaction clusters, thus activating alternative signaling pathways. To test this hypothesis, we established and characterized an in vitro system of transduced immortalized murine macrophages expressing either WT or enzymatically inactive (p.C284A) procaspase-1 fusion reporter proteins. Macrophages with variant p.C284A caspase-1 did not secrete IL-1β and exhibited reduced inflammatory cell death, referred to as pyroptosis. Caspase-1 and apoptosis-associated speck-like protein containing a CARD (ASC) formed cytosolic macromolecular complexes (so-called pyroptosomes) that were significantly increased in number and size in cells carrying the p.C284A caspase-1 variant compared with WT caspase-1. Furthermore, enzymatically inactive caspase-1 interacted with ASC longer and with increased intensity compared with WT caspase-1. Applying live cell imaging, we documented for the first time that pyroptosomes containing enzymatically inactive variant p.C284A caspase-1 spread during cell division. In conclusion, variant p.C284A caspase-1 stabilizes pyroptosome formation, potentially enhancing inflammation by two IL-1β-independent mechanisms: pyroptosomes convey an enhanced inflammatory stimulus through the recruitment of additional proteins (such as RIP2, receptor interacting protein kinase 2), which is further amplified through pyroptosome and cell division. PMID:27402835

  2. Characterization of substances that restore impaired cell division of UV-irradiated E. coli B

    International Nuclear Information System (INIS)

    Substances which restore impaired cell division in UV-irradiated E. coli B were surveyed among various bacteria. The active substance was found only in several genera of Gram-negative bacteria, i.e., Escherichia, Enterobacter, Salmonella and some species of Pseudomonas. The activity in the dialyzed cell extract of E. coli B/r was observed in the presence of β-NAD and was enhanced by Mg2+ and Mn2+. The active substance was very labile, but the activity was protected by 1 mM dithiothreitol in the process of purification. The activity of a fraction recovered through DEAE-cellulose column chromatography was stimulated by the presence of membrane fraction. Upon treatment with lipid-degrading enzymes and proteases, the division-stimulating activity was lost or reduced. It appears that the inactivation by lipase and phospholipase A2 was due to the formation of lysophospholipids and that a proteinous substance participated in the recovery of impaired cell division of UV-irradiated E. coli B

  3. Cell Division, a new open access online forum for and from the cell cycle community

    Directory of Open Access Journals (Sweden)

    Kaldis Philipp

    2006-04-01

    Full Text Available Abstract Cell Division is a new, open access, peer-reviewed online journal that publishes cutting-edge articles, commentaries and reviews on all exciting aspects of cell cycle control in eukaryotes. A major goal of this new journal is to publish timely and significant studies on the aberrations of the cell cycle network that occur in cancer and other diseases.

  4. The effectiveness and problems of utilizing diagrams for secondary school students' learning of cell divisions

    OpenAIRE

    Cheung, Man-lai; 張敏麗

    2014-01-01

    Cell division is a topic widely recognized by biology teachers for its importance in the curriculum. However, it is also regarded by teachers and students as a very difficult topic. Students at different school levels often hold many misconceptions in cell divisions. While diagrams have been so commonly employed in biology textbooks to enhance the textual representation of cell divisions, the researcher of this study questioned about the cognitive role of the diagrams in helping students cons...

  5. From HeLa cell division to infectious diarrhoea

    Energy Technology Data Exchange (ETDEWEB)

    Stephen, J.; Osborne, M.P.; Spencer, A.J.; Warley, A. (Univ. of Birmingham (England))

    1990-09-01

    Hela S3 cells were grown in suspension both randomly and, synchronously using hydroxyurea which blocks cells at the G1/S interface. Cryosections were prepared, freeze-dried and analyzed by X-ray microanalysis. As cells moved into S and through M phases (Na) and (Cl) increased; both returned to normal levels upon re-entering G1 phase. The Na/K ratio was 1:1 in G1 phase. Infection of HeLa S3 cells in G1 phase with vaccinia virus resulted in no change in intracellular (Na). Infection of neonatal mice with murine rotavirus was localized to villus tip enterocytes and gave rise to diarrhoea which was maximal at 72h post-infection (p.i.). Diarrhoea was preceded by ischemia of villi (18-42h p.i.) and villus shortening (maximal at 42h p.i.), and was also coincident with a dramatic regrowth of villi. At 48h p.i. a proliferative zone of electron lucent cells was observed in villus base regions. Cryosections of infected gut, taken before, during, and after infection, together with corresponding age-matched controls, were freeze-dried and analysed by X-ray microanalysis. At 48h p.i. electron lucent villus base cells were shown to be more hydrated, and, to contain higher levels of both Na and Cl and lower levels of P, S, K and Mg than corresponding control cells. These studies increase confidence in the use of X-ray microanalysis in studying biological systems, provide some insight into the process of cell division, and constitute the basis of a new concept of diarrhoeal secretion.27 references.

  6. Positioning of polarity formation by extracellular signaling during asymmetric cell division.

    Science.gov (United States)

    Seirin Lee, Sungrim

    2016-07-01

    Anterior-posterior (AP) polarity formation of cell membrane proteins plays a crucial role in determining cell asymmetry, which ultimately generates cell diversity. In Caenorhabditis elegans, a single fertilized egg cell (P0), its daughter cell (P1), and the germline precursors (P2 and P3 cells) form two exclusive domains of different PAR proteins on the membrane along the anterior-posterior axis. However, the phenomenon of polarity reversal has been observed in which the axis of asymmetric cell division of the P2 and P3 cells is formed in an opposite manner to that of the P0 and P1 cells. The extracellular signal MES-1/SRC-1 has been shown to induce polarity reversal, but the detailed mechanism remains elusive. Here, using a mathematical model, I explore the mechanism by which MES-1/SRC-1 signaling can induce polarity reversal and ultimately affect the process of polarity formation. I show that a positive correlation between SRC-1 and the on-rate of PAR-2 is the essential mechanism underlying polarity reversal, providing a mathematical basis for the orientation of cell polarity patterns. PMID:27086039

  7. Radiomimetic effect of cisplatin on cucumber root development: the relationship between cell division and cell growth

    International Nuclear Information System (INIS)

    Cisplatin [DDP, cis-dichlorodiammine platinum (II)], a strong cytostatic and antineoplastic agent, was tested on seedlings of cucumber Cucumis sativus L. for its general effect on root development and its particular effects on root cell division and cell growth. DDP was characterized as a radiomimetic compound since both DDP (1·3 × 10-5 M) and γ-irradiation (2·5-10 kGy) drastically and irreversibly stopped development of embryonic lateral root primordia (LRPs) in the radicle by inhibiting both mitotic activity and cell growth. In 20% of the LRPs of DDP-treated roots, cells did not divide at all. Dividing cells completed no more than two cell cycles. These effects were specific because when DDP was available to the roots only at the onset of cell division, cell proliferation and cell growth were similar to that produced by constant incubation. Neither DDP nor γ-irradiation affected non-meristematic cell elongation. It was concluded that cell growth of meristematic cells is closely related to cell division. However, non-meristematic cell growth is independent of DNA damage. This suggests DDP as a tool to reveal these autonomous processes in plants development and to detect tissue compartments in mature plant embryos which contain potentially non-meristematic cells. (author)

  8. Resistance-Nodulation-Cell Division-Type Efflux Pump Involved in Aminoglycoside Resistance in Acinetobacter baumannii Strain BM4454

    OpenAIRE

    Magnet, Sophie; Courvalin, Patrice; Lambert, Thierry

    2001-01-01

    Multidrug-resistant strain Acinetobacter baumannii BM4454 was isolated from a patient with a urinary tract infection. The adeB gene, which encodes a resistance-nodulation-cell division (RND) protein, was detected in this strain by PCR with two degenerate oligodeoxynucleotides. Insertional inactivation of adeB in BM4454, which generated BM4454-1, showed that the corresponding protein was responsible for aminoglycoside resistance and was involved in the level of susceptibility to other drugs in...

  9. Asymmetric Inheritance of Mother Versus Daughter Centrosome in Stem Cell Division

    OpenAIRE

    Yamashita, Yukiko M.; Anthony P Mahowald; Perlin, Julie R.; Fuller, Margaret T.

    2007-01-01

    Adult stem cells often divide asymmetrically to produce one self-renewed stem cell and one differentiating cell, thus maintaining both populations. The asymmetric outcome of stem cell divisions can be specified by an oriented spindle and local self-renewal signals from the stem cell niche. Here we show that developmentally programmed asymmetric behavior and inheritance of mother and daughter centrosomes underlies the stereotyped spindle orientation and asymmetric outcome of stem cell division...

  10. The Protective Role of Symmetric Stem Cell Division on the Accumulation of Heritable Damage

    OpenAIRE

    McHale, PT; Lander, AD

    2014-01-01

    Stem cell divisions are either asymmetric-in which one daughter cell remains a stem cell and one does not-or symmetric, in which both daughter cells adopt the same fate, either stem or non-stem. Recent studies show that in many tissues operating under homeostatic conditions stem cell division patterns are strongly biased toward the symmetric outcome, raising the question of whether symmetry confers some benefit. Here, we show that symmetry, via extinction of damaged stem-cell clones, reduces ...

  11. The deletion of bacterial dynamin and flotillin genes results in pleiotrophic effects on cell division, cell growth and in cell shape maintenance

    Directory of Open Access Journals (Sweden)

    Dempwolff Felix

    2012-12-01

    Full Text Available Abstract Background In eukaryotic cells, dynamin and flotillin are involved in processes such as endocytosis and lipid raft formation, respectively. Dynamin is a GTPase that exerts motor-like activity during the pinching off of vesicles, while flotillins are coiled coil rich membrane proteins with no known enzymatic activity. Bacteria also possess orthologs of both classes of proteins, but their function has been unclear. Results We show that deletion of the single dynA or floT genes lead to no phenotype or a mild defect in septum formation in the case of the dynA gene, while dynA floT double mutant cells were highly elongated and irregularly shaped, although the MreB cytoskeleton appeared to be normal. DynA colocalizes with FtsZ, and the dynA deletion strain shows aberrant FtsZ rings in a subpopulation of cells. The mild division defect of the dynA deletion is exacerbated by an additional deletion in ezrA, which affects FtsZ ring formation, and also by the deletion of a late division gene (divIB, indicating that DynA affects several steps in cell division. DynA and mreB deletions generated a synthetic defect in cell shape maintenance, showing that MreB and DynA play non-epistatic functions in cell shape maintenance. TIRF microscopy revealed that FloT forms many dynamic membrane assemblies that frequently colocalize with the division septum. The deletion of dynA did not change the pattern of localization of FloT, and vice versa, showing that the two proteins play non redundant roles in a variety of cellular processes. Expression of dynamin or flotillin T in eukaryotic S2 cells revealed that both proteins assemble at the cell membrane. While FloT formed patch structures, DynA built up tubulated structures extending away from the cells. Conclusions Bacillus subtilis dynamin ortholog DynA plays a role during cell division and in cell shape maintenance. It shows a genetic link with flotillin T, with both proteins playing non-redundant functions at

  12. Uncovering the link between malfunctions in Drosophila neuroblast asymmetric cell division and tumorigenesis

    Directory of Open Access Journals (Sweden)

    Kelsom Corey

    2012-11-01

    Full Text Available Abstract Asymmetric cell division is a developmental process utilized by several organisms. On the most basic level, an asymmetric division produces two daughter cells, each possessing a different identity or fate. Drosophila melanogaster progenitor cells, referred to as neuroblasts, undergo asymmetric division to produce a daughter neuroblast and another cell known as a ganglion mother cell (GMC. There are several features of asymmetric division in Drosophila that make it a very complex process, and these aspects will be discussed at length. The cell fate determinants that play a role in specifying daughter cell fate, as well as the mechanisms behind setting up cortical polarity within neuroblasts, have proved to be essential to ensuring that neurogenesis occurs properly. The role that mitotic spindle orientation plays in coordinating asymmetric division, as well as how cell cycle regulators influence asymmetric division machinery, will also be addressed. Most significantly, malfunctions during asymmetric cell division have shown to be causally linked with neoplastic growth and tumor formation. Therefore, it is imperative that the developmental repercussions as a result of asymmetric cell division gone awry be understood.

  13. Is the cell division cycle gated by a circadian clock? The case of Chlamydomonas reinhardtii

    OpenAIRE

    1995-01-01

    Circadian oscillators are known to regulate the timing of cell division in many organisms. In the case of Chlamydomonas reinhardtii, however, this conclusion has been challenged by several investigators. We have reexamined this issue and find that the division behavior of Chlamydomonas meets all the criteria for circadian rhythmicity: persistence of a cell division rhythm (a) with a period of approximately 24 h under free-running conditions, (b) that is temperature compensated, and (c) which ...

  14. Entropyomics as the Blueprint of the Logic of Normal Cell Division and Malignancy

    Directory of Open Access Journals (Sweden)

    Kambiz Afrasiabi

    2011-01-01

    Full Text Available Problem statement: In this article I propose a blueprint based on one of the most fundamental laws governing the known universe, namely the second law of thermodynamics and I present support for its central role in initiation of mitosis and relationship of the other sub cellular compartments and their organization. Approach: Life is considered to be the most sophisticated antientropy machinery ever born on the face of the universe as far as its power to minimize the speed of rise in entropy is concerned, however we all get old, sick and die because it is not possible to stop the rise in entropy based on the nature of the known universe. Results: Lack of understanding of the scientific foundation of logic of the normal cell division has surrounded us by darkness and has made analysis of an ever increasing and explosive amount of information originating from whole genome sequencing, genomics, exonomics, proteomics and metabolomics more problematic. Clearly this understanding is the prerequisite for understanding of pathological states of cell division including malignancy. Conclusion/Recommendations: The main approach to this problem is calculation of the free energy of the master regulator proteins of the intracellular communication network of the cancer stem cell and its normal counterpart which in turn could get identified by the available mathematical models that could identify master regulator proteins of the intracellular communication network and deciphering the difference by spectrophotometry at a given wavelength of light and identification of higher absorbance in the malignant counterpart and designing epigenetic or homologous recombination mediated methodology using nanotechology as a delivery mechanism targeting transcription of mRNAs which would lead to protein products with a normal free energy for that cell lineage / higher free energy compared with its malignant counterpart and by doing so we could convert the

  15. Nek11 regulates asymmetric cell division during mouse oocyte meiotic maturation.

    Science.gov (United States)

    Guo, Lei; Wang, Zhen-Bo; Wang, Hong-Hui; Zhang, Teng; Qi, Shu-Tao; Ouyang, Ying-Chun; Hou, Yi; Sun, Qing-Yuan

    2016-06-10

    Nek11, a member of the never in mitosis gene A (NIMA) family, is activated in somatic cells associated with G1/S or G2/M arrest. However, its function in meiosis is unknown. In this research, the expression, localization and functions of NEK11 in the mouse oocyte meiotic maturation were examined. Western blotting indicated that NEK11S was the major NEK11 protein in mouse oocyte. MYC-tagged Nek11 mRNA microinjection and immunofluorescent staining showed that NEK11 was localized to the meiotic spindles at MI and MII stage. Knockdown of Nek11 by microinjection of siRNA did not affect germinal vesicle breakdown (GVBD) and the first polar body extrusion, but caused formation of 2-cell-like eggs. These results demonstrate that Nek11 regulates asymmetric cell division during oocyte meiotic maturation. PMID:27150633

  16. The role of GlsA in the evolution of asymmetric cell division in the green alga Volvox carteri.

    Science.gov (United States)

    Cheng, Qian; Fowler, Rachel; Tam, Lai-wa; Edwards, Lisseth; Miller, Stephen M

    2003-07-01

    Volvox carteri, a green alga in the order Volvocales, contains two completely differentiated cell types, small motile somatic cells and large reproductive cells called gonidia, that are set apart from each other during embryogenesis by a series of visibly asymmetric cell divisions. Mutational analysis has revealed a class of genes (gonidialess, gls) that are required specifically for asymmetric divisions in V. carteri, but that are dispensable for symmetric divisions. Previously we cloned one of these genes, glsA, and showed that it encodes a chaperone-like protein (GlsA) that has close orthologs in a diverse set of eukaryotes, ranging from fungi to vertebrates and higher plants. In the present study we set out to explore the role of glsA in the evolution of asymmetric division in the volvocine algae by cloning and characterizing a glsA ortholog from one of the simplest members of the group, Chlamydomonas reinhardtii, which does not undergo asymmetric divisions. This ortholog (which we have named gar1, for glsA related) is predicted to encode a protein that is 70% identical to GlsA overall, and that is most closely related to GlsA in the same domains that are most highly conserved between GlsA and its other known orthologs. We report that a gar1 transgene fully complements the glsA mutation in V. carteri, a result that suggests that asymmetric division probably arose through the modification of a gene whose product interacts with GlsA, but not through a modification of glsA itself. PMID:12743823

  17. A systematic analysis of cell cycle regulators in yeast reveals that most factors act independently of cell size to control initiation of division.

    Directory of Open Access Journals (Sweden)

    Scott A Hoose

    Full Text Available Upstream events that trigger initiation of cell division, at a point called START in yeast, determine the overall rates of cell proliferation. The identity and complete sequence of those events remain unknown. Previous studies relied mainly on cell size changes to identify systematically genes required for the timely completion of START. Here, we evaluated panels of non-essential single gene deletion strains for altered DNA content by flow cytometry. This analysis revealed that most gene deletions that altered cell cycle progression did not change cell size. Our results highlight a strong requirement for ribosomal biogenesis and protein synthesis for initiation of cell division. We also identified numerous factors that have not been previously implicated in cell cycle control mechanisms. We found that CBS, which catalyzes the synthesis of cystathionine from serine and homocysteine, advances START in two ways: by promoting cell growth, which requires CBS's catalytic activity, and by a separate function, which does not require CBS's catalytic activity. CBS defects cause disease in humans, and in animals CBS has vital, non-catalytic, unknown roles. Hence, our results may be relevant for human biology. Taken together, these findings significantly expand the range of factors required for the timely initiation of cell division. The systematic identification of non-essential regulators of cell division we describe will be a valuable resource for analysis of cell cycle progression in yeast and other organisms.

  18. Specific polar subpopulations of astral microtubules control spindle orientation and symmetric neural stem cell division.

    Science.gov (United States)

    Mora-Bermúdez, Felipe; Matsuzaki, Fumio; Huttner, Wieland B

    2014-01-01

    Mitotic spindle orientation is crucial for symmetric vs asymmetric cell division and depends on astral microtubules. Here, we show that distinct subpopulations of astral microtubules exist, which have differential functions in regulating spindle orientation and division symmetry. Specifically, in polarized stem cells of developing mouse neocortex, astral microtubules reaching the apical and basal cell cortex, but not those reaching the central cell cortex, are more abundant in symmetrically than asymmetrically dividing cells and reduce spindle orientation variability. This promotes symmetric divisions by maintaining an apico-basal cleavage plane. The greater abundance of apical/basal astrals depends on a higher concentration, at the basal cell cortex, of LGN, a known spindle-cell cortex linker. Furthermore, newly developed specific microtubule perturbations that selectively decrease apical/basal astrals recapitulate the symmetric-to-asymmetric division switch and suffice to increase neurogenesis in vivo. Thus, our study identifies a novel link between cell polarity, astral microtubules, and spindle orientation in morphogenesis. PMID:24996848

  19. Light can rescue auxin-dependent synchrony of cell division in a tobacco cell line

    Czech Academy of Sciences Publication Activity Database

    Qiao, F.; Petrášek, Jan; Nick, P.

    2010-01-01

    Roč. 61, č. 2 (2010), s. 503-510. ISSN 0022-0957 Institutional research plan: CEZ:AV0Z50380511 Keywords : Auxin transport * cell division * NPA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.818, year: 2010 http://jxb.oxfordjournals.org/content/61/2/503.abstract

  20. Progesterone Receptor Membrane Component 1 (PGRMC1 in cell division: its role in bovine granulosa cells mitosis

    Directory of Open Access Journals (Sweden)

    Laura Terzaghi

    2015-07-01

    Full Text Available The present studies were aimed to assess Progesterone Receptor Membrane Component-1 (PGRMC1 role in regulating bovine granulosa cells (bGC mitosis. First, we performed immunofluorescence studies on in vitro cultured bGC collected from antral follicles, which showed that PGRMC1 localizes to the spindle apparatus in mitotic cells. Then, to evaluate PGRMC1 effect on cell proliferation we silenced its expression with RNA interference technique (RNAi. Quantitative RT-PCR and immunoblotting confirmed down-regulation of PGRMC1 expression, when compared to CTRL-RNAi treated bGC (p<0.05. After 72h of culture, PGRMC1 silencing determined a lower growth rate (p<0.05 and a higher percentage of cells arrested at G2/M phase as assessed by flowcytometry (p<0.05. Accordingly, live imaging studies revealed more aberrant mitosis and a delayed M-phase in PGRMC1-RNAi treated cells compared to CTRL-RNAi group (p<0.05. These data confirmed that PGRMC1 is directly involved in bGC mitosis and ongoing preliminary studies are aimed to elucidate its putative mechanisms of action. Since PGRMC1 is a membrane protein, we hypothesize its possible involvement in vesicular trafficking and endocytosis, which is in turn an important process to assure proper cell division. To assess this hypothesis, we have preliminarily conducted immunofluorescence and in situ proximity ligation assay experiments that showed PGRMC1 co-localization and direct interaction with clathrin. This is important since clathrin is an essential protein for both endosomes formation, and cell division acting directly on the spindle apparatus. Thus our studies set the stage for analysis aimed to further characterize PGRMC1’s mechanism of action in mitotic cell.

  1. Different degree in proteasome malfunction has various effects on root growth possibly through preventing cell division and promoting autophagic vacuolization.

    Directory of Open Access Journals (Sweden)

    Xianyong Sheng

    Full Text Available The ubiquitin/proteasome pathway plays a vital role in plant development. But the effects of proteasome malfunction on root growth, and the mechanism underlying this involvement remains unclear. In the present study, the effects of proteasome inhibitors on Arabidopsis root growth were studied through the analysis of the root length, and meristem size and cell length in maturation zone using FM4-64, and cell-division potential using GFP fusion cyclin B, and accumulation of ubiquitinated proteins using immunofluorescence labeling, and autophagy activity using LysoTracker and MDC. The results indicated that lower concentration of proteasome inhibitors promoted root growth, whereas higher concentration of inhibitors had the opposite effects. The accumulation of cyclin B was linked to MG132-induced decline in meristem size, indicating that proteasome malfunction prevented cell division. Besides, MG132-induced accumulation of the ubiquitinated proteins was associated with the increasing fluorescence signal of LysoTracker and MDC in the elongation zone, revealing a link between the activation of autophagy and proteasome malfunction. These results suggest that weak proteasome malfunction activates moderate autophagy and promotes cell elongation, which compensates the inhibitor-induced reduction of cell division, resulting in long roots. Whereas strong proteasome malfunction induces severe autophagy and disturbs cell elongation, resulting in short roots.

  2. The equatorial position of the metaphase plate ensures symmetric cell divisions.

    Science.gov (United States)

    Tan, Chia Huei; Gasic, Ivana; Huber-Reggi, Sabina P; Dudka, Damian; Barisic, Marin; Maiato, Helder; Meraldi, Patrick

    2015-01-01

    Chromosome alignment in the middle of the bipolar spindle is a hallmark of metazoan cell divisions. When we offset the metaphase plate position by creating an asymmetric centriole distribution on each pole, we find that metaphase plates relocate to the middle of the spindle before anaphase. The spindle assembly checkpoint enables this centering mechanism by providing cells enough time to correct metaphase plate position. The checkpoint responds to unstable kinetochore-microtubule attachments resulting from an imbalance in microtubule stability between the two half-spindles in cells with an asymmetric centriole distribution. Inactivation of the checkpoint prior to metaphase plate centering leads to asymmetric cell divisions and daughter cells of unequal size; in contrast, if the checkpoint is inactivated after the metaphase plate has centered its position, symmetric cell divisions ensue. This indicates that the equatorial position of the metaphase plate is essential for symmetric cell divisions. PMID:26188083

  3. Downregulation of cell division cycle 25 homolog C reduces the radiosensitivity and proliferation activity of esophageal squamous cell carcinoma.

    Science.gov (United States)

    Yin, Yachao; Dou, Xiaoyan; Duan, Shimiao; Zhang, Lei; Xu, Quanjing; Li, Hongwei; Li, Duojie

    2016-09-30

    Radiation therapy is one of the most important methods of contemporary cancer treatment. Cells in the G2 and M phases are more sensitive to radiation therapy, and cell division cycle 25 homolog C (CDC25C) is essential in shifting the cell cycle between these two phases. In this study, the knockdown of CDC25C in human esophageal squamous carcinoma EC9706 cells was mediated by transfecting shRNA against human CDC25C-subcloning into pGV248. The levels of CDC25C mRNA and protein expression were assessed by reverse transcription-polymerase chain reaction (RT-PCR) and western blotting, respectively. Moreover, cell proliferation and radiosensitivity were measured. Stable CDC25C-knockdown EC9706 cell lines were successfully established. Furthermore, the proliferation of both control and CDC25C-shRNA-EC9706 cells was inhibited after the cells were treated with increasing X-ray doses, and the proliferation of the control cells was affected more significantly (p<0.05). Moreover, cell colony formation assays allowed us to reach the same conclusion. Taken together, our experiments demonstrated that the knockdown of CDC25C can reduce both the radiotherapy sensitivity and the proliferation activity of EC9706 cells. Thus, CDC25C might be a potential biomarker for radiotherapy treatment. PMID:27188256

  4. Cell division plane orientation based on tensile stress in Arabidopsis thaliana

    Science.gov (United States)

    Louveaux, Marion; Julien, Jean-Daniel; Mirabet, Vincent; Boudaoud, Arezki; Hamant, Olivier

    2016-01-01

    Cell geometry has long been proposed to play a key role in the orientation of symmetric cell division planes. In particular, the recently proposed Besson–Dumais rule generalizes Errera’s rule and predicts that cells divide along one of the local minima of plane area. However, this rule has been tested only on tissues with rather local spherical shape and homogeneous growth. Here, we tested the application of the Besson–Dumais rule to the divisions occurring in the Arabidopsis shoot apex, which contains domains with anisotropic curvature and differential growth. We found that the Besson–Dumais rule works well in the central part of the apex, but fails to account for cell division planes in the saddle-shaped boundary region. Because curvature anisotropy and differential growth prescribe directional tensile stress in that region, we tested the putative contribution of anisotropic stress fields to cell division plane orientation at the shoot apex. To do so, we compared two division rules: geometrical (new plane along the shortest path) and mechanical (new plane along maximal tension). The mechanical division rule reproduced the enrichment of long planes observed in the boundary region. Experimental perturbation of mechanical stress pattern further supported a contribution of anisotropic tensile stress in division plane orientation. Importantly, simulations of tissues growing in an isotropic stress field, and dividing along maximal tension, provided division plane distributions comparable to those obtained with the geometrical rule. We thus propose that division plane orientation by tensile stress offers a general rule for symmetric cell division in plants. PMID:27436908

  5. Noise and Epigenetic Inheritance of Single-Cell Division Times Influence Population Fitness.

    Science.gov (United States)

    Cerulus, Bram; New, Aaron M; Pougach, Ksenia; Verstrepen, Kevin J

    2016-05-01

    The fitness effect of biological noise remains unclear. For example, even within clonal microbial populations, individual cells grow at different speeds. Although it is known that the individuals' mean growth speed can affect population-level fitness, it is unclear how or whether growth speed heterogeneity itself is subject to natural selection. Here, we show that noisy single-cell division times can significantly affect population-level growth rate. Using time-lapse microscopy to measure the division times of thousands of individual S. cerevisiae cells across different genetic and environmental backgrounds, we find that the length of individual cells' division times can vary substantially between clonal individuals and that sublineages often show epigenetic inheritance of division times. By combining these experimental measurements with mathematical modeling, we find that, for a given mean division time, increasing heterogeneity and epigenetic inheritance of division times increases the population growth rate. Furthermore, we demonstrate that the heterogeneity and epigenetic inheritance of single-cell division times can be linked with variation in the expression of catabolic genes. Taken together, our results reveal how a change in noisy single-cell behaviors can directly influence fitness through dynamics that operate independently of effects caused by changes to the mean. These results not only allow a better understanding of microbial fitness but also help to more accurately predict fitness in other clonal populations, such as tumors. PMID:27068419

  6. ParA and ParB coordinate chromosome segregation with cell elongation and division during Streptomyces sporulation.

    Science.gov (United States)

    Donczew, Magdalena; Mackiewicz, Paweł; Wróbel, Agnieszka; Flärdh, Klas; Zakrzewska-Czerwińska, Jolanta; Jakimowicz, Dagmara

    2016-04-01

    In unicellular bacteria, the ParA and ParB proteins segregate chromosomes and coordinate this process with cell division and chromosome replication. During sporulation of mycelial Streptomyces, ParA and ParB uniformly distribute multiple chromosomes along the filamentous sporogenic hyphal compartment, which then differentiates into a chain of unigenomic spores. However, chromosome segregation must be coordinated with cell elongation and multiple divisions. Here, we addressed the question of whether ParA and ParB are involved in the synchronization of cell-cycle processes during sporulation in Streptomyces To answer this question, we used time-lapse microscopy, which allows the monitoring of growth and division of single sporogenic hyphae. We showed that sporogenic hyphae stop extending at the time of ParA accumulation and Z-ring formation. We demonstrated that both ParA and ParB affect the rate of hyphal extension. Additionally, we showed that ParA promotes the formation of massive nucleoprotein complexes by ParB. We also showed that FtsZ ring assembly is affected by the ParB protein and/or unsegregated DNA. Our results indicate the existence of a checkpoint between the extension and septation of sporogenic hyphae that involves the ParA and ParB proteins. PMID:27248800

  7. Small molecule inhibitor of formin homology 2 domains (SMIFH2 reveals the roles of the formin family of proteins in spindle assembly and asymmetric division in mouse oocytes.

    Directory of Open Access Journals (Sweden)

    Hak-Cheol Kim

    Full Text Available Dynamic actin reorganization is the main driving force for spindle migration and asymmetric cell division in mammalian oocytes. It has been reported that various actin nucleators including Formin-2 are involved in the polarization of the spindle and in asymmetric cell division. In mammals, the formin family is comprised of 15 proteins. However, their individual roles in spindle migration and/or asymmetric division have not been elucidated yet. In this study, we employed a newly developed inhibitor for formin family proteins, small molecule inhibitor of formin homology 2 domains (SMIFH2, to assess the functions of the formin family in mouse oocyte maturation. Treatment with SMIFH2 during in vitro maturation of mouse oocytes inhibited maturation by decreasing cytoplasmic and cortical actin levels. In addition, treatment with SMIFH2, especially at higher concentrations (500 μM, impaired the proper formation of meiotic spindles, indicating that formins play a role in meiotic spindle formation. Knockdown of the mDia2 formins caused a similar decrease in oocyte maturation and abnormal spindle morphology, mimicking the phenotype of SMIFH2-treated cells. Collectively, these results suggested that besides Formin-2, the other proteins of the formin, including mDia family play a role in asymmetric division and meiotic spindle formation in mammalian oocytes.

  8. The protective role of symmetric stem cell division on the accumulation of heritable damage.

    Directory of Open Access Journals (Sweden)

    Peter T McHale

    2014-08-01

    Full Text Available Stem cell divisions are either asymmetric-in which one daughter cell remains a stem cell and one does not-or symmetric, in which both daughter cells adopt the same fate, either stem or non-stem. Recent studies show that in many tissues operating under homeostatic conditions stem cell division patterns are strongly biased toward the symmetric outcome, raising the question of whether symmetry confers some benefit. Here, we show that symmetry, via extinction of damaged stem-cell clones, reduces the lifetime risk of accumulating phenotypically silent heritable damage (mutations or aberrant epigenetic changes in individual stem cells. This effect is greatest in rapidly cycling tissues subject to accelerating rates of damage accumulation over time, a scenario that describes the progression of many cancers. A decrease in the rate of cellular damage accumulation may be an important factor favoring symmetric patterns of stem cell division.

  9. The Protective Role of Symmetric Stem Cell Division on the Accumulation of Heritable Damage

    Science.gov (United States)

    McHale, Peter T.; Lander, Arthur D.

    2014-01-01

    Stem cell divisions are either asymmetric—in which one daughter cell remains a stem cell and one does not—or symmetric, in which both daughter cells adopt the same fate, either stem or non-stem. Recent studies show that in many tissues operating under homeostatic conditions stem cell division patterns are strongly biased toward the symmetric outcome, raising the question of whether symmetry confers some benefit. Here, we show that symmetry, via extinction of damaged stem-cell clones, reduces the lifetime risk of accumulating phenotypically silent heritable damage (mutations or aberrant epigenetic changes) in individual stem cells. This effect is greatest in rapidly cycling tissues subject to accelerating rates of damage accumulation over time, a scenario that describes the progression of many cancers. A decrease in the rate of cellular damage accumulation may be an important factor favoring symmetric patterns of stem cell division. PMID:25121484

  10. Cell division of cycle of Bacillus subtilis: evidence of variability in period D.

    OpenAIRE

    Holmes, M.; Rickert, M; Pierucci, O

    1980-01-01

    In Bacillus subtilis the deoxyribonucleic acid content and the extent of cell division during inhibition of chromosome replication increased as a function of the average cell mass, independent of the growth rate. At each growth rate, mass, deoxyribonucleic acid, and residual division varied in different cultures. The variation is consistent with a large variability in the D period. At growth rates higher than 1.5 doublings per h at 37 degrees C, the change in D accounts for the growth rate de...

  11. Effects of brevetoxins on murine myeloma SP2/O cells: aberrant cellular division.

    Science.gov (United States)

    Han, Thomas K; Derby, Melissa; Martin, Dean F; Wright, Scott D; Dao, My Lien

    2003-01-01

    Massive deaths of manatees (Trichechus manatus latirostris) during the red tide seasons have been attributed to brevetoxins produced by the dinoflagellate Karenia brevis (formerly Ptychodiscus breve and Gymnodinium breve). Although these toxins have been found in macrophages and lymphocytes in the lung, liver, and secondary lymphoid tissues of these animals, the molecular mechanisms of brevetoxicosis have not yet been identified. To investigate the effects of brevetoxins on immune cells, a murine myeloma cell line (SP2/O) was used as a model for in vitro studies. By adding brevetoxins to cultures of the SP2/O cells at concentrations ranging from 20 to 600 ng/ml, an apparent increase in proliferation was observed at around 2 hours post challenge as compared to the unchallenged cell cultures. This was followed by a drop in cell number at around 3 hours, suggesting an aberrant effect of brevetoxins on cellular division, the cells generated at 2 hours being apparently short-lived. In situ immunochemical staining of the SP2/O cells at 1 and 2 hour post challenge showed an accumulation of the toxins in the nucleus. A 21-kDa protein was subsequently isolated from the SP2/O cells as having brevetoxin-binding properties, and immunologically identified as p21, a nuclear factor known to down-regulate cellular proliferation through inhibition of cyclin-dependent kinases. These data are the first on a possible effect of brevetoxins on the cell cycle via binding to p21, a phenomenon that needs to be further investigated and validated in normal immune cells. PMID:12745987

  12. Tumor-initiating label-retaining cancer cells in human gastrointestinal cancers undergo asymmetric cell division.

    Science.gov (United States)

    Xin, Hong-Wu; Hari, Danielle M; Mullinax, John E; Ambe, Chenwi M; Koizumi, Tomotake; Ray, Satyajit; Anderson, Andrew J; Wiegand, Gordon W; Garfield, Susan H; Thorgeirsson, Snorri S; Avital, Itzhak

    2012-04-01

    Label-retaining cells (LRCs) have been proposed to represent adult tissue stem cells. LRCs are hypothesized to result from either slow cycling or asymmetric cell division (ACD). However, the stem cell nature and whether LRC undergo ACD remain controversial. Here, we demonstrate label-retaining cancer cells (LRCCs) in several gastrointestinal (GI) cancers including fresh surgical specimens. Using a novel method for isolation of live LRCC, we demonstrate that a subpopulation of LRCC is actively dividing and exhibits stem cells and pluripotency gene expression profiles. Using real-time confocal microscopic cinematography, we show live LRCC undergoing asymmetric nonrandom chromosomal cosegregation LRC division. Importantly, LRCCs have greater tumor-initiating capacity than non-LRCCs. Based on our data and that cancers develop in tissues that harbor normal-LRC, we propose that LRCC might represent a novel population of GI stem-like cancer cells. LRCC may provide novel mechanistic insights into the biology of cancer and regenerative medicine and present novel targets for cancer treatment. PMID:22331764

  13. Cyclin B synthesis and rapamycin-sensitive regulation of protein synthesis during starfish oocyte meiotic divisions.

    Science.gov (United States)

    Lapasset, Laure; Pradet-Balade, Bérengère; Vergé, Valérie; Lozano, Jean-Claude; Oulhen, Nathalie; Cormier, Patrick; Peaucellier, Gérard

    2008-11-01

    Translation of cyclin mRNAs represents an important event for proper meiotic maturation and post-fertilization mitoses in many species. Translational control of cyclin B mRNA has been described to be achieved through two separate but related mechanisms: translational repression and polyadenylation. In this paper, we evaluated the contribution of global translational regulation by the cap-dependent translation repressor 4E-BP (eukaryotic initiation factor 4E-binding protein) on the cyclin B protein synthesis during meiotic maturation of the starfish oocytes. We used the immunosupressant drug rapamycin, a strong inhibitor of cap-dependent translation, to check for the involvement of this protein synthesis during this physiological process. Rapamycin was found to prevent dissociation of 4E-BP from the initiation factor eIF4E and to suppress correlatively a burst of global protein synthesis occurring at the G2/M transition. The drug had no effect on first meiotic division but defects in meiotic spindle formation prevented second polar body emission, demonstrating that a rapamycin-sensitive pathway is involved in this mechanism. While rapamycin affected the global protein synthesis, the drug altered neither the specific translation of cyclin B mRNA nor the expression of the Mos protein. The expression of these two proteins was correlated with the phosphorylation and the dissociation of the cytoplasmic polyadenylation element-binding protein from eIF4E. PMID:18361417

  14. Study of the mechanism of diatom cell division by means of 29Si isotope tracing

    International Nuclear Information System (INIS)

    Diatoms are delicate unicellular organisms enclosed in a silica frustule, that is made up of two valves. Multiplication of the diatoms occurs by ordinary mitotic cell division. During cell division each cell produces two daughter cells, each of them keeping one of the two valves of the mother cell and producing a new valve by absorbing the silicon present in the environment. The NanoSIMS 50 allows ion imaging to be performed on diatoms in order to determine the site of fixation of silicon. The aim of this study was to observe and compare the mechanism of the construction of the new valve after cell division. To this end, different types of diatoms have been transferred in a culture medium enriched with 29Si and after several days, the distribution of the different isotopes of silicon has been determined by NanoSIMS50 imaging. The construction of new valves has been observed and the isotopic ratio has been determined

  15. System X supercomputer provides super tool for simulation of cell division

    OpenAIRE

    Trulove, Susan

    2007-01-01

    Virginia Tech researchers in computer science and biology have used the university's supercomputer, System X, to create models and algorithms that make it possible to simulate the cell cycle -- the processes leading to cell division. They have demonstrated that the new mathematical models and numerical algorithms provide powerful tools for studying the complex processes going on inside living cells.

  16. From cell differentiation to cell collectives: Bacillus subtilis uses division of labor to migrate.

    Directory of Open Access Journals (Sweden)

    Jordi van Gestel

    2015-04-01

    Full Text Available The organization of cells, emerging from cell-cell interactions, can give rise to collective properties. These properties are adaptive when together cells can face environmental challenges that they separately cannot. One particular challenge that is important for microorganisms is migration. In this study, we show how flagellum-independent migration is driven by the division of labor of two cell types that appear during Bacillus subtilis sliding motility. Cell collectives organize themselves into bundles (called "van Gogh bundles" of tightly aligned cell chains that form filamentous loops at the colony edge. We show, by time-course microscopy, that these loops migrate by pushing themselves away from the colony. The formation of van Gogh bundles depends critically on the synergistic interaction of surfactin-producing and matrix-producing cells. We propose that surfactin-producing cells reduce the friction between cells and their substrate, thereby facilitating matrix-producing cells to form bundles. The folding properties of these bundles determine the rate of colony expansion. Our study illustrates how the simple organization of cells within a community can yield a strong ecological advantage. This is a key factor underlying the diverse origins of multicellularity.

  17. Real-Time Lineage Analysis to Study Cell Division Orientation in the Arabidopsis Shoot Meristem.

    Science.gov (United States)

    Tobin, Cory J; Meyerowitz, Elliot M

    2016-01-01

    Cells in the Arabidopsis shoot apical meristem are small and divide frequently throughout the life-time of the organism making them good candidates for studying the mechanisms of cell division in plants. But tracking these cell divisions requires multiple images to be taken of the same specimen over time which means the specimen must stay alive throughout the process. This chapter provides details on how to prepare plants for live imaging, keep them alive and growing through multiple time points, and how to process the data to extract cell boundary coordinates from three-dimensional images. PMID:26659961

  18. Interaction of hyperthermia and radiation on the induction of division delay in Chinese hamster ovary cells

    International Nuclear Information System (INIS)

    The mitotic selection procedure for cell cycle analysis was used in the investigation of the interaction of hyperthermia and ionizing radiation on the induction and duration of division delay in Chinese hamster ovary cells. Hyperthermia (immersion in a 45 degrees C water bath) produced a blockade of cell cycle progression with a transition point in late G2-early M, approximately at the X-ray transition point (35 min prior to selection). The duration of division delay for heated cells depended on the time of immersion: 24 minutes/minute at 45 degrees C. Radiation-induced division delay occurred at a rate of 45 minutes/gray of X-irradiation. When hyperthermic exposure and X-irradiation were combined with less than 1 minute between treatments, a division delay resulted that was approximately the sum of the delays produced by the individual treatments. As the interval between treatments was increased, the overall division delay also increased beyond that which could be accounted for solely by the postponement of the second treatment. These results indicate that hyperthermia and radiation induce division delay by different mechanisms

  19. Cell division patterns and chromosomal segregation defects in oral cancer stem cells.

    Science.gov (United States)

    Kaseb, Hatem O; Lewis, Dale W; Saunders, William S; Gollin, Susanne M

    2016-09-01

    Oral squamous cell carcinoma (OSCC) is a serious public health problem caused primarily by smoking and alcohol consumption or human papillomavirus. The cancer stem cell (CSC) theory posits that CSCs show unique characteristics, including self-renewal and therapeutic resistance. Examining biomarkers and other features of CSCs is critical to better understanding their biology. To this end, the results show that cellular SOX2 immunostaining correlates with other CSC biomarkers in OSCC cell lines and marks the rare CSC population. To assess whether CSC division patterns are symmetrical, resulting in two CSC, or asymmetrical, leading to one CSC and one cancer cell, cell size and fluorescence intensity of mitotic cells stained with SOX2 were analyzed. Asymmetrical SOX2 distribution in ≈25% of the mitoses analyzed was detected. Chromosomal instability, some of which is caused by chromosome segregation defects (CSDs), is a feature of cancer cells that leads to altered gene copy numbers. We compare chromosomal instability (as measured by CSDs) between CSCs (SOX2+) and non-CSCs (SOX2-) from the same OSCC cell lines. CSDs were more common in non-CSCs (SOX2-) than CSCs (SOX2+) and in symmetrical CSC (SOX2+) mitotic pairs than asymmetrical CSC (SOX2+/SOX2-) mitotic pairs. CSCs showed fewer and different types of CSDs after ionizing radiation treatment than non-CSCs. Overall, these data are the first to demonstrate both symmetrical and asymmetrical cell divisions with CSDs in OSCC CSC. Further, the results suggest that CSCs may undergo altered behavior, including therapeutic resistance as a result of chromosomal instability due to chromosome segregation defects. © 2016 Wiley Periodicals, Inc. PMID:27123539

  20. Temperature gradient stimulation for cell division in C. Elegans Embryos on chip

    OpenAIRE

    Baranek, Sophie; Bezler, Alexandra; Adamczyk, Christian; Gönczy, Pierre; Renaud, Philippe

    2010-01-01

    This paper reports on a new microfluidic device for temperature stimulation of cell in in-vitro culture. Micro-electrodes in a meander shape are embedded into the microfluidic channels to generate either a temperature gradient through the culture chamber or a local heat spot under specific cells. One promising application is the control of cell di- vision rate. Here we present first results of the synchronization of cell division in a two-cell stage embryos of C. Elegans.

  1. Expression of the wild-type p53 antioncogene induces guanine nucleotide-dependent stem cell division kinetics.

    OpenAIRE

    Sherley, J L; Stadler, P B; Johnson, D. R.

    1995-01-01

    The predominant type of cell division in adult mammals is renewal growth. Renewing stem cells in somatic tissues undergo continuous asymmetric divisions. One new daughter cell retains the division potential of the original stem cell, while the other differentiates into a functional constituent of the tissue. Disruptions of this process lead to the development of human cancers. We show that through a guanine nucleotide-dependent mechanism, the p53 antioncogene can induce exponentially dividing...

  2. Structural and Functional Characterizations of SsgB, a Conserved Activator of Developmental Cell Division in Morphologically Complex Actinomycetes

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Qingping; Traag, Bjørn A.; Willemse, Joost; McMullan, Daniel; Miller, Mitchell D.; Elsliger, Marc-André; Abdubek, Polat; Astakhova, Tamara; Axelrod, Herbert L.; Bakolitsa, Constantina; Carlton, Dennis; Chen, Connie; Chiu, Hsiu-Ju; Chruszcz, Maksymilian; Clayton, Thomas; Das, Debanu; Deller, Marc C.; Duan, Lian; Ellrott, Kyle; Ernst, Dustin; Farr, Carol L.; Feuerhelm, Julie; Grant, Joanna C.; Grzechnik, Anna; Grzechnik, Slawomir K.; Han, Gye Won; Jaroszewski, Lukasz; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Kozbial, Piotr; Krishna, S. Sri; Kumar, Abhinav; Marciano, David; Minor, Wladek; Mommaas, A. Mieke; Morse, Andrew T.; Nigoghossian, Edward; Nopakun, Amanda; Okach, Linda; Oommachen, Silvya; Paulsen, Jessica; Puckett, Christina; Reyes, Ron; Rife, Christopher L.; Sefcovic, Natasha; Tien, Henry J.; Trame, Christine B.; van den Bedem, Henry; Wang, Shuren; Weekes, Dana; Hodgson, Keith O.; Wooley, John; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wilson, Ian A.; van Wezel, Gilles P.; (Leiden-MC); (SLAC); (Scripps); (UV); (UCSD); (Burnham)

    2010-01-20

    SsgA-like proteins (SALPs) are a family of homologous cell division-related proteins that occur exclusively in morphologically complex actinomycetes. We show that SsgB, a subfamily of SALPs, is the archetypal SALP that is functionally conserved in all sporulating actinomycetes. Sporulation-specific cell division of Streptomyces coelicolor ssgB mutants is restored by introduction of distant ssgB orthologues from other actinomycetes. Interestingly, the number of septa (and spores) of the complemented null mutants is dictated by the specific ssgB orthologue that is expressed. The crystal structure of the SsgB from Thermobifida fusca was determined at 2.6 {angstrom} resolution and represents the first structure for this family. The structure revealed similarities to a class of eukaryotic 'whirly' single-stranded DNA/RNA-binding proteins. However, the electro-negative surface of the SALPs suggests that neither SsgB nor any of the other SALPs are likely to interact with nucleotide substrates. Instead, we show that a conserved hydrophobic surface is likely to be important for SALP function and suggest that proteins are the likely binding partners.

  3. Symmetric cell division in pseudohyphae of the yeast Saccharomyces cerevisiae.

    OpenAIRE

    Kron, S J; Styles, C. A.; Fink, G R

    1994-01-01

    Laboratory strains of Saccharomyces cerevisiae are dimorphic; in response to nitrogen starvation they switch from a yeast form (YF) to a filamentous pseudohyphal (PH) form. Time-lapse video microscopy of dividing cells reveals that YF and PH cells differ in their cell cycles and budding polarity. The YF cell cycle is controlled at the G1/S transition by the cell-size checkpoint Start. YF cells divide asymmetrically, producing small daughters from full-sized mothers. As a result, mothers and d...

  4. Phylogeography, Salinity Adaptations and Metabolic Potential of the Candidate Division KB1 Bacteria Based on a Partial Single Cell Genome.

    Science.gov (United States)

    Nigro, Lisa M; Hyde, Andrew S; MacGregor, Barbara J; Teske, Andreas

    2016-01-01

    Deep-sea hypersaline anoxic basins and other hypersaline environments contain abundant and diverse microbial life that has adapted to these extreme conditions. The bacterial Candidate Division KB1 represents one of several uncultured groups that have been consistently observed in hypersaline microbial diversity studies. Here we report the phylogeography of KB1, its phylogenetic relationships to Candidate Division OP1 Bacteria, and its potential metabolic and osmotic stress adaptations based on a partial single cell amplified genome of KB1 from Orca Basin, the largest hypersaline seafloor brine basin in the Gulf of Mexico. Our results are consistent with the hypothesis - previously developed based on (14)C incorporation experiments with mixed-species enrichments from Mediterranean seafloor brines - that KB1 has adapted its proteins to elevated intracellular salinity, but at the same time KB1 apparently imports glycine betaine; this compatible solute is potentially not limited to osmoregulation but could also serve as a carbon and energy source. PMID:27597842

  5. LIN-39/Hox triggers cell division and represses EFF-1/fusogen-dependent vulval cell fusion

    OpenAIRE

    Shemer, Gidi; Podbilewicz, Benjamin

    2002-01-01

    General mechanisms by which Hox genes establish cell fates are known. However, a few Hox effectors mediating cell behaviors have been identified. Here we found the first effector of LIN-39/HoxD4/Dfd in Caenorhabditis elegans. In specific vulval precursor cells (VPCs), LIN-39 represses early and late expression of EFF-1, a membrane protein essential for cell fusion. Repression of eff-1 is also achieved by the activity of CEH-20/Exd/Pbx, a known cofactor of Hox proteins. Unfused VPCs in lin-39(...

  6. A P-Loop NTPase Regulates Quiescent Center Cell Division and Distal Stem Cell Identity through the Regulation of ROS Homeostasis in Arabidopsis Root.

    Science.gov (United States)

    Yu, Qianqian; Tian, Huiyu; Yue, Kun; Liu, Jiajia; Zhang, Bing; Li, Xugang; Ding, Zhaojun

    2016-09-01

    Reactive oxygen species (ROS) are recognized as important regulators of cell division and differentiation. The Arabidopsis thaliana P-loop NTPase encoded by APP1 affects root stem cell niche identity through its control of local ROS homeostasis. The disruption of APP1 is accompanied by a reduction in ROS level, a rise in the rate of cell division in the quiescent center (QC) and the promotion of root distal stem cell (DSC) differentiation. Both the higher level of ROS induced in the app1 mutant by exposure to methyl viologen (MV), and treatment with hydrogen peroxide (H2O2) rescued the mutant phenotype, implying that both the increased rate of cell division in the QC and the enhancement in root DSC differentiation can be attributed to a low level of ROS. APP1 is expressed in the root apical meristem cell mitochondria, and its product is associated with ATP hydrolase activity. The key transcription factors, which are defining root distal stem niche, such as SCARECROW (SCR) and SHORT ROOT (SHR) are both significantly down-regulated at both the transcriptional and protein level in the app1 mutant, indicating that SHR and SCR are important downstream targets of APP1-regulated ROS signaling to control the identity of root QC and DSCs. PMID:27583367

  7. Scaling laws governing stochastic growth and division of single bacterial cells

    CERN Document Server

    Iyer-Biswas, Srividya; Henry, Jonathan T; Lo, Klevin; Burov, Stanislav; Lin, Yihan; Crooks, Gavin E; Crosson, Sean; Dinner, Aaron R; Scherer, Norbert F

    2014-01-01

    Uncovering the quantitative laws that govern the growth and division of single cells remains a major challenge. Using a unique combination of technologies that yields unprecedented statistical precision, we find that the sizes of individual Caulobacter crescentus cells increase exponentially in time. We also establish that they divide upon reaching a critical multiple ($\\approx$1.8) of their initial sizes, rather than an absolute size. We show that when the temperature is varied, the growth and division timescales scale proportionally with each other over the physiological temperature range. Strikingly, the cell-size and division-time distributions can both be rescaled by their mean values such that the condition-specific distributions collapse to universal curves. We account for these observations with a minimal stochastic model that is based on an autocatalytic cycle. It predicts the scalings, as well as specific functional forms for the universal curves. Our experimental and theoretical analysis reveals a ...

  8. Cell division licensing in the multi-chromosomal Vibrio cholerae bacterium.

    Science.gov (United States)

    Galli, Elisa; Poidevin, Mickaël; Le Bars, Romain; Desfontaines, Jean-Michel; Muresan, Leila; Paly, Evelyne; Yamaichi, Yoshiharu; Barre, François-Xavier

    2016-01-01

    Cell division must be coordinated with chromosome replication and segregation to ensure the faithful transmission of genetic information during proliferation. In most bacteria, assembly of the division apparatus, the divisome, starts with the polymerization of a tubulin homologue, FtsZ, into a ring-like structure at mid-cell, the Z-ring(1). It typically occurs at half of the cell cycle when most of the replication and segregation cycle of the unique chromosome they generally harbour is achieved(2). The chromosome itself participates in the regulation of cell division, at least in part because it serves as a scaffold to position FtsZ polymerization antagonists(3). However, about 10% of bacteria have more than one chromosome(4), which raises questions about the way they license cell division(3). For instance, the genome of Vibrio cholerae, the agent of cholera, is divided between a 3 Mbp replicon that originates from the chromosome of its mono-chromosomal ancestor, Chr1, and a 1 Mbp plasmid-derived replicon, Chr2 (ref. 5). Here, we show that Chr2 harbours binding motifs for an inhibitor of Z-ring formation, which helps accurately position the V. cholerae divisome at mid-cell and postpones its assembly to the very end of the cell cycle. PMID:27562255

  9. Asymmetric cell division and its role in cell fate determination in the green alga Tetraselmis indica

    Indian Academy of Sciences (India)

    Mani Arora; Arga Chandrashekar Anil; Karl Burgess; Jane Delany; Ehsan Mesbahi

    2015-12-01

    The prasinophytes (early diverging Chlorophyta), consisting of simple unicellular green algae, occupy a critical position at the base of the green algal tree of life, with some of its representatives viewed as the cell form most similar to the first green alga, the `ancestral green flagellate'. Relatively large-celled unicellular eukaryotic phytoflagellates (such as Tetraselmis and Scherffelia), traditionally placed in Prasinophyceae but now considered as members of Chlorodendrophyceae (core Chlorophyta), have retained some primitive characteristics of prasinophytes. These organisms share several ultrastructural features with the other core chlorophytes (Trebouxiophyceae, Ulvophyceae and Chlorophyceae). However, the role of Chlorodendrophycean algae as the evolutionary link between cellular individuality and cellular cooperation has been largely unstudied. Here, we show that clonal populations of a unicellular chlorophyte, Tetraselmis indica, consist of morphologically and ultrastructurally variant cells which arise through asymmetric cell division. These cells also differ in their physiological properties. The structural and physiological differences in the clonal cell population correlate to a certain extent with the longevity and function of cells.

  10. Pseudomonas aeruginosa Transmigrates at Epithelial Cell-Cell Junctions, Exploiting Sites of Cell Division and Senescent Cell Extrusion.

    Directory of Open Access Journals (Sweden)

    Guillaume Golovkine

    2016-01-01

    Full Text Available To achieve systemic infection, bacterial pathogens must overcome the critical and challenging step of transmigration across epithelial barriers. This is particularly true for opportunistic pathogens such as Pseudomonas aeruginosa, an agent which causes nosocomial infections. Despite extensive study, details on the mechanisms used by this bacterium to transmigrate across epithelial tissues, as well as the entry sites it uses, remain speculative. Here, using real-time microscopy and a model epithelial barrier, we show that P. aeruginosa employs a paracellular transmigration route, taking advantage of altered cell-cell junctions at sites of cell division or when senescent cells are expelled from the cell layer. Once a bacterium transmigrates, it is followed by a cohort of bacteria using the same entry point. The basal compartment is then invaded radially from the initial penetration site. Effective transmigration and propagation require type 4 pili, the type 3 secretion system (T3SS and a flagellum, although flagellum-deficient bacteria can occasionally invade the basal compartment from wounded areas. In the basal compartment, the bacteria inject the T3SS toxins into host cells, disrupting the cytoskeleton and focal contacts to allow their progression under the cells. Thus, P. aeruginosa exploits intrinsic host cell processes to breach the epithelium and invade the subcellular compartment.

  11. 3′ UTR-Dependent, miR-92-Mediated Restriction of Tis21 Expression Maintains Asymmetric Neural Stem Cell Division to Ensure Proper Neocortex Size

    Directory of Open Access Journals (Sweden)

    Ji-Feng Fei

    2014-04-01

    Full Text Available Mammalian neocortex size primarily reflects the number and mode of divisions of neural stem and progenitor cells. Cortical stem cells (apical progenitors switching from symmetric divisions, which expand their population, to asymmetric divisions, which generate downstream neuronal progenitors (basal progenitors, start expressing Tis21, a so-called antiproliferative/prodifferentiative gene. Tis21 encodes a small (17.5 kDa, functionally poorly characterized protein and a relatively large (2 kb, highly conserved 3′ UTR. Here, we show that mice lacking the Tis21 3′ UTR develop a microcephalic neocortex with fewer neurons, notably in the upper layers. This reflects a progressive decrease in basal progenitors, which in turn is due to a fraction of apical progenitors prematurely switching from asymmetric self-renewing to symmetric self-consuming divisions. This switch is caused by the markedly increased Tis21 protein level resulting from lack of microRNA-, notably miR-92-, dependent restriction of Tis21 expression. Our data show that a premature onset of consumptive neural stem cell divisions can lead to microcephaly.

  12. Vegetative Cell Division and Nuclear Translocation in Three Algae Species of Netrium (Zygnematales, Chlorophyta

    Directory of Open Access Journals (Sweden)

    DIAN HENDRAYANTI

    2006-03-01

    Full Text Available Three species of Netrium oblongum, N. digitus v. latum, and N. interruptum were studied for their mode in the vegetative cell division and nuclear translocation during mitosis using light and fluorescence microscopy. The process of cell division in the three species began with the prominent constriction at the chloroplast in both semicells about half way from the apex. The constriction of chloroplast was mostly visible in N. digitus v. latum. Soon after nucleus divided, septum was formed across the cell and cytokinesis occurred. Observation with fluorescence microscope showed that the movement of nucleus moved back into the center of daughter cells was not always synchronous. Division of chloroplast in N. oblongum and N. digitus v. latum were different with that of N. interruptum. Chloroplast division in two former species occured following the movement of the nucleus down semicell. However, in N. interruptum, chloroplast divided later after nucleus occupied the position at the center of the daughter cells. Cell restoration started after the completion of mitosis and cytokinesis.

  13. Electromagnetic field influences on cell surface potential and cell division in saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    The effect of electromagnetic field on cell surface potential and cell division were studied in s.cerevisiae. The strains used were, GM3 (a/gal 10,trp1, ura4, met 8, ade 5,7,les1, ilvl,arol D, suc-mal, cupr.)and ural (a/urap+w-c 321, R E 221, R) an electromagnetic field (h) .O.I.T, cell resistance (R) increased from 0.158 MΩ to 0.200 M Ω through 5 min. The magnetic field (MF) were switching off. The resistance spontaneously increased reaching 1.000 M Ω at the 9 Th min. However, slowly decrease occurred and reaching 0.560 M Omega at the 15 Th min. By using the MF after 15 min., the resistance value reaching 0.180 M OMEGA, through 15-25 min and cell potential (V) ranged between 130-240 mV. Cell culture, of two strains (same mating type) was used, the resistance, R., was 4000 M Ω and V; 600 mV with two cycles min, R; reached 3200 M Ω. On further cycle of (H) led to a huge sudden decrease of R; 0.176 M Ω the cell numbers were depended, upon the cell potential, due to the application of (H). For the first strain used, cell number decreased from 2x106 cells/ml to 1.5x106 cells/ml and from 2.1x108 cells/ml to 1.7x108 cells/ml after 5 min exposure to (H) for culture incubated at 30 degree on log and stationary phases respectively. While, the cell number in ural was decreased from 3.5x106 cells/ml and from 1.78x108 cells/ ml. to 1.71x108 cells/ml through 5 min exposure to (H) for culture incubated at 30 degree on log and stationary phases respectively

  14. The asymmetric segregation of damaged proteins is stem cell-type dependent.

    Science.gov (United States)

    Bufalino, Mary Rose; DeVeale, Brian; van der Kooy, Derek

    2013-05-13

    Asymmetric segregation of damaged proteins (DPs) during mitosis has been linked in yeast and bacteria to the protection of one cell from aging. Recent evidence suggests that stem cells may use a similar mechanism; however, to date there is no in vivo evidence demonstrating this effect in healthy adult stem cells. We report that stem cells in larval (neuroblast) and adult (female germline and intestinal stem cell) Drosophila melanogaster asymmetrically segregate DPs, such as proteins with the difficult-to-degrade and age-associated 2,4-hydroxynonenal (HNE) modification. Surprisingly, of the cells analyzed only the intestinal stem cell protects itself by segregating HNE to differentiating progeny, whereas the neuroblast and germline stem cells retain HNE during division. This led us to suggest that chronological life span, and not cell type, determines the amount of DPs a cell receives during division. Furthermore, we reveal a role for both niche-dependent and -independent mechanisms of asymmetric DP division. PMID:23649805

  15. Investigating the Molecular Mechanism of TSO1 Function in Arabidopsis cell division and meristem development

    Energy Technology Data Exchange (ETDEWEB)

    Zhongchi Liu

    2004-10-01

    Unlike animals, plants are constantly exposed to environmental mutagens including ultraviolet light and reactive oxygen species. Further, plant cells are totipotent with highly plastic developmental programs. An understanding of molecular mechanisms underlying the ability of plants to monitor and repair its DNA and to eliminate damaged cells are of great importance. Previously we have identified two genes, TSO1 and TSO2, from a flowering plant Arabidopsis thaliana. Mutations in these two genes cause callus-like flowers, fasciated shoot apical meristems, and abnormal cell division, indicating that TSO1 and TSO2 may encode important cell cycle regulators. Previous funding from DOE led to the molecular cloning of TSO1, which was shown to encode a novel nuclear protein with two CXC domains suspected to bind DNA. This DOE grant has allowed us to characterize and isolate TSO2 that encodes the small subunit of the ribonucleotide reductase (RNR). RNR comprises two large subunits (R1) an d two small subunits (R2), catalyzes a rate-limiting step in the production of deoxyribonucleotides needed for DNA replication and repair. Previous studies in yeast and mammals indicated that defective RNR often led to cell cycle arrest, growth retardation and p53-dependent apoptosis while abnormally elevated RNR activities led to higher mutation rates. Subsequently, we identified two additional R2 genes, R2A and R2B in the Arabidopsis genome. Using reverse genetics, mutations in R2A and R2B were isolated, and double and triple mutants among the three R2 genes (TSO2, R2A and R2B) were constructed and analyzed. We showed that Arabidopsis tso2 mutants, with reduced dNTP levels, were more sensitive to UV-C. While r2a or r2b single mutants did not exhibit any phenotypes, tso2 r2b double mutants were embryonic lethal and tso2 r2a double mutants were seedling lethal indicating redundant functions among the three R2 genes. Furthermore, tso2 r2a double mutants exhibited increased DNA dam age

  16. Fission yeast Nod1 is a component of cortical nodes involved in cell size control and division site placement.

    Directory of Open Access Journals (Sweden)

    Isabelle Jourdain

    Full Text Available Most cells enter mitosis once they have reached a defined size. In the fission yeast Schizosaccharomyces pombe, mitotic entry is orchestrated by a geometry-sensing mechanism that involves the Cdk1/Cdc2-inhibiting Wee1 kinase. The factors upstream of Wee1 gather together in interphase to form a characteristic medial and cortical belt of nodes. Nodes are also considered to be precursors of the cytokinesis contractile actomyosin ring (CAR. Here we describe a new component of the interphase nodes and cytokinesis rings, which we named Nod1. Consistent with its role in cell size control at division, nod1Δ cells were elongated and epistatic with regulators of Wee1. Through biochemical and localisation studies, we placed Nod1 in a complex with the Rho-guanine nucleotide exchange factor Gef2. Nod1 and Gef2 mutually recruited each other in nodes and Nod1 also assembles Gef2 in rings. Like gef2Δ, nod1Δ cells showed a mild displacement of their division plane and this phenotype was severely exacerbated when the parallel Polo kinase pathway was also compromised. We conclude that Nod1 specifies the division site by localising Gef2 to the mitotic cell middle. Previous work showed that Gef2 in turn anchors factors that control the spatio-temporal recruitment of the actin nucleation machinery. It is believed that the actin filaments originated from the nodes pull nodes together into a single contractile ring. Surprisingly however, we found that node proteins could form pre-ring helical filaments in a cdc12-112 mutant in which nucleation of the actin ring is impaired. Furthermore, the deletion of either nod1 or gef2 created an un-expected situation where different ring components were recruited sequentially rather than simultaneously. At later stages of cytokinesis, these various rings appeared inter-fitted rather than merged. This study brings a new slant to the understanding of CAR assembly and function.

  17. Do Online Labs Work? An Assessment of an Online Lab on Cell Division

    Science.gov (United States)

    Gilman, Sharon L.

    2006-01-01

    Some studies show students successfully learning science through online courses. This study compared students doing an online and in-class lab exercise on cell division. Online students performed slightly but significantly better on a follow-up content quiz, however, about half those expressed a strong preference for in-class lab work.

  18. Intercellular Variability in Protein Levels from Stochastic Expression and Noisy Cell Cycle Processes

    Science.gov (United States)

    Soltani, Mohammad; Vargas-Garcia, Cesar A.; Antunes, Duarte; Singh, Abhyudai

    2016-01-01

    Inside individual cells, expression of genes is inherently stochastic and manifests as cell-to-cell variability or noise in protein copy numbers. Since proteins half-lives can be comparable to the cell-cycle length, randomness in cell-division times generates additional intercellular variability in protein levels. Moreover, as many mRNA/protein species are expressed at low-copy numbers, errors incurred in partitioning of molecules between two daughter cells are significant. We derive analytical formulas for the total noise in protein levels when the cell-cycle duration follows a general class of probability distributions. Using a novel hybrid approach the total noise is decomposed into components arising from i) stochastic expression; ii) partitioning errors at the time of cell division and iii) random cell-division events. These formulas reveal that random cell-division times not only generate additional extrinsic noise, but also critically affect the mean protein copy numbers and intrinsic noise components. Counter intuitively, in some parameter regimes, noise in protein levels can decrease as cell-division times become more stochastic. Computations are extended to consider genome duplication, where transcription rate is increased at a random point in the cell cycle. We systematically investigate how the timing of genome duplication influences different protein noise components. Intriguingly, results show that noise contribution from stochastic expression is minimized at an optimal genome-duplication time. Our theoretical results motivate new experimental methods for decomposing protein noise levels from synchronized and asynchronized single-cell expression data. Characterizing the contributions of individual noise mechanisms will lead to precise estimates of gene expression parameters and techniques for altering stochasticity to change phenotype of individual cells. PMID:27536771

  19. Building the perfect parasite: cell division in apicomplexa.

    Directory of Open Access Journals (Sweden)

    Boris Striepen

    2007-06-01

    Full Text Available Apicomplexans are pathogens responsible for malaria, toxoplasmosis, and crytposporidiosis in humans, and a wide range of livestock diseases. These unicellular eukaryotes are stealthy invaders, sheltering from the immune response in the cells of their hosts, while at the same time tapping into these cells as source of nutrients. The complexity and beauty of the structures formed during their intracellular development have made apicomplexans the darling of electron microscopists. Dramatic technological progress over the last decade has transformed apicomplexans into respectable genetic model organisms. Extensive genomic resources are now available for many apicomplexan species. At the same time, parasite transfection has enabled researchers to test the function of specific genes through reverse and forward genetic approaches with increasing sophistication. Transfection also introduced the use of fluorescent reporters, opening the field to dynamic real time microscopic observation. Parasite cell biologists have used these tools to take a fresh look at a classic problem: how do apicomplexans build the perfect invasion machine, the zoite, and how is this process fine-tuned to fit the specific niche of each pathogen in this ancient and very diverse group? This work has unearthed a treasure trove of novel structures and mechanisms that are the focus of this review.

  20. The ClpP protease homologue is required for the transmission traits and cell division of the pathogen Legionella pneumophila

    Directory of Open Access Journals (Sweden)

    Zhang Qin-fen

    2010-02-01

    Full Text Available Abstract Background Legionella pneumophila, the intracellular bacterial pathogen that causes Legionnaires' disease, exhibit characteristic transmission traits such as elevated stress tolerance, shortened length and virulence during the transition from the replication phase to the transmission phase. ClpP, the catalytic core of the Clp proteolytic complex, is widely involved in many cellular processes via the regulation of intracellular protein quality. Results In this study, we showed that ClpP was required for optimal growth of L. pneumophila at high temperatures and under several other stress conditions. We also observed that cells devoid of clpP exhibited cell elongation, incomplete cell division and compromised colony formation. Furthermore, we found that the clpP-deleted mutant was more resistant to sodium stress and failed to proliferate in the amoebae host Acanthamoeba castellanii. Conclusions The data present in this study illustrate that the ClpP protease homologue plays an important role in the expression of transmission traits and cell division of L. pneumophila, and further suggest a putative role of ClpP in virulence regulation.

  1. Partitioning and Exocytosis of Secretory Granules during Division of PC12 Cells

    Directory of Open Access Journals (Sweden)

    Nickolay Vassilev Bukoreshtliev

    2012-01-01

    Full Text Available The biogenesis, maturation, and exocytosis of secretory granules in interphase cells have been well documented, whereas the distribution and exocytosis of these hormone-storing organelles during cell division have received little attention. By combining ultrastructural analyses and time-lapse microscopy, we here show that, in dividing PC12 cells, the prominent peripheral localization of secretory granules is retained during prophase but clearly reduced during prometaphase, ending up with only few peripherally localized secretory granules in metaphase cells. During anaphase and telophase, secretory granules exhibited a pronounced movement towards the cell midzone and, evidently, their tracks colocalized with spindle microtubules. During cytokinesis, secretory granules were excluded from the midbody and accumulated at the bases of the intercellular bridge. Furthermore, by measuring exocytosis at the single granule level, we showed, that during all stages of cell division, secretory granules were competent for regulated exocytosis. In conclusion, our data shed new light on the complex molecular machinery of secretory granule redistribution during cell division, which facilitates their release from the F-actin-rich cortex and active transport along spindle microtubules.

  2. Scaling laws governing stochastic growth and division of single bacterial cells.

    Science.gov (United States)

    Iyer-Biswas, Srividya; Wright, Charles S; Henry, Jonathan T; Lo, Klevin; Burov, Stanislav; Lin, Yihan; Crooks, Gavin E; Crosson, Sean; Dinner, Aaron R; Scherer, Norbert F

    2014-11-11

    Uncovering the quantitative laws that govern the growth and division of single cells remains a major challenge. Using a unique combination of technologies that yields unprecedented statistical precision, we find that the sizes of individual Caulobacter crescentus cells increase exponentially in time. We also establish that they divide upon reaching a critical multiple (≈ 1.8) of their initial sizes, rather than an absolute size. We show that when the temperature is varied, the growth and division timescales scale proportionally with each other over the physiological temperature range. Strikingly, the cell-size and division-time distributions can both be rescaled by their mean values such that the condition-specific distributions collapse to universal curves. We account for these observations with a minimal stochastic model that is based on an autocatalytic cycle. It predicts the scalings, as well as specific functional forms for the universal curves. Our experimental and theoretical analysis reveals a simple physical principle governing these complex biological processes: a single temperature-dependent scale of cellular time governs the stochastic dynamics of growth and division in balanced growth conditions. PMID:25349411

  3. Cell division in Escherichia coli BS-12 is hypersensitive to deoxyribonucleic acid damage by ultraviolet light

    International Nuclear Information System (INIS)

    Escherichia coli BS-12 uvrA lon is hypersensitive to ultraviolet light. On minimal agar plates at densities in excess of about 10(7) bacteria per plate, as few as one or two photoreversible pyrimidine dimers in the entire genome are sufficient to cause inhibition of cell division. Most of the resulting filaments are unable to divide or form a viable colony. Inhibition of cell division appears to be a rapid consequence of replication of deoxyribonucleic acid containing a pyrimidine dimer. Photoreversibility of the inhibition of cell division persists indefinitely, indicating that the continued presence of the pyrimidine dimers (or the continued generation of daughter strand gaps) is necessary to maintain the division-inhibited state. In view of the kinetics for the production of filamentation by ultraviolet light and the extremely low average inducing fluence (0.03 J/m2), it is concluded that the initiating signal is not the same as that causing other inducible phenomena such as prophage induction or Weigle reactivation

  4. Tension-oriented cell divisions limit anisotropic tissue tension in epithelial spreading during zebrafish epiboly

    OpenAIRE

    Campinho, Pedro; Behrndt, Martin; Ranft, Jonas; Risler, Thomas; Minc, Nicolas; Heisenberg, Carl-Philipp

    2015-01-01

    Epithelial spreading is a common and fundamental aspect of various developmental and disease-related processes such as epithelial closure and wound healing. A key challenge for epithelial tissues undergoing spreading is to increase their surface area without disrupting epithelial integrity. Here we show that orienting cell divisions by tension constitutes an efficient mechanism by which the enveloping cell layer (EVL) releases anisotropic tension while undergoing spreading during zebrafish ep...

  5. LGH00031, a novel ortho-quinonoid inhibitor of cell division cycle 25B, inhibits human cancer cells via ROS generation

    Institute of Scientific and Technical Information of China (English)

    Yu-bo ZHOU; Xu FENG; Li-na WANG; Jun-qing DU; Yue-yang ZHOU; Hai-ping YU; Yi ZANG; Jing-ya LI; Jia LI

    2009-01-01

    Aim: To discover novel cell division cycle 25 (CDC25) B inhibitors and elucidate the mechanisms of inhibition in cancer cells. Methods: Cell growth inhibition was detected by MTT assay, the cell cycle was analyzed by flow cytometry, and protein expression and phosphorylation was examined by Western blot analysis. Results: LGH00031 inhibited CDC25B irreversibly in vitro in a dose-dependent manner, and impaired the proliferation of tumor cell lines. In synchronized HeLa cells, LGH00031 delayed the cell cycle progression at the G2/M phase. LGH00031 increased cyclin-dependent kinase 1 (CDK1) tyrosine 15 phosphorylation and cyclin B1 protein level. The activity of LGH00031 against CDC25B in vitro relied on the existence of 1, 4-dithiothreitol (DTT) or dihydrolipoic acid and oxygen. The oxygen free radical scavenger catalase and superoxide dismutase reduced the inactivation of CDC25 by LGH00031, confirming that reactive oxygen species (ROS) mediate the inactivation process in vitro. LGH00031 accelerated cellular ROS production in a dose-dependent manner, and N-acetyl cysteine (NAC) markedly decreased the ROS production induced by LGH00031.Correspondingly, the LGH00031-induced decrease in cell viability and cell cycle arrest, cyclin B1 protein level, and phosphorylation of CDK1 tyrosine 15 were also rescued by NAC that decreased ROS pro-duction. Conclusion: The activity of LGH00031 at the molecular and cellular level is mediated by ROS.

  6. Fission yeast cells undergo nuclear division in the absence of spindle microtubules.

    Directory of Open Access Journals (Sweden)

    Stefania Castagnetti

    Full Text Available Mitosis in eukaryotic cells employs spindle microtubules to drive accurate chromosome segregation at cell division. Cells lacking spindle microtubules arrest in mitosis due to a spindle checkpoint that delays mitotic progression until all chromosomes have achieved stable bipolar attachment to spindle microtubules. In fission yeast, mitosis occurs within an intact nuclear membrane with the mitotic spindle elongating between the spindle pole bodies. We show here that in fission yeast interference with mitotic spindle formation delays mitosis only briefly and cells proceed to an unusual nuclear division process we term nuclear fission, during which cells perform some chromosome segregation and efficiently enter S-phase of the next cell cycle. Nuclear fission is blocked if spindle pole body maturation or sister chromatid separation cannot take place or if actin polymerization is inhibited. We suggest that this process exhibits vestiges of a primitive nuclear division process independent of spindle microtubules, possibly reflecting an evolutionary intermediate state between bacterial and Archeal chromosome segregation where the nucleoid divides without a spindle and a microtubule spindle-based eukaryotic mitosis.

  7. DNA synthesis and cell division in the adult primate brain

    International Nuclear Information System (INIS)

    It is generally accepted that the adult human brain is incapable of producing new neuron. Even cursory examination of neurologic, neuropathologic, or neurobiological textbooks published during the past 50 years will testify that this belief is deeply entrenched. In his classification of cell populations on the basis of their proliferative behavior, Leblond regarded neurons of the central nervous system as belonging to a category of static, nonrenewing epithelial tissue incapable of expanding or replenishing itself. This belief, however needs to re reexamined for two major reasons: First, as reviewed below, a number of reports have provided evidence of neurogenesis in adult brain of several vertebrate species. Second, the capacity for neurogenesis in the adult primate central nervous system has never been examined by modern methods. In this article the author described recent results from an extensive autoradiographic analysis performed on twelve rhesus monkeys injected with the specific DNA precursor [3H] thymidine at ages ranging from 6 postnatal months to 17 years

  8. Transfer of a eubacteria-type cell division site-determining factor CrMinD gene to the nucleus from the chloroplast genome in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    LIU WeiZhong; HU Yong; ZHANG RunJie; ZHOU WeiWei; ZHU JiaYing; LIU XiangLin; HE YiKun

    2007-01-01

    MinD is a ubiquitous ATPase that plays a crucial role in selection of the division site in eubacteria, chloroplasts, and probably Archaea. In four green algae, Mesostigma viride, Nephroselmis olivacea, Chlorella vulgaris and Prototheca wickerhamii, MinD homologues are encoded in the plastid genome. However, in Arabidopsis, MinD is a nucleus-encoded, chloroplast-targeted protein involved in chloroplast division, which suggests that MinD has been transferred to the nucleus in higher land plants. Yet the lateral gene transfer (LGT) of MinD from plastid to nucleus during plastid evolution remains poorly understood. Here, we identified a nucleus-encoded MinD homologue from unicellular green alga Chlamydomonas reinhardtii, a basal species in the green plant lineage. Overexpression of CrMinD in wild type E. coli inhibited cell division and resulted in the filamentous cell formation, clearly demonstrated the conservation of the MinD protein during the evolution of photosynthetic eukaryotes. The transient expression of CrMinD-egfp confirmed the role of CrMinD protein in the regulation of plastid division. Searching all the published plastid genomic sequences of land plants, no MinD homologues were found, which suggests that the transfer of MinD from plastid to nucleus might have occurred before the evolution of land plants.

  9. Cell segmentation for division rate estimation in computerized video time-lapse microscopy

    Science.gov (United States)

    He, Weijun; Wang, Xiaoxu; Metaxas, Dimitris; Mathew, Robin; White, Eileen

    2007-02-01

    The automated estimation of cell division rate plays an important role in the evaluation of a gene function in high throughput biomedical research. Using Computerized Video Time-Lapse (CVTL) microcopy , it is possible to follow a large number of cells in their physiological conditions for several generations. However analysis of this large volume data is complicated due to cell to cell contacts in a high density population. We approach this problem by segmenting out cells or cell clusters through a learning method. The feature of a pixel is represented by the intensity and gradient information in a small surrounding sub-window. Curve evolution techniques are used to accurately find the cell or cell cluster boundary. With the assumption that the average cell size is the same in each frame, we can use the cell area to estimate the cell division rate. Our segmentation results are compared to manually-defined ground truth. Both recall and precision measures for segmentation accuracy are above 95%.

  10. Asymmetric division triggers cell-specific gene expression through coupled capture and stabilization of a phosphatase

    OpenAIRE

    Bradshaw, Niels; Losick, Richard

    2015-01-01

    Formation of a division septum near a randomly chosen pole during sporulation in Bacillus subtilis creates unequal sized daughter cells with dissimilar programs of gene expression. An unanswered question is how polar septation activates a transcription factor (σF) selectively in the small cell. We present evidence that the upstream regulator of σF, the phosphatase SpoIIE, is compartmentalized in the small cell by transfer from the polar septum to the adjacent cell pole where SpoIIE is protect...

  11. AHP6 inhibits cytokinin signaling to regulate the orientation of pericycle cell division during lateral root initiation.

    Directory of Open Access Journals (Sweden)

    Sofia Moreira

    Full Text Available In Arabidopsis thaliana, lateral roots (LRs initiate from anticlinal cell divisions of pericycle founder cells. The formation of LR primordia is regulated antagonistically by the phytohormones cytokinin and auxin. It has previously been shown that cytokinin has an inhibitory effect on the patterning events occurring during LR formation. However, the molecular players involved in cytokinin repression are still unknown. In a similar manner to protoxylem formation in Arabidopsis roots, in which AHP6 (ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN 6 acts as a cytokinin inhibitor, we reveal that AHP6 also functions as a cytokinin repressor during early stages of LR development. We show that AHP6 is expressed at different developmental stages during LR formation and is required for the correct orientation of cell divisions at the onset of LR development. Moreover, we demonstrate that AHP6 influences the localization of the auxin efflux carrier PIN1, which is necessary for patterning the LR primordia. In summary, we show that the inhibition of cytokinin signaling through AHP6 is required to establish the correct pattern during LR initiation.

  12. Cell division in the unicellular microalga Dunaliella viridis depends on phosphorylation of extracellular signal-regulated kinases (ERKs).

    Science.gov (United States)

    Jiménez, Carlos; Cossío, Belén R; Rivard, Christopher J; Berl, Tomás; Capasso, Juan M

    2007-01-01

    In mammalian cells, MAPKs are involved in both stress response (JNK and p38 pathways) and cell proliferation and differentiation [extracellular signal-regulated kinase (ERK)] through protein kinase cascades. Exposure of Dunaliella viridis cell cultures to PD98059, a very specific inhibitor of the ERK signalling pathway, resulted in a total arrest of cell proliferation and a complete dephosphorylation of ERK. As shown by flow cytometry analysis of propidium iodide-stained cells, PD98059 stopped mitosis at the G(2) phase after the S phase has been completed. Multiple physiological parameters such as cell motility and reducing power generation (NADPH) clearly indicate that the treated cells are wholly viable. Exposure of D. viridis to environmental stresses that impair cell division, such as hyperosmotic shock, nitrogen starvation, or sublethal UV irradiation, caused a marked decrease in the phospho-ERK levels as detected by western blot. Two 400 bp polynucleotides from D. viridis with high homologies to published sequences of ERK1 and ERK2 were cloned, sequenced, and submitted to GenBank. Northern blot analysis revealed two mRNA bands of approximately 1.9 kb, consistent with the expected size of ERK proteins ( approximately 40 kDa). Sequence analysis showed that they contained several mitogen-activated protein kinase (MAPK) conserved domains, including II, III, VIb, VII, and the double phosphorylation motif. Interestingly, in D. viridis, this motif was T*DY* instead of the canonic T*EY*. Based on this finding, ERK plant sequences can be divided into two groups, one termed the T*DY* branch and the other termed the T*EY* branch. The molecular and functional data presented here suggest that ERK is a very ancient signalling pathway and that it was already present in the last common ancestor of all eukaryotic cells. PMID:17220513

  13. Physical association between a novel plasma-membrane structure and centrosome orients cell division

    Science.gov (United States)

    Negishi, Takefumi; Miyazaki, Naoyuki; Murata, Kazuyoshi; Yasuo, Hitoyoshi; Ueno, Naoto

    2016-01-01

    In the last mitotic division of the epidermal lineage in the ascidian embryo, the cells divide stereotypically along the anterior-posterior axis. During interphase, we found that a unique membrane structure invaginates from the posterior to the centre of the cell, in a microtubule-dependent manner. The invagination projects toward centrioles on the apical side of the nucleus and associates with one of them. Further, a cilium forms on the posterior side of the cell and its basal body remains associated with the invagination. A laser ablation experiment suggests that the invagination is under tensile force and promotes the posterior positioning of the centrosome. Finally, we showed that the orientation of the invaginations is coupled with the polarized dynamics of centrosome movements and the orientation of cell division. Based on these findings, we propose a model whereby this novel membrane structure orchestrates centrosome positioning and thus the orientation of cell division axis. DOI: http://dx.doi.org/10.7554/eLife.16550.001 PMID:27502556

  14. Mechanism of murine epidermal maintenance: Cell division and the Voter Model

    CERN Document Server

    Klein, Allon M; Jones, Philip H; Simons, Benjamin D

    2007-01-01

    This paper presents an interesting experimental example of voter-model statistics in biology. In recent work on mouse tail-skin, where proliferating cells are confined to a two-dimensional layer, we showed that cells proliferate and differentiate according to a simple stochastic model of cell division involving just one type of proliferating cell that may divide both symmetrically and asymmetrically. Curiously, these simple rules provide excellent predictions of the cell population dynamics without having to address their spatial distribution. Yet, if the spatial behaviour of cells is addressed by allowing cells to diffuse at random, one deduces that density fluctuations destroy tissue confluence, implying some hidden degree of spatial regulation in the physical system. To infer the mechanism of spatial regulation, we consider a two-dimensional model of cell fate that preserves the overall population dynamics. By identifying the resulting behaviour with a three-species variation of the "Voter" model, we predi...

  15. Individuality and universality in the growth-division laws of single E. coli cells

    Science.gov (United States)

    Kennard, Andrew S.; Osella, Matteo; Javer, Avelino; Grilli, Jacopo; Nghe, Philippe; Tans, Sander J.; Cicuta, Pietro; Cosentino Lagomarsino, Marco

    2016-01-01

    The mean size of exponentially dividing Escherichia coli cells in different nutrient conditions is known to depend on the mean growth rate only. However, the joint fluctuations relating cell size, doubling time, and individual growth rate are only starting to be characterized. Recent studies in bacteria reported a universal trend where the spread in both size and doubling times is a linear function of the population means of these variables. Here we combine experiments and theory and use scaling concepts to elucidate the constraints posed by the second observation on the division control mechanism and on the joint fluctuations of sizes and doubling times. We found that scaling relations based on the means collapse both size and doubling-time distributions across different conditions and explain how the shape of their joint fluctuations deviates from the means. Our data on these joint fluctuations highlight the importance of cell individuality: Single cells do not follow the dependence observed for the means between size and either growth rate or inverse doubling time. Our calculations show that these results emerge from a broad class of division control mechanisms requiring a certain scaling form of the "division hazard rate function," which defines the probability rate of dividing as a function of measurable parameters. This "model free" approach gives a rationale for the universal body-size distributions observed in microbial ecosystems across many microbial species, presumably dividing with multiple mechanisms. Additionally, our experiments show a crossover between fast and slow growth in the relation between individual-cell growth rate and division time, which can be understood in terms of different regimes of genome replication control.

  16. Flat leaf formation realized by cell-division control and mutual recessive gene regulation.

    Science.gov (United States)

    Hayakawa, Yoshinori; Tachikawa, Masashi; Mochizuki, Atsushi

    2016-09-01

    Most of the land plants generally have dorsoventrally flat leaves, maximizing the surface area of both upper (adaxial) side and lower (abaxial) side. The former is specialized for light capturing for photosynthesis and the latter is specialized for gas exchange. From findings of molecular genetics, it has been considered that the coupled dynamics between tissue morphogenesis and gene regulation for cell identity is responsible for making flat leaves. The hypothesis claims that a flat leaf is generated under two assumptions, (i) two mutually recessive groups of genes specify adaxial and abaxial sides of a leaf, (ii) cell divisions are induced at the limited region in the leaf margin where both of two groups are expressed. We examined the plausibility and possibility of this hypothesis from the dynamical point of view. We studied a mathematical model where two processes are coupled, tissue morphogenesis induced by cell division and deformation, and dynamics of gene regulations. From the analysis of the model we found that the classically believed hypothesis is not sufficient to generate flat leaves with high probability. We examined several different modifications and revision of the model. Then we found that a simple additional rule of polarized cell division facilitates flat leaf formation. The result of our analysis gives prediction of possible mechanism, which can be easily verified in experiments. PMID:27287339

  17. Cell division interference in newly fertilized ovules induces stenospermocarpy in cross-pollinated citrus fruit.

    Science.gov (United States)

    Mesejo, Carlos; Muñoz-Fambuena, Natalia; Reig, Carmina; Martínez-Fuentes, Amparo; Agustí, Manuel

    2014-08-01

    Seedlessness is a highly desirable characteristic in fresh fruits. However, post-fertilization seed abortion of cross-pollinated citrus fruit is uncommon. The factors regulating stenospermocarpy in citrus are unknown. In this research, we induced stenospermocarpy interfering in newly fertilized ovule cell division. The research also elucidates the most sensitive stage for ovule/seed abortion in citrus. Experiments were conducted with 'Afourer' mandarin that cross-pollinates with several cultivars and species. Cross-pollinated fruitlets were treated with maleic hydrazide (MH), a systemic growth regulator that specifically interferes in cell division. MH reduced ovule growth rate, the number of cell layers in nucella and inhibited embryo sac expansion; moreover, the treatment increased callose accumulation in nucella and surrounding the embryo sac. Fruits developed an early-aborted seed type with an immature, soft and edible seed coat. Seed number (-80%) and seed weight (-46%) were reduced in mature fruits. MH also hampered cell division in ovary walls, mesocarp and endocarp, thus reducing daily fruitlet growth and increasing fruit abscission. Stenospermocarpy could only be induced for a short period of time in the progamic phase of fertilization, specifically, when ovules are ready to be fertilized (7 days after anthesis) to early stages of embryo sac development (14 days after anthesis). PMID:25017163

  18. A Negative Feedback Loop Controlling bHLH Complexes Is Involved in Vascular Cell Division and Differentiation in the Root Apical Meristem.

    Science.gov (United States)

    Katayama, Hirofumi; Iwamoto, Kuninori; Kariya, Yuka; Asakawa, Tomohiro; Kan, Toshiyuki; Fukuda, Hiroo; Ohashi-Ito, Kyoko

    2015-12-01

    Controlling cell division and differentiation in meristems is essential for proper plant growth. Two bHLH heterodimers consisting of LONESOME HIGHWAY (LHW) and TARGET OF MONOPTEROS 5 (TMO5)/TMO5-LIKE1 (T5L1) regulate periclinal cell division in vascular cells in the root apical meristem (RAM). In this study, we further investigated the functions of LHW-T5L1, finding that in addition to controlling cell division, this complex regulates xylem differentiation in the RAM via a novel negative regulatory system. LHW-T5L1 upregulated the thermospermine synthase gene ACAULIS5 (ACL5), as well as SUPPRESSOR OF ACAULIS5 LIKE3 (SACL3), which encodes a bHLH protein, in the RAM. The SACL3 promoter sequence contains a conserved upstream open reading frame (uORF), which blocked translation of the main SACL3 ORF in the absence of thermospermine. Thermospermine eliminated the negative effect of uORF and enhanced SACL3 production. Further genetic and molecular biological analyses indicated that ACL5 and SACL3 suppress the function of LHW-T5L1 through a protein-protein interaction between LHW and SACL3. Finally, we showed that a negative feedback loop consisting of LHW-T5L1, ACL5, SACL3, and LHW-SACL3 contributes to maintain RAM size and proper root growth. These findings suggest that a negative feedback loop regulates the LHW-T5L1 output level to coordinate cell division and differentiation in a cell-autonomous manner. PMID:26616019

  19. Scaffolding during the cell cycle by A-kinase anchoring proteins

    NARCIS (Netherlands)

    Han, B; Poppinga, W J; Schmidt, M

    2015-01-01

    Cell division relies on coordinated regulation of the cell cycle. A process including a well-defined series of strictly regulated molecular mechanisms involving cyclin-dependent kinases, retinoblastoma protein, and polo-like kinases. Dysfunctions in cell cycle regulation are associated with disease

  20. Role of the Number of Microtubules in Chromosome Segregation during Cell Division

    CERN Document Server

    Bertalan, Zsolt; La Porta, Caterina A M; Zapperi, Stefano

    2015-01-01

    Faithful segregation of genetic material during cell division requires alignment of chromosomes between two spindle poles and attachment of their kinetochores to each of the poles. Failure of these complex dynamical processes leads to chromosomal instability (CIN), a characteristic feature of several diseases including cancer. While a multitude of biological factors regulating chromosome congression and bi-orientation have been identified, it is still unclear how they are integrated so that coherent chromosome motion emerges from a large collection of random and deterministic processes. Here we address this issue by a three dimensional computational model of motor-driven chromosome congression and bi-orientation during mitosis. Our model reveals that successful cell division requires control of the total number of microtubules: if this number is too small bi-orientation fails, while if it is too large not all the chromosomes are able to congress. The optimal number of microtubules predicted by our model compa...

  1. Automatic detection of cell divisions (mitosis) in live-imaging microscopy images using Convolutional Neural Networks.

    Science.gov (United States)

    Shkolyar, Anat; Gefen, Amit; Benayahu, Dafna; Greenspan, Hayit

    2015-08-01

    We propose a semi-automated pipeline for the detection of possible cell divisions in live-imaging microscopy and the classification of these mitosis candidates using a Convolutional Neural Network (CNN). We use time-lapse images of NIH3T3 scratch assay cultures, extract patches around bright candidate regions that then undergo segmentation and binarization, followed by a classification of the binary patches into either containing or not containing cell division. The classification is performed by training a Convolutional Neural Network on a specially constructed database. We show strong results of AUC = 0.91 and F-score = 0.89, competitive with state-of-the-art methods in this field. PMID:26736369

  2. Interplay of migratory and division forces as a generic mechanism for stem cell patterns

    Science.gov (United States)

    Hannezo, Edouard; Coucke, Alice; Joanny, Jean-François

    2016-02-01

    In many adult tissues, stem cells and differentiated cells are not homogeneously distributed: stem cells are arranged in periodic "niches," and differentiated cells are constantly produced and migrate out of these niches. In this article, we provide a general theoretical framework to study mixtures of dividing and actively migrating particles, which we apply to biological tissues. We show in particular that the interplay between the stresses arising from active cell migration and stem cell division give rise to robust stem cell patterns. The instability of the tissue leads to spatial patterns which are either steady or oscillating in time. The wavelength of the instability has an order of magnitude consistent with the biological observations. We also discuss the implications of these results for future in vitro and in vivo experiments.

  3. An interplay of migratory and division forces as a generic mechanism for stem cell patterns

    CERN Document Server

    Hannezo, Edouard; Joanny, Jean-François

    2015-01-01

    In many adult tissues, stem cells and differentiated cells are not homogeneously distributed : stem cells are arranged in periodic "niches", and differentiated cells are constantly produced and migrate out of these niches. In this article, we provide a general theoretical framework to study mixtures of dividing and actively migrating particles, which we apply to biological tissues. We show in particular that the interplay between the stresses arising from active cell migration and stem cell division give rise to robust stem cell patterns. The instability of the tissue leads to spatial patterns which are either steady or oscillating in time. The wavelength of the instability has an order of magnitude consistent with the biological observations. We also discuss the implications of these results for future in vitro and in vivo experiments.

  4. Timing of Tissue-specific Cell Division Requires a Differential Onset of Zygotic Transcription during Metazoan Embryogenesis.

    Science.gov (United States)

    Wong, Ming-Kin; Guan, Daogang; Ng, Kaoru Hon Chun; Ho, Vincy Wing Sze; An, Xiaomeng; Li, Runsheng; Ren, Xiaoliang; Zhao, Zhongying

    2016-06-10

    Metazoan development demands not only precise cell fate differentiation but also accurate timing of cell division to ensure proper development. How cell divisions are temporally coordinated during development is poorly understood. Caenorhabditis elegans embryogenesis provides an excellent opportunity to study this coordination due to its invariant development and widespread division asynchronies. One of the most pronounced asynchronies is a significant delay of cell division in two endoderm progenitor cells, Ea and Ep, hereafter referred to as E2, relative to its cousins that mainly develop into mesoderm organs and tissues. To unravel the genetic control over the endoderm-specific E2 division timing, a total of 822 essential and conserved genes were knocked down using RNAi followed by quantification of cell cycle lengths using in toto imaging of C. elegans embryogenesis and automated lineage. Intriguingly, knockdown of numerous genes encoding the components of general transcription pathway or its regulatory factors leads to a significant reduction in the E2 cell cycle length but an increase in cell cycle length of the remaining cells, indicating a differential requirement of transcription for division timing between the two. Analysis of lineage-specific RNA-seq data demonstrates an earlier onset of transcription in endoderm than in other germ layers, the timing of which coincides with the birth of E2, supporting the notion that the endoderm-specific delay in E2 division timing demands robust zygotic transcription. The reduction in E2 cell cycle length is frequently associated with cell migration defect and gastrulation failure. The results suggest that a tissue-specific transcriptional activation is required to coordinate fate differentiation, division timing, and cell migration to ensure proper development. PMID:27056332

  5. Regulation of cell division and expansion by sugar and auxin signaling

    Directory of Open Access Journals (Sweden)

    Lu eWang

    2013-05-01

    Full Text Available Plant growth and development are modulated by concerted actions of a variety of signaling molecules. In recent years, evidence has emerged on the roles of sugar and auxin signals in diverse aspects of plant growth and development. Here, based on recent progress of genetic analyses and gene expression profiling studies, we summarize the functional similarities, diversities and their interactions of sugar and auxin signals in regulating two major processes of plant development: cell division and cell expansion. We focus on roles of sugar and auxin signaling in both vegetative and reproductive tissues including developing seed.

  6. Arabidopsis brassinosteroid biosynthetic mutant dwarf7-1 exhibits slower rates of cell division and shoot induction

    Directory of Open Access Journals (Sweden)

    Schulz Burkhard

    2010-12-01

    Full Text Available Abstract Background Plant growth depends on both cell division and cell expansion. Plant hormones, including brassinosteroids (BRs, are central to the control of these two cellular processes. Despite clear evidence that BRs regulate cell elongation, their roles in cell division have remained elusive. Results Here, we report results emphasizing the importance of BRs in cell division. An Arabidopsis BR biosynthetic mutant, dwarf7-1, displayed various characteristics attributable to slower cell division rates. We found that the DWARF4 gene which encodes for an enzyme catalyzing a rate-determining step in the BR biosynthetic pathways, is highly expressed in the actively dividing callus, suggesting that BR biosynthesis is necessary for dividing cells. Furthermore, dwf7-1 showed noticeably slower rates of callus growth and shoot induction relative to wild-type control. Flow cytometric analyses of the nuclei derived from either calli or intact roots revealed that the cell division index, which was represented as the ratio of cells at the G2/M vs. G1 phases, was smaller in dwf7-1 plants. Finally, we found that the expression levels of the genes involved in cell division and shoot induction, such as PROLIFERATING CELL NUCLEAR ANTIGEN2 (PCNA2 and ENHANCER OF SHOOT REGENERATION2 (ESR2, were also lower in dwf7-1 as compared with wild type. Conclusions Taken together, results of callus induction, shoot regeneration, flow cytometry, and semi-quantitative RT-PCR analysis suggest that BRs play important roles in both cell division and cell differentiation in Arabidopsis.

  7. Resistance-nodulation-cell division-type efflux pump involved in aminoglycoside resistance in Acinetobacter baumannii strain BM4454.

    Science.gov (United States)

    Magnet, S; Courvalin, P; Lambert, T

    2001-12-01

    Multidrug-resistant strain Acinetobacter baumannii BM4454 was isolated from a patient with a urinary tract infection. The adeB gene, which encodes a resistance-nodulation-cell division (RND) protein, was detected in this strain by PCR with two degenerate oligodeoxynucleotides. Insertional inactivation of adeB in BM4454, which generated BM4454-1, showed that the corresponding protein was responsible for aminoglycoside resistance and was involved in the level of susceptibility to other drugs including fluoroquinolones, tetracyclines, chloramphenicol, erythromycin, trimethoprim, and ethidium bromide. Study of ethidium bromide accumulation in BM4454 and BM4454-1, in the presence or in the absence of carbonyl cyanide m-chlorophenylhydrazone, demonstrated that AdeB was responsible for the decrease in intracellular ethidium bromide levels in a proton motive force-dependent manner. The adeB gene was part of a cluster that included adeA and adeC which encodes proteins homologous to membrane fusion and outer membrane proteins of RND-type three-component efflux systems, respectively. The products of two upstream open reading frames encoding a putative two-component regulatory system might be involved in the regulation of expression of the adeABC gene cluster. PMID:11709311

  8. A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Damodaran, Shima P; Eberhard, Stephan; Boitard, Laurent; Rodriguez, Jairo Garnica; Wang, Yuxing; Bremond, Nicolas; Baudry, Jean; Bibette, Jérôme; Wollman, Francis-André

    2015-01-01

    To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers). These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes. PMID:25760649

  9. A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Shima P Damodaran

    Full Text Available To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers and a significant subpopulation of slowly dividing cells (slow-growers. These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

  10. Altered mRNA cap recognition activity of initiation factor 4E in the yeast cell cycle division mutant cdc33.

    OpenAIRE

    Altmann, M; Trachsel, H

    1989-01-01

    The mutation in the S. cerevisiae cell cycle division mutant cdc33 consists of a single G to A transition in the open reading frame encoding translation initiation factor 4E (eIF-4E). This leads to the substitution of glycine 113 by aspartic acid close to tryptophane 115 in the protein. This mutation reduces cap binding activity of eIF-4E as measured by binding of eIF-4E to m7GDP agarose columns and slows down overall protein synthesis at the non-permissive temperature. Comparison of the cdc3...

  11. Effect of microgravity environment on cell wall regeneration, cell divisions, growth, and differentiation of plants from protoplasts (7-IML-1)

    Science.gov (United States)

    Rasmussen, Ole

    1992-01-01

    The primary goal of this project is to investigate if microgravity has any influence on growth and differentiation of protoplasts. Formation of new cell walls on rapeseed protoplasts takes place within the first 24 hours after isolation. Cell division can be observed after 2-4 days and formation of cell aggregates after 5-7 days. Therefore, it is possible during the 7 day IML-1 Mission to investigate if cell wall formation, cell division, and cell differentiation are influenced by microgravity. Protoplasts of rapeseeds and carrot will be prepared shortly before launch and injected into 0.6 ml polyethylene bags. Eight bags are placed in an aluminum block inside the ESA Type 1 container. The containers are placed at 4 C in PTCU's and transferred to orbiter mid-deck. At 4 C all cell processes are slowed down, including cell wall formation. Latest access to the shuttle will be 12 hours before launch. In orbit the containers will be transferred from the PTC box to the 22 C Biorack incubator. The installation of a 1 g centrifuge in Biorack will make it possible to distinguish between effects of near weightlessness and effects caused by cosmic radiation and other space flight factors including vibrations. Parallel control experiments will be carried out on the ground. Other aspects of the experiment are discussed.

  12. Tau protein function in living cells

    OpenAIRE

    1986-01-01

    Tau protein from mammalian brain promotes microtubule polymerization in vitro and is induced during nerve cell differentiation. However, the effects of tau or any other microtubule-associated protein on tubulin assembly within cells are presently unknown. We have tested tau protein activity in vivo by microinjection into a cell type that has no endogenous tau protein. Immunofluorescence shows that tau protein microinjected into fibroblast cells associates specifically with microtubules. The i...

  13. Characterization of the minimum domain required for targeting budding yeast myosin II to the site of cell division

    Directory of Open Access Journals (Sweden)

    Tolliday Nicola J

    2006-06-01

    Full Text Available Abstract Background All eukaryotes with the exception of plants use an actomyosin ring to generate a constriction force at the site of cell division (cleavage furrow during mitosis and meiosis. The structure and filament forming abilities located in the C-terminal or tail region of one of the main components, myosin II, are important for localising the molecule to the contractile ring (CR during cytokinesis. However, it remains poorly understood how myosin II is recruited to the site of cell division and how this recruitment relates to myosin filament assembly. Significant conservation between species of the components involved in cytokinesis, including those of the CR, allows the use of easily genetically manipulated organisms, such as budding yeast (Saccharomyces cerevisiae, in the study of cytokinesis. Budding yeast has a single myosin II protein, named Myo1. Unlike most other class II myosins, the tail of Myo1 has an irregular coiled coil. In this report we use molecular genetics, biochemistry and live cell imaging to characterize the minimum localisation domain (MLD of budding yeast Myo1. Results We show that the MLD is a small region in the centre of the tail of Myo1 and that it is both necessary and sufficient for localisation of Myo1 to the yeast bud neck, the pre-determined site of cell division. Hydrodynamic measurements of the MLD, purified from bacteria or yeast, show that it is likely to exist as a trimer. We also examine the importance of a small region of low coiled coil forming probability within the MLD, which we call the hinge region. Removal of the hinge region prevents contraction of the CR. Using fluorescence recovery after photobleaching (FRAP, we show that GFP-tagged MLD is slightly more dynamic than the GFP-tagged full length molecule but less dynamic than the GFP-tagged Myo1 construct lacking the hinge region. Conclusion Our results define the intrinsic determinant for the localization of budding yeast myosin II and show

  14. The cyanobacterial cell division factor Ftn6 contains an N-terminal DnaD-like domain

    Directory of Open Access Journals (Sweden)

    Saguez Cyril

    2009-08-01

    Full Text Available Abstract Background DNA replication and cell cycle as well as their relationship have been extensively studied in the two model organisms E. coli and B. subtilis. By contrast, little is known about these processes in cyanobacteria, even though they are crucial to the biosphere, in utilizing solar energy to renew the oxygenic atmosphere and in producing the biomass for the food chain. Recent studies have allowed the identification of several cell division factors that are specifics to cyanobacteria. Among them, Ftn6 has been proposed to function in the recruitment of the crucial FtsZ proteins to the septum or the subsequent Z-ring assembly and possibly in chromosome segregation. Results In this study, we identified an as yet undescribed domain located in the conserved N-terminal region of Ftn6. This 77 amino-acids-long domain, designated here as FND (Ftn6 N-Terminal Domain, exhibits striking sequence and structural similarities with the DNA-interacting module, listed in the PFAM database as the DnaD-like domain (pfam04271. We took advantage of the sequence similarities between FND and the DnaD-like domains to construct a homology 3D-model of the Ftn6 FND domain from the model cyanobacterium Synechocystis PCC6803. Mapping of the conserved residues exposed onto the FND surface allowed us to identify a highly conserved area that could be engaged in Ftn6-specific interactions. Conclusion Overall, similarities between FND and DnaD-like domains as well as previously reported observations on Ftn6 suggest that FND may function as a DNA-interacting module thereby providing an as yet missing link between DNA replication and cell division in cyanobacteria. Consistently, we also showed that Ftn6 is involved in tolerance to DNA damages generated by UV rays.

  15. Reprogramming cells with synthetic proteins.

    Science.gov (United States)

    Yang, Xiaoxiao; Malik, Vikas; Jauch, Ralf

    2015-01-01

    Conversion of one cell type into another cell type by forcibly expressing specific cocktails of transcription factors (TFs) has demonstrated that cell fates are not fixed and that cellular differentiation can be a two-way street with many intersections. These experiments also illustrated the sweeping potential of TFs to "read" genetically hardwired regulatory information even in cells where they are not normally expressed and to access and open up tightly packed chromatin to execute gene expression programs. Cellular reprogramming enables the modeling of diseases in a dish, to test the efficacy and toxicity of drugs in patient-derived cells and ultimately, could enable cell-based therapies to cure degenerative diseases. Yet, producing terminally differentiated cells that fully resemble their in vivocounterparts in sufficient quantities is still an unmet clinical need. While efforts are being made to reprogram cells nongenetically by using drug-like molecules, defined TF cocktails still dominate reprogramming protocols. Therefore, the optimization of TFs by protein engineering has emerged as a strategy to enhance reprogramming to produce functional, stable and safe cells for regenerative biomedicine. Engineering approaches focused on Oct4, MyoD, Sox17, Nanog and Mef2c and range from chimeric TFs with added transactivation domains, designer transcription activator-like effectors to activate endogenous TFs to reprogramming TFs with rationally engineered DNA recognition principles. Possibly, applying the complete toolkit of protein design to cellular reprogramming can help to remove the hurdles that, thus far, impeded the clinical use of cells derived from reprogramming technologies. PMID:25652623

  16. Reprogramming cells with synthetic proteins

    Directory of Open Access Journals (Sweden)

    Xiaoxiao Yang

    2015-06-01

    Full Text Available Conversion of one cell type into another cell type by forcibly expressing specific cocktails of transcription factors (TFs has demonstrated that cell fates are not fixed and that cellular differentiation can be a two-way street with many intersections. These experiments also illustrated the sweeping potential of TFs to "read" genetically hardwired regulatory information even in cells where they are not normally expressed and to access and open up tightly packed chromatin to execute gene expression programs. Cellular reprogramming enables the modeling of diseases in a dish, to test the efficacy and toxicity of drugs in patient-derived cells and ultimately, could enable cell-based therapies to cure degenerative diseases. Yet, producing terminally differentiated cells that fully resemble their in vivocounterparts in sufficient quantities is still an unmet clinical need. While efforts are being made to reprogram cells nongenetically by using drug-like molecules, defined TF cocktails still dominate reprogramming protocols. Therefore, the optimization of TFs by protein engineering has emerged as a strategy to enhance reprogramming to produce functional, stable and safe cells for regenerative biomedicine. Engineering approaches focused on Oct4, MyoD, Sox17, Nanog and Mef2c and range from chimeric TFs with added transactivation domains, designer transcription activator-like effectors to activate endogenous TFs to reprogramming TFs with rationally engineered DNA recognition principles. Possibly, applying the complete toolkit of protein design to cellular reprogramming can help to remove the hurdles that, thus far, impeded the clinical use of cells derived from reprogramming technologies.

  17. Mini-F plasmid genes that couple host cell division to plasmid proliferation.

    OpenAIRE

    Ogura, T; Hiraga, S

    1983-01-01

    A mechanism for stable maintenance of plasmids, besides the replication and partition mechanisms, has been found to be specified by genes of a mini-F plasmid. An oriC plasmid carrying both a mini-F segment necessary for partition [coordinates 46.4-49.4 kilobase pairs (kb) on the F map] and another segment (42.9-43.6 kb), designated ccd (coupled cell division), is more stably maintained than are oriC plasmids carrying only the partition segment; the stability is comparable to that of the paren...

  18. Cdc42 and Rab8a are critical for intestinal stem cell division, survival, and differentiation in mice

    DEFF Research Database (Denmark)

    Sakamori, Ryotaro; Das, Soumyashree; Yu, Shiyan; Feng, Shanshan; Stypulkowski, Ewa; Guan, Yinzheng; Douard, Veronique; Tang, Waixing; Ferraris, Ronaldo P; Harada, Akihiro; Brakebusch, Cord; Guo, Wei; Gao, Nan

    2012-01-01

    reminiscent of human microvillus inclusion disease (MVID), a devastating congenital intestinal disorder that results in severe nutrient deprivation. Further analysis revealed that Cdc42-deficient stem cells had cell division defects, reduced capacity for clonal expansion and differentiation into Paneth cells...

  19. Role of SufI (FtsP) in cell division of Escherichia coli: evidence for its involvement in stabilizing the assembly of the divisome.

    Science.gov (United States)

    Samaluru, Harish; SaiSree, L; Reddy, Manjula

    2007-11-01

    The function of SufI, a well-studied substrate of the TatABC translocase in Escherichia coli, is not known. It was earlier implicated in cell division, based on the finding that multiple copies of sufI suppressed the phenotypes of cells with mutations in ftsI (ftsI23), which encodes a divisomal transpeptidase. Recently, sufI was identified as both a multicopy suppressor gene and a synthetic lethal mutant of ftsEX, which codes for a division-specific putative ABC transporter. In this study, we show that sufI is essential for the viability of E. coli cells subjected to various forms of stress, including oxidative stress and DNA damage. The sufI mutant also exhibits sulA-independent filamentation, indicating a role in cell division. The phenotypes of the sufI mutant are suppressed by factors that stabilize FtsZ ring assembly, such as increased expression of cell division proteins FtsQAZ or FtsN or the presence of the gain-of-function ftsA* (FtsA R286W) mutation, suggesting that SufI is a divisomal protein required during stress conditions. In support of this, multicopy sufI suppressed the divisional defects of mutants carrying the ftsA12, ftsQ1, or ftsK44 allele but not those of mutants carrying ftsZ84. Most of the division-defective mutants, in particular those carrying a DeltaftsEX or ftsI23 allele, exhibited sensitivity to oxidative stress or DNA damage, and this sensitivity was also abolished by multiple copies of SufI. All of these data suggest that SufI is a division component involved in protecting or stabilizing the divisomal assembly under conditions of stress. Since sufI fulfils the requirements to be designated an fts gene, we propose that it be renamed ftsP. PMID:17766410

  20. Sonic hedgehog signaling regulates mode of cell division of early cerebral cortex progenitors and increases astrogliogenesis

    Directory of Open Access Journals (Sweden)

    Geissy LL Araújo

    2014-03-01

    Full Text Available The morphogen Sonic Hedgehog (SHH plays a critical role in the development of different tissues. In the central nervous system, SHH is well known to contribute to the patterning of the spinal cord and separation of the brain hemispheres. In addition, it has recently been shown that SHH signaling also contributes to the patterning of the telencephalon and establishment of adult neurogenic niches. In this work, we investigated whether SHH signaling influences the behavior of neural progenitors isolated from the dorsal telencephalon, which generate excitatory neurons and macroglial cells in vitro. We observed that SHH increases proliferation of cortical progenitors and generation of astrocytes, whereas blocking SHH signaling with cyclopamine has opposite effects. In both cases, generation of neurons did not seem to be affected. However, cell survival was broadly affected by blockade of SHH signaling. SHH effects were related to three different cell phenomena: mode of cell division, cell cycle length and cell growth. Together, our data in vitro demonstrate that SHH signaling controls cell behaviors that are important for proliferation of cerebral cortex progenitors, as well as differentiation and survival of neurons and astroglial cells.

  1. The Relationship between Cell Number, Division Behavior and Developmental Potential of Cleavage Stage Human Embryos: A Time-Lapse Study

    Science.gov (United States)

    Gong, Fei; Lu, Changfu; Zhang, Shuoping; Lu, Guangxiu; Lin, Ge

    2016-01-01

    Day 3 cleavage embryo transfer is routine in many assisted reproductive technology centers today. Embryos are usually selected according to cell number, cell symmetry and fragmentation for transfer. Many studies have showed the relationship between cell number and embryo developmental potential. However, there is limited understanding of embryo division behavior and their association with embryo cell number and developmental potential. A retrospective and observational study was conducted to investigate how different division behaviors affect cell number and developmental potential of day 3 embryos by time-lapse imaging. Based on cell number at day 3, the embryos (from 104 IVF/intracytoplasmic sperm injection (ICSI) treatment cycles, n = 799) were classified as follows: less than 5 cells (10C; n = 42). Division behavior, morphokinetic parameters and blastocyst formation rate were analyzed in 5 groups of day 3 embryos with different cell numbers. In 10C embryos increased compared to 7–8C embryos (45.8%, 33.3% vs. 11.1%, respectively). In ≥5C embryos, FR and DC significantly reduced developmental potential, whereas 10C). In NB embryos, the cell cycle elongation or shortening was the main cause for abnormally low or high cell number, respectively. After excluding embryos with abnormal division behaviors, the developmental potential, implantation rate and live birth rate of day 3 embryos increased with cell number. PMID:27077739

  2. A pulse-chase strategy for EdU labelling assay is able to rapidly quantify cell division orientation.

    Science.gov (United States)

    Yin, Xiaofeng; Tsukaya, Hirokazu

    2016-09-01

    Measurement of the direction of cell division is an important, yet difficult, task to analyse how a plant organ acquires its final shape from an initially small group of cells. We introduce a method that rapidly and easily quantifies cell division direction and is applicable to all plant species. A pulse-chase strategy for 5-ethynyl-2'-deoxyuridine (EdU) labelling assay was established and was shown to be successful for leaves of Arabidopsis thaliana (Arabidopsis) and Juncus prismatocarpus. By optimization of the pulse and chase periods, most of the signals obtained were sets of daughter nuclei. For Arabidopsis, the optimal time was a 45-min pulse and a 7-h chase. For J. prismatocarpus, the optimal time was a 2-h pulse and a 13.5-h chase. The positions of the daughter nuclei were used to quantify cell division direction in the Arabidopsis leaf primordia. Overall, cell division along the proximal-distal axis was more frequent than along the medial-lateral axis. In petiole, major vein, minor vein and margin areas, the major cell division direction seemed to be coincident with the direction of auxin flow. The advantages of our method over the few methods used previously are discussed. We anticipate that it will provide opportunities to study plant development in the near future. PMID:27121010

  3. Influence of hydroxyurea on cell divisions and microtubular cytoskeleton in Allium cepa root meristem

    Directory of Open Access Journals (Sweden)

    H. Q. Zhang

    2014-02-01

    Full Text Available In onion roots, hydroxyurea (HU causes a gradual depression of mitotic activity which ceases after 24-36 hrs. The effect is reversible; divisions begin after several hours of recovery and after 12-14 hrs about 90% cells undergo mitosis. Mitotic activity commences in the distal region of the apical meristem, and as a wave it spreads towards the apex. In the roots treated with HU for a short time, the tubulin immunofluorescence method reveals normal arrays of microtubules (MTs. After 36 hrs of HU treatment there are only cortical and endocytoplasmatic MTs. In the recovering roots, preprophase bands (PB mitotic spindles and phragmoplasts appear. Some PBs are split into two parallel rings. These abnormal PBs mostly occur in elongated cells. Apart from this, HU does not appear to have any significant influence on microtubular organization.

  4. Toward Spatially Regulated Division of Protocells: Insights into the E. coli Min System from in Vitro Studies

    OpenAIRE

    Simon Kretschmer; Petra Schwille

    2014-01-01

    For reconstruction of controlled cell division in a minimal cell model, or protocell , a positioning mechanism that spatially regulates division is indispensable. In Escherichia coli , the Min protein s oscillate from pole to pole to determine the division site by inhibition of the primary divisome protein FtsZ anywhere but in the cell middle. Remarkably, when reconstituted under defined conditions in vitro , the Min proteins self - organize into spatiotem poral patterns in the presence of a ...

  5. Par1b links lumen polarity with LGN-NuMA positioning for distinct epithelial cell division phenotypes

    NARCIS (Netherlands)

    Lazaro-Dieguez, Francisco; Cohen, David; Fernandez, Dawn; Hodgson, Louis; van IJzendoorn, Sven C. D.; Muesch, Anne

    2013-01-01

    Columnar epithelia establish their luminal domains and their mitotic spindles parallel to the basal surface and undergo symmetric cell divisions in which the cleavage furrow bisects the apical domain. Hepatocyte lumina interrupt the lateral domain of neighboring cells perpendicular to two basal doma

  6. Role of FtsEX in cell division of Escherichia coli: viability of ftsEX mutants is dependent on functional SufI or high osmotic strength.

    Science.gov (United States)

    Reddy, Manjula

    2007-01-01

    In Escherichia coli, at least 12 proteins, FtsZ, ZipA, FtsA, FtsE/X, FtsK, FtsQ, FtsL, FtsB, FtsW, FtsI, FtsN, and AmiC, are known to localize to the septal ring in an interdependent and sequential pathway to coordinate the septum formation at the midcell. The FtsEX complex is the latest recruit of this pathway, and unlike other division proteins, it is shown to be essential only on low-salt media. In this study, it is shown that ftsEX null mutations are not only salt remedial but also osmoremedial, which suggests that FtsEX may not be involved in salt transport as previously thought. Increased coexpression of cell division proteins FtsQ-FtsA-FtsZ or FtsN alone restored the growth defects of ftsEX mutants. ftsEX deletion exacerbated the defects of most of the mutants affected in Z ring localization and septal assembly; however, the ftsZ84 allele was a weak suppressor of ftsEX. The viability of ftsEX mutants in high-osmolarity conditions was shown to be dependent on the presence of a periplasmic protein, SufI, a substrate of twin-arginine translocase. In addition, SufI in multiple copies could substitute for the functions of FtsEX. Taken together, these results suggest that FtsE and FtsX are absolutely required for the process of cell division in conditions of low osmotic strength for the stability of the septal ring assembly and that, during high-osmolarity conditions, the FtsEX and SufI functions are redundant for this essential process. PMID:17071757

  7. Dominant-Negative Effects of Adult-Onset Huntingtin Mutations Alter the Division of Human Embryonic Stem Cells-Derived Neural Cells

    Science.gov (United States)

    Lopes, Carla; Aubert, Sophie; Bourgois-Rocha, Fany; Barnat, Monia; Rego, Ana Cristina; Déglon, Nicole

    2016-01-01

    Mutations of the huntingtin protein (HTT) gene underlie both adult-onset and juvenile forms of Huntington’s disease (HD). HTT modulates mitotic spindle orientation and cell fate in mouse cortical progenitors from the ventricular zone. Using human embryonic stem cells (hESC) characterized as carrying mutations associated with adult-onset disease during pre-implantation genetic diagnosis, we investigated the influence of human HTT and of an adult-onset HD mutation on mitotic spindle orientation in human neural stem cells (NSCs) derived from hESCs. The RNAi-mediated silencing of both HTT alleles in neural stem cells derived from hESCs disrupted spindle orientation and led to the mislocalization of dynein, the p150Glued subunit of dynactin and the large nuclear mitotic apparatus (NuMA) protein. We also investigated the effect of the adult-onset HD mutation on the role of HTT during spindle orientation in NSCs derived from HD-hESCs. By combining SNP-targeting allele-specific silencing and gain-of-function approaches, we showed that a 46-glutamine expansion in human HTT was sufficient for a dominant-negative effect on spindle orientation and changes in the distribution within the spindle pole and the cell cortex of dynein, p150Glued and NuMA in neural cells. Thus, neural derivatives of disease-specific human pluripotent stem cells constitute a relevant biological resource for exploring the impact of adult-onset HD mutations of the HTT gene on the division of neural progenitors, with potential applications in HD drug discovery targeting HTT-dynein-p150Glued complex interactions. PMID:26863614

  8. The cell birth marker BrdU does not affect recruitment of subsequent cell divisions in the adult avian brain.

    Science.gov (United States)

    Cattan, Anat; Ayali, Amir; Barnea, Anat

    2015-01-01

    BrdU is commonly used to quantify neurogenesis but also causes mutation and has mitogenic, transcriptional, and translational effects. In mammalian studies, attention had been given to its dosage, but in birds such examination was not conducted. Our previous study suggested that BrdU might affect subsequent cell divisions and neuronal recruitment in the brain. Furthermore, this effect seemed to increase with time from treatment. Accordingly, we examined whether BrdU might alter neurogenesis in the adult avian brain. We compared recruitment of [(3)H]-thymidine(+) neurons in brains of zebra finches (Taeniopygia guttata) when no BrdU was involved and when BrdU was given 1 or 3 months prior to [(3)H]-thymidine. In nidopallium caudale, HVC, and hippocampus, no differences were found between groups in densities and percentages of [(3)H]-thymidine(+) neurons. The number of silver grains per [(3)H]-thymidine(+) neuronal nucleus and their distribution were similar across groups. Additionally, time did not affect the results. The results indicate that the commonly used dosage of BrdU in birds has no long-term effects on subsequent cell divisions and neuronal recruitment. This conclusion is also important in neuronal replacement experiments, where BrdU and another cell birth marker are given, with relatively long intervals between them. PMID:25759813

  9. ALIX and ESCRT-III Coordinately Control Cytokinetic Abscission during Germline Stem Cell Division In Vivo

    Science.gov (United States)

    Eikenes, Åsmund H.; Malerød, Lene; Christensen, Anette Lie; Steen, Chloé B.; Mathieu, Juliette; Nezis, Ioannis P.; Liestøl, Knut; Huynh, Jean-René; Stenmark, Harald; Haglund, Kaisa

    2015-01-01

    Abscission is the final step of cytokinesis that involves the cleavage of the intercellular bridge connecting the two daughter cells. Recent studies have given novel insight into the spatiotemporal regulation and molecular mechanisms controlling abscission in cultured yeast and human cells. The mechanisms of abscission in living metazoan tissues are however not well understood. Here we show that ALIX and the ESCRT-III component Shrub are required for completion of abscission during Drosophila female germline stem cell (fGSC) division. Loss of ALIX or Shrub function in fGSCs leads to delayed abscission and the consequent formation of stem cysts in which chains of daughter cells remain interconnected to the fGSC via midbody rings and fusome. We demonstrate that ALIX and Shrub interact and that they co-localize at midbody rings and midbodies during cytokinetic abscission in fGSCs. Mechanistically, we show that the direct interaction between ALIX and Shrub is required to ensure cytokinesis completion with normal kinetics in fGSCs. We conclude that ALIX and ESCRT-III coordinately control abscission in Drosophila fGSCs and that their complex formation is required for accurate abscission timing in GSCs in vivo. PMID:25635693

  10. Single-cell analysis reveals a novel uncultivated magnetotactic bacterium within the candidate division OP3.

    Science.gov (United States)

    Kolinko, Sebastian; Jogler, Christian; Katzmann, Emanuel; Wanner, Gerhard; Peplies, Jörg; Schüler, Dirk

    2012-07-01

    Magnetotactic bacteria (MTB) are a diverse group of prokaryotes that orient along magnetic fields using membrane-coated magnetic nanocrystals of magnetite (Fe(3) O(4) ) or greigite (Fe(3) S(4) ), the magnetosomes. Previous phylogenetic analysis of MTB has been limited to few cultivated species and most abundant members of natural populations, which were assigned to Proteobacteria and the Nitrospirae phyla. Here, we describe a single cell-based approach that allowed the targeted phylogenetic and ultrastructural analysis of the magnetotactic bacterium SKK-01, which was low abundant in sediments of Lake Chiemsee. Morphologically conspicuous single cells of SKK-01 were micromanipulated from magnetically collected multi-species MTB populations, which was followed by whole genome amplification and ultrastructural analysis of sorted cells. Besides intracellular sulphur inclusions, the large ovoid cells of SKK-01 harbour ∼175 bullet-shaped magnetosomes arranged in multiple chains that consist of magnetite as revealed by TEM and EDX analysis. Sequence analysis of 16 and 23S rRNA genes from amplified genomic DNA as well as fluorescence in situ hybridization assigned SKK-01 to the candidate division OP3, which so far lacks any cultivated representatives. SKK-01 represents the first morphotype that can be assigned to the OP3 group as well as the first magnetotactic member of the PVC superphylum. PMID:22003954

  11. Adaptive drug resistance mediated by root-nodulation-cell division efflux pumps.

    Science.gov (United States)

    Daniels, C; Ramos, J L

    2009-01-01

    Bacterial resistance to antibiotics is a major therapeutic problem. Bacteria use the same mechanisms for developing resistance to antibiotics as they do for developing resistance to biocide compounds present in some cleaning and personal care products. Root-nodulation-cell division (RND) family efflux pumps are a common means of multidrug resistance, and induction of their expression can explain the observed cross-resistance found between antibiotics and biocides in laboratory strains. Hence, there is a relationship between the active chemicals used in household products, organic solvents and antibiotics. The widespread use of biocide-containing modern-day household products may promote the development of microbial resistance and, in particular, cross-resistance to antibiotics. PMID:19220351

  12. Human disc cells in monolayer vs 3D culture: cell shape, division and matrix formation

    Directory of Open Access Journals (Sweden)

    Hanley Edward N

    2000-10-01

    Full Text Available Abstract Background The relationship between cell shape, proliferation, and extracellular matrix (ECM production, important aspects of cell behavior, is examined in a little-studied cell type, the human annulus cell from the intervertebral disc, during monolayer vs three-dimensional (3D culture. Results Three experimental studies showed that cells respond specifically to culture microenvironments by changes in cell shape, mitosis and ECM production: 1 Cell passages showed extensive immunohistochemical evidence of Type I and II collagens only in 3D culture. Chondroitin sulfate and keratan sulfate were abundant in both monolayer and 3D cultures. 2 Cells showed significantly greater proliferation in monolayer in the presence of platelet-derived growth factor compared to cells in 3D. 3 Cells on Matrigel™-coated monolayer substrates became rounded and formed nodular colonies, a finding absent during monolayer growth. Conclusions The cell's in vivo interactions with the ECM can regulate shape, gene expression and other cell functions. The shape of the annulus cell changes markedly during life: the young, healthy disc contains spindle shaped cells and abundant collagen. With aging and degeneration, many cells assume a strikingly different appearance, become rounded and are surrounded by unusual accumulations of ECM products. In vitro manipulation of disc cells provides an experimental window for testing how disc cells from given individuals respond when they are grown in environments which direct cells to have either spindle- or rounded-shapes. In vitro assessment of the response of such cells to platelet-derived growth factor and to Matrigel™ showed a continued influence of cell shape even in the presence of a growth factor stimulus. These findings contribute new information to the important issue of the influence of cell shape on cell behavior.

  13. From cell differentiation to cell collectives : Bacillus subtilis uses division of labor to migrate

    NARCIS (Netherlands)

    van Gestel, Jordi; Vlamakis, Hera; Kolter, Roberto

    2015-01-01

    The organization of cells, emerging from cell-cell interactions, can give rise to collective properties. These properties are adaptive when together cells can face environmental challenges that they separately cannot. One particular challenge that is important for microorganisms is migration. In thi

  14. Detecting protein-protein interactions in living cells

    DEFF Research Database (Denmark)

    Gottschalk, Marie; Bach, Anders; Hansen, Jakob Lerche;

    2009-01-01

    to the endogenous C-terminal peptide of the NMDA receptor, as evaluated by a cell-free protein-protein interaction assay. However, it is important to address both membrane permeability and effect in living cells. Therefore a bioluminescence resonance energy transfer (BRET) assay was established, where the C...

  15. Characterization of a null allelic mutant of the rice NAL1 gene reveals its role in regulating cell division.

    Directory of Open Access Journals (Sweden)

    Dan Jiang

    Full Text Available Leaf morphology is closely associated with cell division. In rice, mutations in Narrow leaf 1 (NAL1 show narrow leaf phenotypes. Previous studies have shown that NAL1 plays a role in regulating vein patterning and increasing grain yield in indica cultivars, but its role in leaf growth and development remains unknown. In this report, we characterized two allelic mutants of NARROW LEAF1 (NAL1, nal1-2 and nal1-3, both of which showed a 50% reduction in leaf width and length, as well as a dwarf culm. Longitudinal and transverse histological analyses of leaves and internodes revealed that cell division was suppressed in the anticlinal orientation but enhanced in the periclinal orientation in the mutants, while cell size remained unaltered. In addition to defects in cell proliferation, the mutants showed abnormal midrib in leaves. Map-based cloning revealed that nal1-2 is a null allelic mutant of NAL1 since both the whole promoter and a 404-bp fragment in the first exon of NAL1 were deleted, and that a 6-bp fragment was deleted in the mutant nal1-3. We demonstrated that NAL1 functions in the regulation of cell division as early as during leaf primordia initiation. The altered transcript level of G1- and S-phase-specific genes suggested that NAL1 affects cell cycle regulation. Heterogeneous expression of NAL1 in fission yeast (Schizosaccharomyces pombe further supported that NAL1 affects cell division. These results suggest that NAL1 controls leaf width and plant height through its effects on cell division.

  16. A SCARECROW-RETINOBLASTOMA protein network controls protective quiescence in the Arabidopsis root stem cell organizer.

    OpenAIRE

    Alfredo Cruz-Ramírez; Sara Díaz-Triviño; Guy Wachsman; Yujuan Du; Mario Arteága-Vázquez; Hongtao Zhang; Rene Benjamins; Ikram Blilou; Neef, Anne B.; Vicki Chandler; Ben Scheres

    2013-01-01

    Author Summary In the plant Arabidposis thaliana, root meristems (in the growing tip of the root) contain slowly dividing cells that act as an organizing center for the root stem cells that surround them. This centre is called the quiescent centre (QC). In this study, we show that the slow rate of division in the QC is regulated by the interaction between two proteins: Retinoblastoma homolog (RBR) and SCARECROW (SCR), a transcription factor that controls stem cell maintenance. RBR and SCR reg...

  17. Scaffolding during the cell cycle by A-kinase anchoring proteins

    OpenAIRE

    Han, B.; Poppinga, W J; Schmidt, M.

    2015-01-01

    Cell division relies on coordinated regulation of the cell cycle. A process including a well-defined series of strictly regulated molecular mechanisms involving cyclin-dependent kinases, retinoblastoma protein, and polo-like kinases. Dysfunctions in cell cycle regulation are associated with disease such as cancer, diabetes, and neurodegeneration. Compartmentalization of cellular signaling is a common strategy used to ensure the accuracy and efficiency of cellular responses. Compartmentalizati...

  18. Effect of gamma-irradiation and colchicine on cell division and differentiation of xylem elements in citrus limon juice vesicle cultures

    International Nuclear Information System (INIS)

    The effects of varying doses of gamma irradiation on cell division and cytodifferentiation of tracheary elements in cultured juice vesicles of Citrus limon (L) Burmann var. Assam lemon were investigated. Low radiation doses stimulated cell division and differentiation of xylem fibres, sclereids and tracheids in explants given up to 10 Gy of gamma rays. Although cell division and cytodifferentiation of fibers and sclereids occurred in explants exposed to 150 dose of Gy radiation, the intensity of differentiation was much less than that induced by 10 Gy radiation dose. Amongst the differential elements, tracheids were more sensitive to radiation than fibres and sclereids. The requirement of cell division for differentiation of xylem cells was also studied by using different concentrations of colchicine in Citrus limon juice vesicle cultures. It was found that the low concentrations of colchicine permitted normal cell division and also resulted in normal differentiation of xylem cells; higher colchicine concentration, however, inhibited cell division as well as differentiation and resulted in an abnormal differentiation of tracheary element. A positive correlation between intensity of nucleic acid staining and cell division in both the above-mentioned experiments was qualitatively confirmed by Azur B staining test of nucleic acid. Thus, it was concluded that juice vesicle parenchyma cells go through nucleic acid synthesis, followed by cell division before differentiation. (author)

  19. Intracellular photoreceptive site for blue light-induced cell division in protonemata of the fern Adiantum [Pteridophyta]: Further analyses by polarized light irradiation and cell centrifugation

    International Nuclear Information System (INIS)

    The intracellular localization of the photoreceptive site for blue light-induced cell division in single-celled protonemata of Adiantum capillus-veneris L. was investigated using polarized light irradiation and protonemal cell centrifugation. The response to irradiation with polarized blue light showed no dependence on the direction of light polarization. However, centrifugation of the protonemata followed by microbeam irradiation showed that the site of blue light perception could be displaced together with the nucleus. Centrifugal treatment changed the distribution of intracellular organelles at the time of light exposure and basipetally displaced the nucleus about 90μm. This treatment had no effect on the induction of cell division with blue light if the protonemata were centrifuged again acropetally after the light treatment. Microbeam (30×30 μm2) irradiation with blue light of the apical 45–75 βm region, the receptive site of blue light in non-centrifuged cell, did not induce cell division. However, cell division was induced by irradiation of the nucleus-containing region, indicating that the photoreceptive site was displaced together with the nucleus by the centrifugation. These results suggest that the blue light receptor regulating cell division in Adiantum protonemata is not likely to be located on the plasma membrane. (author)

  20. Sensory mother cell division is specifically affected in a Cyclin-A mutant of Drosophila melanogaster.

    OpenAIRE

    Ueda, R; Togashi, S; Takahisa, M; Tsurumura, S; Mikuni, M; Kondo, K.(Yamagata University, Yamagata, 992-8510, Japan); Miyake, T

    1992-01-01

    Cyclin proteins are one of the important components of the mechanism regulating mitosis in eukaryotic cells. We isolated a Drosophila Cyclin-A mutant in which the progenitor cells of the peripheral nervous system (the sensory mother cells) do not divide properly, causing the loss and other abnormalities of mechanosensory organs in the adult fly. Sequence analysis of the mutant genome reveals that a P element is inserted into the first intron of the Cyclin-A gene. A 13 kb wild-type genomic DNA...

  1. Temperature stress promotes cell division arrest in Xanthomonas citri subsp. citri.

    Science.gov (United States)

    Sumares, Júlia A P; Morão, Luana Galvão; Martins, Paula M M; Martins, Daniela A B; Gomes, Eleni; Belasque, José; Ferreira, Henrique

    2016-04-01

    Citrus canker is an economically important disease that affects orange production in some of the most important producing areas around the world. It represents a great threat to the Brazilian and North American citriculture, particularly to the states of São Paulo and Florida, which together correspond to the biggest orange juice producers in the world. The etiological agent of this disease is the Gram-negative bacterium Xanthomonas citri subsp. citri (Xcc), which grows optimally in laboratory cultures at ~30°C. To investigate how temperatures differing from 30°C influence the development of Xcc, we subjected the bacterium to thermal stresses, and afterward scored its recovery capability. In addition, we analyzed cell morphology and some markers of essential cellular processes that could indicate the extent of the heat-induced damage. We found that the exposure of Xcc to 37°C for a period of 6 h led to a cell cycle arrest at the division stage. Thermal stress might have also interfered with the DNA replication and/or the chromosome segregation apparatuses, since cells displayed an increased number of sister origins side-by-side within rods. Additionally, Xcc treated at 37°C was still able to induce citrus canker symptoms, showing that thermal stress did not affect the ability of Xcc to colonize the host citrus. At 40-42°C, Xcc lost viability and became unable to induce disease symptoms in citrus. Our results provide evidence about essential cellular mechanisms perturbed by temperature, and can be potentially explored as a new method for Xanthomonas citri synchronization in cell cycle studies, as well as for the sanitation of plant material. PMID:26663580

  2. Ciprofloxacin Derivatives Affect Parasite Cell Division and Increase the Survival of Mice Infected with Toxoplasma gondii.

    Directory of Open Access Journals (Sweden)

    Erica S Martins-Duarte

    Full Text Available Toxoplasmosis, caused by the protozoan Toxoplasma gondii, is a worldwide disease whose clinical manifestations include encephalitis and congenital malformations in newborns. Previously, we described the synthesis of new ethyl-ester derivatives of the antibiotic ciprofloxacin with ~40-fold increased activity against T. gondii in vitro, compared with the original compound. Cipro derivatives are expected to target the parasite's DNA gyrase complex in the apicoplast. The activity of these compounds in vivo, as well as their mode of action, remained thus far uncharacterized. Here, we examined the activity of the Cipro derivatives in vivo, in a model of acute murine toxoplasmosis. In addition, we investigated the cellular effects T. gondii tachyzoites in vitro, by immunofluorescence and transmission electron microscopy (TEM. When compared with Cipro treatment, 7-day treatments with Cipro derivatives increased mouse survival significantly, with 13-25% of mice surviving for up to 60 days post-infection (vs. complete lethality 10 days post-infection, with Cipro treatment. Light microscopy examination early (6 and 24h post-infection revealed that 6-h treatments with Cipro derivatives inhibited the initial event of parasite cell division inside host cells, in an irreversible manner. By TEM and immunofluorescence, the main cellular effects observed after treatment with Cipro derivatives and Cipro were cell scission inhibition--with the appearance of 'tethered' parasites--malformation of the inner membrane complex, and apicoplast enlargement and missegregation. Interestingly, tethered daughter cells resulting from Cipro derivatives, and also Cipro, treatment did not show MORN1 cap or centrocone localization. The biological activity of Cipro derivatives against C. parvum, an apicomplexan species that lacks the apicoplast, is, approximately, 50 fold lower than that in T. gondii tachyzoites, supporting that these compounds targets the apicoplast. Our results

  3. Versatile protein tagging in cells with split fluorescent protein

    OpenAIRE

    Kamiyama, Daichi; Sekine, Sayaka; Barsi-Rhyne, Benjamin; Hu, Jeffrey; Chen, Baohui; Gilbert, Luke A.; Ishikawa, Hiroaki; Leonetti, Manuel D.; Marshall, Wallace F.; Jonathan S Weissman; Huang, Bo

    2016-01-01

    In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respec...

  4. Structural determinants underlying the temperature-sensitive nature of a Galpha mutant in asymmetric cell division of Caenorhabditis elegans.

    Science.gov (United States)

    Johnston, Christopher A; Afshar, Katayoun; Snyder, Jason T; Tall, Gregory G; Gönczy, Pierre; Siderovski, David P; Willard, Francis S

    2008-08-01

    Heterotrimeric G-proteins are integral to a conserved regulatory module that influences metazoan asymmetric cell division (ACD). In the Caenorhabditis elegans zygote, GOA-1 (Galpha(o)) and GPA-16 (Galpha(i)) are involved in generating forces that pull on astral microtubules and position the spindle asymmetrically. GPA-16 function has been analyzed in vivo owing notably to a temperature-sensitive allele gpa-16(it143), which, at the restrictive temperature, results in spindle orientation defects in early embryos. Here we identify the structural basis of gpa-16(it143), which encodes a point mutation (G202D) in the switch II region of GPA-16. Using Galpha(i1)(G202D) as a model in biochemical analyses, we demonstrate that high temperature induces instability of the mutant Galpha. At the permissive temperature, the mutant Galpha was stable upon GTP binding, but switch II rearrangement was compromised, as were activation state-selective interactions with regulators involved in ACD, including GoLoco motifs, RGS proteins, and RIC-8. We solved the crystal structure of the mutant Galpha bound to GDP, which indicates a unique switch II conformation as well as steric constraints that suggest activated GPA-16(it143) is destabilized relative to wild type. Spindle severing in gpa-16(it143) embryos revealed that pulling forces are symmetric and markedly diminished at the restrictive temperature. Interestingly, pulling forces are asymmetric and generally similar in magnitude to wild type at the permissive temperature despite defects in the structure of GPA-16(it143). These normal pulling forces in gpa-16(it143) embryos at the permissive temperature were attributable to GOA-1 function, underscoring a complex interplay of Galpha subunit function in ACD. PMID:18519563

  5. Assessment of three Resistance-Nodulation-Cell Division drug efflux transporters of Burkholderia cenocepacia in intrinsic antibiotic resistance

    Directory of Open Access Journals (Sweden)

    Venturi Vittorio

    2009-09-01

    Full Text Available Abstract Background Burkholderia cenocepacia are opportunistic Gram-negative bacteria that can cause chronic pulmonary infections in patients with cystic fibrosis. These bacteria demonstrate a high-level of intrinsic antibiotic resistance to most clinically useful antibiotics complicating treatment. We previously identified 14 genes encoding putative Resistance-Nodulation-Cell Division (RND efflux pumps in the genome of B. cenocepacia J2315, but the contribution of these pumps to the intrinsic drug resistance of this bacterium remains unclear. Results To investigate the contribution of efflux pumps to intrinsic drug resistance of B. cenocepacia J2315, we deleted 3 operons encoding the putative RND transporters RND-1, RND-3, and RND-4 containing the genes BCAS0591-BCAS0593, BCAL1674-BCAL1676, and BCAL2822-BCAL2820. Each deletion included the genes encoding the RND transporter itself and those encoding predicted periplasmic proteins and outer membrane pores. In addition, the deletion of rnd-3 also included BCAL1672, encoding a putative TetR regulator. The B. cenocepacia rnd-3 and rnd-4 mutants demonstrated increased sensitivity to inhibitory compounds, suggesting an involvement of these proteins in drug resistance. Moreover, the rnd-3 and rnd-4 mutants demonstrated reduced accumulation of N-acyl homoserine lactones in the growth medium. In contrast, deletion of the rnd-1 operon had no detectable phenotypes under the conditions assayed. Conclusion Two of the three inactivated RND efflux pumps in B. cenocepacia J2315 contribute to the high level of intrinsic resistance of this strain to some antibiotics and other inhibitory compounds. Furthermore, these efflux systems also mediate accumulation in the growth medium of quorum sensing molecules that have been shown to contribute to infection. A systematic study of RND efflux systems in B. cenocepacia is required to provide a full picture of intrinsic antibiotic resistance in this opportunistic

  6. Different Degree in Proteasome Malfunction Has Various Effects on Root Growth Possibly through Preventing Cell Division and Promoting Autophagic Vacuolization

    OpenAIRE

    Xianyong Sheng; Qian Wei; Liping Jiang; Xue Li; Yuan Gao; Li Wang

    2012-01-01

    The ubiquitin/proteasome pathway plays a vital role in plant development. But the effects of proteasome malfunction on root growth, and the mechanism underlying this involvement remains unclear. In the present study, the effects of proteasome inhibitors on Arabidopsis root growth were studied through the analysis of the root length, and meristem size and cell length in maturation zone using FM4-64, and cell-division potential using GFP fusion cyclin B, and accumulation of ubiquitinated protei...

  7. Function and regulation of Aurora/Ipl1p kinase family in cell division

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    During mitosis, the parent cell distributes its genetic materials equally into two daughter cells through chromosome segregation, a complex movements orchestrated by mitotic kinases and its effector proteins.Faithful chromosome segregation and cytokinesis ensure that each daughter cell receives a full copy of genetic materials of parent cell. Defects in these processes can lead to aneuploidy or polyploidy. Aurora/Ipl1p fanily,a class of conserved serine/threonine kinases, plays key roles in chromosome segregation and cytokinesis.This article highlights the function and regulation of Aurora/Ipl1p family in mitosis and provides potential links between aberrant regulation of Aurora/Ipl1p kinases and pathogenesis of human cancer.

  8. Brassinazole resistant 1 (BZR1)-dependent brassinosteroid signalling pathway leads to ectopic activation of quiescent cell division and suppresses columella stem cell differentiation.

    Science.gov (United States)

    Lee, Hak-Soo; Kim, Yoon; Pham, Giang; Kim, Ju Won; Song, Ji-Hye; Lee, Yew; Hwang, Yong-Sic; Roux, Stanley J; Kim, Soo-Hwan

    2015-08-01

    Previous publications have shown that BRI1 EMS suppressor 1 (BES1), a positive regulator of the brassinosteroid (BR) signalling pathway, enhances cell divisions in the quiescent centre (QC) and stimulates columella stem cell differentiation. Here, it is demonstrated that BZR1, a BES1 homologue, also promotes cell divisions in the QC, but it suppresses columella stem cell differentiation, opposite to the action of BES1. In addition, BR and its BZR1-mediated signalling pathway are shown to alter the expression/subcellular distribution of pin-formed (PINs), which may result in changes in auxin movement. BR promotes intense nuclear accumulation of BZR1 in the root tip area, and the binding of BZR1 to the promoters of several root development-regulating genes, modulating their expression in the root stem cell niche area. These BZR1-mediated signalling cascades may account for both the ectopic activation of QC cell divisions as well as the suppression of the columella stem cell differentiation. They could also inhibit auxin-dependent distal stem cell differentiation by antagonizing the auxin/WOX5-dependent pathway. In conclusion, BZR1-/BES1-mediated BR signalling pathways show differential effects on the maintenance of root apical meristem activities: they stimulate ectopic QC division while they show opposite effects on the differentiation of distal columella stem cells in a BR concentration- and BZR1-/BES1-dependent manner. PMID:26136267

  9. Asymmetric cell division and its role in cell fate determination in the green alga Tetraselmis indica

    Digital Repository Service at National Institute of Oceanography (India)

    Arora, M.; Anil, A.C.; Burgess, K.; Delany, J.E.; Mesbahi, E.

    The prasinophytes (early diverging Chlorophyta), consisting of simple unicellular green algae, occupy a critical position at the base of the green algal tree of life, with some of its representatives viewed as the cell form most similar to the first...

  10. Emp is a component of the nuclear matrix of mammalian cells and undergoes dynamic rearrangements during cell division

    International Nuclear Information System (INIS)

    Emp, originally detected in erythroblastic islands, is expressed in numerous cell types and tissues suggesting a functionality not limited to hematopoiesis. To study the function of Emp in non-hematopoietic cells, an epitope-tagged recombinant human Emp was expressed in HEK cells. Preliminary studies revealed that Emp partitioned into both the nuclear and Triton X-100-insoluble cytoskeletal fractions in approximately a 4:1 ratio. In this study, we report investigations of Emp in the nucleus. Sequential extractions of interphase nuclei showed that recombinant Emp was present predominantly in the nuclear matrix. Immunofluorescence microscopy showed that Emp was present in typical nuclear speckles enriched with the spliceosome assembly factor SC35 and partially co-localized with actin staining. Coimmunoprecipitation and GST-pull-down assays confirmed the apparent close association of Emp with nuclear actin. During mitosis, Emp was detected at the mitotic spindle/spindle poles, as well as in the contractile ring during cytokinesis. These results suggest that Emp undergoes dynamic rearrangements within the nuclear architecture that are correlated with cell division

  11. A self-associating protein critical for chromosome attachment, division, and polar organization in caulobacter

    DEFF Research Database (Denmark)

    Ebersbach, Gitte; Briegel, Ariane; Jensen, Grant J; Jacobs-Wagner, Christine; Charbon, Gitte Ebersbach

    2008-01-01

    Cell polarization is an integral part of many unrelated bacterial processes. How intrinsic cell polarization is achieved is poorly understood. Here, we provide evidence that Caulobacter crescentus uses a multimeric pole-organizing factor (PopZ) that serves as a hub to concurrently achieve several...... suggests that localization of PopZ largely relies on PopZ multimerization in chromosome-free regions, consistent with a self-organizing mechanism.......Cell polarization is an integral part of many unrelated bacterial processes. How intrinsic cell polarization is achieved is poorly understood. Here, we provide evidence that Caulobacter crescentus uses a multimeric pole-organizing factor (PopZ) that serves as a hub to concurrently achieve several...

  12. Cell division factors from crown gall tumors: a strategy for structural elucidation

    International Nuclear Information System (INIS)

    Mitogenic compounds present in extracts of Vinca rosea crown gall tumor tissue were investigated. An isolation procedure, consisting of solvent partitions and reverse phase chromatography, has yielded a group of isomeric compounds which show activity in the tobacco pith bioassay. Initial characterizations revealed an unsaturated base, a sugar residue, a β-linked glucose, an allylic alcohol, and two methyl groups. A two part strategy of mass spectrometry (MS) in combination with proton nuclear magnetic resonance (1H NMR) was envisioned. The aglycone structure would be determined by MS and the regiochemical relationships among the structural units would be defined by 1H NMR data. The utility of this approach was demonstrated by the structure assignment of a specific inhibitor of β-D-glucuronidase, 2(S)-carboxy-3(R),4(R),5(S)-trihydroxypiperidine. The relative stereochemistry of the hydroxyls was revealed by 1H NMR and the absolute configuration was deduced by a comparison of Cotton effects with a model compound. The use of 1H NMR to establish regiochemical relationships was investigated. Terpenes containing quaternary carbons and methyl groups were excellent models for the regiochemical problems presented by the mitogenic factors. This 1H NMR spectroscopy has been applied to the cell division factor structure problem. These data, with information from two dimensional nOe experiments, have defined some of the regio-relationships among the structural units present in the isolated factors

  13. Cell division cycle 25 homolog c effects on low-dose hyper-radiosensitivity and induced radioresistance at elevated dosage in A549 cells

    International Nuclear Information System (INIS)

    The underlying mechanisms behind both low-dose hyper-radiosensitivity (HRS) and induced radioresistance (IRR), generally occurring at elevated radiation levels, remain unclear; however, elucidation of the relationship between cell cycle division 25 homolog c (Cdc25c) phosphatase and HRS/IRR may provide important insights into this process. Two cell lines with disparate HRS status, A549 and SiHa cells, were selected as cell models for comparison of dose-dependent Cdc25c phosphatase expression subsequent to low-dose irradiation. Knockdown of Cdc25c in A549 cells was mediated by transfection with a pGCsi-RAN-U6neo vector containing hairpin siRNA sequences. S216-phosphorylated Cdc25c protein [p-Cdc25c (Ser216)], cell survival and mitotic ratio were measured by western blot, colony-forming assay and histone H3 phosphorylation analysis. Variant p-Cdc25c (Ser216) expression was observed in the two cell lines after irradiation. The p-Cdc25c (Ser216) expression noted in SiHa cells after administration of 0-1 Gy radiation was similar to the radioresistance model; however, in A549 cells, the dose response for the phosphorylation of the Cdc25c Ser216 residue overlapped the level required to overcome the HRS response. Furthermore, Cdc25c repression prior to low-dose radiation induced more distinct HRS and prevented the development of IRR. The dose required to overcome the HRS response coincided with the effect of early G2-phase checkpoint arrest in A549 cells (approximately 0.3 Gy), and Cdc25c knockdown in A549 cells (approximately 0.5 Gy) corresponded to the phosphorylation of the Cdc25c Ser216 residue. Resultant data confirmed that dose-dependent Cdc25c phosphatase does effectively act as an early G2-phase checkpoint, thus indicating mechanistic importance in the HRS to IRR transition in A549 cells. (author)

  14. The simulation model of growth and cell divisions for the root apex with an apical cell in application to Azolla pinnata.

    Science.gov (United States)

    Piekarska-Stachowiak, Anna; Nakielski, Jerzy

    2013-12-01

    In contrast to seed plants, the roots of most ferns have a single apical cell which is the ultimate source of all cells in the root. The apical cell has a tetrahedral shape and divides asymmetrically. The root cap derives from the distal division face, while merophytes derived from three proximal division faces contribute to the root proper. The merophytes are produced sequentially forming three sectors along a helix around the root axis. During development, they divide and differentiate in a predictable pattern. Such growth causes cell pattern of the root apex to be remarkably regular and self-perpetuating. The nature of this regularity remains unknown. This paper shows the 2D simulation model for growth of the root apex with the apical cell in application to Azolla pinnata. The field of growth rates of the organ, prescribed by the model, is of a tensor type (symplastic growth) and cells divide taking principal growth directions into account. The simulations show how the cell pattern in a longitudinal section of the apex develops in time. The virtual root apex grows realistically and its cell pattern is similar to that observed in anatomical sections. The simulations indicate that the cell pattern regularity results from cell divisions which are oriented with respect to principal growth directions. Such divisions are essential for maintenance of peri-anticlinal arrangement of cell walls and coordinated growth of merophytes during the development. The highly specific division program that takes place in merophytes prior to differentiation seems to be regulated at the cellular level. PMID:23989670

  15. Nanoscale imaging of the growth and division of bacterial cells on planar substrates with the atomic force microscope

    Energy Technology Data Exchange (ETDEWEB)

    Van Der Hofstadt, M. [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Hüttener, M.; Juárez, A. [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Departament de Microbiologia, Universitat de Barcelona, Avinguda Diagonal 645, 08028 Barcelona (Spain); Gomila, G., E-mail: ggomila@ibecbarcelona.eu [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Departament d' Electronica, Universitat de Barcelona, C/ Marti i Franqués 1, 08028 Barcelona (Spain)

    2015-07-15

    With the use of the atomic force microscope (AFM), the Nanomicrobiology field has advanced drastically. Due to the complexity of imaging living bacterial processes in their natural growing environments, improvements have come to a standstill. Here we show the in situ nanoscale imaging of the growth and division of single bacterial cells on planar substrates with the atomic force microscope. To achieve this, we minimized the lateral shear forces responsible for the detachment of weakly adsorbed bacteria on planar substrates with the use of the so called dynamic jumping mode with very soft cantilever probes. With this approach, gentle imaging conditions can be maintained for long periods of time, enabling the continuous imaging of the bacterial cell growth and division, even on planar substrates. Present results offer the possibility to observe living processes of untrapped bacteria weakly attached to planar substrates. - Highlights: • Gelatine coatings used to weakly attach bacterial cells onto planar substrates. • Use of the dynamic jumping mode as a non-perturbing bacterial imaging mode. • Nanoscale resolution imaging of unperturbed single living bacterial cells. • Growth and division of single bacteria cells on planar substrates observed.

  16. Deliberate ROS production and auxin synergistically trigger the asymmetrical division generating the subsidiary cells in Zea mays stomatal complexes.

    Science.gov (United States)

    Livanos, Pantelis; Galatis, Basil; Apostolakos, Panagiotis

    2016-07-01

    Subsidiary cell generation in Poaceae is an outstanding example of local intercellular stimulation. An inductive stimulus emanates from the guard cell mother cells (GMCs) towards their laterally adjacent subsidiary cell mother cells (SMCs) and triggers the asymmetrical division of the latter. Indole-3-acetic acid (IAA) immunolocalization in Zea mays protoderm confirmed that the GMCs function as local sources of auxin and revealed that auxin is polarly accumulated between GMCs and SMCs in a timely-dependent manner. Besides, staining techniques showed that reactive oxygen species (ROS) exhibit a closely similar, also time-dependent, pattern of appearance suggesting ROS implication in subsidiary cell formation. This phenomenon was further investigated by using the specific NADPH-oxidase inhibitor diphenylene iodonium, the ROS scavenger N-acetyl-cysteine, menadione which leads to ROS overproduction, and H2O2. Treatments with diphenylene iodonium, N-acetyl-cysteine, and menadione specifically blocked SMC polarization and asymmetrical division. In contrast, H2O2 promoted the establishment of SMC polarity and subsequently subsidiary cell formation in "younger" protodermal areas. Surprisingly, H2O2 favored the asymmetrical division of the intervening cells of the stomatal rows leading to the creation of extra apical subsidiary cells. Moreover, H2O2 altered IAA localization, whereas synthetic auxin analogue 1-napthaleneacetic acid enhanced ROS accumulation. Combined treatments with ROS modulators along with 1-napthaleneacetic acid or 2,3,5-triiodobenzoic acid, an auxin efflux inhibitor, confirmed the crosstalk between ROS and auxin functioning during subsidiary cell generation. Collectively, our results demonstrate that ROS are critical partners of auxin during development of Z. mays stomatal complexes. The interplay between auxin and ROS seems to be spatially and temporarily regulated. PMID:26250135

  17. Metabolic behavior of cell surface biotinylated proteins

    International Nuclear Information System (INIS)

    The turnover of proteins on the surface of cultured mammalian cells was measured by a new approach. Reactive free amino or sulfhydryl groups on surface-accessible proteins were derivatized with biotinyl reagents and the proteins solubilized from culture dishes with detergent. Solubilized, biotinylated proteins were then adsorbed onto streptavidin-agarose, released with sodium dodecyl sulfate and mercaptoethanol, and separated on polyacrylamide gels. Biotin-epsilon-aminocaproic acid N-hydroxysuccinimide ester (BNHS) or N-biotinoyl-N'-(maleimidohexanoyl)hydrazine (BM) were the derivatizing agents. Only 10-12 bands were adsorbed onto streptavidin-agarose from undervatized cells or from derivatized cells treated with free avidin at 4 degrees C. Two-dimensional isoelectric focusing-sodium dodecyl sulfate gel electrophoresis resolved greater than 100 BNHS-derivatized proteins and greater than 40 BM-derivatized proteins. There appeared to be little overlap between the two groups of derivatized proteins. Short-term pulse-chase studies showed an accumulation of label into both groups of biotinylated proteins up until 1-2 h of chase and a rapid decrease over the next 1-5 h. Delayed appearance of labeled protein at the cell surface was attributed to transit time from site of synthesis. The unexpected and unexplained rapid disappearance of pulse-labeled proteins from the cell surface was invariant for all two-dimensionally resolved proteins and was sensitive to temperature reduction to 18 degrees C. Long-term pulse-chase experiments beginning 4-8 h after the initiation of chase showed the disappearance of derivatized proteins to be a simple first-order process having a half-life of 115 h in the case of BNHS-derivatized proteins and 30 h in the case of BM-derivatized proteins

  18. Asymmetric Wnt Pathway Signaling Facilitates Stem Cell-Like Divisions via the Nonreceptor Tyrosine Kinase FRK-1 in Caenorhabditis elegans.

    Science.gov (United States)

    Mila, Danielle; Calderon, Adriana; Baldwin, Austin T; Moore, Kelsey M; Watson, McLane; Phillips, Bryan T; Putzke, Aaron P

    2015-11-01

    Asymmetric cell division is critical during development, as it influences processes such as cell fate specification and cell migration. We have characterized FRK-1, a homolog of the mammalian Fer nonreceptor tyrosine kinase, and found it to be required for differentiation and maintenance of epithelial cell types, including the stem cell-like seam cells of the hypodermis. A genomic knockout of frk-1, allele ok760, results in severely uncoordinated larvae that arrest at the L1 stage and have an excess number of lateral hypodermal cells that appear to have lost asymmetry in the stem cell-like divisions of the seam cell lineage. frk-1(ok760) mutants show that there are excess lateral hypodermal cells that are abnormally shaped and smaller in size compared to wild type, a defect that could be rescued only in a manner dependent on the kinase activity of FRK-1. Additionally, we observed a significant change in the expression of heterochronic regulators in frk-1(ok760) mutants. However, frk-1(ok760) mutants do not express late, nonseam hypodermal GFP markers, suggesting the seam cells do not precociously differentiate as adult-hyp7 cells. Finally, our data also demonstrate a clear role for FRK-1 in seam cell proliferation, as eliminating FRK-1 during the L3-L4 transition results in supernumerary seam cell nuclei that are dependent on asymmetric Wnt signaling. Specifically, we observe aberrant POP-1 and WRM-1 localization that is dependent on the presence of FRK-1 and APR-1. Overall, our data suggest a requirement for FRK-1 in maintaining the identity and proliferation of seam cells primarily through an interaction with the asymmetric Wnt pathway. PMID:26358719

  19. Functional dynamics of cell surface membrane proteins

    Science.gov (United States)

    Nishida, Noritaka; Osawa, Masanori; Takeuchi, Koh; Imai, Shunsuke; Stampoulis, Pavlos; Kofuku, Yutaka; Ueda, Takumi; Shimada, Ichio

    2014-04-01

    Cell surface receptors are integral membrane proteins that receive external stimuli, and transmit signals across plasma membranes. In the conventional view of receptor activation, ligand binding to the extracellular side of the receptor induces conformational changes, which convert the structure of the receptor into an active conformation. However, recent NMR studies of cell surface membrane proteins have revealed that their structures are more dynamic than previously envisioned, and they fluctuate between multiple conformations in an equilibrium on various timescales. In addition, NMR analyses, along with biochemical and cell biological experiments indicated that such dynamical properties are critical for the proper functions of the receptors. In this review, we will describe several NMR studies that revealed direct linkage between the structural dynamics and the functions of the cell surface membrane proteins, such as G-protein coupled receptors (GPCRs), ion channels, membrane transporters, and cell adhesion molecules.

  20. Inhibition of deoxyribonucleic acid gyrase: effects on nucleic acid synthesis and cell division in Escherichia coli K-12.

    OpenAIRE

    Fairweather, N F; Orr, E; Holland, I B

    1980-01-01

    Mutants of Escherichia coli resistant to the antibiotic clorobiocin are also coumermycin resistant, and the mutation to resistance in at least one mutant was mapped near gyrB. We conclude, therefore, that clorobiocin inhibits deoxyribonucleic acid gyrase, and the drug was used to probe the role of this enzyme in vivo. Deozyribonucleic acid synthesis was preferentially inhibited but not completely blocked by the antibiotic. Transcription and cell division were also markedly affected. However, ...

  1. Spatial and Temporal Quantitative Analysis of Cell Division and Elongation Rate in Growing Wheat Leaves under Saline Conditions

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Leaf growth in grasses is determined by the cell division and elongation rates, with the duration of cell elongation being one of the processes that is the most sensitive to salinity. Our objective was to investigate the distribution profiles of cell production, cell length and the duration of cell elongation in the growing zone of the wheat leaf during the steady growth phase. Plants were grown in loamy soil with or without 120 mmol/L NaCl in a growth chamber, and harvested at day 3 after leaf 4 emerged. Results show that the elongation rate of leaf 4 was reduced by 120 mmol/L NaCl during the steady growth phase. The distribution profile of the lengths of abaxial epidermal cells of leaf 4 during the steady growth stage shows a sigmoidal pattern along the leaf axis for both treatments. Although salinity did not affect or even increased the length of the epidermal cells in some locations in the growth zone compared to the control treatment, the final length of the epidermal cells was reduced by 14% at 120 mmol/L NaCl. Thus, we concluded that the observed reduction in the leaf elongation rate derived in part from the reduced cell division rate and either the shortened cell elongation zone or shortened duration of cell elongation. This suggests that more attention should be paid to the effects of salinity on those properties of cell production and the period of cell maturation that are related to the properties of cell wall.

  2. A Systematic Analysis of Cell Cycle Regulators in Yeast Reveals That Most Factors Act Independently of Cell Size to Control Initiation of Division

    OpenAIRE

    Scott A Hoose; Jeremy A Rawlings; Kelly, Michelle M.; M Camille Leitch; Ababneh, Qotaiba O; Robles, Juan P.; David Taylor; Hoover, Evelyn M.; Bethel Hailu; McEnery, Kayla A.; S Sabina Downing; Deepika Kaushal; Yi Chen; Alex Rife; Kirtan A Brahmbhatt

    2012-01-01

    Upstream events that trigger initiation of cell division, at a point called START in yeast, determine the overall rates of cell proliferation. The identity and complete sequence of those events remain unknown. Previous studies relied mainly on cell size changes to identify systematically genes required for the timely completion of START. Here, we evaluated panels of non-essential single gene deletion strains for altered DNA content by flow cytometry. This analysis revealed that most gene dele...

  3. Protein variants in human cells: enumeration by protein indexing

    International Nuclear Information System (INIS)

    In any attempt to construct a catalog of the proteins of a given species, the genetic heterogeneity of natural plant and animal populations makes it necessary to consider variants of each protein. Thus in compiling a Human Protein Index using high resolution two-dimensional electrophoresis as a separating technique, protein variants differing from the wild type by charge or by polypeptide length will be recognized as different and must be accounted for. Results of several recent investigations have argued for average heterozygosities of approximately 1% for human cellular proteins examined by two-dimensional electrophoresis, while the results of classical biochemical genetics lead to higher results. The ability to observe genetic variation in large numbers of proteins can be valuable in several contexts. More than 2000 human genetic diseases have been identified, the vast majority of which have not as yet been associated with a defect in a particular protein. The present study of 63 human fibroblast cell lines was initiated in order to determine the level of genetic variation at many loci and to see whether known genetic diseases were associated with any obvious variant proteins that might be expressed in fibroblasts in culture. The main result is a group of ten new putative variants available in permanent cell lines

  4. A new loss-of-function allele 28y reveals a role of ARGONAUTE1 in limiting asymmetric division of stomatal lineage ground cell

    Institute of Scientific and Technical Information of China (English)

    Kezhen Yangy; Min Jiangy; Jie Le

    2014-01-01

    In Arabidopsis thaliana L., stomata are produced through a series of divisions including asymmetric and symmetric divisions. Asymmetric entry division of meristemoid mother cellproduces two daughter cells, the smal er meristemoid and the larger sister cell, a stomatal lineage ground cell(SLGC). Stomatal lineage ground cells can differentiate into epidermal pavement cells but have the potential to divide asymmetrical y, spacing divisions, to create satel ite meristemoids. Peptide ligands and TOO MANY MOUTHS (TMM) and ERECTA family receptors regulate the initiation of stomatal lineages, activity, and orientation of spacing divisions. Here, we reported that a natural mutant 28y displayed an increased stomatal density and index. Using map-based cloning, we identified mutation in ARGONAUTE1 (AGO1) as the cause of 28y phenotypes. Time-lapse tracing of stomatal lineage cells reveals that stomatal overproduction in 28y is caused by the excessive asymmetric spacing division of SLGCs.Further genetic results demonstrated that AGO1 acts down-stream of TMM and negatively regulates the SPCH transcripts, but in a brassinosteroid-independent manner. Upregulation of AGAMOUS-LIKE16 (AGL16) in 28y mutants suggests that AGO1 is required to restrict AGL16-mediated stomatal spacing divisions, an miRNA pathway in addition to ligand-receptor signaling modules.

  5. Evolution of the chloroplast division machinery

    Institute of Scientific and Technical Information of China (English)

    Hongbo GAO; Fuli GAO

    2011-01-01

    Chloroplasts are photosynthetic organelles derived from endosymbiotic cyanobacteria during evolution.Dramatic changes occurred during the process of the formation and evolution of chloroplasts,including the large-scale gene transfer from chloroplast to nucleus.However,there are still many essential characters remaining.For the chloroplast division machinery,FtsZ proteins,Ftn2,SulA and part of the division site positioning system- MinD and MinE are still conserved.New or at least partially new proteins,such as FtsZ family proteins FtsZl and ARC3,ARC6H,ARC5,PDV1,PDV2 and MCD1,were introduced for the division of chloroplasts during evolution.Some bacterial cell division proteins,such as FtsA,MreB,Ftn6,FtsW and Ftsl,probably lost their function or were gradually lost.Thus,the chloroplast division machinery is a dynamically evolving structure with both conservation and innovation.

  6. Sea urchin akt activity is Runx-dependent and required for post-cleavage stage cell division

    KAUST Repository

    Robertson, Anthony J.

    2013-03-25

    In animal development following the initial cleavage stage of embryogenesis, the cell cycle becomes dependent on intercellular signaling and controlled by the genomically encoded ontogenetic program. Runx transcription factors are critical regulators of metazoan developmental signaling, and we have shown that the sea urchin Runx gene runt-1, which is globally expressed during early embryogenesis, functions in support of blastula stage cell proliferation and expression of the mitogenic genes pkc1, cyclinD, and several wnts. To obtain a more comprehensive list of early runt-1 regulatory targets, we screened a Strongylocentrotus purpuratus microarray to identify genes mis-expressed in mid-blastula stage runt-1 morphants. This analysis showed that loss of Runx function perturbs the expression of multiple genes involved in cell division, including the pro-growth and survival kinase Akt (PKB), which is significantly underexpressed in runt-1 morphants. Further genomic analysis revealed that Akt is encoded by two genes in the S. purpuratus genome, akt-1 and akt-2, both of which contain numerous canonical Runx target sequences. The transcripts of both genes accumulate several fold during blastula stage, contingent on runt-1 expression. Inhibiting Akt expression or activity causes blastula stage cell cycle arrest, whereas overexpression of akt-1 mRNA rescues cell proliferation in runt-1 morphants. These results indicate that post-cleavage stage cell division requires Runx-dependent expression of akt.

  7. Parkin suppresses Drp1-independent mitochondrial division.

    Science.gov (United States)

    Roy, Madhuparna; Itoh, Kie; Iijima, Miho; Sesaki, Hiromi

    2016-07-01

    The cycle of mitochondrial division and fusion disconnect and reconnect individual mitochondria in cells to remodel this energy-producing organelle. Although dynamin-related protein 1 (Drp1) plays a major role in mitochondrial division in cells, a reduced level of mitochondrial division still persists even in the absence of Drp1. It is unknown how much Drp1-mediated mitochondrial division accounts for the connectivity of mitochondria. The role of a Parkinson's disease-associated protein-parkin, which biochemically and genetically interacts with Drp1-in mitochondrial connectivity also remains poorly understood. Here, we quantified the number and connectivity of mitochondria using mitochondria-targeted photoactivatable GFP in cells. We show that the loss of Drp1 increases the connectivity of mitochondria by 15-fold in mouse embryonic fibroblasts (MEFs). While a single loss of parkin does not affect the connectivity of mitochondria, the connectivity of mitochondria significantly decreased compared with a single loss of Drp1 when parkin was lost in the absence of Drp1. Furthermore, the loss of parkin decreased the frequency of depolarization of the mitochondrial inner membrane that is caused by increased mitochondrial connectivity in Drp1-knockout MEFs. Therefore, our data suggest that parkin negatively regulates Drp1-indendent mitochondrial division. PMID:27181353

  8. ftsZ gene and plastid division

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Plastid is one of the most important cellular organelles, the normal division process of plastid is essential for the differentiation and development of plant cells. For a long time, morphological observations and genetic analyses to special mutants are the major research fields of plastid division, but the molecular mechanisms underlying plastid division are largely unknown. Because of the endosymbiotic origin, plastid division might have mechanisms in common with those involved in bacterial cell division. It has been proved that several prokaryotic cell division genes also participate in the plastid division. Recently, the mechanisms of prokaryotic cell division have been well documented, which provides a valuable paradigm for understanding the plastid division mechanisms. In plants, the functional analyses of ftsZ, a key gene involved both in bacteria and plastid division, have established the solid foundation for people to understand the plastid division in molecular level. In this paper we will make a review for the research history and progress of plastid division.

  9. Disorganization of cell division of methicillin-resistant Staphylococcus aureus by methanolic extract from Phyllanthus columnaris stem bark

    Energy Technology Data Exchange (ETDEWEB)

    Adnalizawati, A. Siti Noor; Nazlina, I. [School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor (Malaysia); Yaacob, W. A. [School of Chemical Sciences and Food Technology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor (Malaysia)

    2013-11-27

    The in vitro activity of methanolic extract from Phyllanthus columnaris stem bark was studied against Methicillin-resistant Staphylococcus aureus (MRSA) ATCC 43300 and MRSA BM1 (clinical strain) using time-kill curves in conjunction with scanning and transmission electron microscopy. The extract showed more markedly bactericidal activity in MRSA BM1 clinical strain within less than 4 h by 6.25-12.5 mg/mL and within 6 h by 1.56 mg/mL. Scanning electron microscopy of MRSA BM1 revealed distortion of cell whilst transmission electron microscopy revealed disruption in cell wall division.

  10. Disorganization of cell division of methicillin-resistant Staphylococcus aureus by methanolic extract from Phyllanthus columnaris stem bark

    Science.gov (United States)

    Adnalizawati, A. Siti Noor; Nazlina, I.; Yaacob, W. A.

    2013-11-01

    The in vitro activity of methanolic extract from Phyllanthus columnaris stem bark was studied against Methicillin-resistant Staphylococcus aureus (MRSA) ATCC 43300 and MRSA BM1 (clinical strain) using time-kill curves in conjunction with scanning and transmission electron microscopy. The extract showed more markedly bactericidal activity in MRSA BM1 clinical strain within less than 4 h by 6.25-12.5 mg/mL and within 6 h by 1.56 mg/mL. Scanning electron microscopy of MRSA BM1 revealed distortion of cell whilst transmission electron microscopy revealed disruption in cell wall division.

  11. Cell death in the central division of the medial preoptic nucleus of male and female lamb fetuses

    OpenAIRE

    Reddy, Radhika C.; Scheldrup, Melissa; Meaker, Mary; Stormshak, Fred; Estill, Charles T.; Roselli, Charles E.

    2014-01-01

    The medial preoptic area of the adult sheep contains an ovine sexually dimorphic nucleus (oSDN) that is larger and has more neurons in males than in females. In the lamb fetus, the nascent oSDN occupies the central division of the medial preoptic nucleus (MPNc) and consists of a cluster of cells that is organized by the action of testosterone during gestational days 60 to 90 of a 147 day term pregnancy. The current study sought to determine whether programmed cell death cont...

  12. Disorganization of cell division of methicillin-resistant Staphylococcus aureus by methanolic extract from Phyllanthus columnaris stem bark

    International Nuclear Information System (INIS)

    The in vitro activity of methanolic extract from Phyllanthus columnaris stem bark was studied against Methicillin-resistant Staphylococcus aureus (MRSA) ATCC 43300 and MRSA BM1 (clinical strain) using time-kill curves in conjunction with scanning and transmission electron microscopy. The extract showed more markedly bactericidal activity in MRSA BM1 clinical strain within less than 4 h by 6.25-12.5 mg/mL and within 6 h by 1.56 mg/mL. Scanning electron microscopy of MRSA BM1 revealed distortion of cell whilst transmission electron microscopy revealed disruption in cell wall division

  13. Targeting Cell Cycle Proteins in Breast Cancer Cells with siRNA by Using Lipid-Substituted Polyethylenimines.

    Science.gov (United States)

    Parmar, Manoj B; Aliabadi, Hamidreza Montazeri; Mahdipoor, Parvin; Kucharski, Cezary; Maranchuk, Robert; Hugh, Judith C; Uludağ, Hasan

    2015-01-01

    The cell cycle proteins are key regulators of cell cycle progression whose deregulation is one of the causes of breast cancer. RNA interference (RNAi) is an endogenous mechanism to regulate gene expression and it could serve as the basis of regulating aberrant proteins including cell cycle proteins. Since the delivery of small interfering RNA (siRNA) is a main barrier for implementation of RNAi therapy, we explored the potential of a non-viral delivery system, 2.0 kDa polyethylenimines substituted with linoleic acid and caprylic acid, for this purpose. Using a library of siRNAs against cell cycle proteins, we identified cell division cycle protein 20 (CDC20), a recombinase RAD51, and serine-threonine protein kinase CHEK1 as effective targets for breast cancer therapy, and demonstrated their therapeutic potential in breast cancer MDA-MB-435, MDA-MB-231, and MCF7 cells with respect to another well-studied cell cycle protein, kinesin spindle protein. We also explored the efficacy of dicer-substrate siRNA (DsiRNA) against CDC20, RAD51, and CHEK1, where a particular DsiRNA against CDC20 showed an exceptionally high inhibition of cell growth in vitro. There was no apparent effect of silencing selected cell cycle proteins on the potency of the chemotherapy drug doxorubicin. The efficacy of DsiRNA against CDC20 was subsequently assessed in a xenograft model, which indicated a reduced tumor growth as a result of CDC20 DsiRNA therapy. The presented study highlighted specific cell cycle protein targets critical for breast cancer therapy, and provided a polymeric delivery system for their effective down-regulation. PMID:25763370

  14. RNA helicase Belle (DDX3) is essential for male germline stem cell maintenance and division in Drosophila.

    Science.gov (United States)

    Kotov, Alexei A; Olenkina, Oxana M; Kibanov, Mikhail V; Olenina, Ludmila V

    2016-06-01

    The present study showed that RNA helicase Belle (DDX3) was required intrinsically for mitotic progression and survival of germline stem cells (GSCs) and spermatogonial cells in the Drosophila melanogaster testes. We found that deficiency of Belle in the male germline resulted in a strong germ cell loss phenotype. Early germ cells are lost through cell death, whereas somatic hub and cyst cell populations are maintained. The observed phenotype is related to that of the human Sertoli Cell-Only Syndrome caused by the loss of DBY (DDX3) expression in the human testes and results in a complete lack of germ cells with preservation of somatic Sertoli cells. We found the hallmarks of mitotic G2 delay in early germ cells of the larval testes of bel mutants. Both mitotic cyclins, A and B, are markedly reduced in the gonads of bel mutants. Transcription levels of cycB and cycA decrease significantly in the testes of hypomorph bel mutants. Overexpression of Cyclin B in the germline partially rescues germ cell survival, mitotic progression and fertility in the bel-RNAi knockdown testes. Taken together, these results suggest that a role of Belle in GSC maintenance and regulation of early germ cell divisions is associated with the expression control of mitotic cyclins. PMID:26876306

  15. Systemic Control of Cell Division and Endoreduplication by NAA and BAP by Modulating CDKs in Root Tip Cells of Allium cepa

    Directory of Open Access Journals (Sweden)

    Jigna G. Tank

    2014-01-01

    Full Text Available Molecular mechanism regulated by auxin and cytokinin during endoreduplication, cell division, and elongation process is studied by using Allium cepa roots as a model system. The activity of CDK genes modulated by auxin and cytokinin during cell division, elongation, and endoreduplication process is explained in this research work. To study the significance of auxin and cytokinin in the management of cell division and endoreduplication process in plant meristematic cells at molecular level endoreduplication was developed in root tips of Allium cepa by giving colchicine treatment. There were inhibition of vegetative growth, formation of c-tumor at root tip, and development of endoreduplicated cells after colchicine treatment. This c-tumor was further treated with NAA and BAP to reinitiate vegetative growth in roots. BAP gave positive response in reinitiation of vegetative growth of roots from center of c-tumor. However, NAA gave negative response in reinitiation of vegetative growth of roots from c-tumor. Further, CDKs gene expression analysis from normal, endoreduplicated, and phytohormone (NAA or BAP treated root tip was done and remarkable changes in transcription level of CDK genes in normal, endoreduplicated, and phytohormones treated cells were observed.

  16. Cell-to-cell propagation of infectious cytosolic protein aggregates

    Science.gov (United States)

    Hofmann, Julia P.; Denner, Philip; Nussbaum-Krammer, Carmen; Kuhn, Peer-Hendrik; Suhre, Michael H.; Scheibel, Thomas; Lichtenthaler, Stefan F.; Schätzl, Hermann M.; Bano, Daniele; Vorberg, Ina M.

    2013-01-01

    Prions are self-templating protein conformers that replicate by recruitment and conversion of homotypic proteins into growing protein aggregates. Originally identified as causative agents of transmissible spongiform encephalopathies, increasing evidence now suggests that prion-like phenomena are more common in nature than previously anticipated. In contrast to fungal prions that replicate in the cytoplasm, propagation of mammalian prions derived from the precursor protein PrP is confined to the cell membrane or endocytic vesicles. Here we demonstrate that cytosolic protein aggregates can also behave as infectious entities in mammalian cells. When expressed in the mammalian cytosol, protein aggregates derived from the prion domain NM of yeast translation termination factor Sup35 persistently propagate and invade neighboring cells, thereby inducing a self-perpetuating aggregation state of NM. Cell contact is required for efficient infection. Aggregates can also be induced in primary astrocytes, neurons, and organotypic cultures, demonstrating that this phenomenon is not specific to immortalized cells. Our data have important implications for understanding prion-like phenomena of protein aggregates associated with human diseases and for the growing number of amyloidogenic proteins discovered in mammals. PMID:23509289

  17. Origins of Protein Functions in Cells

    Science.gov (United States)

    Seelig, Burchard; Pohorille, Andrzej

    2011-01-01

    In modern organisms proteins perform a majority of cellular functions, such as chemical catalysis, energy transduction and transport of material across cell walls. Although great strides have been made towards understanding protein evolution, a meaningful extrapolation from contemporary proteins to their earliest ancestors is virtually impossible. In an alternative approach, the origin of water-soluble proteins was probed through the synthesis and in vitro evolution of very large libraries of random amino acid sequences. In combination with computer modeling and simulations, these experiments allow us to address a number of fundamental questions about the origins of proteins. Can functionality emerge from random sequences of proteins? How did the initial repertoire of functional proteins diversify to facilitate new functions? Did this diversification proceed primarily through drawing novel functionalities from random sequences or through evolution of already existing proto-enzymes? Did protein evolution start from a pool of proteins defined by a frozen accident and other collections of proteins could start a different evolutionary pathway? Although we do not have definitive answers to these questions yet, important clues have been uncovered. In one example (Keefe and Szostak, 2001), novel ATP binding proteins were identified that appear to be unrelated in both sequence and structure to any known ATP binding proteins. One of these proteins was subsequently redesigned computationally to bind GTP through introducing several mutations that introduce targeted structural changes to the protein, improve its binding to guanine and prevent water from accessing the active center. This study facilitates further investigations of individual evolutionary steps that lead to a change of function in primordial proteins. In a second study (Seelig and Szostak, 2007), novel enzymes were generated that can join two pieces of RNA in a reaction for which no natural enzymes are known

  18. Single-cell time-lapse analysis of depletion of the universally conserved essential protein YgjD

    Directory of Open Access Journals (Sweden)

    Ackermann Martin

    2011-05-01

    Full Text Available Abstract Background The essential Escherichia coli gene ygjD belongs to a universally conserved group of genes whose function has been the focus of a number of recent studies. Here, we put ygjD under control of an inducible promoter, and used time-lapse microscopy and single cell analysis to investigate the phenotypic consequences of the depletion of YgjD protein from growing cells. Results We show that loss of YgjD leads to a marked decrease in cell size and termination of cell division. The transition towards smaller size occurs in a controlled manner: cell elongation and cell division remain coupled, but cell size at division decreases. We also find evidence that depletion of YgjD leads to the synthesis of the intracellular signaling molecule (pppGpp, inducing a cellular reaction resembling the stringent response. Concomitant deletion of the relA and spoT genes - leading to a strain that is uncapable of synthesizing (pppGpp - abrogates the decrease in cell size, but does not prevent termination of cell division upon YgjD depletion. Conclusions Depletion of YgjD protein from growing cells leads to a decrease in cell size that is contingent on (pppGpp, and to a termination of cell division. The combination of single-cell timelapse microscopy and statistical analysis can give detailed insights into the phenotypic consequences of the loss of essential genes, and can thus serve as a new tool to study the function of essential genes.

  19. An immunosupressive protein supporting embryonic implantation and having an important role in cell proliferation: The Early Pregnancy Factor

    OpenAIRE

    Özbek, Elvan; Cengiz, Nureddin; Yanar, Sevinç

    2013-01-01

    Early pregnancy factor (EPF), identified as an immunosuppresive protein, can be detected in maternal serum within 6-24 hours of fertilization and is present for at least the first two trimester of pregnancy. Besides pregnancy, EPF is secreted by tumor cells, normal growing cells and active platelets during cell growth and division. It is also required for tumor proliferation and tissue renewal. Seventy per cent of the amino acid sequence of EPF derived from human platelets was determined and ...

  20. Compartmentalization and Cell Division through Molecular Discreteness and Crowding in a Catalytic Reaction Network

    Directory of Open Access Journals (Sweden)

    Atsushi Kamimura

    2014-10-01

    Full Text Available Explanation of the emergence of primitive cellular structures from a set of chemical reactions is necessary to unveil the origin of life and to experimentally synthesize protocells. By simulating a cellular automaton model with a two-species hypercycle, we demonstrate the reproduction of a localized cluster; that is, a protocell with a growth-division process emerges when the replication and degradation speeds of one species are respectively slower than those of the other species, because of overcrowding of molecules as a natural outcome of the replication. The protocell exhibits synchrony between its division process and replication of the minority molecule. We discuss the effects of the crowding molecule on the formation of primitive structures. The generality of this result is demonstrated through the extension of our model to a hypercycle with three molecular species, where a localized layered structure of molecules continues to divide, triggered by the replication of a minority molecule at the center.

  1. Versatile protein tagging in cells with split fluorescent protein.

    Science.gov (United States)

    Kamiyama, Daichi; Sekine, Sayaka; Barsi-Rhyne, Benjamin; Hu, Jeffrey; Chen, Baohui; Gilbert, Luke A; Ishikawa, Hiroaki; Leonetti, Manuel D; Marshall, Wallace F; Weissman, Jonathan S; Huang, Bo

    2016-01-01

    In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respectively. The small size of FP11-tags enables a cost-effective and scalable way to insert them into endogenous genomic loci via CRISPR-mediated homology-directed repair. Tandem arrangement FP11-tags allows proportional enhancement of fluorescence signal in tracking intraflagellar transport particles, or reduction of photobleaching for live microtubule imaging. Finally, we show the utility of tandem GFP11-tag in scaffolding protein oligomerization. These experiments illustrate the versatility of FP11-tag as a labelling tool as well as a multimerization-control tool for both imaging and non-imaging applications. PMID:26988139

  2. Compartmentalization and Cell Division through Molecular Discreteness and Crowding in a Catalytic Reaction Network

    OpenAIRE

    Atsushi Kamimura; Kunihiko Kaneko

    2014-01-01

    Explanation of the emergence of primitive cellular structures from a set of chemical reactions is necessary to unveil the origin of life and to experimentally synthesize protocells. By simulating a cellular automaton model with a two-species hypercycle, we demonstrate the reproduction of a localized cluster; that is, a protocell with a growth-division process emerges when the replication and degradation speeds of one species are respectively slower than those of the other species, because of ...

  3. Effect of chronic fractionated low-dose gamma irradiation on division potential of human embryonic cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, Masami; Suzuki, Masao; Suzuki, Keiji; Watanabe, Kimiko (Yokohama City Univ. (Japan). Faculty of Medicine); Nakano, Kazushiro

    1991-12-01

    We investigated the in vitro phenotypic transformation of human embryo (HE) cells that were repeatedly irradiated (7.5 cGy once a week) throughout their life-span. Irradiation was repeated until the cells had accumulated 195 cGy (equivalent to the 26th passage). Samples of cells were assayed for survival by colony formation, as well as for mutation at the hypoxanthine guanine phosphoribosyl transferase (HGPRT) locus and for transformation by focus formation. The life-span (mean number of population doublings) of multiply irradiated cells with a total dose of 97.5 cGy was slightly but significantly prolonged over that of controls. After HE cells had accumulated 195 cGy, the maximum number of divisions increased to 130-160% of the number in non-irradiated control cells. Transformed foci were not observed until cells had accumulated 97.5 cGy, and then increased with the increasing accumulation of radiation. However, no cells showed immortality or expressed a malignant phenotype in vitro. (author).

  4. SUN Regulates Vegetative and Reproductive Organ Shape by Changing Cell Division Patterns1[C][W][OA

    Science.gov (United States)

    Wu, Shan; Xiao, Han; Cabrera, Antonio; Meulia, Tea; van der Knaap, Esther

    2011-01-01

    One of the major genes controlling the elongated fruit shape of tomato (Solanum lycopersicum) is SUN. In this study, we explored the roles of SUN in vegetative and reproductive development using near isogenic lines (NILs) that differ at the sun locus, and SUN overexpressors in both the wild species LA1589 (Solanum pimpinellifolium) and the cultivar Sun1642 background. Our results demonstrate that SUN controls tomato shape through redistribution of mass that is mediated by increased cell division in the longitudinal and decreased cell division in the transverse direction of the fruit. The expression of SUN is positively correlated with slender phenotypes in cotyledon, leaflet, and floral organs, an elongated ovary, and negatively correlated with seed weight. Overexpression of SUN leads to more extreme phenotypes than those shown in the NILs and include thinner leaf rachises and stems, twisted leaf rachises, increased serrations of the leaflets, and dramatically increased elongation at the proximal end of the ovary and fruit. In situ hybridizations of the NILs showed that SUN is expressed throughout the ovary and young fruit, particularly in the vascular tissues and placenta surface, and in the ovules and developing seed. The phenotypic effects resulting from high expression of SUN suggest that the gene is involved in several plant developmental processes. PMID:21921117

  5. Rice OsRAD21-2 is Expressed in Actively Dividing Tissues and its Ectopic Expression in Yeast Results in Aberrant Cell Division and Growth

    Institute of Scientific and Technical Information of China (English)

    Chunyan Gong; Tang Li; Qi Li; Longfeng Yan; Tai Wang

    2011-01-01

    Rad21 and its meiotic counterpart Rec8,the key components of the cohesin complex,are essential for sister chromatid cohesion and chromosome segregation in mitosis and meiosis,respectively.In contrast to yeast and vertebrates,which have only two RAD21/REC8 genes,the rice genome encodes four Rad21/Rec8 proteins.Here,we report on the cloning and characterization of OsRAD21-2 from rice (Oryza sativa L.).Phylogenetic analysis of the full-length amino acids showed that OsRad21-2 was grouped into the plant-specific Rad21 subfamily.Semi-quantitative reverse transcription-polymerase chain reaction revealed OsRAD21-2 preferentially expressed in premeiotic flowers.Further RNA in situ hybridization analysis and promoter::β-glucuronidase staining indicated that OsRAD21-2 was mainly expressed in actively dividing tissues including premeiotic stamen,stem intercalary meristem,leaf meristem,and root pericycle.Ectopic expression of OsRAD21-2 in fission yeast resulted in cell growth delay and morphological abnormality.Flow cytometric analysis revealed that the OsRAD21-2-expressed cells were arrested in G2 phase.Our results suggest that OsRad21-2 functions in regulation of cell division and growth.

  6. The ABC's of Cell Division: Regulation of Peptidoglycan Amidase Activity during Cytokinesis in Escherichia coli

    OpenAIRE

    Yang, Desiree Choy

    2012-01-01

    The bacterial cell wall, composed of peptidoglycan (PG), is an essential component of the cell envelope. This macromolecular structure fortifies the cell membrane, determines cell shape, and helps prevent osmotic lysis. The synthesis and remodeling/recycling of this polymer is mediated by PG synthases and hydrolases, respectively. Proper control of the PG hydrolases is particularly important since misregulation of these enzymes can lead to lethal breaches in the cell wall. Surprisingly, howev...

  7. CDP1, a novel component of chloroplast division site positioning system in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Min Zhang; Yong Hu; Jingjing Jia; Dapeng Li; Runjie Zhang; Hongbo Gao; Yikun He

    2009-01-01

    Chloroplasts are plant-specific organelles that evolved from endosymbiotic cyanobacteria. They divide through binary fission. Selection of the chloroplast division site is pivotal for the symmetric chloroplast division. In E. coli, positioning of the division site at the midpoint of the cell is regulated by dynamic oscillation of the Min system, which includes MinC, MinD and MinE. Homologs of Mind and MinE in plants are involved in chloroplast division. The homolog of MinC still has not been identified in higher plants. However, an FtsZ-like protein, ARC3, was found to be involved in chloroplast division site positioning. Here, we report that chloroplast division site positioning 1 (AtCDP1) is a novel chloroplast division protein involved in chloroplast division site placement in Arabidopsis. AtCDP1 was dis-covered by screening an Arabidopsis cDNA expression library in bacteria for colonies with a cell division phenotype. AtCDP1 is exclusively expressed in young green tissues in Arabidopsis. Elongated chloroplasts with multiple division sites were observed in the loss-of-function cdpl mutant. Overexpression of AtCDPI caused a chloroplast division phe-notype too. Protein interaction assays suggested that AtCDP1 may mediate the chloroplast division site positioning through the interaction with ARC3. Overall, our results indicate that AtCDP1 is a novel component of the chloroplast division site positioning system, and the working mechanism of this system is different from that of the traditional MinCDE system in prokaryotic cells.

  8. Lyme disease and relapsing fever Borrelia elongate through zones of peptidoglycan synthesis that mark division sites of daughter cells.

    Science.gov (United States)

    Jutras, Brandon Lyon; Scott, Molly; Parry, Bradley; Biboy, Jacob; Gray, Joe; Vollmer, Waldemar; Jacobs-Wagner, Christine

    2016-08-16

    Agents that cause Lyme disease, relapsing fever, leptospirosis, and syphilis belong to the phylum Spirochaetae-a unique lineage of bacteria most known for their long, spiral morphology. Despite the relevance to human health, little is known about the most fundamental aspects of spirochete growth. Here, using quantitative microscopy to track peptidoglycan cell-wall synthesis, we found that the Lyme disease spirochete Borrelia burgdorferi displays a complex pattern of growth. B. burgdorferi elongates from discrete zones that are both spatially and temporally regulated. In addition, some peptidoglycan incorporation occurs along the cell body, with the notable exception of a large region at the poles. Newborn cells inherit a highly active zone of peptidoglycan synthesis at midcell that contributes to elongation for most of the cell cycle. Concomitant with the initiation of nucleoid separation and cell constriction, second and third zones of elongation are established at the 1/4 and 3/4 cellular positions, marking future sites of division for the subsequent generation. Positioning of elongation zones along the cell is robust to cell length variations and is relatively precise over long distances (>30 µm), suggesting that cells ‟sense" relative, as opposed to absolute, cell length to establish zones of peptidoglycan synthesis. The transition from one to three zones of peptidoglycan growth during the cell cycle is also observed in relapsing fever Borrelia. However, this mode of growth does not extend to representative species from other spirochetal genera, suggesting that this distinctive growth mode represents an evolutionary divide in the spirochete phylum. PMID:27506799

  9. Enteral peptide formulas inhibit radiation induced enteritis and apoptosis in intestinal epithelial cells and suppress the expression and function of Alzheimer's and cell division control gene products

    International Nuclear Information System (INIS)

    Studies have shown that patients receiving enteral peptide formulas prior to irradiation have a significantly reduced incidence of enteritis and express a profound increase in intestinal cellularity. Two conceptual approaches were taken to describe this response. First was the evaluation in changes in programmed intestinal cell death and secondly the evaluation of a gene product controlling cell division cycling. This study provided a relationship between the ratio of cell death to cell formulations. The results indicate that in the canine and murine models, irradiation induces expression of the Alzheimer's gene in intestinal crypt cells, while the incidence of apoptosis in apical cells is significantly increased. The use of peptide enteral formulations suppresses the expression of the Alzheimer's gene in crypt cells, while apoptosis is eliminated in the apical cells of the intestine. Concomitantly, enteral peptide formulations suppress the function of the CK-II gene product in the basal and baso-lateral cells of the intestine. These data indicate that although the mitotic index is significantly reduced in enterocytes, this phenomenon alone is not sufficient to account for the peptide-induced radio-resistance of the intestine. The data also indicate a significant reduction of normal apoptosis in the upper lateral and apical cells of the intestinal villi. Thus, the ratio of cell death to cell replacement is significantly decreased resulting in an increase in villus height and hypertrophy of the apical villus cells. Thus, peptide solutions should be considered as an adjunct treatment both in radio- and chemotherapy

  10. Asymmetric division of cyst stem cells in Drosophila testis is ensured by anaphase spindle repositioning

    OpenAIRE

    Cheng, Jun; Tiyaboonchai, Amita; Yamashita, Yukiko M.; Hunt, Alan J.

    2011-01-01

    Many stem cells divide asymmetrically to balance self-renewal and differentiation. In Drosophila testes, two stem cell populations, germline stem cells (GSCs) and somatic cyst stem cells (CySCs), cohere and regulate one another. Here, we report that CySCs divide asymmetrically through repositioning the mitotic spindle around anaphase. CySC spindle repositioning requires functional centrosomes, Dynein and the actin-membrane linker Moesin. Anaphase spindle repositioning is required to achieve h...

  11. AUREOCHROME1a-mediated induction of the diatom-specific cyclin dsCYC2 controls the onset of cell division in diatoms (Phaeodactylum tricornutum)

    OpenAIRE

    Huysman, Marie; Fortunato, Antonio E; Matthijs, Michiel; Costa, Benjamin Schellenberger; Vanderhaeghen, Rudy; Van Den Daele, Hilde; Sachse, Matthias; Inzé, Dirk; Bowler, Chris; Kroth, Peter G.; Wilhelm, Christian; Falciatore, Angela; Vyverman, Wim; De Veylder, Lieven

    2013-01-01

    Cell division in photosynthetic organisms is tightly regulated by light. Although the light dependency of the onset of the cell cycle has been well characterized in various phototrophs, little is known about the cellular signaling cascades connecting light perception to cell cycle activation and progression. Here, we demonstrate that diatom-specific cyclin 2 (dsCYC2) in Phaeodactylum tricornutum displays a transcriptional peak within 15 min after light exposure, long before the onset of cell ...

  12. Dynamic FtsA and FtsZ localization and outer membrane alterations during polar growth and cell division in Agrobacterium tumefaciens

    OpenAIRE

    Zupan, John R.; Cameron, Todd A.; Anderson-Furgeson, James; Zambryski, Patricia C.

    2013-01-01

    Growth and cell division in rod-shaped bacteria have been primarily studied in species that grow predominantly by peptidoglycan (PG) synthesis along the length of the cell. Rhizobiales species, however, predominantly grow by PG synthesis at a single pole. Here we characterize the dynamic localization of several Agrobacterium tumefaciens components during the cell cycle. First, the lipophilic dye FM 4-64 predominantly stains the outer membranes of old poles versus growing poles. In cells about...

  13. Role of Heat Shock Proteins in Stem Cell Behavior

    OpenAIRE

    Fan, Guo-Chang

    2012-01-01

    Stress response is well appreciated to induce the expression of heat shock proteins (Hsps) in the cell. Numerous studies have demonstrated that Hsps function as molecular chaperones in the stabilization of intracellular proteins, repairing damaged proteins, and assisting in protein translocation. Various kinds of stem cells (embryonic stem cells, adult stem cells, or induced pluripotent stem cells) have to maintain their stemness and, under certain circumstances, undergo stress. Therefore, Hs...

  14. Algorithm development and simulation outcomes for hypoxic head and neck cancer radiotherapy using a Monte Carlo cell division model

    International Nuclear Information System (INIS)

    Full text: A temporal Monte Carlo tumour model, 'Hyp-RT'. sim ulating hypoxic head and neck cancer has been updated and extended to model radiothcrapy. The aim is to providc a convenient radiobio logical tool for clinicians to evaluate radiotherapy treatment schedules based on many individual tumour properties including oxygenation. FORTRAN95 and JA YA havc been utilised to develop the efficient algorithm, which can propagate 108 cells. Epithelial cell kill is affected by dose, oxygenation and proliferativc status. Accelerated repopulation (AR) has been modelled by increasing the symmetrical stem cell division probability, and reoxygenation (ROx) has been modelled using random incremental boosts of oxygen to the cell po ulation throughout therapy. Results The stem cell percentage and the degree of hypoxia dominate tumour growth rate. For conventional radiotherapy. 15-25% more dose was required for a hypox ic versus oxic tumours, depending on the time of AR onset (0-3 weeks after thc start of treatment). ROx of hypoxic tumours resulted in tumoUJ: sensitisation and therefore a dose reduction, of up to 35%, varying with the time of onset. Fig. I shows results for all combinations of AR and ROx onset times for the moderate hypoxia case. Conclusions In hypoxic tumours, accelerated repopulation and reoxy genation affect ccll kill in the same manner as when the effects are modelled individually. however the degree of the effect is altered and therefore the combined result is difficult to predict. providing evidence for the usefulness of computer models. Simulations have quantitatively

  15. Spindle formation and microtubule organization during first division in reconstructed rat embryos produced by somatic cell nuclear transfer.

    Science.gov (United States)

    Tomioka, Ikuo; Mizutani, Eiji; Yoshida, Tomoyuki; Sugawara, Atsushi; Inai, Kentaro; Sasada, Hiroshi; Sato, Eimei

    2007-08-01

    The present study was conducted to demonstrate the spindle formation and behavior of chromosomes and microtubules during first division in reconstructed rat embryos produced by somatic cell nuclear transfer (SCNT) with cumulus cell nuclei. To demonstrate the effect of oocyte aging after ovulation on the cleavage of SCNT embryos, micromanipulation was carried out 11, 15 and 18 h after injection of hCG. SCNT oocytes were activated by incubation in culture medium supplemented with 5 microM ionomycin for 5 min followed by treatment with 2 mM 6-dimethylaminopurine (6-DMAP) in mR1ECM for 2-3 h. For immunocytochemical observation, the SCNT embryos were incubated with monoclonal anti-alpha-tubulin antibody and then fluorescein isothiocyanate (FITC)-conjugated goat anti-mouse IgG. Cleavage rates were significantly higher for oocytes collected after 15 and 18 h rather than for those collected 11 h after injection of hCG (56 and 53%, respectively vs. 28%; P<0.05). Premature chromosome condensation occurred before activation of the SCNT oocytes, but adequate spindle formation was only rarely observed. The distribution of microtubules in SCNT embryos after activation was different from those of fertilized and parthenogenic oocytes, i.e., a dense microtubule organization shaped like a ring was observed. Eighteen to 20 h post-activation, most SCNT embryos were in the 2-cell stage, but no nucleoli were clearly visible, which was quite different from the fertilized oocytes. In addition, first division with and without small cellular bodies containing DNA was observed in the rat SCNT embryos in some cases. The present study suggests that reorganization of transferred nuclei in rat SCNT embryos may be inadequate in terms of formation of the mitotic assembly and nucleolar reorganization. PMID:17446658

  16. Asymmetric protein localization in planar cell polarity

    Science.gov (United States)

    Peng, Ying; Axelrod, Jeffrey D.

    2016-01-01

    The polarization of epithelial cells along an axis orthogonal to their apical-basal axis is increasingly recognized for roles in a variety of developmental events and physiological functions. While now studied in many model organisms, mechanistic understanding is rooted in intensive investigations of Planar Cell Polarity (PCP) in Drosophila. Consensus has emerged that two molecular modules, referred to here as the global and core modules, operate upstream of effector proteins to produce morphological PCP. Proteins of the core module develop subcellular asymmetry, accumulating in two groups on opposite sides of cells, consistent with proposed functions in producing cell polarity and in communicating that polarity between neighboring cells. Less clear are the molecular and cell biological mechanisms underlying core module function in the generation and communication of subcellular asymmetry, and the relationship between the global and core modules. In this review, we discuss these two unresolved questions, highlighting important studies and potentially enlightening avenues for further investigation. It is likely that results from Drosophila will continue to inform our views of the growing list of examples of PCP in vertebrate systems. PMID:23140624

  17. Biological Evaluation of Single Cell Protein

    International Nuclear Information System (INIS)

    In this study, the nutritional value of single cell protein (SCP) was evaluated as a non conventional protein source produced by fermenting fungal local strains of Trichoderma longibrachiatum, Aspergillus niger, Aspergillus terreus and Penicillium funiculosum with alkali treated sugar cane bagasse. Amino acid analysis revealed that the produced SCP contains essential and non essential amino acids. Male mice were fed on normal (basal) diet which contains 18% conventional protein and served as control group. In the second (T1) and the third (T2) group, the animals were fed on a diet in which 15% and 30% of conventional protein source were replaced by SCP, respectively. At intervals of 15, 30, 45 and 60 days, mice were sacrificed and the blood samples were collected for the biochemical evaluation. The daily averages of body weight were significantly higher with group T2 than group T1. Where as, the kidney weights in groups (T1) and (T2) were significantly increased as compared with control. A non significant difference between the tested groups in the enzyme activities of AST, ALT and GSH content of liver tissue were recorded. While, cholesterol and triglycerides contents showed a significant decrease in both (T1) and (T2) groups as compared with control. The recorded values of the serum hormone (T4), ALP activities, albumin and A/G ratio did not changed by the previous treatments. Serum levels of total protein, urea, creatinine and uric acid were higher for groups (T1) and (T2) than the control group. In conclusion, partial substitution of soy bean protein in mice diet with single cell protein (15%) improved the mice growth without any adverse effects on some of the physiological functions tested

  18. The Garlic Allelochemical Diallyl Disulfide Affects Tomato Root Growth by Influencing Cell Division, Phytohormone Balance and Expansin Gene Expression

    Science.gov (United States)

    Cheng, Fang; Cheng, Zhihui; Meng, Huanwen; Tang, Xiangwei

    2016-01-01

    Diallyl disulfide (DADS) is a volatile organosulfur compound derived from garlic (Allium sativum L.), and it is known as an allelochemical responsible for the strong allelopathic potential of garlic. The anticancer properties of DADS have been studied in experimental animals and various types of cancer cells, but to date, little is known about its mode of action as an allelochemical at the cytological level. The current research presents further studies on the effects of DADS on tomato (Solanum lycopersicum L.) seed germination, root growth, mitotic index, and cell size in root meristem, as well as the phytohormone levels and expression profile of auxin biosynthesis genes (FZYs), auxin transport genes (SlPINs), and expansin genes (EXPs) in tomato root. The results showed a biphasic, dose-dependent effect on tomato seed germination and root growth under different DADS concentrations. Lower concentrations (0.01–0.62 mM) of DADS significantly promoted root growth, whereas higher levels (6.20–20.67 mM) showed inhibitory effects. Cytological observations showed that the cell length of root meristem was increased and that the mitotic activity of meristematic cells in seedling root tips was enhanced at lower concentrations of DADS. In contrast, DADS at higher concentrations inhibited root growth by affecting both the length and division activity of meristematic cells. However, the cell width of the root meristem was not affected. Additionally, DADS increased the IAA and ZR contents of seedling roots in a dose-dependent manner. The influence on IAA content may be mediated by the up-regulation of FZYs and PINs. Further investigation into the underlying mechanism revealed that the expression levels of tomato EXPs were significantly affected by DADS. The expression levels of EXPB2 and beta-expansin precursor were increased after 3 d, and those of EXP1, EXPB3 and EXLB1 were increased after 5 d of DADS treatment (0.41 mM). This result suggests that tomato root growth may be

  19. The Garlic Allelochemical Diallyl Disulfide Affects Tomato Root Growth by Influencing Cell Division, Phytohormone Balance and Expansin Gene Expression.

    Science.gov (United States)

    Cheng, Fang; Cheng, Zhihui; Meng, Huanwen; Tang, Xiangwei

    2016-01-01

    Diallyl disulfide (DADS) is a volatile organosulfur compound derived from garlic (Allium sativum L.), and it is known as an allelochemical responsible for the strong allelopathic potential of garlic. The anticancer properties of DADS have been studied in experimental animals and various types of cancer cells, but to date, little is known about its mode of action as an allelochemical at the cytological level. The current research presents further studies on the effects of DADS on tomato (Solanum lycopersicum L.) seed germination, root growth, mitotic index, and cell size in root meristem, as well as the phytohormone levels and expression profile of auxin biosynthesis genes (FZYs), auxin transport genes (SlPINs), and expansin genes (EXPs) in tomato root. The results showed a biphasic, dose-dependent effect on tomato seed germination and root growth under different DADS concentrations. Lower concentrations (0.01-0.62 mM) of DADS significantly promoted root growth, whereas higher levels (6.20-20.67 mM) showed inhibitory effects. Cytological observations showed that the cell length of root meristem was increased and that the mitotic activity of meristematic cells in seedling root tips was enhanced at lower concentrations of DADS. In contrast, DADS at higher concentrations inhibited root growth by affecting both the length and division activity of meristematic cells. However, the cell width of the root meristem was not affected. Additionally, DADS increased the IAA and ZR contents of seedling roots in a dose-dependent manner. The influence on IAA content may be mediated by the up-regulation of FZYs and PINs. Further investigation into the underlying mechanism revealed that the expression levels of tomato EXPs were significantly affected by DADS. The expression levels of EXPB2 and beta-expansin precursor were increased after 3 d, and those of EXP1, EXPB3 and EXLB1 were increased after 5 d of DADS treatment (0.41 mM). This result suggests that tomato root growth may be

  20. From models to pathogens: how much have we learned about Streptococcus pneumoniae cell division?

    Czech Academy of Sciences Publication Activity Database

    Massida, O.; Nováková, Linda; Vollner, W.

    2013-01-01

    Roč. 15, č. 12 (2013), s. 3133-3157. ISSN 1462-2912 R&D Projects: GA ČR GAP302/12/0256; GA ČR GAP207/12/1568 Institutional support: RVO:61388971 Keywords : SERINE/THREONINE PROTEIN-KINASE * PENICILLIN-BINDING PROTEINS * YYCF RESPONSE-REGULATOR Subject RIV: EE - Microbiology, Virology Impact factor: 6.240, year: 2013

  1. Mutations in the Borrelia burgdorferi Flagellar Type III Secretion System Genes fliH and fliI Profoundly Affect Spirochete Flagellar Assembly, Morphology, Motility, Structure, and Cell Division

    Science.gov (United States)

    Gao, Lihui; Zhao, Xiaowei; Liu, Jun; Norris, Steven J.

    2015-01-01

    ABSTRACT The Lyme disease spirochete Borrelia burgdorferi migrates to distant sites in the tick vectors and mammalian hosts through robust motility and chemotaxis activities. FliH and FliI are two cytoplasmic proteins that play important roles in the type III secretion system (T3SS)-mediated export and assembly of flagellar structural proteins. However, detailed analyses of the roles of FliH and FliI in B. burgdorferi have not been reported. In this study, fliH and fliI transposon mutants were utilized to dissect the mechanism of the Borrelia type III secretion system. The fliH and fliI mutants exhibited rod-shaped or string-like morphology, greatly reduced motility, division defects (resulting in elongated organisms with incomplete division points), and noninfectivity in mice by needle inoculation. Mutants in fliH and fliI were incapable of translational motion in 1% methylcellulose or soft agar. Inactivation of either fliH or fliI resulted in the loss of the FliH-FliI complex from otherwise intact flagellar motors, as determined by cryo-electron tomography (cryo-ET). Flagellar assemblies were still present in the mutant cells, albeit in lower numbers than in wild-type cells and with truncated flagella. Genetic complementation of fliH and fliI mutants in trans restored their wild-type morphology, motility, and flagellar motor structure; however, full-length flagella and infectivity were not recovered in these complemented mutants. Based on these results, disruption of either fliH or fliI in B. burgdorferi results in a severe defect in flagellar structure and function and cell division but does not completely block the export and assembly of flagellar hook and filament proteins. PMID:25968649

  2. Mapping the MinE Site Involved in Interaction with the MinD Division Site Selection Protein of Escherichia coli

    OpenAIRE

    Ma, Lu-Yan; King, Glenn; Rothfield, Lawrence

    2003-01-01

    Interactions between the MinD and MinE proteins are required for proper placement of the Escherichia coli division septum. The site within MinE that is required for interaction with MinD was mapped by studying the effects of site-directed minE mutations on MinD-MinE interactions in yeast two-hybrid and three-hybrid experiments. This confirmed that the MinE N-terminal domain is responsible for the interaction of MinE with MinD. Mutations that interfered with the interaction defined an extended...

  3. Cell wall proteins: a new insight through proteomics

    OpenAIRE

    Jamet, Elisabeth; Canut, Hervé; Boudart, Georges; Pont-Lezica, Rafael F

    2006-01-01

    Cell wall proteins are essential constituents of plant cell walls; they are involved in modifications of cell wall components, wall structure, signaling and interactions with plasma membrane proteins at the cell surface. The application of proteomic approaches to the cell wall compartment raises important questions: are there technical problems specific to cell wall proteomics? What kinds of proteins can be found in Arabidopsis walls? Are some of them unexpected? What sort of post-translation...

  4. Role of SufI (FtsP) in Cell Division of Escherichia coli: Evidence for Its Involvement in Stabilizing the Assembly of the Divisome▿

    OpenAIRE

    Samaluru, Harish; SaiSree, L.; Reddy, Manjula

    2007-01-01

    The function of SufI, a well-studied substrate of the TatABC translocase in Escherichia coli, is not known. It was earlier implicated in cell division, based on the finding that multiple copies of sufI suppressed the phenotypes of cells with mutations in ftsI (ftsI23), which encodes a divisomal transpeptidase. Recently, sufI was identified as both a multicopy suppressor gene and a synthetic lethal mutant of ftsEX, which codes for a division-specific putative ABC transporter. In this study, we...

  5. A parasitic nematode releases cytokinin that controls cell division and orchestrates feeding site formation in host plants

    Science.gov (United States)

    Siddique, Shahid; Radakovic, Zoran S.; De La Torre, Carola M.; Chronis, Demosthenis; Novák, Ondřej; Ramireddy, Eswarayya; Holbein, Julia; Matera, Christiane; Hütten, Marion; Gutbrod, Philipp; Anjam, Muhammad Shahzad; Rozanska, Elzbieta; Habash, Samer; Elashry, Abdelnaser; Sobczak, Miroslaw; Kakimoto, Tatsuo; Strnad, Miroslav; Schmülling, Thomas; Mitchum, Melissa G.; Grundler, Florian M. W.

    2015-01-01

    Sedentary plant-parasitic cyst nematodes are biotrophs that cause significant losses in agriculture. Parasitism is based on modifications of host root cells that lead to the formation of a hypermetabolic feeding site (a syncytium) from which nematodes withdraw nutrients. The host cell cycle is activated in an initial cell selected by the nematode for feeding, followed by activation of neighboring cells and subsequent expansion of feeding site through fusion of hundreds of cells. It is generally assumed that nematodes manipulate production and signaling of the plant hormone cytokinin to activate cell division. In fact, nematodes have been shown to produce cytokinin in vitro; however, whether the hormone is secreted into host plants and plays a role in parasitism remained unknown. Here, we analyzed the spatiotemporal activation of cytokinin signaling during interaction between the cyst nematode, Heterodera schachtii, and Arabidopsis using cytokinin-responsive promoter:reporter lines. Our results showed that cytokinin signaling is activated not only in the syncytium but also in neighboring cells to be incorporated into syncytium. An analysis of nematode infection on mutants that are deficient in cytokinin or cytokinin signaling revealed a significant decrease in susceptibility of these plants to nematodes. Further, we identified a cytokinin-synthesizing isopentenyltransferase gene in H. schachtii and show that silencing of this gene in nematodes leads to a significant decrease in virulence due to a reduced expansion of feeding sites. Our findings demonstrate the ability of a plant-parasitic nematode to synthesize a functional plant hormone to manipulate the host system and establish a long-term parasitic interaction. PMID:26417108

  6. Highly Transient Molecular Interactions Underlie the Stability of Kinetochore–Microtubule Attachment During Cell Division

    Science.gov (United States)

    Zaytsev, Anatoly V.; Ataullakhanov, Fazly I.; Grishchuk, Ekaterina L.

    2013-01-01

    Chromosome segregation during mitosis is mediated by spindle microtubules that attach to chromosomal kinetochores with strong yet labile links. The exact molecular composition of the kinetochore–microtubule interface is not known but microtubules are thought to bind to kinetochores via the specialized microtubule-binding sites, which contain multiple microtubule-binding proteins. During prometaphase the lifetime of microtubule attachments is short but in metaphase it increases 3-fold, presumably owing to dephosphorylation of the microtubule-binding proteins that increases their affinity. Here, we use mathematical modeling to examine in quantitative and systematic manner the general relationships between the molecular properties of microtubule-binding proteins and the resulting stability of microtubule attachment to the protein-containing kinetochore site. We show that when the protein connections are stochastic, the physiological rate of microtubule turnover is achieved only if these molecular interactions are very transient, each lasting fraction of a second. This “microscopic” time is almost four orders of magnitude shorter than the characteristic time of kinetochore–microtubule attachment. Cooperativity of the microtubule-binding events further increases the disparity of these time scales. Furthermore, for all values of kinetic parameters the microtubule stability is very sensitive to the minor changes in the molecular constants. Such sensitivity of the lifetime of microtubule attachment to the kinetics and cooperativity of molecular interactions at the microtubule-binding site may hinder the accurate regulation of kinetochore–microtubule stability during mitotic progression, and it necessitates detailed experimental examination of the microtubule-binding properties of kinetochore-localized proteins. PMID:24376473

  7. Highly Transient Molecular Interactions Underlie the Stability of Kinetochore-Microtubule Attachment During Cell Division.

    Science.gov (United States)

    Zaytsev, Anatoly V; Ataullakhanov, Fazly I; Grishchuk, Ekaterina L

    2013-12-13

    Chromosome segregation during mitosis is mediated by spindle microtubules that attach to chromosomal kinetochores with strong yet labile links. The exact molecular composition of the kinetochore-microtubule interface is not known but microtubules are thought to bind to kinetochores via the specialized microtubule-binding sites, which contain multiple microtubule-binding proteins. During prometaphase the lifetime of microtubule attachments is short but in metaphase it increases 3-fold, presumably owing to dephosphorylation of the microtubule-binding proteins that increases their affinity. Here, we use mathematical modeling to examine in quantitative and systematic manner the general relationships between the molecular properties of microtubule-binding proteins and the resulting stability of microtubule attachment to the protein-containing kinetochore site. We show that when the protein connections are stochastic, the physiological rate of microtubule turnover is achieved only if these molecular interactions are very transient, each lasting fraction of a second. This "microscopic" time is almost four orders of magnitude shorter than the characteristic time of kinetochore-microtubule attachment. Cooperativity of the microtubule-binding events further increases the disparity of these time scales. Furthermore, for all values of kinetic parameters the microtubule stability is very sensitive to the minor changes in the molecular constants. Such sensitivity of the lifetime of microtubule attachment to the kinetics and cooperativity of molecular interactions at the microtubule-binding site may hinder the accurate regulation of kinetochore-microtubule stability during mitotic progression, and it necessitates detailed experimental examination of the microtubule-binding properties of kinetochore-localized proteins. PMID:24376473

  8. Hsp70 protects mitotic cells against heat-induced centrosome damage and division abnormalities

    NARCIS (Netherlands)

    Hut, HMJ; Kampinga, HH; Sibon, OCM

    2005-01-01

    The effect of heat shock on centrosomes has been mainly studied in interphase cells. Centrosomes play a key role in proper segregation of DNA during mitosis. However, the direct effect and consequences of heat shock on mitotic cells and a possible cellular defense system against proteotoxic stress d

  9. Protein Dynamics in Individual Human Cells: Experiment and Theory

    OpenAIRE

    Cohen, Ariel Aharon; Kalisky, Tomer; Mayo, Avi; Geva-Zatorsky, Naama; Danon, Tamar; Issaeva, Irina; Perzov, Natalie; Sigal, Alex; Alon, Uri; Isalan, Mark; Kopito, Ronen; Milo, Ron

    2009-01-01

    A current challenge in biology is to understand the dynamics of protein circuits in living human cells. Can one define and test equations for the dynamics and variability of a protein over time? Here, we address this experimentally and theoretically, by means of accurate time-resolved measurements of endogenously tagged proteins in individual human cells. As a model system, we choose three stable proteins displaying cell-cycle–dependant dynamics. We find that protein accumulation with time pe...

  10. Overview of Production of Protein Using Cell-free Systems

    OpenAIRE

    Gao, Fei Philip

    2014-01-01

    One of the most important steps in protein research is production of the target protein. Cell based systems are mature tools that have long been used to express recombinant proteins by manipulation of the expression organisms. However, it is often challenging to find suitable cell systems that allow for rapid screening of conditions and constructs to produce properly folded, functional proteins in a cost effective manner. As a result, cell-free protein production emerged as an attractive alte...

  11. ASPM regulates symmetric stem cell division by tuning Cyclin E ubiquitination

    Science.gov (United States)

    Capecchi, Mario R.; Pozner, Amir

    2016-01-01

    We generate a mouse model for the human microcephaly syndrome by mutating the ASPM locus, and demonstrate a premature exhaustion of the neuronal progenitor pool due to dysfunctional self-renewal processes. Earlier studies have linked ASPM mutant progenitor excessive cell cycle exit to a mitotic orientation defect. Here, we demonstrate a mitotic orientation-independent effect of ASPM on cell cycle duration. We pinpoint the cell fate-determining factor to the length of time spent in early G1 before traversing the restriction point. Characterization of the molecular mechanism reveals an interaction between ASPM and the Cdk2/Cyclin E complex, regulating the Cyclin activity by modulating its ubiquitination, phosphorylation and localization into the nucleus, before the cell is fated to transverse the restriction point. Thus, we reveal a novel function of ASPM in mediating the tightly coordinated Ubiquitin- Cyclin E- Retinoblastoma- E2F bistable-signalling pathway controlling restriction point progression and stem cell maintenance. PMID:26581405

  12. Redefining Langerhans Cell Histiocytosis as a Myeloid Dysplasia and Identifying B | Division of Cancer Prevention

    Science.gov (United States)

    DESCRIPTION (provided by applicant): Redefining Langerhans Cell Histiocytosis as a Myeloid Dysplasia and Identifying Biomarkers for Early Detection and Risk Assessment. This application addresses Program Announcement PA-09-197: Biomarkers for Early Detection of Hematopoietic Malignancies (R01). The overall aim of this project is to identify novel biomarkers that may be used to diagnose and treat patients with Langerhans Cell Histiocytosis (LCH). LCH occurs with similar frequency as other rare malignancies including Hodgkin's lymphoma and AML. |

  13. A hypothesis to explain division site selection in Escherichia coli by combining nucleoid occlusion and Min.

    Science.gov (United States)

    Norris, Vic; Woldringh, Conrad; Mileykovskaya, Eugenia

    2004-03-12

    The positioning of the site of cell division in Escherichia coli results, it is generally believed, from the operation of nucleoid occlusion in combination with the Min system. Nucleoid occlusion prevents division over the nucleoids and directs it by default to the mid-cell region between segregating nucleoids or to polar regions while the Min system prevents division in polar regions. Unresolved questions include how these systems interact to control the earliest known event in division, the assembly at the membrane of the tubulin-like protein, FtsZ, and, more importantly, what exactly constitutes a division site. Evidence exists that (1) the coupled transcription, translation and insertion of proteins into membrane (transertion), can structure the cytoplasmic membrane into phospholipid domains, (2) the MinD protein can convert vesicles into tubes and (3) a variety of membranous structures can be observed at mid-cell. These data support a model in which transertion from the segregating daughter chromosomes leads to the formation of a distinct proteolipid domain between them at mid-cell; the composition of this domain allows phospholipid tubes to extend like fingers into the cytoplasm; these tubes then become the substrate for the dynamic assembly and disassembly of FtsZ which converts them into the invaginating fold responsible for division; the Min system inhibits division at unwanted sites and times by removing these tubes especially at the cell poles. PMID:15013745

  14. Arabidopsis R-SNARE proteins VAMP721 and VAMP722 are required for cell plate formation.

    Directory of Open Access Journals (Sweden)

    Liang Zhang

    Full Text Available BACKGROUND: Cell plate formation during plant cytokinesis is facilitated by SNARE complex-mediated vesicle fusion at the cell-division plane. However, our knowledge regarding R-SNARE components of membrane fusion machinery for cell plate formation remains quite limited. METHODOLOGY/PRINCIPAL FINDINGS: We report the in vivo function of Arabidopsis VAMP721 and VAMP722, two closely sequence-related R-SNAREs, in cell plate formation. Double homozygous vamp721vamp722 mutant seedlings showed lethal dwarf phenotypes and were characterized by rudimentary roots, cotyledons and hypocotyls. Furthermore, cell wall stubs and incomplete cytokinesis were frequently observed in vamp721vamp722 seedlings. Confocal images revealed that green fluorescent protein-tagged VAMP721 and VAMP722 were preferentially localized to the expanding cell plates in dividing cells. Drug treatments and co-localization analyses demonstrated that punctuate organelles labeled with VAMP721 and VAMP722 represented early endosomes overlapped with VHA-a1-labeled TGN, which were distinct from Golgi stacks and prevacuolar compartments. In addition, protein traffic to the plasma membrane, but not to the vacuole, was severely disrupted in vamp721vamp722 seedlings by subcellular localization of marker proteins. CONCLUSION/SIGNIFICANCE: These observations suggest that VAMP721 and VAMP722 are involved in secretory trafficking to the plasma membrane via TGN/early endosomal compartment, which contributes substantially to cell plate formation during plant cytokinesis.

  15. Viability of adhered bacterial cells: tracking MinD protein oscillations

    Science.gov (United States)

    Barrett, Matt; Colville, Keegan; Schultz-Nielsen, Chris; Jericho, Manfred; Dutcher, John

    2010-03-01

    To study bacterial cells using atomic force microscopy, it is necessary to immobilize the cells on a substrate. Because bacterial cells and common substrates such as glass and mica have a net negative charge, positively charged polymers such as poly-L-lysine (PLL) and polyethyleneimine (PEI) are commonly used as adhesion layers. However, the use of adhesion polymers could stress the cell and even render it inviable. Viable E. coli cells use oscillations of Min proteins along the axis of the rod-shaped cells to ensure accurate cell division. By tagging MinD proteins with GFP, oscillations can be observed using fluorescence microscopy. For a healthy cell in an ideal environment, the oscillation period is measured to be ˜40 s. Prior experiments have shown that PLL increases the oscillation period significantly (up to 80%). In the present study, we have used epifluorescence and total internal reflection fluorescence (TIRF) to track MinD protein oscillations in E. coli bacteria adhered to a variety of positively charged polymers on mica as a function of polymer surface coverage.

  16. Eukaryotic checkpoints are absent in the cell division cycle of Entamoeba histolytica

    Indian Academy of Sciences (India)

    Sulagna Banerjee; Suchismita Das; Anuradha Lohia

    2002-11-01

    Fidelity in transmission of genetic characters is ensured by the faithful duplication of the genome, followed by equal segregation of the genetic material in the progeny. Thus, alternation of DNA duplication (S-phase) and chromosome segregation during the M-phase are hallmarks of most well studied eukaryotes. Several rounds of genome reduplication before chromosome segregation upsets this cycle and leads to polyploidy. Polyploidy is often witnessed in cells prior to differentiation, in embryonic cells or in diseases such as cancer. Studies on the protozoan parasite, Entamoeba histolytica suggest that in its proliferative phase, this organism may accumulate polyploid cells. It has also been shown that although this organism contains sequence homologs of genes which are known to control the cell cycle of most eukaryotes, these genes may be structurally altered and their equivalent function yet to be demonstrated in amoeba. The available information suggests that surveillance mechanisms or ‘checkpoints’ which are known to regulate the eukaryotic cell cycle may be absent or altered in E. histolytica.

  17. In vivo robustness analysis of cell division cycle genes in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Hisao Moriya

    2006-07-01

    Full Text Available Intracellular biochemical parameters, such as the expression level of gene products, are considered to be optimized so that a biological system, including the parameters, works effectively. Those parameters should have some permissible range so that the systems have robustness against perturbations, such as noise in gene expression. However, little is known about the permissible range in real cells because there has been no experimental technique to test it. In this study, we developed a genetic screening method, named "genetic tug-of-war" (gTOW that evaluates upper limit copy numbers of genes in a model eukaryote Saccharomyces cerevisiae, and we applied it for 30 cell-cycle related genes (CDC genes. The experiment provided unique quantitative data that could be used to argue the system-level properties of the cell cycle such as robustness and fragility. The data were used to evaluate the current computational model, and refinements to the model were suggested.

  18. AUREOCHROME1a-Mediated Induction of the Diatom-Specific Cyclin dsCYC2 Controls the Onset of Cell Division in Diatoms (Phaeodactylum tricornutum)[W

    Science.gov (United States)

    Huysman, Marie J.J.; Fortunato, Antonio E.; Matthijs, Michiel; Costa, Benjamin Schellenberger; Vanderhaeghen, Rudy; Van den Daele, Hilde; Sachse, Matthias; Inzé, Dirk; Bowler, Chris; Kroth, Peter G.; Wilhelm, Christian; Falciatore, Angela; Vyverman, Wim; De Veylder, Lieven

    2013-01-01

    Cell division in photosynthetic organisms is tightly regulated by light. Although the light dependency of the onset of the cell cycle has been well characterized in various phototrophs, little is known about the cellular signaling cascades connecting light perception to cell cycle activation and progression. Here, we demonstrate that diatom-specific cyclin 2 (dsCYC2) in Phaeodactylum tricornutum displays a transcriptional peak within 15 min after light exposure, long before the onset of cell division. The product of dsCYC2 binds to the cyclin-dependent kinase CDKA1 and can complement G1 cyclin-deficient yeast. Consistent with the role of dsCYC2 in controlling a G1-to-S light-dependent cell cycle checkpoint, dsCYC2 silencing decreases the rate of cell division in diatoms exposed to light-dark cycles but not to constant light. Transcriptional induction of dsCYC2 is triggered by blue light in a fluence rate-dependent manner. Consistent with this, dsCYC2 is a transcriptional target of the blue light sensor AUREOCHROME1a, which functions synergistically with the basic leucine zipper (bZIP) transcription factor bZIP10 to induce dsCYC2 transcription. The functional characterization of a cyclin whose transcription is controlled by light and whose activity connects light signaling to cell cycle progression contributes significantly to our understanding of the molecular mechanisms underlying light-dependent cell cycle onset in diatoms. PMID:23292736

  19. Treatment with IL-2 and IL-12 inhibits tumour cell division in SL2 lymphoma

    NARCIS (Netherlands)

    Masztalerz, A; Van Luyn, M; Werner, N; Molema, G; Everse, LA; Den Otter, W

    2004-01-01

    We examined which mechanism plays a dominant role in the rejection of solid SL2 lymphoma treated with locally applied IL-2 and /or IL-12. This treatment resulted in about 80% cures. There was a moderate influx of leukocytes in the tissue surrounding tumours; yet these cells failed to invade the soli

  20. The special case of hepatocytes : unique tissue architecture calls for a distinct mode of cell division

    NARCIS (Netherlands)

    Slim, Christiaan L; van IJzendoorn, Sven C D; Lázaro-Diéguez, Francisco; Müsch, Anne

    2014-01-01

    Columnar epithelia (e.g., kidney, intestine) and hepatocytes embody the two major organizational phenotypes of non-stratified epithelial cells. Columnar epithelia establish their apical and basal domains at opposing poles and organize in monolayered cysts and tubules, in which their apical surfaces

  1. Factors Influencing Academic Performance of Students Enrolled in a Lower Division Cell Biology Core Course

    Science.gov (United States)

    Soto, Julio G.; Anand, Sulekha

    2009-01-01

    Students' performance in two semesters of our Cell Biology course was examined for this study. Teaching strategies, behaviors, and pre-course variables were analyzed with respect to students' performance. Pre-semester and post-semester surveys were administered to ascertain students' perceptions about class difficulty, amount of study and effort…

  2. CYCP2;1 integrates genetic and nutritional information to promote meristem cell division in Arabidopsis

    Czech Academy of Sciences Publication Activity Database

    Peng, L.; Skylar, A.; Chang, P.L.; Bišová, Kateřina; Wu, X.

    2014-01-01

    Roč. 393, č. 2 (2014), s. 160-170. ISSN 0012-1606 R&D Projects: GA AV ČR M200201205 Grant ostatní: NSF(US) MCB-1122213 Institutional support: RVO:61388971 Keywords : cell cycle * arabidopsis * meristem Subject RIV: EE - Microbiology, Virology Impact factor: 3.547, year: 2014

  3. Effects of the Scientific Argumentation Based Learning Process on Teaching the Unit of Cell Division and Inheritance to Eighth Grade Students

    Science.gov (United States)

    Balci, Ceyda; Yenice, Nilgun

    2016-01-01

    The aim of this study is to analyse the effects of scientific argumentation based learning process on the eighth grade students' achievement in the unit of "cell division and inheritance". It also deals with the effects of this process on their comprehension about the nature of scientific knowledge, their willingness to take part in…

  4. Development and Application of a Two-Tier Multiple-Choice Diagnostic Test for High School Students' Understanding of Cell Division and Reproduction

    Science.gov (United States)

    Sesli, Ertugrul; Kara, Yilmaz

    2012-01-01

    This study involved the development and application of a two-tier diagnostic test for measuring students' understanding of cell division and reproduction. The instrument development procedure had three general steps: defining the content boundaries of the test, collecting information on students' misconceptions, and instrument development.…

  5. Biosynthesis and release of proteins by isolated pulmonary Clara cells

    International Nuclear Information System (INIS)

    The major proteins synthesized and released by Clara cells were identified and compared with those synthesized and released by mixed lung cells. Highly purified Clara cells (85.9 +/- 2.4%) and mixed lung cells (Clara cells 4%, Type II cells 33%, granulocytes 18%, macrophages 2.7%, ciliated cells 1.2%) were isolated from rabbit lungs, incubated with Ham's F12 medium in collagen/fibronectin-coated plastic culture dishes in the presence of 35S-methionine for periods of 4 and 18 hrs. Radiolabelled proteins were isolated from the cells and from the culture medium, electrophoresed on polyacrylamide gels in the presence of SDS under reducing conditions, and then autoradiographed. After 4 and 18 hr of incubation of the Clara cells the major radiolabelled cell-associated proteins were those with molecular weights of 6, 48, and 180 Kd. The major radiolabelled proteins released by Clara cells into the medium after 4 hrs of incubation had molecular weights of 6, 48, and 180 Kd, accounting for 42, 16, and 10%, respectively, of the total extracellular protein-associated radioactivity. After 18 hr of incubation the 6 and 48 Kd proteins represented 30 and 18% of the total released radioactivity, and the relative amount of the 180 Kd protein had decreased to 3%. With the mixed lung cells, the major proteins released into the medium had molecular weights of 6 and 48 Kd. Under nonreducing conditions the 6 Kd protein released by Clara cells had an apparent molecular weight of 12 Kd. Labelling isolated Clara cells with a mixture of 14C-amino acids also identified this low molecular weight protein as the major secretory product of the Clara cell. The 6 Kd protein did not label when the cells were incubated with 14C-glucosamine indicating that it was not a glycoprotein. Data demonstrate the release of several proteins from isolated Clara cells but the major protein had a M.W. of 6 Kd

  6. Single-cell protein from waste cellulose

    Science.gov (United States)

    Dunlap, C. E.; Callihan, C. D.

    1973-01-01

    The recycle, reuse, or reclamation of single cell protein from liquid and solid agricultural waste fibers by a fermentation process is reported. It is shown that cellulose comprises the bulk of the fibers at 50% to 55% of the dry weight of the refuse and that its biodegradability is of prime importance in the choice of a substrate. The application of sodium hydroxide followed by heat and pressure serves to de-polymerize and disrupt lignin structure while swelling the cellulose to increase water uptake and pore volume. Some of the lignin, hemi-celluloses, ash, and cellulose of the material is hydrolized and solubilized. Introduction of microorganisms to the substrate fibers mixed with nutrients produces continuous fermentation of cellulose for further protein extraction and purification.

  7. Robustness of the division symmetry in Escherichia coli and functional consequences of symmetry breaking

    International Nuclear Information System (INIS)

    The morphological symmetry of the division process of Escherichia coli is well-known. Recent studies verified that, in optimal growth conditions, most divisions are symmetric, although there are exceptions. We investigate whether such morphological asymmetries in division introduce functional asymmetries between sister cells, and assess the robustness of the symmetry in division to mild chemical stresses and sub-optimal temperatures. First, we show that the difference in size between daughter cells at birth is positively correlated to the difference between the numbers of fluorescent protein complexes inherited from the parent cell. Next, we show that the degree of symmetry in division observed in optimal conditions is robust to mild acidic shift and to mild oxidative stress, but not to sub-optimal temperatures, in that the variance of the difference between the sizes of sister cells at birth is minimized at 37 °C. This increased variance affects the functionality of the cells in that, at sub-optimal temperatures, larger/smaller cells arising from asymmetric divisions exhibit faster/slower division times than the mean population division time, respectively. On the other hand, cells dividing faster do not do so at the cost of morphological symmetry in division. Finally we show that at suboptimal temperatures the mean distance between the nucleoids increases, explaining the increased variance in division. We conclude that the functionality of E. coli cells is not immune to morphological asymmetries at birth, and that the effectiveness of the mechanism responsible for ensuring the symmetry in division weakens at sub-optimal temperatures. (paper)

  8. Genes adopt non-optimal codon usage to generate cell cycle-dependent oscillations in protein levels

    DEFF Research Database (Denmark)

    Frenkel-Morgenstern, Milana; Danon, Tamar; Christian, Thomas;

    2012-01-01

    The cell cycle is a temporal program that regulates DNA synthesis and cell division. When we compared the codon usage of cell cycle-regulated genes with that of other genes, we discovered that there is a significant preference for non-optimal codons. Moreover, genes encoding proteins that cycle at...... the protein level exhibit non-optimal codon preferences. Remarkably, cell cycle-regulated genes expressed in different phases display different codon preferences. Here, we show empirically that transfer RNA (tRNA) expression is indeed highest in the G2 phase of the cell cycle, consistent with the non......-optimal codon usage of genes expressed at this time, and lowest toward the end of G1, reflecting the optimal codon usage of G1 genes. Accordingly, protein levels of human glycyl-, threonyl-, and glutamyl-prolyl tRNA synthetases were found to oscillate, peaking in G2/M phase. In light of our findings, we propose...

  9. A Cell-Based Protein-Protein Interaction Method Using a Permuted Luciferase Reporter

    OpenAIRE

    Eishingdrelo, Haifeng; Cai, Jidong; Weissensee, Paul; Sharma, Praveen; Tocci, Michael J; Wright, Paul S

    2011-01-01

    We have developed a novel cell-based protein-protein interaction assay method. The method relies on conversion of an inactive permuted luciferase containing a Tobacco Etch Virus protease (TEV) cleavage sequence fused onto protein (A) to an active luciferase upon interaction and cleavage by another protein (B) fused with the TEV protease. We demonstrate assay applicability for ligand-induced protein-protein interactions including G-protein coupled receptors, receptor tyrosine kinases and nucle...

  10. DNA replication initiation, doubling of rate of phospholipid synthesis, and cell division in Escherichia coli.

    OpenAIRE

    Joseleau-Petit, D; Képès, F; Peutat, L; D'Ari, R; Képès, A

    1987-01-01

    In synchronized culture of Escherichia coli, the specific arrest of phospholipid synthesis (brought about by glycerol starvation in an appropriate mutant) did not affect the rate of ongoing DNA synthesis but prevented the initiation of new rounds. The initiation block did not depend on cell age at the time of glycerol removal, which could be before, during, or after the doubling in the rate of phospholipid synthesis (DROPS) and as little as 10 min before the expected initiation. We conclude t...

  11. Osteogenic differentiation of hypertrophic chondrocytes involves asymmetric cell divisions and apoptosis

    OpenAIRE

    1995-01-01

    We have investigated the early cellular events that take place during the change in lineage commitment from hypertrophic chondrocytes to osteoblast-like cells. We have induced this osteogenic differentiation by cutting through the hypertrophic cartilage of embryonic chick femurs and culturing the explants. Immunocytochemical characterization, [3H]thymidine pulse-chase labeling, in situ nick translation or end labeling of DNA breaks were combined with ultrastructural studies to characterize th...

  12. Quantification of Protein Levels in Single Living Cells

    Directory of Open Access Journals (Sweden)

    Chiu-An Lo

    2015-12-01

    Full Text Available Accurate measurement of the amount of specific protein a cell produces is important for investigating basic molecular processes. We have developed a technique that allows for quantitation of protein levels in single cells in vivo. This protein quantitation ratioing (PQR technique uses a genetic tag that produces a stoichiometric ratio of a fluorescent protein reporter and the protein of interest during protein translation. The fluorescence intensity is proportional to the number of molecules produced of the protein of interest and is used to determine the relative amount of protein within the cell. We use PQR to quantify protein expression of different genes using quantitative imaging, electrophysiology, and phenotype. We use genome editing to insert Protein Quantitation Reporters into endogenous genomic loci in three different genomes for quantitation of endogenous protein levels. The PQR technique will allow for a wide range of quantitative experiments examining gene-to-phenotype relationships with greater accuracy.

  13. Cell-cycle specific expression of a small proline-rich protein in Chinese hamster ovary cells

    International Nuclear Information System (INIS)

    Squamous metaplasia of the bronchial epithelium is generally believed to be involved in the neoplastic progression toward squamous cell carcinomas. Thus, it is important to understand the mechanisms controlling this type of differentiation. The induction of two families of cDNAs encoding a small proline-rich protein (sPRP), sprI and sprII, was first identified in human keratinocytes exhibiting squamous differentiation. cDNAs similar to sprI have also been identified in cultured tracheal epithelial cells undergoing squamous differentiation. The first step during the squamous differentiation process is the inhibition of cell growth; it has also been noted that a sPRP mRNA in Chinese hamster ovary (CHO) cells is induced 10-fold just before the cultures reach confluence. Thus, sPRP may stop cell division in cells undergoing squamous differentation. In support of this possibility are the recent investigations correlating expression of sPRP with cell morphology. Specific immunoreactivity to sPRP, using affinity-purified antibodies, showed a strong immunostaining in cells with a round configuration, while less staining was observed in other cells. The major part of the CHO population showed no immunoreactivity. One interpretation of this observation is that the expression of sPRP may be cell-cyle regulated. The purpose of this investigation was to determine the phase of the cell cycle where induced synthesis of sPRP mRNA occurs

  14. Lymph Node Metastases and Prognosis in Left Upper Division Non-Small Cell Lung Cancers: The Impact of Interlobar Lymph Node Metastasis.

    Directory of Open Access Journals (Sweden)

    Hiroaki Kuroda

    Full Text Available Left upper division segmentectomy is one of the major pulmonary procedures; however, it is sometimes difficult to completely dissect interlobar lymph nodes. We attempted to clarify the prognostic importance of hilar and mediastinal nodes, especially of interlobar lymph nodes, in patients with primary non-small cell lung cancer (NSCLC located in the left upper division.We retrospectively studied patients with primary left upper lobe NSCLC undergoing surgical pulmonary resection (at least lobectomy with radical lymphadenectomy. The representative evaluation of therapeutic value from the lymph node dissection was determined using Sasako's method. This analysis was calculated by multiplying the frequency of metastasis to the station and the 5-year survival rate of the patients with metastasis to the station.We enrolled 417 patients (237 men, 180 women. Tumors were located in the lingular lobe and at the upper division of left upper lobe in 69 and 348 patients, respectively. The pathological nodal statuses were pN0 in 263 patients, pN1 in 70 patients, and pN2 in 84 patients. Lymph nodes #11 and #7 were significantly correlated with differences in node involvement in patients with left upper lobe NSCLC. Among those with left upper division NSCLC, the 5-year overall survival in pN1 was 31.5% for #10, 39.3% for #11, and 50.4% for #12U. The involvement of node #11 was 1.89-fold higher in the anterior segment than that in the apicoposterior segment. The therapeutic index of estimated benefit from lymph node dissection for #11 was 3.38, #4L was 1.93, and the aortopulmonary window was 4.86 in primary left upper division NSCLC.Interlobar node involvement is not rare in left upper division NSCLC, occurring in >20% cases. Furthermore, dissection of interlobar nodes was found to be beneficial in patients with left upper division NSCLC.

  15. Calreticulin: Roles in Cell-Surface Protein Expression

    Directory of Open Access Journals (Sweden)

    Yue Jiang

    2014-09-01

    Full Text Available In order to perform their designated functions, proteins require precise subcellular localizations. For cell-surface proteins, such as receptors and channels, they are able to transduce signals only when properly targeted to the cell membrane. Calreticulin is a multi-functional chaperone protein involved in protein folding, maturation, and trafficking. However, evidence has been accumulating that calreticulin can also negatively regulate the surface expression of certain receptors and channels. In these instances, depletion of calreticulin enhances cell-surface expression and function. In this review, we discuss the role of calreticulin with a focus on its negative effects on the expression of cell-surface proteins.

  16. Surviving protein quality control catastrophes--from cells to organisms.

    Science.gov (United States)

    Schneider, Kim; Bertolotti, Anne

    2015-11-01

    Organisms have evolved mechanisms to cope with and adapt to unexpected challenges and harsh conditions. Unfolded or misfolded proteins represent a threat for cells and organisms, and the deposition of misfolded proteins is a defining feature of many age-related human diseases, including the increasingly prevalent neurodegenerative diseases. These protein misfolding diseases are devastating and currently cannot be cured, but are hopefully not incurable. In fact, the aggregation-prone and potentially harmful proteins at the origins of protein misfolding diseases are expressed throughout life, whereas the diseases are late onset. This reveals that cells and organisms are normally resilient to disease-causing proteins and survive the threat of misfolded proteins up to a point. This Commentary will outline the limits of the cellular resilience to protein misfolding, and discuss the possibility of pushing these limits to help cells and organisms to survive the threat of misfolding proteins and to avoid protein quality control catastrophes. PMID:26483388

  17. VP22 herpes simplex virus protein can transduce proteins into stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Gabanyi, I.; Lojudice, F.H.; Kossugue, P.M. [Centro de Terapia Celular e Molecular, Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP (Brazil); Rebelato, E. [Departamento de Fisiologia e Biofísica, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP (Brazil); Demasi, M.A.; Sogayar, M.C. [Centro de Terapia Celular e Molecular, Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP (Brazil)

    2013-02-01

    The type I herpes simplex virus VP22 tegument protein is abundant and well known for its ability to translocate proteins from one cell to the other. In spite of some reports questioning its ability to translocate proteins by attributing the results observed to fixation artifacts or simple attachment to the cell membrane, VP22 has been used to deliver several proteins into different cell types, triggering the expected cell response. However, the question of the ability of VP22 to enter stem cells has not been addressed. We investigated whether VP22 could be used as a tool to be applied in stem cell research and differentiation due to its capacity to internalize other proteins without altering the cell genome. We generated a VP22.eGFP construct to evaluate whether VP22 could be internalized and carry another protein with it into two different types of stem cells, namely adult human dental pulp stem cells and mouse embryonic stem cells. We generated a VP22.eGFP fusion protein and demonstrated that, in fact, it enters stem cells. Therefore, this system may be used as a tool to deliver various proteins into stem cells, allowing stem cell research, differentiation and the generation of induced pluripotent stem cells in the absence of genome alterations.

  18. VP22 herpes simplex virus protein can transduce proteins into stem cells

    International Nuclear Information System (INIS)

    The type I herpes simplex virus VP22 tegument protein is abundant and well known for its ability to translocate proteins from one cell to the other. In spite of some reports questioning its ability to translocate proteins by attributing the results observed to fixation artifacts or simple attachment to the cell membrane, VP22 has been used to deliver several proteins into different cell types, triggering the expected cell response. However, the question of the ability of VP22 to enter stem cells has not been addressed. We investigated whether VP22 could be used as a tool to be applied in stem cell research and differentiation due to its capacity to internalize other proteins without altering the cell genome. We generated a VP22.eGFP construct to evaluate whether VP22 could be internalized and carry another protein with it into two different types of stem cells, namely adult human dental pulp stem cells and mouse embryonic stem cells. We generated a VP22.eGFP fusion protein and demonstrated that, in fact, it enters stem cells. Therefore, this system may be used as a tool to deliver various proteins into stem cells, allowing stem cell research, differentiation and the generation of induced pluripotent stem cells in the absence of genome alterations

  19. A trisubstituted benzimidazole cell division inhibitor with efficacy against Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Susan E Knudson

    Full Text Available Trisubstituted benzimidazoles have demonstrated potency against Gram-positive and Gram-negative bacterial pathogens. Previously, a library of novel trisubstituted benzimidazoles was constructed for high throughput screening, and compounds were identified that exhibited potency against M. tuberculosis H37Rv and clinical isolates, and were not toxic to Vero cells. A new series of 2-cyclohexyl-5-acylamino-6-N, N-dimethylaminobenzimidazoles derivatives has been developed based on SAR studies. Screening identified compounds with potency against M. tuberculosis. A lead compound from this series, SB-P17G-A20, was discovered to have an MIC of 0.16 µg/mL and demonstrated efficacy in the TB murine acute model of infection based on the reduction of bacterial load in the lungs and spleen by 1.73 ± 0.24 Log10 CFU and 2.68 ± Log10 CFU, respectively, when delivered at 50 mg/kg by intraperitoneal injection (IP twice daily (bid. The activity of SB-P17G-A20 was determined to be concentration dependent and to have excellent stability in mouse and human plasma, and liver microsomes. Together, these studies demonstrate that SB-P17G-A20 has potency against M. tuberculosis clinical strains with varying susceptibility and efficacy in animal models of infection, and that trisubstituted benzimidazoles continue to be a platform for the development of novel inhibitors with efficacy.

  20. Effective formation of the segregation-competent complex determines successful partitioning of the bovine papillomavirus genome during cell division.

    Science.gov (United States)

    Silla, Toomas; Männik, Andres; Ustav, Mart

    2010-11-01

    Effective segregation of the bovine papillomavirus type 1 (BPV1), Epstein-Barr virus (EBV), and Kaposi's sarcoma-associated human herpesvirus type 8 (KSHV) genomes into daughter cells is mediated by a single viral protein that tethers viral genomes to host mitotic chromosomes. The linker proteins that mediate BPV1, EBV, and KSHV segregation are E2, LANA1, and EBNA1, respectively. The N-terminal transactivation domain of BPV1 E2 is responsible for chromatin attachment and subsequent viral genome segregation. Because E2 transcriptional activation and chromatin attachment functions are not mutually exclusive, we aimed to determine the requirement of these activities during segregation by analyzing chimeric E2 proteins. This approach allowed us to separate the two activities. Our data showed that attachment of the segregation protein to chromatin is not sufficient for proper segregation. Rather, formation of a segregation-competent complex which carries multiple copies of the segregation protein is required. Complementation studies of E2 functional domains indicated that chromatin attachment and transactivation functions must act in concert to ensure proper plasmid segregation. These data indicate that there are specific interactions between linker molecules and transcription factors/complexes that greatly increase segregation-competent complex formation. We also showed, using hybrid E2 molecules, that restored segregation function does not involve interactions with Brd4. PMID:20810736

  1. Denaturation of membrane proteins and hyperthermic cell killing

    OpenAIRE

    Burgman, Paulus Wilhelmus Johannes Jozef

    1993-01-01

    Summarizing: heat induced denaturation of membrane proteins is probably related to hyperthermic cell killing. Induced resistance of heat sensitive proteins seems to be involved in the development of thermotolerance. Although many questions remain still to be answered, it appears that HSP72, when bound to membrane proteins, is capable of providing heat resistance to these proteins. ... Zie: Summary

  2. Learning Cell Biology as a Team: A Project-Based Approach to Upper-Division Cell Biology

    Science.gov (United States)

    Wright, Robin; Boggs, James

    2002-01-01

    To help students develop successful strategies for learning how to learn and communicate complex information in cell biology, we developed a quarter-long cell biology class based on team projects. Each team researches a particular human disease and presents information about the cellular structure or process affected by the disease, the cellular…

  3. Fluorescence lifetime imaging microscopy (FLIM) to quantify protein-protein interactions inside cells.

    Science.gov (United States)

    Duncan, R R

    2006-11-01

    Recent developments in cellular imaging spectroscopy now permit the minimally invasive study of protein dynamics inside living cells. These advances are of interest to cell biologists, as proteins rarely act in isolation, but rather in concert with others in forming cellular machinery. Until recently, all protein interactions had to be determined in vitro using biochemical approaches: this biochemical legacy has provided cell biologists with the basis to test defined protein-protein interactions not only inside cells, but now also with high spatial resolution. These techniques can detect and quantify protein behaviours down to the single-molecule level, all inside living cells. More recent developments in TCSPC (time-correlated single-photon counting) imaging are now also driving towards being able to determine protein interaction rates with similar spatial resolution, and together, these experimental advances allow investigators to perform biochemical experiments inside living cells. PMID:17052173

  4. Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is due to altered cell division, phytohormone balance and expansin gene expression.

    Science.gov (United States)

    Soltys, Dorota; Rudzińska-Langwald, Anna; Gniazdowska, Agnieszka; Wiśniewska, Anita; Bogatek, Renata

    2012-11-01

    Cyanamide (CA) has been reported as a natural compound produced by hairy vetch (Vicia villosa Roth.) and it was shown also to be an allelochemical, responsible for strong allelopathic potential in this species. CA phytotoxicity has been demonstrated on various plant species, but to date little is known about its mode of action at cellular level. Treatment of tomato (Solanum lycopersicum L.) roots with CA (1.2 mM) resulted in inhibition of growth accompanied by alterations in cell division, and imbalance of plant hormone (ethylene and auxin) homeostasis. Moreover, the phytotoxic effect of CA was also manifested by modifications in expansin gene expression, especially in expansins responsible for cell wall remodeling after the cytokinesis (LeEXPA9, LeEXPA18). Based on these results the phytotoxic activity of CA on growth of roots of tomato seedlings is likely due to alterations associated with cell division. PMID:22847024

  5. CEH-20/Pbx and UNC-62/Meis function upstream of rnt-1/Runx to regulate asymmetric divisions of the C. elegans stem-like seam cells

    Directory of Open Access Journals (Sweden)

    Samantha Hughes

    2013-06-01

    Caenorhabditis elegans seam cells divide in the stem-like mode throughout larval development, with the ability to both self-renew and produce daughters that differentiate. Seam cells typically divide asymmetrically, giving rise to an anterior daughter that fuses with the hypodermis and a posterior daughter that proliferates further. Previously we have identified rnt-1 (a homologue of the mammalian cancer-associated stem cell regulator Runx as being an important regulator of seam development, acting to promote proliferation; rnt-1 mutants have fewer seam cells whereas overexpressing rnt-1 causes seam cell hyperplasia. We isolated the interacting CEH-20/Pbx and UNC-62/Meis TALE-class transcription factors during a genome-wide RNAi screen for novel regulators of seam cell number. Animals lacking wild type CEH-20 or UNC-62 display seam cell hyperplasia, largely restricted to the anterior of the worm, whereas double mutants have many additional seam cells along the length of the animal. The cellular basis of the hyperplasia involves the symmetrisation of normally asymmetric seam cell divisions towards the proliferative stem-like fate. The hyperplasia is completely suppressed in rnt-1 mutants, and rnt-1 is upregulated in ceh-20 and unc-62 mutants, suggesting that CEH-20 and UNC-62 function upstream of rnt-1 to limit proliferative potential to the appropriate daughter cell. In further support of this we find that CEH-20 is asymmetrically localised in seam daughters following an asymmetric division, being predominantly restricted to anterior nuclei whose fate is to differentiate. Thus, ceh-20 and unc-62 encode crucial regulators of seam cell division asymmetry, acting via rnt-1 to regulate the balance between proliferation and differentiation.

  6. Proteins upregulated by mild and severe hypoxia in squamous cell carcinomas in vitro identified by proteomics

    International Nuclear Information System (INIS)

    found to be upregulated in FaDuDD at both 1% and 0% oxygen. Conclusions: The upregulated proteins observed in this study are involved in different cellular processes, as regulators of both cell metabolism and stress response, and in cell migration and cell division. All of which may contribute to cell survival and adaptation during oxygen starvation.

  7. A comparison between protein profiles of B cell subpopulations and mantle cell lymphoma cells

    Directory of Open Access Journals (Sweden)

    Lehtiö Janne

    2009-11-01

    Full Text Available Abstract Background B-cell lymphomas are thought to reflect different stages of B-cell maturation. Based on cytogenetics and molecular markers, mantle cell lymphoma (MCL is presumed to derive predominantly from naïve, pre-germinal centre (pre-GC B lymphocytes. The aim of this study was to develop a method to investigate the similarity between MCL cells and different B-cell compartments on a protein expression level. Methods Subpopulations of B cells representing the germinal centre (GC, the pre-GC mantle zone and the post-GC marginal zone were isolated from tonsils using automated magnetic cell sorting (AutoMACS of cells based on their expression of CD27 and IgD. Protein profiling of the B cell subsets, of cell lines representing different lymphomas and of primary MCL samples was performed using top-down proteomics profiling by surface-enhanced laser detection/ionization time-of-flight mass spectrometry (SELDI-TOF-MS. Results Quantitative MS data of significant protein peaks (p-value Conclusion AutoMACS sorting generates sufficient purity to enable a comparison between protein profiles of B cell subpopulations and malignant B lymphocytes applying SELDI-TOF-MS. Further validation with an increased number of patient samples and identification of differentially expressed proteins would enable a search for possible treatment targets that are expressed during the early development of MCL.

  8. Pro-B-cell-specific transcription and proapoptotic function of protein kinase Ceta.

    Science.gov (United States)

    Morrow, T A; Muljo, S A; Zhang, J; Hardwick, J M; Schlissel, M S

    1999-08-01

    Using a subtractive cloning scheme on cDNA prepared from primary pro-B and pre-B cells, we identified several genes whose products regulate apoptosis. We further characterized one of these genes, encoding protein kinase Ceta (PKCeta). PKCeta transcripts were readily detected in pro-B cells but were absent in pre-B cells. Although both a full-length and a truncated form of PKCeta were detectable in bone marrow pro-B cells, transition to the pre-B-cell stage was associated with increased relative levels of truncated PKCeta. We found that PKCeta is proteolyzed in apoptotic lymphocytes, generating a kinase-active fragment identical to the truncated form which is capable of inducing apoptosis when expressed in a pro-B cell line. Caspase-3 can generate an identical PKCeta cleavage product in vitro, and caspase inhibitors prevent the generation of this product during apoptosis in transfected cell lines. Inducible overexpression of either the full-length or truncated form of PKCeta results in cell cycle arrest at the G(1)/S transition. These results suggest that the expression and proteolytic activation of PKCeta play an important role in the regulation of cell division and cell death during early B-cell development. PMID:10409750

  9. Interaction of Proteins Identified in Human Thyroid Cells

    Directory of Open Access Journals (Sweden)

    Jessica Pietsch

    2013-01-01

    Full Text Available Influence of gravity forces on the regulation of protein expression by healthy and malignant thyroid cells was studied with the aim to identify protein interactions. Western blot analyses of a limited number of proteins suggested a time-dependent regulation of protein expression by simulated microgravity. After applying free flow isoelectric focusing and mass spectrometry to search for differently expressed proteins by thyroid cells exposed to simulated microgravity for three days, a considerable number of candidates for gravi-sensitive proteins were detected. In order to show how proteins sensitive to microgravity could directly influence other proteins, we investigated all polypeptide chains identified with Mascot scores above 100, looking for groups of interacting proteins. Hence, UniProtKB entry numbers of all detected proteins were entered into the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING and processed. The program indicated that we had detected various groups of interacting proteins in each of the three cell lines studied. The major groups of interacting proteins play a role in pathways of carbohydrate and protein metabolism, regulation of cell growth and cell membrane structuring. Analyzing these groups, networks of interaction could be established which show how a punctual influence of simulated microgravity may propagate via various members of interaction chains.

  10. Contribution of the Ade Resistance-Nodulation-Cell Division-Type Efflux Pumps to Fitness and Pathogenesis of Acinetobacter baumannii

    Science.gov (United States)

    Yoon, Eun-Jeong; Balloy, Viviane; Fiette, Laurence; Chignard, Michel; Courvalin, Patrice

    2016-01-01

    ABSTRACT Overexpression of chromosomal resistance-nodulation-cell division (RND)-type efflux systems with broad substrate specificity contributes to multidrug resistance (MDR) in Acinetobacter baumannii. We have shown that modulation of expression of the structural genes for the efflux systems AdeABC and AdeIJK confers MDR and results in numerous alterations of membrane-associated cellular functions, in particular biofilm formation. However, the contribution of these RND pumps to cell fitness and virulence has not yet been studied. The biological cost of an antibiotic resistance mechanism is a key parameter in determining its stability and dissemination. From an entirely sequenced susceptible clinical isolate, we have generated a set of isogenic derivatives having single point mutations resulting in overexpression of each efflux system or with every pump deleted by allelic replacement. We found that overproduction of the pumps results in a significant decrease in fitness of the bacterial host when measured by competition experiments in vitro. Fitness and virulence were also evaluated in vivo both in systemic and pulmonary infection models in immunocompetent mice. A diminished competitiveness of the AdeABC-overexpressing mutant was observed only after intraperitoneal inoculation, but not after intranasal inoculation, the latter mimicking the most frequent type of human A. baumannii infection. However, in mice infected intranasally, this mutant was more virulent and stimulated an enhanced neutrophil activation in the lungs. Altogether, these data account for the observation that adeABC overexpression is common in MDR A. baumannii frequently found in ventilator-associated pneumonia. PMID:27247231

  11. Microcarrier culture of lepidopteran cell lines: implications for growth and recombinant protein production.

    Science.gov (United States)

    Ikonomou, Laertis; Drugmand, Jean-Christophe; Bastin, Georges; Schneider, Yves-Jacques; Agathos, Spiros N

    2002-01-01

    Several microcarrier systems were screened with Sf-9 and High-Five cell lines as to their ability to support cell growth and recombinant (beta-galactosidase) protein production. Growth of both cell lines on compact microcarriers, such as Cytodex-1 and glass beads, was minimal, as cells detached easily from the microcarrier surface and grew as single cells in the medium. Cell growth was also problematic on Cytopore-1 and -2 porous microcarriers. Cells remained attached for several days inside the microcarrier pores, but no cell division and proliferation were observed. On the contrary, insect cells grew well in the interior of Fibra-Cel disks mainly as aggregates at points of fiber intersection, reaching final (plateau) densities of about 4 x 10(6) (Sf-9) and 2.7 x 10(6) (High-Five) cells mL(-1) (8 x 10(6) and 5.5 x 10(6) cells per cm(2) of projected disk area, respectively). Their growth was described well by the logistic equation, which takes into account possible inhibition effects. Beta-Galactosidase (beta-gal) production of Sf-9 cells on Fibra-Cel disks (infected at 3.3 x 10(6) cells mL(-1)) was prolonged (192 h), and specific protein production was similar to that of high-density free cell infection. Cultispher-S microcarriers were found to be a very efficient system for the growth of High-Five cells, whereas no growth of Sf-9 cells took place for the same system. Concentrations of about 9 x 10(6) cells mL(-1) were reached within 120 h, with cell growth in both microcarriers and aggregates, appearance of cellular bridges between microcarriers and aggregates, and eventual formation of macroaggregates incorporating several microcarriers. Specific protein productions after beta-gal baculovirus infection at increasing cell concentrations were almost constant, thus leading to elevated volumetric protein production: final beta-gal titers of 946, 1728, and 1484 U mL(-1) were obtained for infection densities of 3.4, 7.2, and 8.9 x 10(6) cells mL(-1), respectively

  12. Protein dynamics in individual human cells: experiment and theory.

    Directory of Open Access Journals (Sweden)

    Ariel Aharon Cohen

    Full Text Available A current challenge in biology is to understand the dynamics of protein circuits in living human cells. Can one define and test equations for the dynamics and variability of a protein over time? Here, we address this experimentally and theoretically, by means of accurate time-resolved measurements of endogenously tagged proteins in individual human cells. As a model system, we choose three stable proteins displaying cell-cycle-dependant dynamics. We find that protein accumulation with time per cell is quadratic for proteins with long mRNA life times and approximately linear for a protein with short mRNA lifetime. Both behaviors correspond to a classical model of transcription and translation. A stochastic model, in which genes slowly switch between ON and OFF states, captures measured cell-cell variability. The data suggests, in accordance with the model, that switching to the gene ON state is exponentially distributed and that the cell-cell distribution of protein levels can be approximated by a Gamma distribution throughout the cell cycle. These results suggest that relatively simple models may describe protein dynamics in individual human cells.

  13. Effects of the Scientific Argumentation Based Learning Process on Teaching the Unit of Cell Division and Inheritance to Eighth Grade Students

    OpenAIRE

    Balci, Ceyda; Yenice, Nilgun

    2015-01-01

    The aim of this study is to analyse the effects of scientific argumentation based learning process on the eighth grade students’ achievement in the unit of “cell division and inheritance”. It also deals with the effects of this process on their comprehension about the nature of scientific knowledge, their willingness to take part in discussions and their attitude towards the course of science and technology. The study employed the design of pretest-post test matched control group design which...

  14. Toward Spatially Regulated Division of Protocells: Insights into the E. coli Min System from in Vitro Studies.

    Science.gov (United States)

    Kretschmer, Simon; Schwille, Petra

    2014-01-01

    For reconstruction of controlled cell division in a minimal cell model, or protocell, a positioning mechanism that spatially regulates division is indispensable. In Escherichia coli, the Min proteins oscillate from pole to pole to determine the division site by inhibition of the primary divisome protein FtsZ anywhere but in the cell middle. Remarkably, when reconstituted under defined conditions in vitro, the Min proteins self-organize into spatiotemporal patterns in the presence of a lipid membrane and ATP. We review recent progress made in studying the Min system in vitro, particularly focusing on the effects of various physicochemical parameters and boundary conditions on pattern formation. Furthermore, we discuss implications and challenges for utilizing the Min system for division site placement in protocells. PMID:25513760

  15. Toward Spatially Regulated Division of Protocells: Insights into the E. coli Min System from in Vitro Studies

    Directory of Open Access Journals (Sweden)

    Simon Kretschmer

    2014-12-01

    Full Text Available For reconstruction of controlled cell division in a minimal cell model, or protocell, a positioning mechanism that spatially regulates division is indispensable. In Escherichia coli, the Min proteins oscillate from pole to pole to determine the division site by inhibition of the primary divisome protein FtsZ anywhere but in the cell middle. Remarkably, when reconstituted under defined conditions in vitro, the Min proteins self-organize into spatiotemporal patterns in the presence of a lipid membrane and ATP. We review recent progress made in studying the Min system in vitro, particularly focusing on the effects of various physicochemical parameters and boundary conditions on pattern formation. Furthermore, we discuss implications and challenges for utilizing the Min system for division site placement in protocells.

  16. Formation of multinuclear cells induced by dimethyl sulfoxide: inhibition of cytokinesis and occurrence of novel nuclear division in dictyostelium cells

    OpenAIRE

    Fukui, Y.

    1980-01-01

    Our previous studies showed that 10 percent dimethyl sulfoxide (DMSO) induces the formation of actin microfilament bundles in the cell nucleus together with the dislocation of cortical microfilaments from the plasma membrane. The present study investigated the effects of DMSO on diverse activities mediated by cellular microfilaments as the second step toward assessing potential differences between nuclear and cytoplasmic actins of dictyostelium mucoroides. DMSO was found to reversibly inhibit...

  17. Wounding coordinately induces cell wall protein, cell cycle and pectin methyl esterase genes involved in tuber closing layer and wound periderm development.

    Science.gov (United States)

    Neubauer, Jonathan D; Lulai, Edward C; Thompson, Asunta L; Suttle, Jeffrey C; Bolton, Melvin D

    2012-04-15

    Little is known about the coordinate induction of genes that may be involved in agriculturally important wound-healing events. In this study, wound-healing events were determined together with wound-induced expression profiles of selected cell cycle, cell wall protein, and pectin methyl esterase genes using two diverse potato genotypes and two harvests (NDTX4271-5R and Russet Burbank tubers; 2008 and 2009 harvests). By 5 d after wounding, the closing layer and a nascent phellogen had formed. Phellogen cell divisions generated phellem layers until cessation of cell division at 28 d after wounding for both genotypes and harvests. Cell cycle genes encoding epidermal growth factor binding protein (StEBP), cyclin-dependent kinase B (StCDKB) and cyclin-dependent kinase regulatory subunit (StCKS1At) were induced by 1 d after wounding; these expressions coordinated with related phellogen formation and the induction and cessation of phellem cell formation. Genes encoding the structural cell wall proteins extensin (StExt1) and extensin-like (StExtlk) were dramatically up-regulated by 1-5 d after wounding, suggesting involvement with closing layer and later phellem cell layer formation. Wounding up-regulated pectin methyl esterase genes (StPME and StPrePME); StPME expression increased during closing layer and phellem cell formation, whereas maximum expression of StPrePME occurred at 5-14 d after wounding, implicating involvement in later modifications for closing layer and phellem cell formation. The coordinate induction and expression profile of StTLRP, a gene encoding a cell wall strengthening "tyrosine-and lysine-rich protein," suggested a role in the formation of the closing layer followed by phellem cell generation and maturation. Collectively, the genes monitored were wound-inducible and their expression profiles markedly coordinated with closing layer formation and the index for phellogen layer meristematic activity during wound periderm development; results were more

  18. Protein translation machinery holds a key for transition of planktonic cells to biofilm state in Enterococcus faecalis: A proteomic approach.

    Science.gov (United States)

    Qayyum, Shariq; Sharma, Divakar; Bisht, Deepa; Khan, Asad U

    2016-06-10

    Enterococcus faecalis is a member of human gut microflora causing nosocomial infection involving biofilm formation. Ethyl methyl sulfonate induced mutants were analysed using crystal violet assay, SEM and CLSM microscopy which confirmed AK-E12 as biofilm efficient and AK-F6 as biofilm deficient mutants. Growth curve pattern revealed AK-E12 was fast growing whereas, AK-F6 was found slow growing mutant. 2D-Electrophorosis and MALDI-TOF analysis revealed over and underexpression of many translation-elongation associated proteins in mutants compared to wild type. Protein translation elongation factor G, translation elongation factor Tu and ribosomal subunit interface proteins were underexpressed and UTP-glucose-1-phosphate uridylyl transferase and cell division protein divIVA were overexpressed in AK-E12 as compared to wild type. In AK-F6, except 10 kDa chaperonin which was over-expressed other selected proteins were found to be suppressed. RT-PCR confirmed proteomic data except for the translation elongation factor G which showed contradictory data of proteome expression in AK-E12. Protein-protein interaction networks were constructed using STRING 10.0 which demonstrated strong connection of translation-elongation proteins with other proteins. Hence, it concludes from the data that translation elongation factors are important in transition of planktonic cells to biofilm cells in Enterococcus faecalis. PMID:27144316

  19. Controlled expression of enhanced green fluorescent protein and hepatitis B virus precore protein in mammalian cells

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    A novel tetracycline regulation expression system was used to regulate the expression of enhanced green fluorescent protein (EGFP) and hepatitis B virus precore protein in the mammalian cell lines with lipofectAMINE. Flow cytometry assays showed that application of the system resulted in about 18-fold induction of EGFP expression in CHO cell lines and 5-fold induction in SSMC-7721 cells and about 2-fold in the HEK293 cells. Furthermore, the effective use of this system for the controlled expression of HBV precore protein gene in hepatocellular carcinoma cells was tested.

  20. Gangliosides in cell recognition and membrane protein regulation

    OpenAIRE

    Lopez, Pablo H. H.; Schnaar, Ronald L.

    2009-01-01

    Gangliosides, sialic acid-bearing glycosphingolipids, are expressed on all vertebrate cells, and are the major glycans on nerve cells. They are anchored to the plasma membrane through their ceramide lipids with their varied glycans extending into the extracellular space. Through sugar-specific interactions with glycan binding proteins on apposing cells, gangliosides function as receptors in cell-cell recognition, regulating natural killer cell cytotoxicity via Siglec-7 binding, myelin-axon in...

  1. Comparison of cell wall proteins of Saccharomyces cerevisiae as anchors for cell surface expression of heterologous proteins.

    OpenAIRE

    van der Vaart, J. M.; te Biesebeke, R; Chapman, J.W.; Toschka, H Y; Klis, F M; Verrips, C. T.

    1997-01-01

    The carboxyl-terminal regions of five cell wall proteins (Cwp1p, Cwp2p, Ag alpha 1p, Tip1p, and Flo1p) and three potential cell wall proteins (Sed1p, YCR89w, and Tir1p) all proved capable of immobilizing alpha-galactosidase in the cell wall of Saccharomyces cerevisiae. The fraction of the total amount of fusion protein that was localized to the cell wall varied depending on the anchor domain used. The highest proportion of cell wall incorporation was achieved with Cwp2p, Ag alpha 1p, or Sed1p...

  2. Using Fluorescent Protein Fusions to Study Protein Subcellular Localization and Dynamics in Plant Cells.

    Science.gov (United States)

    Cui, Yong; Gao, Caiji; Zhao, Qiong; Jiang, Liwen

    2016-01-01

    Studies of protein subcellular localization and dynamics are helpful in understanding the cellular functions of proteins in an organism. In the past decade, the use of green fluorescent protein (GFP) as a fusion tag has dramatically extended our knowledge in this field. Transient expression and stable transformation of GFP-tagged proteins have been wildly used to study protein localization in vivo in different systems. Although GFP-based tags provide a fast and convenient way to characterize protein properties in living cells, several reports have demonstrated that GFP fusions might not accurately reflect the localization of the native protein as GFP tags may alter the protein properties. To facilitate proper usage of GFP tags in plant cell biology study, we describe detailed protocols to identify possible inhibitory effects of fluorescent tags on protein subcellular localization and to determine if a fluorescently tagged protein is localized to the correct subcellular compartment. Using Arabidopsis Endomembrane protein 12 (EMP12) as an example, we first show the possible inhibitory effect of GFP tags on proper protein localization and then describe the immunofluorescence labeling method to verify the correct localization of GFP fusion proteins. Next, a method is presented using the ImageJ program with the Pearson-Spearman correlation (PSC) colocalization plug-in for statistical quantification of colocalization ratios of two fluorophores. Finally we provide a detailed method for protein dynamics studies using spinning disk confocal microscopy in Arabidopsis cells. PMID:27515077

  3. Putative RopGAPs impact division plane selection and interact with kinesin-12 POK1.

    Science.gov (United States)

    Stöckle, Dorothee; Herrmann, Arvid; Lipka, Elisabeth; Lauster, Theresa; Gavidia, Richard; Zimmermann, Steffi; Müller, Sabine

    2016-01-01

    Cell shape is defined by the surrounding cell walls in plants. Thus, spatial control over cell division planes and cell expansion polarity are essential to maintain cell morphology. In eukaryotes, cell polarity and expansion are controlled by Rho GTPase signalling, regulating cytoskeletal reorganization and vesicle trafficking(1). However, until now, Rho signalling was not implicated in mitotic events in plants. Here, we report a pair of putative Rho GTPase activating proteins (RhoGAPs) that interact with the mitosis-specific kinesin-12 POK1, a core component of the cortical division zone/site (CDZ/CDS) that is required for division plane maintenance in Arabidopsis(2-4). The designated pleckstrin homology GAPs (PHGAPs) are cytoplasmic and plasma membrane associated in interphase, but during mitosis they additionally localize to the CDZ/CDS in a POK-dependent manner. In contrast to pok1 pok2 mutants, phgap1 phgap2 double mutants show moderate cell wall positioning defects as a consequence of inaccurate positioning of the cortical division zone marker POK1. We conclude that loss of PHGAP function interferes with division plane selection in proliferative cell divisions. PMID:27501519

  4. Cell division arrest by gamma-irradiation in photoautotrophic suspension culture of Euphorbia characias: maintenance of photosynthetic capacity and overaccumulation of sucrose

    International Nuclear Information System (INIS)

    Gamma-irradiation (250 Gy) applied to photoautotrophic cell suspensions of Euphorbia characias L. in the exponential growth phase led to the arrest of cell division and to a subsequent overaccumulation of sucrose and dry matter. From the fourth day of culture, the chlorophyll content and gross photosynthesis were not depressed by gamma-treatment nor by sugar accumulation. In both cultures, no difference was observed between oxygen uptake in the light at CO2 saturating concentration and in the dark, suggesting that no change in energy-dissipative reactions took place after irradiation. A slight increase in oxygen uptake in both light and dark was observed in irradiated cells during the first four days. However, in the absence of limiting factors, the photosynthetic capacities of the dividing and irradiated non-dividing photoautotrophic cells were identical but higher than that of the non-dividing cells in the stationary growth phase. This suggests that gamma-irradiation arrests cell division by a mechanism different to that occurring in stationary-phase cultures. This may be of value in investigating the metabolism of secondary products. (author)

  5. Multidimensional profiling of cell surface proteins and nuclear markers

    Energy Technology Data Exchange (ETDEWEB)

    Han, Ju; Chang, Hang; Andarawewa, Kumari; Yaswen, Paul; Helen Barcellos-Hoff, Mary; Parvin, Bahram

    2009-01-30

    Cell membrane proteins play an important role in tissue architecture and cell-cell communication. We hypothesize that segmentation and multidimensional characterization of the distribution of cell membrane proteins, on a cell-by-cell basis, enable improved classification of treatment groups and identify important characteristics that can otherwise be hidden. We have developed a series of computational steps to (i) delineate cell membrane protein signals and associate them with a specific nucleus; (ii) compute a coupled representation of the multiplexed DNA content with membrane proteins; (iii) rank computed features associated with such a multidimensional representation; (iv) visualize selected features for comparative evaluation through heatmaps; and (v) discriminate between treatment groups in an optimal fashion. The novelty of our method is in the segmentation of the membrane signal and the multidimensional representation of phenotypic signature on a cell-by-cell basis. To test the utility of this method, the proposed computational steps were applied to images of cells that have been irradiated with different radiation qualities in the presence and absence of other small molecules. These samples are labeled for their DNA content and E-cadherin membrane proteins. We demonstrate that multidimensional representations of cell-by-cell phenotypes improve predictive and visualization capabilities among different treatment groups, and identify hidden variables.

  6. AMP-activated protein kinase induces actin cytoskeleton reorganization in epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Miranda, Lisa; Carpentier, Sarah; Platek, Anna; Hussain, Nusrat; Gueuning, Marie-Agnes; Vertommen, Didier; Ozkan, Yurda; Sid, Brice; Hue, Louis; Courtoy, Pierre J.; Rider, Mark H. [de Duve Institute, Universite catholique de Louvain, Avenue Hippocrate, B-1200 Brussels (Belgium); Horman, Sandrine, E-mail: sandrine.horman@uclouvain.be [Institute of Experimental and Clinical Research - Pole of Cardiovascular Research, Universite catholique de Louvain, Avenue Hippocrate, B-1200 Brussels (Belgium)

    2010-06-04

    AMP-activated protein kinase (AMPK), a known regulator of cellular and systemic energy balance, is now recognized to control cell division, cell polarity and cell migration, all of which depend on the actin cytoskeleton. Here we report the effects of A769662, a pharmacological activator of AMPK, on cytoskeletal organization and signalling in epithelial Madin-Darby canine kidney (MDCK) cells. We show that AMPK activation induced shortening or radiation of stress fibers, uncoupling from paxillin and predominance of cortical F-actin. In parallel, Rho-kinase downstream targets, namely myosin regulatory light chain and cofilin, were phosphorylated. These effects resembled the morphological changes in MDCK cells exposed to hyperosmotic shock, which led to Ca{sup 2+}-dependent AMPK activation via calmodulin-dependent protein kinase kinase-{beta}(CaMKK{beta}), a known upstream kinase of AMPK. Indeed, hypertonicity-induced AMPK activation was markedly reduced by the STO-609 CaMKK{beta} inhibitor, as was the increase in MLC and cofilin phosphorylation. We suggest that AMPK links osmotic stress to the reorganization of the actin cytoskeleton.

  7. Cyanamide mode of action during inhibition of onion (Allium cepa L.) root growth involves disturbances in cell division and cytoskeleton formation.

    Science.gov (United States)

    Soltys, Dorota; Rudzińska-Langwald, Anna; Kurek, Wojciech; Gniazdowska, Agnieszka; Sliwinska, Elwira; Bogatek, Renata

    2011-09-01

    Cyanamide is an allelochemical produced by hairy vetch (Vicia villosa Roth.). Its phyotoxic effect on plant growth was examined on roots of onion (Allium cepa L.) bulbs. Water solution of cyanamide (2-10 mM) restricted growth of onion roots in a dose-dependent manner. Treatment of onion roots with cyanamide resulted in a decrease in root growth rate accompanied by a decrease in accumulation of fresh and dry weight. The inhibitory effect of cyanamide was reversed by its removal from the environment, but full recovery was observed only for tissue treated with this chemical at low concentration (2-6 mM). Cytological observations of root tip cells suggest that disturbances in cell division may explain the strong cyanamide allelopathic activity. Moreover, in cyanamide-treated onion the following changes were detected: reduction of mitotic cells, inhibition of proliferation of meristematic cells and cell cycle, and modifications of cytoskeleton arrangement. PMID:21573814

  8. Porcine circovirus-2 capsid protein induces cell death in PK15 cells

    International Nuclear Information System (INIS)

    Studies have shown that Porcine circovirus (PCV)-2 induces apoptosis in PK15 cells. Here we report that cell death is induced in PCV2b-infected PK15 cells that express Capsid (Cap) protein and this effect is enhanced in interferon gamma (IFN-γ)-treated cells. We further show that transient PCV2a and 2b-Cap protein expression induces cell death in PK15 cells at rate similar to PCV2 infection, regardless of Cap protein localization. These data suggest that Cap protein may have the capacity to trigger different signaling pathways involved in cell death. Although further investigation is needed to gain deeper insights into the nature of the pathways involved in Cap-induced cell death, this study provides evidence that PCV2-induced cell death in kidney epithelial PK15 cells can be mapped to the Cap protein and establishes the need for future research regarding the role of Cap-induced cell death in PCV2 pathogenesis. - Highlights: • IFN-γ enhances PCV2 replication that leads to cell death in PK15 cells. • IFN-γ enhances nuclear localization of the PCV2 Capsid protein. • Transient PCV2a and 2b-Capsid protein expression induces cell death. • Cell death is not dictated by specific Capsid protein sub-localization

  9. Porcine circovirus-2 capsid protein induces cell death in PK15 cells

    Energy Technology Data Exchange (ETDEWEB)

    Walia, Rupali; Dardari, Rkia, E-mail: rdardari@ucalgary.ca; Chaiyakul, Mark; Czub, Markus

    2014-11-15

    Studies have shown that Porcine circovirus (PCV)-2 induces apoptosis in PK15 cells. Here we report that cell death is induced in PCV2b-infected PK15 cells that express Capsid (Cap) protein and this effect is enhanced in interferon gamma (IFN-γ)-treated cells. We further show that transient PCV2a and 2b-Cap protein expression induces cell death in PK15 cells at rate similar to PCV2 infection, regardless of Cap protein localization. These data suggest that Cap protein may have the capacity to trigger different signaling pathways involved in cell death. Although further investigation is needed to gain deeper insights into the nature of the pathways involved in Cap-induced cell death, this study provides evidence that PCV2-induced cell death in kidney epithelial PK15 cells can be mapped to the Cap protein and establishes the need for future research regarding the role of Cap-induced cell death in PCV2 pathogenesis. - Highlights: • IFN-γ enhances PCV2 replication that leads to cell death in PK15 cells. • IFN-γ enhances nuclear localization of the PCV2 Capsid protein. • Transient PCV2a and 2b-Capsid protein expression induces cell death. • Cell death is not dictated by specific Capsid protein sub-localization.

  10. A family of snail-related zinc finger proteins regulates two distinct and parallel mechanisms that mediate Drosophila neuroblast asymmetric divisions.

    Science.gov (United States)

    Cai, Y; Chia, W; Yang, X

    2001-04-01

    Three snail family genes snail, escargot and worniu, encode related zinc finger transcription factors that mediate Drosophila central nervous system (CNS) development. We show that simultaneous removal of all three genes causes defective neuroblast asymmetric divisions; inscuteable transcription/translation is delayed/suppressed in the segmented CNS. Further more, defects in localization of cell fate determinants and orientation of the mitotic spindle in dividing neuroblasts are much stronger than those associated with inscuteable loss of function. In inscuteable neuroblasts, cell fate determinants are mislocalized during prophase and metaphase, yet during anaphase and telophase the great majority of mutant neuroblasts localize these determinants as cortical crescents overlying one of the spindle poles. This phenomenon, known as 'telophase rescue', does not occur in the absence of the snail family genes; moreover, in contrast to inscuteable mutants, mitotic spindle orientation is completely randomized. Our data provide further evidence for the existence of two distinct asymmetry-controlling mechanisms in neuroblasts both of which require snail family gene function: an inscuteable-dependent mechanism that functions throughout mitosis and an inscuteable-independent mechanism that acts during anaphase/telophase. PMID:11285234

  11. PERSONNEL DIVISION BECOMES HUMAN RESOURCES DIVISION

    CERN Multimedia

    Division des ressources humaines

    2000-01-01

    In the years to come, CERN faces big challenges in the planning and use of human resources. At this moment, Personnel (PE) Division is being reorganised to prepare for new tasks and priorities. In order to accentuate the purposes of the operation, the name of the division has been changed into Human Resources (HR) Division, with effect from 1st January 2000. Human Resources DivisionTel.73222

  12. Chimera proteins with affinity for membranes and microtubule tips polarize in the membrane of fission yeast cells.

    Science.gov (United States)

    Recouvreux, Pierre; Sokolowski, Thomas R; Grammoustianou, Aristea; Ten Wolde, Pieter Rein; Dogterom, Marileen

    2016-02-16

    Cell polarity refers to a functional spatial organization of proteins that is crucial for the control of essential cellular processes such as growth and division. To establish polarity, cells rely on elaborate regulation networks that control the distribution of proteins at the cell membrane. In fission yeast cells, a microtubule-dependent network has been identified that polarizes the distribution of signaling proteins that restricts growth to cell ends and targets the cytokinetic machinery to the middle of the cell. Although many molecular components have been shown to play a role in this network, it remains unknown which molecular functionalities are minimally required to establish a polarized protein distribution in this system. Here we show that a membrane-binding protein fragment, which distributes homogeneously in wild-type fission yeast cells, can be made to concentrate at cell ends by attaching it to a cytoplasmic microtubule end-binding protein. This concentration results in a polarized pattern of chimera proteins with a spatial extension that is very reminiscent of natural polarity patterns in fission yeast. However, chimera levels fluctuate in response to microtubule dynamics, and disruption of microtubules leads to disappearance of the pattern. Numerical simulations confirm that the combined functionality of membrane anchoring and microtubule tip affinity is in principle sufficient to create polarized patterns. Our chimera protein may thus represent a simple molecular functionality that is able to polarize the membrane, onto which additional layers of molecular complexity may be built to provide the temporal robustness that is typical of natural polarity patterns. PMID:26831106

  13. Protein expression analyses at the single cell level.

    Science.gov (United States)

    Ohno, Masae; Karagiannis, Peter; Taniguchi, Yuichi

    2014-01-01

    The central dogma of molecular biology explains how genetic information is converted into its end product, proteins, which are responsible for the phenotypic state of the cell. Along with the protein type, the phenotypic state depends on the protein copy number. Therefore, quantification of the protein expression in a single cell is critical for quantitative characterization of the phenotypic states. Protein expression is typically a dynamic and stochastic phenomenon that cannot be well described by standard experimental methods. As an alternative, fluorescence imaging is being explored for the study of protein expression, because of its high sensitivity and high throughput. Here we review key recent progresses in fluorescence imaging-based methods and discuss their application to proteome analysis at the single cell level. PMID:25197931

  14. Protein Expression Analyses at the Single Cell Level

    Directory of Open Access Journals (Sweden)

    Masae Ohno

    2014-09-01

    Full Text Available The central dogma of molecular biology explains how genetic information is converted into its end product, proteins, which are responsible for the phenotypic state of the cell. Along with the protein type, the phenotypic state depends on the protein copy number. Therefore, quantification of the protein expression in a single cell is critical for quantitative characterization of the phenotypic states. Protein expression is typically a dynamic and stochastic phenomenon that cannot be well described by standard experimental methods. As an alternative, fluorescence imaging is being explored for the study of protein expression, because of its high sensitivity and high throughput. Here we review key recent progresses in fluorescence imaging-based methods and discuss their application to proteome analysis at the single cell level.

  15. Bone Morphogenetic Protein 4 Mediates Human Embryonic Germ Cell Derivation

    OpenAIRE

    Hiller, Marc; Liu, Cyndi; Blumenthal, Paul D; John D Gearhart; Kerr, Candace L.

    2010-01-01

    Human primordial germ cells (PGCs) have proven to be a source of pluripotent stem cells called embryonic germ cells (EGCs). Unlike embryonic stem cells, virtually little is known regarding the factors that regulate EGC survival and maintenance. In mice, the growth factor bone morphogenetic protein 4 (BMP4) has been shown to be required for maintaining mouse embryonic stem cells, and disruptions in this gene lead to defects in mouse PGC specification. Here, we sought to determine whether recom...

  16. Relationships between cell cycle regulator gene copy numbers and protein expression levels in Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Ayako Chino

    Full Text Available We previously determined the copy number limits of overexpression for cell division cycle (cdc regulatory genes in the fission yeast Schizosaccharomyces pombe using the "genetic tug-of-war" (gTOW method. In this study, we measured the levels of tandem affinity purification (TAP-tagged target proteins when their copy numbers are increased in gTOW. Twenty analyzed genes showed roughly linear correlations between increased protein levels and gene copy numbers, which suggested a general lack of compensation for gene dosage in S. pombe. Cdc16 and Sid2 protein levels but not their mRNA levels were much lower than that expected by their copy numbers, which suggested the existence of a post-transcriptional down regulation of these genes. The cyclin Cig1 protein level and its mRNA level were much higher than that expected by its copy numbers, which suggested a positive feedback mechanism for its expression. A higher Cdc10 protein level and its mRNA level, probably due to cloning its gene into a plasmid, indicated that Cdc10 regulation was more robust than that previously predicted.

  17. Relationships between cell cycle regulator gene copy numbers and protein expression levels in Schizosaccharomyces pombe.

    Science.gov (United States)

    Chino, Ayako; Makanae, Koji; Moriya, Hisao

    2013-01-01

    We previously determined the copy number limits of overexpression for cell division cycle (cdc) regulatory genes in the fission yeast Schizosaccharomyces pombe using the "genetic tug-of-war" (gTOW) method. In this study, we measured the levels of tandem affinity purification (TAP)-tagged target proteins when their copy numbers are increased in gTOW. Twenty analyzed genes showed roughly linear correlations between increased protein levels and gene copy numbers, which suggested a general lack of compensation for gene dosage in S. pombe. Cdc16 and Sid2 protein levels but not their mRNA levels were much lower than that expected by their copy numbers, which suggested the existence of a post-transcriptional down regulation of these genes. The cyclin Cig1 protein level and its mRNA level were much higher than that expected by its copy numbers, which suggested a positive feedback mechanism for its expression. A higher Cdc10 protein level and its mRNA level, probably due to cloning its gene into a plasmid, indicated that Cdc10 regulation was more robust than that previously predicted. PMID:24019917

  18. Cell-specific monitoring of protein synthesis in vivo.

    Directory of Open Access Journals (Sweden)

    Nikos Kourtis

    Full Text Available Analysis of general and specific protein synthesis provides important information, relevant to cellular physiology and function. However, existing methodologies, involving metabolic labelling by incorporation of radioactive amino acids into nascent polypeptides, cannot be applied to monitor protein synthesis in specific cells or tissues, in live specimens. We have developed a novel approach for monitoring protein synthesis in specific cells or tissues, in vivo. Fluorescent reporter proteins such as GFP are expressed in specific cells and tissues of interest or throughout animals using appropriate promoters. Protein synthesis rates are assessed by following fluorescence recovery after partial photobleaching of the fluorophore at targeted sites. We evaluate the method by examining protein synthesis rates in diverse cell types of live, wild type or mRNA translation-defective Caenorhabditis elegans animals. Because it is non-invasive, our approach allows monitoring of protein synthesis in single cells or tissues with intrinsically different protein synthesis rates. Furthermore, it can be readily implemented in other organisms or cell culture systems.

  19. Effects of sense and antisense centromere/kinetochore complex protein-B (CENP-B) in cell cycle regulation

    Institute of Scientific and Technical Information of China (English)

    LUO Song; LIN Haiyan; QI Jianguo; WANG Yongchao

    2005-01-01

    This paper investigates the effects of sense and antisense centromere/kinetochore complex protein-B (CENP-B) in cell cycle regulation. Full-length cenpb cDNA was subcloned into pBI-EGFP eukaryotic expression vector in both sense and antisense orientation. HeLa-Tet-Off cells were transfected with sense or antisense cenpb vectors. Sense transfection of HeLa-Tet-Off cells resulted in the formation of a large centromere/kinetochore complex, and apoptosis of cells following several times of cell division. A stable antisense cenpb transfected cell line, named HACPB, was obtained. The centromere/kinetochore complex of HACPB cells became smaller than control HeLa-Tet-Off cells and scattered, and the expression of CENP-B was down-regulated. In addition, delayed cell cycle progression, inhibited malignant phenotype, restrained ability of tumor formation in nude mice, and delayed entry from G2/M phase into next G1 phase were observed in HACPB cells. Furthermore, the expression of cyclin-dependent kinases (CDKs), cyclins, and CDK inhibitors (CKIs) were modulated during different phases of the cell cycle. CENP-B is an essential protein for the maintenance of the structure and function of centromere/kinetochore complex, and plays important roles in cell cycle regulation.

  20. 14-3-3 proteins in guard cell signaling

    Directory of Open Access Journals (Sweden)

    Valérie eCotelle

    2016-01-01

    Full Text Available Guard cells are specialized cells located at the leaf surface delimiting pores which control gas exchanges between the plant and the atmosphere. To optimize the CO2 uptake necessary for photosynthesis while minimizing water loss, guard cells integrate environmental signals to adjust stomatal aperture. The size of the stomatal pore is regulated by movements of the guard cells driven by variations in their volume and turgor. As guard cells perceive and transduce a wide array of environmental cues, they provide an ideal system to elucidate early events of plant signaling. Reversible protein phosphorylation events are known to play a crucial role in the regulation of stomatal movements. However, in some cases, phosphorylation alone is not sufficient to achieve complete protein regulation, but is necessary to mediate the binding of interactors that modulate protein function. Among the phosphopeptide-binding proteins, the 14-3-3 proteins are the best characterized in plants. The 14-3-3s are found as multiple isoforms in eukaryotes and have been shown to be involved in the regulation of stomatal movements. In this review, we describe the current knowledge about 14-3-3 roles in the regulation of their binding partners in guard cells: receptors, ion pumps, channels, protein kinases and some of their substrates. Regulation of these targets by 14-3-3 proteins is discussed and related to their function in guard cells during stomatal movements in response to abiotic or biotic stresses.

  1. 14-3-3 Proteins in Guard Cell Signaling

    Science.gov (United States)

    Cotelle, Valérie; Leonhardt, Nathalie

    2016-01-01

    Guard cells are specialized cells located at the leaf surface delimiting pores which control gas exchanges between the plant and the atmosphere. To optimize the CO2 uptake necessary for photosynthesis while minimizing water loss, guard cells integrate environmental signals to adjust stomatal aperture. The size of the stomatal pore is regulated by movements of the guard cells driven by variations in their volume and turgor. As guard cells perceive and transduce a wide array of environmental cues, they provide an ideal system to elucidate early events of plant signaling. Reversible protein phosphorylation events are known to play a crucial role in the regulation of stomatal movements. However, in some cases, phosphorylation alone is not sufficient to achieve complete protein regulation, but is necessary to mediate the binding of interactors that modulate protein function. Among the phosphopeptide-binding proteins, the 14-3-3 proteins are the best characterized in plants. The 14-3-3s are found as multiple isoforms in eukaryotes and have been shown to be involved in the regulation of stomatal movements. In this review, we describe the current knowledge about 14-3-3 roles in the regulation of their binding partners in guard cells: receptors, ion pumps, channels, protein kinases, and some of their substrates. Regulation of these targets by 14-3-3 proteins is discussed and related to their function in guard cells during stomatal movements in response to abiotic or biotic stresses. PMID:26858725

  2. Role of heat shock proteins in cell apoptosis

    Directory of Open Access Journals (Sweden)

    Arleta Kaźmierczuk

    2010-06-01

    Full Text Available Apoptosis is, apart from necrosis and autophagy, one of the possible cell death mechanisms eliminating needless, not normal or infected cells. This process ensures quantitative and qualitative cell control of organisms. Apoptosis is tightly regulated, it requires both activation of a large number of genes and energy input. Up-to-date two main apoptotic pathways have been recognized – external/receptor and internal, processed with the participation of mitochondria. Heat shock proteins HSPs, the molecules known from their chaperone activity and molecular conservatism, play essential functions in the course of apoptosis. Among that proteins family, i.e. HSP100, 90, 70, 60, 40 and small molecular (sHSP, there are agents mainly protective against programmed cell death. However, in some conditions some of these proteins may promote apoptosis. This review describes different key apoptotic proteins interacting with main members of HSP family and the consequence of these events for cell survival or apoptosis.

  3. Effects on cell growth processes (mitosis, synthesis of nucleic acids and of proteins). Chapter 7

    International Nuclear Information System (INIS)

    A review is presented of reports of the interference of -SH radioprotective agents with cell division and with the processes of nucleic acid and protein synthesis which are a prerequisite for mitosis. Mitotic activity is inhibited to the same extent in mammalian tissues as in cultures of animal and plant cells and bacteria. With cultured cells, the toxicity and the antimitotic activity have been found to be at their highest level for intermediate concentrations of the compound and to decrease for higher and lower concentrations. Inhibition of the synthesis of nucleic acids by -SH radioprotective substances has been observed with cultures of cells and bacteria and in mammalian tissues. In vitro interactions with the structures of free DNA and nucleoprotein have also been studied. The extent to which such complexes between the protective agent and DNA or nucleoprotein occur in vivo is not known. A depression of protein synthesis has been observed, and participates in the more general inhibition of growth processes. Possible mechanisms of these effects are discussed. (U.K.)

  4. Green Fluorescent Protein-Tagged Retroviral Envelope Protein for Analysis of Virus-Cell Interactions

    Science.gov (United States)

    Spitzer, Dirk; Dittmar, Kurt E. J.; Rohde, Manfred; Hauser, Hansjörg; Wirth, Dagmar

    2003-01-01

    Fluorescent retroviral envelope (Env) proteins were developed for direct visualization of viral particles. By fusing the enhanced green fluorescent protein (eGFP) to the N terminus of the amphotropic 4070A envelope protein, extracellular presentation of eGFP was achieved. Viruses incorporated the modified Env protein and efficiently infected cells. We used the GFP-tagged viruses for staining retrovirus receptor-positive cells, thereby circumventing indirect labeling techniques. By generating cells which conditionally expressed the GFP-tagged Env protein, we could confirm an inverse correlation between retroviral Env expression and infectivity (superinfection). eGFP-tagged virus particles are suitable for monitoring the dynamics of virus-cell interactions. PMID:12719600

  5. Interaction of Protein and Cell with Different Chitosan Membranes

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Interaction between proteins, cells and biomaterial surfaces is commonly observed and often used to measure biocompatibility of biomaterials.In this investigation, three kinds of biomaterials derived from chitosan were prepared.The surface wettability of these polymers, interaction of protein with material surface, and their effects on cell adhesion and growth were studied.The results show that the surface contact angle and surface charge of biomaterials have a close bearing on protein adsorption as well as cell adhesion and growth, indicating that through different chemical modifications, chitosan can be made into different kinds of biomedical materials to satisfy various needs.

  6. Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer

    Science.gov (United States)

    Estruch, Sara B.; Fisher, Simon E.

    2014-01-01

    Assays based on Bioluminescence Resonance Energy Transfer (BRET) provide a sensitive and reliable means to monitor protein-protein interactions in live cells. BRET is the non-radiative transfer of energy from a 'donor' luciferase enzyme to an 'acceptor' fluorescent protein. In the most common configuration of this assay, the donor is Renilla reniformis luciferase and the acceptor is Yellow Fluorescent Protein (YFP). Because the efficiency of energy transfer is strongly distance-dependent, observation of the BRET phenomenon requires that the donor and acceptor be in close proximity. To test for an interaction between two proteins of interest in cultured mammalian cells, one protein is expressed as a fusion with luciferase and the second as a fusion with YFP. An interaction between the two proteins of interest may bring the donor and acceptor sufficiently close for energy transfer to occur. Compared to other techniques for investigating protein-protein interactions, the BRET assay is sensitive, requires little hands-on time and few reagents, and is able to detect interactions which are weak, transient, or dependent on the biochemical environment found within a live cell. It is therefore an ideal approach for confirming putative interactions suggested by yeast two-hybrid or mass spectrometry proteomics studies, and in addition it is well-suited for mapping interacting regions, assessing the effect of post-translational modifications on protein-protein interactions, and evaluating the impact of mutations identified in patient DNA. PMID:24893771

  7. Use of abnormal preprophase bands to decipher division plane determination

    Science.gov (United States)

    Granger, C.; Cyr, R.

    2001-01-01

    Many premitotic plant cells possess a cortical preprophase band of microtubules and actin filaments that encircles the nucleus. In vacuolated cells, the preprophase band is visibly connected to the nucleus by a cytoplasmic raft of actin filaments and microtubules termed the phragmosome. Typically, the location of the preprophase band and phragmosome corresponds to, and thus is thought to influence, the location of the cell division plane. To better understand the function of the preprophase band and phragmosome in orienting division, we used a green fluorescent protein-based microtubule reporter protein to observe mitosis in living tobacco bright yellow 2 cells possessing unusual preprophase bands. Observations of mitosis in these unusual cells support the involvement of the preprophase band/phragmosome in properly positioning the preprophase nucleus, influencing spindle orientation such that the cytokinetic phragmoplast initially grows in an appropriate direction, and delineating a region in the cell cortex that attracts microtubules and directs later stages of phragmoplast growth. Thus, the preprophase band/phragmosome appears to perform several interrelated functions to orient the division plane. However, functional information associated with the preprophase band is not always used or needed and there appears to be an age or distance-dependent character to the information. Cells treated with the anti-actin drug, latrunculin B, are still able to position the preprophase nucleus suggesting that microtubules may play a dominant role in premitotic positioning. Furthermore, in treated cells, spindle location and phragmoplast insertion are frequently abnormal suggesting that actin plays a significant role in nuclear anchoring and phragmoplast guidance. Thus, the microtubule and actin components of the preprophase band/phragmosome execute complementary activities to ensure proper orientation of the division plane.

  8. Monitoring G protein activation in cells with BRET

    Science.gov (United States)

    Masuho, Ikuo; Martemyanov, Kirill A.; Lambert, Nevin A.

    2016-01-01

    Summary Live-cell assays based on fluorescence and luminescence are now indispensable tools for the study of G protein signaling. Assays based on fluorescence and bioluminescence resonance energy transfer (FRET and BRET) have been particularly valuable for monitoring changes in second messengers, protein-protein interactions, and protein conformation. Here we describe a BRET assay that monitors the release of free Gβγ dimers after activation of heterotrimers containing Gα subunits from all four G protein subfamilies. This assay provides useful kinetic and pharmacological information with reasonably high throughput using standard laboratory equipment. PMID:26260597

  9. Cell-to-cell propagation of infectious cytosolic protein aggregates

    OpenAIRE

    Hofmann, Julia P.; Denner, Philip; Nussbaum-Krammer, Carmen; Kuhn, Peer-Hendrik; Suhre, Michael H.; Scheibel, Thomas; Lichtenthaler, Stefan F; Schätzl, Hermann M; Bano, Daniele; Vorberg, Ina M.

    2013-01-01

    Prions are self-templating protein conformers that replicate by recruitment and conversion of homotypic proteins into growing protein aggregates. Originally identified as causative agents of transmissible spongiform encephalopathies, increasing evidence now suggests that prion-like phenomena are more common in nature than previously anticipated. In contrast to fungal prions that replicate in the cytoplasm, propagation of mammalian prions derived from the precursor protein PrP is confined to t...

  10. Role of FtsEX in Cell Division of Escherichia coli: Viability of ftsEX Mutants Is Dependent on Functional SufI or High Osmotic Strength▿

    OpenAIRE

    Reddy, Manjula

    2006-01-01

    In Escherichia coli, at least 12 proteins, FtsZ, ZipA, FtsA, FtsE/X, FtsK, FtsQ, FtsL, FtsB, FtsW, FtsI, FtsN, and AmiC, are known to localize to the septal ring in an interdependent and sequential pathway to coordinate the septum formation at the midcell. The FtsEX complex is the latest recruit of this pathway, and unlike other division proteins, it is shown to be essential only on low-salt media. In this study, it is shown that ftsEX null mutations are not only salt remedial but also osmore...

  11. Multiscale molecular simulations of proteins in cell-like conditions

    Science.gov (United States)

    Samiotakis, Antonios

    Proteins are the workhorses of all living organisms, performing a broad range of functions in the crowded cellular interior. However, little is known about how proteins function in cell-like conditions since most studies focus in dilute aqueous environments. In order to address this problem we incorporated molecular simulations and coarse-grained models that capture the protein dynamics in the cellular interior. We study the macromolecular crowding effects of cell-like environments on protein Borrelia Burgdorferi VlsE (variable major protein-like sequence-expressed), an aspherical membrane protein, and the enzyme Phosphoglycerate kinase. We show that protein conformation can be significantly perturbed under crowded cell-like conditions which, in turn, can have dramatic effects to the proteins' function. In addition, we look into the effects of mutations in the folding pathways of the topologically frustrated protein apoflavodoxin while correlation with experiments is also achieved. We further developed a multiscale simulation scheme that combines the sampling efficiency of low-resolution models with the detail of all-atomistic simulations. An algorithm that reconstructs all-atomistic conformations from coarse-grained representations was developed, in addition to an energy function that accounts for chemical interference based on the Boltzamn inversion method. The multiscale simulation scheme manages to sample all-atomistic structures of the protein Trp-cage that match very well with experiments. The folding kinetic behavior of Trp-cage was also studied in the combined presence of urea denaturant and macromolecular crowding.

  12. Protein (Viridiplantae): 308807062 [

    Lifescience Database Archive (English)

    Full Text Available XP_003080842.1 33090:2448 3041:440 1035538:347 13792:347 70447:323 70448:86 FTSH1_SYNY3 Cell ... div ... ision protein ftsH homolog 1 dbj|BAA10230.1| cell ... division prot (ISS) Ostreococcus tauri MRAHFRASVRA ...

  13. Activated protein C modulates the proinflammatory activity of dendritic cells

    Directory of Open Access Journals (Sweden)

    Matsumoto T

    2015-05-01

    Full Text Available Takahiro Matsumoto,1,2* Yuki Matsushima,1* Masaaki Toda,1 Ziaurahman Roeen,1 Corina N D'Alessandro-Gabazza,1,5 Josephine A Hinneh,1 Etsuko Harada,1,3 Taro Yasuma,4 Yutaka Yano,4 Masahito Urawa,1,5 Tetsu Kobayashi,5 Osamu Taguchi,5 Esteban C Gabazza1 1Department of Immunology, Mie University Graduate School of Medicine, Tsu, Mie Prefecture, 2BONAC Corporation, BIO Factory 4F, Fukuoka, 3Iwade Research Institute of Mycology, 4Department of Endocrinology, Diabetes and Metabolism, 5Department of Pulmonary and Critical Care Medicine, Mie University Graduate School of Medicine, Tsu, Mie Prefecture, Japan *These authors contributed equally to this work Background: Previous studies have demonstrated the beneficial activity of activated protein C in allergic diseases including bronchial asthma and rhinitis. However, the exact mechanism of action of activated protein C in allergies is unclear. In this study, we hypothesized that pharmacological doses of activated protein C can modulate allergic inflammation by inhibiting dendritic cells. Materials and methods: Dendritic cells were prepared using murine bone marrow progenitor cells and human peripheral monocytes. Bronchial asthma was induced in mice that received intratracheal instillation of ovalbumin-pulsed dendritic cells. Results: Activated protein C significantly increased the differentiation of tolerogenic plasmacytoid dendritic cells and the secretion of type I interferons, but it significantly reduced lipopolysaccharide-mediated maturation and the secretion of inflammatory cytokines in myeloid dendritic cells. Activated protein C also inhibited maturation and the secretion of inflammatory cytokines in monocyte-derived dendritic cells. Activated protein C-treated dendritic cells were less effective when differentiating naïve CD4 T-cells from Th1 or Th2 cells, and the cellular effect of activated protein C was mediated by its receptors. Mice that received adoptive transfer of activated protein C

  14. A Cell Lysis and Protein Purification - Single Molecule Assay Devices for Evaluation of Genetically Engineered Proteins

    Science.gov (United States)

    Nakyama, Tetsuya; Tabata, Kazuhito; Noji, Hiroyuki; Yokokawa, Ryuji

    We have developed two devices applicable to evaluate genetically engineered proteins in single molecule assay: on-chip cell lysis device, and protein purification - assay device. A motor protein, F1-ATPase expressed in E.coli, was focused in this report as a target protein. Cell lysis was simply performed by applying pulse voltage between Au electrodes patterned by photolithography, and its efficiency was determined by absorptiometry. The subsequent processes, purification and assay of extracted proteins, were demonstrated in order to detect F1-ATPase and to evaluate its activity. The specific bonding between his-tag in F1-ATPase and Ni-NTA coated on a glass surface was utilized for the purification process. After immobilization of F1-ATPase, avidin-coated microspheres and adenosine tri-phosphate (ATP) solution were infused sequentially to assay the protein. Microsphere rotation was realized by activity of F1-ATPase corresponding to ATP hydrolysis. Results show that the cell lysis device, at the optimum condition, extracts enough amount of protein for single molecule assay. Once cell lysate was injected to the purification - assay device, proteins were diffused in the lateral direction in a Y-shape microchannel. The gradient of protein concentratioin provides an optimal concentration for the assay i.e. the highest density of rotating beads. Density of rotating beads is also affected by the initial concentration of protein injected to the device. The optimum concentration was achieved by our cell lysis device not by the conventional method by ultrasonic wave. Rotation speed was analyzed for several microspheres assayed in the purification - assay device, and the results were compatible to that of conventional assay in which F1-ATPase was purified in bulk scale. In conclusion, we have demonstrated on-chip cell lysis and assay appropriate for the sequential analysis without any pretreatment. On-chip devices replacing conventional bioanalytical methods will be

  15. PBP1a-deficiency causes major defects in cell division, growth and biofilm formation by Streptococcus mutans.

    Directory of Open Access Journals (Sweden)

    Zezhang T Wen

    Full Text Available Streptococcus mutans, a key etiological agent of human dental caries, lives almost exclusively on the tooth surface in plaque biofilms and is known for its ability to survive and respond to various environmental insults, including low pH, and antimicrobial agents from other microbes and oral care products. In this study, a penicillin-binding protein (PBP1a-deficient mutant, strain JB467, was generated by allelic replacement mutagenesis and analyzed for the effects of such a deficiency on S. mutans' stress tolerance response and biofilm formation. Our results so far have shown that PBP1a-deficiency in S. mutans affects growth of the deficient mutant, especially at acidic and alkaline pHs. As compared to the wild-type, UA159, the PBP1a-deficient mutant, JB467, had a reduced growth rate at pH 6.2 and did not grow at all at pH 8.2. Unlike the wild-type, the inclusion of paraquat in growth medium, especially at 2 mM or above, significantly reduced the growth rate of the mutant. Acid killing assays showed that the mutant was 15-fold more sensitive to pH 2.8 than the wild-type after 30 minutes. In a hydrogen peroxide killing assay, the mutant was 16-fold more susceptible to hydrogen peroxide (0.2%, w/v after 90 minutes than the wild-type. Relative to the wild-type, the mutant also had an aberrant autolysis rate, indicative of compromises in cell envelope integrity. As analyzed using on 96-well plate model and spectrophotometry, biofilm formation by the mutant was decreased significantly, as compared to the wild-type. Consistently, Field Emission-SEM analysis also showed that the PBP1a-deficient mutant had limited capacity to form biofilms. TEM analysis showed that PBP1a mutant existed primarily in long rod-like cells and cells with multiple septa, as compared to the coccal wild-type. The results presented here highlight the importance of pbp1a in cell morphology, stress tolerance, and biofilm formation in S. mutans.

  16. Molecular evolution in bacteria: cell division Evolução molecular em bactérias: divisão celular

    OpenAIRE

    Trevors, J. T.

    1998-01-01

    Molecular evolution in bacteria is examined with an emphasis on the self-assembly of cells capable of primitive division and growth during early molecular evolution. Also, the possibility that some type of encapsulation structure preceeded biochemical pathways and the assembly of genetic material is examined. These aspects will be considered from an evolutionary perspective.A evolução molecular em bactérias é examinada com ênfase na auto-organização de uma célula capaz de divisão primitiva e ...

  17. The fate of Müller’s glia following experimental retinal detachment: nuclear migration, cell division, and subretinal glial scar formation

    OpenAIRE

    Lewis, Geoffrey P.; Chapin, Ethan A.; Luna, Gabriel; Linberg, Kenneth A.; Fisher, Steven K.

    2010-01-01

    Purpose To study the fate of Müller’s glia following experimental retinal detachment, using a “pulse/chase” paradigm of bromodeoxyuridine (BrdU) labeling for the purpose of understanding the role of Müller cell division in subretinal scar formation. Methods Experimental retinal detachments were created in pigmented rabbit eyes, and 3 days later 10 µg of BrdU was injected intravitreally. The retinas were harvested 4 h after the BrdU was administered (i.e., day 3) or on days 4, 7, and 21 post d...

  18. Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is due to altered cell division, phytohormone balance and expansin gene expression

    OpenAIRE

    Soltys, Dorota; Rudzińska-Langwald, Anna; Gniazdowska, Agnieszka; Wiśniewska, Anita; Bogatek, Renata

    2012-01-01

    Cyanamide (CA) has been reported as a natural compound produced by hairy vetch (Vicia villosa Roth.) and it was shown also to be an allelochemical, responsible for strong allelopathic potential in this species. CA phytotoxicity has been demonstrated on various plant species, but to date little is known about its mode of action at cellular level. Treatment of tomato (Solanum lycopersicum L.) roots with CA (1.2 mM) resulted in inhibition of growth accompanied by alterations in cell division, an...

  19. Thermal protein denaturation and protein aggregation in cells made thermotolerant by various chemicals: role of heat shock proteins.

    Science.gov (United States)

    Kampinga, H H; Brunsting, J F; Stege, G J; Burgman, P W; Konings, A W

    1995-08-01

    Thermotolerance (TT) induced by sodium arsenite (A-TT: 100 microM, 1 h, 37 degrees C) was compared to heat-induced thermotolerance (H-TT: 15 min, 44 degrees C) using HeLa S3 cells. All four pretreatments led to comparable levels of thermotolerance and also induced resistance to arsenite-, ethanol-, and diamide-induced toxicity (clonogenic ability). Stress-induced expression of the major heat shock proteins (hsp27, hsc70(p73), hsp70(p72), and hsp90) was generally highest in H-TT cells and lowest in A-TT cells. Interestingly, the four types of TT cells showed distinct differences in certain aspects of resistance against thermal protein damage. Thermal protein denaturation and aggregation determined in isolated cellular membrane fractions was found to be attenuated when they were isolated from H-TT and A-TT cells but not when isolated from E-TT and D-TT cells. The heat resistance in the proteins of the membrane fraction corresponded with elevated levels of hsp70(p72) associated with the isolated membrane fractions. In the nuclear fraction, only marginal (not significant) attenuation of the formation of protein aggregates (as determined by TX-100 (in)solubility) was observed. However, the postheat recovery from heat-induced protein aggregation in the nucleus was faster in H-TT, E-TT, and D-TT cells, but not in A-TT cells. Despite the fact that elevated levels of hsp27, hsp70(p73), and hsp70(p72) were found in the TX-100 insoluble nuclear fraction derived from all TT cells, no correlation was found with the degree of resistance in terms of the accelerated recovery from nuclear protein aggregation. The only correlation between accelerated recovery from nuclear protein aggregates was that with total cellular levels of hsp27. The data indicate that heat-induced loss of clonogenic ability may be a multitarget rather than a single target event. A threshold of damage may exist in cells after exposure to heat; multiple sets of proteins in (different compartments of) the cell

  20. COMPARATIVE PRODUCTION OF SINGLE CELL PROTEIN FROM FISH PROTEIN ISOLATE WASTAGE AND ULTRA FILTERED CHEESE WHEY

    OpenAIRE

    Soroush Haghighi-Manesh; Marzieh Moosavi-Nasab; Somaye Farhoodi

    2013-01-01

    Fish protein isolate wastage and ultra filtered cheese whey were used as substrates for fermentation by Kluyveromyces marxianus to produce single cell protein, under batch and aerobic condition in which pH and temperature were adjusted to 4.5 and 35°C. The produced biomass was analyzed for protein content in different periods of time during fermentation. About 82% and 75% of total protein was produced in the first 18 h of 96 h fermentation of ultra filtered cheese whey and protein isolate was...

  1. The tumor suppressor APC differentially regulates multiple β-catenins through the function of axin and CKIα during C. elegans asymmetric stem cell divisions.

    Science.gov (United States)

    Baldwin, Austin T; Phillips, Bryan T

    2014-06-15

    The APC tumor suppressor regulates diverse stem cell processes including gene regulation through Wnt-β-catenin signaling and chromosome stability through microtubule interactions, but how the disparate functions of APC are controlled is not well understood. Acting as part of a Wnt-β-catenin pathway that controls asymmetric cell division, Caenorhabditis elegans APC, APR-1, promotes asymmetric nuclear export of the β-catenin WRM-1 by asymmetrically stabilizing microtubules. Wnt function also depends on a second β-catenin, SYS-1, which binds to the C. elegans TCF POP-1 to activate gene expression. Here, we show that APR-1 regulates SYS-1 levels in asymmetric stem cell division, in addition to its known role in lowering nuclear levels of WRM-1. We demonstrate that SYS-1 is also negatively regulated by the C. elegans homolog of casein kinase 1α (CKIα), KIN-19. We show that KIN-19 restricts APR-1 localization, thereby regulating nuclear WRM-1. Finally, the polarity of APR-1 cortical localization is controlled by PRY-1 (C. elegans Axin), such that PRY-1 controls the polarity of both SYS-1 and WRM-1 asymmetries. We propose a model whereby Wnt signaling, through CKIα, regulates the function of two distinct pools of APC - one APC pool negatively regulates SYS-1, whereas the second pool stabilizes microtubules and promotes WRM-1 nuclear export. PMID:24762815

  2. Human neuronal cell protein responses to Nipah virus infection

    Directory of Open Access Journals (Sweden)

    Hassan Sharifah

    2007-06-01

    Full Text Available Abstract Background Nipah virus (NiV, a recently discovered zoonotic virus infects and replicates in several human cell types. Its replication in human neuronal cells, however, is less efficient in comparison to other fully susceptible cells. In the present study, the SK-N-MC human neuronal cell protein response to NiV infection is examined using proteomic approaches. Results Method for separation of the NiV-infected human neuronal cell proteins using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE was established. At least 800 protein spots were resolved of which seven were unique, six were significantly up-regulated and eight were significantly down-regulated. Six of these altered proteins were identified using mass spectrometry (MS and confirmed using MS/MS. The heterogenous nuclear ribonucleoprotein (hnRNP F, guanine nucleotide binding protein (G protein, voltage-dependent anion channel 2 (VDAC2 and cytochrome bc1 were present in abundance in the NiV-infected SK-N-MC cells in contrast to hnRNPs H and H2 that were significantly down-regulated. Conclusion Several human neuronal cell proteins that are differentially expressed following NiV infection are identified. The proteins are associated with various cellular functions and their abundance reflects their significance in the cytopathologic responses to the infection and the regulation of NiV replication. The potential importance of the ratio of hnRNP F, and hnRNPs H and H2 in regulation of NiV replication, the association of the mitochondrial protein with the cytopathologic responses to the infection and induction of apoptosis are highlighted.

  3. RPE cell surface proteins in normal and dystrophic rats

    International Nuclear Information System (INIS)

    Membrane-bound proteins in plasma membrane enriched fractions from cultured rat RPE were analyzed by two-dimensional gel electrophoresis. Membrane proteins were characterized on three increasingly specific levels. Total protein was visualized by silver staining. A maximum of 102 separate proteins were counted in silver-stained gels. Glycoproteins were labeled with 3H-glucosamine or 3H-fucose and detected by autoradiography. Thirty-eight fucose-labeled and 61-71 glucosamine-labeled proteins were identified. All of the fucose-labeled proteins were labeled with glucosamine-derived radioactivity. Proteins exposed at the cell surface were labeled by lactoperoxidase-catalyzed radioiodination prior to preparation of membranes for two-dimensional analysis. Forty separate 125I-labeled surface proteins were resolved by two-dimensional electrophoresis/autoradiography. Comparison with the glycoprotein map showed that a number of these surface labeled proteins were glycoproteins. Two-dimensional maps of total protein, fucose-labeled, and glucosamine-labeled glycoproteins, and 125I-labeled surface proteins of membranes from dystrophic (RCS rdy-p+) and normal (Long Evans or RCS rdy+p+) RPE were compared. No differences in the total protein or surface-labeled proteins were observed. However, the results suggest that a 183K glycoprotein is more heavily glycosylated with glucosamine and fucose in normal RPE membranes as compared to membranes from dystrophic RPE

  4. Buffalo milk: proteins electrophoretic profile and somatic cell count

    OpenAIRE

    S. Mattii; B. Tommei; Pasquini, M.

    2011-01-01

    Water buffalo milk differs from the cow’s milk for greater fat and protein content, very important features in cheese making. Proteins, casein and whey-proteins in particular, are the most important factors determining cheese yield. Several previous research discussed the rule of SCC in cow milk production (Varisco, 1999) and the close relationship existing between cow’s milk cheese yield and somatic cell count (Barbano, 2000). In particular the inverse correlation between cheese ...

  5. Protein Profile of Exosomes from Trabecular Meshwork Cells

    OpenAIRE

    Stamer, WD; Hoffman, EA; Luther, JM; Hachey, DL; Schey, KL

    2011-01-01

    To better understand the role of exosomes in the trabecular meshwork (TM), the site of intraocular pressure control, the exosome proteome from primary cultures of human TM cell monolayers was analyzed. Exosomes were purified from urine and conditioned media from primary cultures of human TM cell monolayers and subjected to two dimensional HPLC separation and MS/MS analyses using the MudPIT strategy. Spectra were searched against a human protein database using Sequest. Protein profiles were co...

  6. Visualization and targeted disruption of protein interactions in living cells.

    Science.gov (United States)

    Herce, Henry D; Deng, Wen; Helma, Jonas; Leonhardt, Heinrich; Cardoso, M Cristina

    2013-01-01

    Protein-protein interactions are the basis of all processes in living cells, but most studies of these interactions rely on biochemical in vitro assays. Here we present a simple and versatile fluorescent-three-hybrid (F3H) strategy to visualize and target protein-protein interactions. A high-affinity nanobody anchors a GFP-fusion protein of interest at a defined cellular structure and the enrichment of red-labelled interacting proteins is measured at these sites. With this approach, we visualize the p53-HDM2 interaction in living cells and directly monitor the disruption of this interaction by Nutlin 3, a drug developed to boost p53 activity in cancer therapy. We further use this approach to develop a cell-permeable vector that releases a highly specific peptide disrupting the p53 and HDM2 interaction. The availability of multiple anchor sites and the simple optical readout of this nanobody-based capture assay enable systematic and versatile analyses of protein-protein interactions in practically any cell type and species. PMID:24154492

  7. Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae.

    Science.gov (United States)

    Zhao, Gang; Chen, Yuping; Carey, Lucas; Futcher, Bruce

    2016-05-19

    Cyclin-dependent kinases (CDKs) control cell division in eukaryotes by phosphorylating proteins involved in division. But successful proliferation requires co-ordination between division and cellular growth in mass. Previous proteomic studies suggested that metabolic proteins, as well as cell division proteins, could potentially be substrates of cyclin-dependent kinases. Here we focus on two metabolic enzymes of the yeast S. cerevisiae, neutral trehalase (Nth1) and glycogen phosphorylase (Gph1), and show that their activities are likely directly controlled by CDK activity, thus allowing co-ordinate regulation of carbohydrate metabolism with cell division processes. In this case, co-ordinate regulation may optimize the decision to undertake a final cell division as nutrients are being exhausted. Co-regulation of cell division processes and metabolic processes by CDK activity may be a general phenomenon important for co-ordinating the cell cycle with growth. PMID:27203179

  8. Surface Functionalization for Protein and Cell Patterning

    Science.gov (United States)

    Colpo, Pascal; Ruiz, Ana; Ceriotti, Laura; Rossi, François

    The interaction of biological systems with synthetic material surfaces is an important issue for many biological applications such as implanted devices, tissue engineering, cell-based sensors and assays, and more generally biologic studies performed ex vivo. To ensure reliable outcomes, the main challenge resides in the ability to design and develop surfaces or artificial micro-environment that mimic 'natural environment' in interacting with biomolecules and cells without altering their function and phenotype. At this effect, microfabrication, surface chemistry and material science play a pivotal role in the design of advanced in-vitro systems for cell culture applications. In this chapter, we discuss and describe different techniques enabling the control of cell-surface interactions, including the description of some techniques for immobilization of ligands for controlling cell-surface interactions and some methodologies for the creation of well confined cell rich areas.

  9. Eight weeks of pre- and postexercise whey protein supplementation increases lean body mass and improves performance in Division III collegiate female basketball players.

    Science.gov (United States)

    Taylor, Lemuel W; Wilborn, Colin; Roberts, Michael D; White, Andrew; Dugan, Kristen

    2016-03-01

    We examined if 8 weeks of whey protein (WP) supplementation improved body composition and performance measures in NCAA Division III female basketball players. Subjects were assigned to consume 24 g WP (n = 8; age, 20 ± 2 years; height, 170 ± 6 cm; weight, 66.0 ± 3.1 kg) or 24 g of maltodextrin (MD) (n = 6; age, 21 ± 3 years; height, 169 ± 6 cm; weight, 68.2 ± 7.6 kg) immediately prior to and following training (4 days/week anaerobic and resistance training) for 8 weeks. Prior to (T1) and 8 weeks following supplementation (T2), subjects underwent dual X-ray absorptiometry body composition assessment as well as performance tests. The WP group gained lean mass from T1 to T2 (+1.4 kg, p = 0.003) whereas the MD group trended to gain lean mass (+0.4 kg, p = 0.095). The WP group also lost fat mass from T1 to T2 (-1.0 kg, p = 0.003) whereas the MD group did not (-0.5 kg, p = 0.41). The WP group presented greater gains in 1-repetition maximum (1RM) bench press (+4.9 kg) compared with the MD group (+2.3 kg) (p < 0.05). Moreover, the WP group improved agility from T1 to T2 (p = 0.001) whereas the MD group did not (p = 0.38). Both groups equally increased leg press 1RM, vertical jump, and broad jump performances. This study demonstrates that 8 weeks of WP supplementation improves body composition and select performance variables in previously trained female athletes. PMID:26842665

  10. Monitoring protein synthesis in single live cancer cells.

    Science.gov (United States)

    Tu, Chengyi; Santo, Loredana; Mishima, Yuko; Raje, Noopur; Smilansky, Zeev; Zoldan, Janet

    2016-05-16

    Protein synthesis is generally under sophisticated and dynamic regulation to meet the ever-changing demands of a cell. Global up or down-regulation of protein synthesis and the shift of protein synthesis location (as shown, for example, during cellular stress or viral infection) are recognized as cellular responses to environmental changes such as nutrient/oxygen deprivation or to alterations such as pathological mutations in cancer cells. Monitoring protein synthesis in single live cells can be a powerful tool for cancer research. Here we employed a microfluidic platform to perform high throughput delivery of fluorescent labeled tRNAs into multiple myeloma cells with high transfection efficiency (∼45%) and high viability (>80%). We show that the delivered tRNAs were actively recruited to the ER for protein synthesis and that treatment with puromycin effectively disrupted this process. Interestingly, we observed the scattered distribution of tRNAs in cells undergoing mitosis, which has not been previously reported. Fluorescence lifetime analysis detected extensive FRET signals generated from tRNAs labeled as FRET pairs, further confirming that the delivered tRNAs were used by active ribosomes for protein translation. Our work demonstrates that the microfluidic delivery of FRET labeled tRNAs into living cancer cells can provide new insights into basic cancer metabolism and has the potential to serve as a platform for drug screening, diagnostics, or personalized medication. PMID:26956582

  11. Markov mean properties for cell death-related protein classification.

    Science.gov (United States)

    Fernandez-Lozano, Carlos; Gestal, Marcos; González-Díaz, Humberto; Dorado, Julián; Pazos, Alejandro; Munteanu, Cristian R

    2014-05-21

    The cell death (CD) is a dynamic biological function involved in physiological and pathological processes. Due to the complexity of CD, there is a demand for fast theoretical methods that can help to find new CD molecular targets. The current work presents the first classification model to predict CD-related proteins based on Markov Mean Properties. These protein descriptors have been calculated with the MInD-Prot tool using the topological information of the amino acid contact networks of the 2423 protein chains, five atom physicochemical properties and the protein 3D regions. The Machine Learning algorithms from Weka were used to find the best classification model for CD-related protein chains using all 20 attributes. The most accurate algorithm to solve this problem was K*. After several feature subset methods, the best model found is based on only 11 variables and is characterized by the Area Under the Receiver Operating Characteristic Curve (AUROC) of 0.992 and the true positive rate (TP Rate) of 88.2% (validation set). 7409 protein chains labeled with "unknown function" in the PDB Databank were analyzed with the best model in order to predict the CD-related biological activity. Thus, several proteins have been predicted to have CD-related function in Homo sapiens: 3DRX-involved in virus-host interaction biological process, protein homooligomerization; 4DWF-involved in cell differentiation, chromatin modification, DNA damage response, protein stabilization; 1IUR-involved in ATP binding, chaperone binding; 1J7D-involved in DNA double-strand break processing, histone ubiquitination, nucleotide-binding oligomerization; 1UTU-linked with DNA repair, regulation of transcription; 3EEC-participating to the cellular membrane organization, egress of virus within host cell, class mediator resulting in cell cycle arrest, negative regulation of ubiquitin-protein ligase activity involved in mitotic cell cycle and apoptotic process. Other proteins from bacteria predicted as

  12. β-catenin: A multi-functional protein’s role at the centrosome and implications for a broader role in cell division

    OpenAIRE

    Mbom, Bertrade C.; Nelson, W. James; Barth, Angela

    2013-01-01

    Beta-catenin is a multifunctional protein with critical roles in cell-cell adhesion, Wnt-signaling and the centrosome cycle. Whereas the roles of β-catenin in cell-cell adhesion and Wnt-signaling have been studied extensively, the mechanism(s) involving β-catenin in centrosome functions are poorly understood. β-Catenin localizes to centrosomes and promotes mitotic progression. NIMA-related protein kinase 2 (Nek2), which stimulates centrosome separation, binds to and phosphorylates β-catenin. ...

  13. Recombinant fusion protein of albumin-retinol binding protein inactivates stellate cells

    International Nuclear Information System (INIS)

    Highlights: ► We designed novel recombinant albumin-RBP fusion proteins. ► Expression of fusion proteins inactivates pancreatic stellate cells (PSCs). ► Fusion proteins are successfully internalized into and inactivate PSCs. ► RBP moiety mediates cell specific uptake of fusion protein. -- Abstract: Quiescent pancreatic- (PSCs) and hepatic- (HSCs) stellate cells store vitamin A (retinol) in lipid droplets via retinol binding protein (RBP) receptor and, when activated by profibrogenic stimuli, they transform into myofibroblast-like cells which play a key role in the fibrogenesis. Despite extensive investigations, there is, however, currently no appropriate therapy available for tissue fibrosis. We previously showed that the expression of albumin, composed of three homologous domains (I–III), inhibits stellate cell activation, which requires its high-affinity fatty acid-binding sites asymmetrically distributed in domain I and III. To attain stellate cell-specific uptake, albumin (domain I/III) was coupled to RBP; RBP-albumindomainIII (R-III) and albumindomainI-RBP-albuminIII (I-R-III). To assess the biological activity of fusion proteins, cultured PSCs were used. Like wild type albumin, expression of R-III or I-R-III in PSCs after passage 2 (activated PSCs) induced phenotypic reversal from activated to fat-storing cells. On the other hand, R-III and I-R-III, but not albumin, secreted from transfected 293 cells were successfully internalized into and inactivated PSCs. FPLC-purified R-III was found to be internalized into PSCs via caveolae-mediated endocytosis, and its efficient cellular uptake was also observed in HSCs and podocytes among several cell lines tested. Moreover, tissue distribution of intravenously injected R-III was closely similar to that of RBP. Therefore, our data suggest that albumin-RBP fusion protein comprises of stellate cell inactivation-inducing moiety and targeting moiety, which may lead to the development of effective anti

  14. Single-cell protein secretomic signatures as potential correlates to tumor cell lineage evolution and cell-cell interaction

    Directory of Open Access Journals (Sweden)

    Minsuk eKwak

    2013-02-01

    Full Text Available Secreted proteins including cytokines, chemokines and growth factors represent important functional regulators mediating a range of cellular behavior and cell-cell paracrine/autocrine signaling, e.g. in the immunological system, tumor microenvironment or stem cell niche. Detection of these proteins is of great value not only in basic cell biology but also for diagnosis and therapeutic monitoring of human diseases such as cancer. However, due to co-production of multiple effector proteins from a single cell, referred to as polyfunctionality, it is biologically informative to measure a panel of secreted proteins, or secretomic signature, at the level of single cells. Recent evidence further indicates that a genetically-identical cell population can give rise to diverse phenotypic differences. It is known that cytokines, for example, in the immune system define the effector functions and lineage differentiation of immune cells. In this Perspective Article, we hypothesize that protein secretion profile may represent a universal measure to identify the definitive correlate in the larger context of cellular functions to dissect cellular heterogeneity and evolutionary lineage relationship in human cancer.

  15. Protein expression profile in the differentiation of rat bone marrow stromal cells into Schwann cell-like cells

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    During the last decade,increasing evidence suggested that bone marrow stromal cells(MSCs) have the potential to differentiate into neural lineages.Many studies have reported that MSCs showed morphological changes and expressed a limited number of neural proteins under experimental conditions.However,no proteomic studies on MSCs differentiated into Schwann cell-like cells have been reported.In this study,we isolated MSCs from adult Sprague-Dawley rat femur and tibia bone marrows and induced the cells in vitro under specific conditions.By using two-dimensional gel electrophoresis(2-DE),we compared the protein profiles of MSCs before and after induced differentiation.We obtained 792 protein spots in the protein profile by 2-DE,and found that 74 spots changed significantly before and after the differentiation using PDQuest software,with 43 up-regulated and 31 down-regulated.We analyzed these 74 spots by a matrix assisted laser desorption ionization-time of flight mass spectrometry(MALDI-TOF-MS) and by database searching,and found that they could be grouped into various classes,including cytoskeleton and structure proteins,growth factors,metabolic proteins,chaperone proteins,receptor proteins,cell cycle proteins,calcium binding proteins,and other proteins.These proteins also include neural and glial proteins,such as BDNF,CNTF and GFAP.The results may provide valuable proteomic information about the differentiation of MSCs into Schwann cell-like cells.

  16. The progression of the intra-erythrocytic cell cycle of Plasmodium falciparum and the role of the centriolar plaques in asynchronous mitotic division during schizogony

    DEFF Research Database (Denmark)

    Arnot, David E; Ronander, Elena; Bengtsson, Dominique C

    2011-01-01

    The cell division cycle and mitosis of intra-erythrocytic (IE) Plasmodium falciparum are poorly understood aspects of parasite development which affect malaria molecular pathogenesis. Specifically, the timing of the multiple gap (G), DNA synthesis (S) and chromosome separation (M) phases of...... parasite mitosis are not well defined, nor whether genome divisions are immediately followed by cleavage of the nuclear envelope. Curiously, daughter merozoite numbers do not follow the geometric expansion expected from equal numbers of binary divisions, an outcome difficult to explain using the standard...

  17. NSA2, a novel nucleolus protein regulates cell proliferation and cell cycle

    International Nuclear Information System (INIS)

    NSA2 (Nop seven-associated 2) was previously identified in a high throughput screen of novel human genes associated with cell proliferation, and the NSA2 protein is evolutionarily conserved across different species. In this study, we revealed that NSA2 is broadly expressed in human tissues and cultured cell lines, and located in the nucleolus of the cell. Both of the putative nuclear localization signals (NLSs) of NSA2, also overlapped with nucleolar localization signals (NoLSs), are capable of directing nucleolar accumulation. Moreover, over-expression of the NSA2 protein promoted cell growth in different cell lines and regulated the G1/S transition in the cell cycle. SiRNA silencing of the NSA2 transcript attenuated the cell growth and dramatically blocked the cell cycle in G1/S transition. Our results demonstrated that NSA2 is a nucleolar protein involved in cell proliferation and cell cycle regulation.

  18. Placental expression of estrogen receptor beta and its hormone binding variant – comparison with estrogen receptor alpha and a role for estrogen receptors in asymmetric division and differentiation of estrogen-dependent cells

    Directory of Open Access Journals (Sweden)

    Henley Donald C

    2003-04-01

    Full Text Available Abstract During human pregnancy, the production of 17-beta-estradiol (E2 rises steadily to eighty fold at term, and placenta has been found to specifically bind estrogens. We have recently demonstrated the expression of estrogen receptor alpha (ER-alpha protein in human placenta and its localization in villous cytotrophoblast (CT, vascular pericytes, and amniotic fibroblasts. In vitro, E2 stimulated development of large syncytiotrophoblast (ST aggregates. In the present study we utilized ER-beta affinity purified polyclonal (N19:sc6820 and ER-alpha monoclonal (clone h-151 antibodies. Western blot analysis revealed a single ~52 kDa ER-beta band in chorionic villi (CV protein extracts. In CV, strong cytoplasmic ER-beta immunoreactivity was confined to ST. Dual color immunohistochemistry revealed asymmetric segregation of ER-alpha in dividing villous CT cells. Prior to separation, the cell nuclei more distant from ST exhibited high ER-alpha, while cell nuclei associated with ST showed diminution of ER-alpha and appearance of ER-beta. In trophoblast cultures, development of ST aggregates was associated with diminution of ER-alpha and appearance of ER-beta immunoreactivity. ER-beta was also detected in endothelial cells, amniotic epithelial cells and fibroblasts, extravillous trophoblast (nuclear and cytoplasmic and decidual cells (cytoplasmic only. In addition, CFK-E12 (E12 and CWK-F12 (F12 monoclonal antibodies, which recognize ~64 kDa ER-beta with hormone binding domain, showed nuclear-specific reactivity with villous ST, extravillous trophoblast, and amniotic epithelium and fibroblasts. Western blot analysis indicated abundant expression of a ~64 kDa ER-beta variant in trophoblast cultures, significantly higher when compared to the chorionic villi and freshly isolated trophoblast cell protein extracts. This is the first report on ER-beta expression in human placenta and cultured trophoblast. Our data indicate that during trophoblast

  19. Proteins from human retinal pigment epithelial cells: evidence that a major protein is actin

    International Nuclear Information System (INIS)

    Two-dimensional gel electrophoresis coupled to fluorography was used to obtain the major 35S-methionine labeled protein patterns of cultured human retinal pigment epithelial cells (RPE) and fibroblasts. Comparisons of these profiles showed that: a) primary and subcultures of RPE were quite similar, each showing about 200 different proteins, b) the RPE protein patterns contained at least eight major acidic proteins (from 31 to 96 kilodaltons) not readily seen in fibroblasts; c) the fibroblast pattern showed three well-labeled, very acidic proteins, one of which (58 kilodaltons) appeared to be unique to fibroblasts; and d) a major complex of acidic protein of 43 kilodaltons with isoelectric points of 5.6 to 5.9 was a common protein in RPE and fibroblasts. These latter macromolecules were found to coelectrophorese with purified chicken muscle actin. Ultrastructural studies of cultured RPE supported the presence of actin microfilaments that were demonstrated specifically by labeling with fluorescent phallotoxin

  20. Cell-free protein production for NMR studies.

    Science.gov (United States)

    Takeda, Mitsuhiro; Kainosho, Masatsune

    2012-01-01

    The cell-free expression system using an Escherichia coli extract is a practical method for producing isotope-labeled proteins. The advantage of the cell-free system over cellular expression is that any isotope-labeled amino acid can be incorporated into the target protein with minimal scrambling, thus providing opportunities for advanced isotope labeling of proteins. We have modified the standard protocol for E. coli cell-free expression to cope with two problems specific to NMR sample preparation. First, endogenous amino acids present in the E. coli S30 extract lead to dilution of the added isotope. To minimize the content of the remaining amino acids, a gel filtration step is included in the preparation of the E. coli extract. Second, proteins produced by the cell-free system are not necessarily homogeneous due to incomplete processing of the N-terminal formyl-methionine residue, which complicates NMR spectra. Therefore, the protein of interest is engineered to contain a cleavable N-terminal histidine-tag, which generates a homogeneous protein after the digestion of the tag. Here, we describe the protocol for modified E. coli cell-free expression. PMID:22167669

  1. Visualization and targeted disruption of protein interactions in living cells

    OpenAIRE

    Herce, Henry D.; Deng, Wen; Helma, Jonas; Leonhardt, Heinrich; Cardoso, M. Cristina

    2013-01-01

    Protein–protein interactions are the basis of all processes in living cells, but most studies of these interactions rely on biochemical in vitro assays. Here we present a simple and versatile fluorescent-three-hybrid (F3H) strategy to visualize and target protein–protein interactions. A high-affinity nanobody anchors a GFP-fusion protein of interest at a defined cellular structure and the enrichment of red-labelled interacting proteins is measured at these sites. With this approach, we visual...

  2. Nanoparticles-cell association predicted by protein corona fingerprints

    Science.gov (United States)

    Palchetti, S.; Digiacomo, L.; Pozzi, D.; Peruzzi, G.; Micarelli, E.; Mahmoudi, M.; Caracciolo, G.

    2016-06-01

    In a physiological environment (e.g., blood and interstitial fluids) nanoparticles (NPs) will bind proteins shaping a ``protein corona'' layer. The long-lived protein layer tightly bound to the NP surface is referred to as the hard corona (HC) and encodes information that controls NP bioactivity (e.g. cellular association, cellular signaling pathways, biodistribution, and toxicity). Decrypting this complex code has become a priority to predict the NP biological outcomes. Here, we use a library of 16 lipid NPs of varying size (Ø ~ 100-250 nm) and surface chemistry (unmodified and PEGylated) to investigate the relationships between NP physicochemical properties (nanoparticle size, aggregation state and surface charge), protein corona fingerprints (PCFs), and NP-cell association. We found out that none of the NPs' physicochemical properties alone was exclusively able to account for association with human cervical cancer cell line (HeLa). For the entire library of NPs, a total of 436 distinct serum proteins were detected. We developed a predictive-validation modeling that provides a means of assessing the relative significance of the identified corona proteins. Interestingly, a minor fraction of the HC, which consists of only 8 PCFs were identified as main promoters of NP association with HeLa cells. Remarkably, identified PCFs have several receptors with high level of expression on the plasma membrane of HeLa cells.In a physiological environment (e.g., blood and interstitial fluids) nanoparticles (NPs) will bind proteins shaping a ``protein corona'' layer. The long-lived protein layer tightly bound to the NP surface is referred to as the hard corona (HC) and encodes information that controls NP bioactivity (e.g. cellular association, cellular signaling pathways, biodistribution, and toxicity). Decrypting this complex code has become a priority to predict the NP biological outcomes. Here, we use a library of 16 lipid NPs of varying size (Ø ~ 100-250 nm) and surface

  3. Substrates of protein kinases involved in cell signal transduction

    International Nuclear Information System (INIS)

    In this study substrates for protein-tyrosine kinases and protein kinase C are examined to gain a better understanding of the conditions of their phosphorylation, their functions, and their potential involvement in intracellular signaling pathways. The tissue, cell type, and intracellular distributions of two protein-tyrosine kinase substrates, termed p36 and p81, are determined by immunoblotting of murine tissues, indirect immunofluorescence and immunoperoxidase staining of frozen rat tissue sections, and biochemical fractionation and indirect immunofluorescence staining of tissue culture cells. Both p36 and p81 are constitutively phosphorylated to low levels in tissue culture cells. In 32P-labeled A431 cells, pp81 contains both phosphoserine and phosphothreonine. Following brief epidermal growth factor treatment of A431 cells, pp81 is more heavily phosphorylated on threonine and approximately 10% of p81 molecules become phosphorylated on tyrosine. Treatment of A431 cells with the potent tumor promoter and protein kinase C activator, 12-O-tetradecanoylphorbol-13-acetate (TPA), does not alter the phosphorylation state of p81. However, TPA treatment of A431 cells and certain other cell types leads to augmented serine phosphorylation of p36

  4. Sphingolipid trafficking and protein sorting in epithelial cells

    NARCIS (Netherlands)

    Slimane, TA; Hoekstra, D

    2002-01-01

    Sphingolipids represent a minor, but highly dynamic subclass of lipids in all eukaryotic cells. They are involved in functions that range from structural protection to signal transduction and protein sorting, and participate in lipid raft assembly. In polarized epithelial cells, which display an asy

  5. Timing the generation of distinct retinal cells by homeobox proteins.

    Directory of Open Access Journals (Sweden)

    Sarah Decembrini

    2006-09-01

    Full Text Available The reason why different types of vertebrate nerve cells are generated in a particular sequence is still poorly understood. In the vertebrate retina, homeobox genes play a crucial role in establishing different cell identities. Here we provide evidence of a cellular clock that sequentially activates distinct homeobox genes in embryonic retinal cells, linking the identity of a retinal cell to its time of generation. By in situ expression analysis, we found that the three Xenopus homeobox genes Xotx5b, Xvsx1, and Xotx2 are initially transcribed but not translated in early retinal progenitors. Their translation requires cell cycle progression and is sequentially activated in photoreceptors (Xotx5b and bipolar cells (Xvsx1 and Xotx2. Furthermore, by in vivo lipofection of "sensors" in which green fluorescent protein translation is under control of the 3' untranslated region (UTR, we found that the 3' UTRs of Xotx5b, Xvsx1, and Xotx2 are sufficient to drive a spatiotemporal pattern of translation matching that of the corresponding proteins and consistent with the time of generation of photoreceptors (Xotx5b and bipolar cells (Xvsx1 and Xotx2. The block of cell cycle progression of single early retinal progenitors impairs their differentiation as photoreceptors and bipolar cells, but is rescued by the lipofection of Xotx5b and Xvsx1 coding sequences, respectively. This is the first evidence to our knowledge that vertebrate homeobox proteins can work as effectors of a cellular clock to establish distinct cell identities.

  6. Protein Expression Profiling in the Spectrum of Renal Cell Carcinomas

    Directory of Open Access Journals (Sweden)

    Vladimir A Valera, Elsa Li-Ning-T, Beatriz A Walter, David D. Roberts, W M Linehan, Maria J Merino

    2010-01-01

    Full Text Available In this study, we aimed to evaluate the protein expression profile of a spectrum of renal cell carcinomas (RCC to find potential biomarkers for disease onset and progression and therefore, prospective therapeutic targets. A 2D-gel based proteomic analysis was used to outline differences in protein levels among different subtypes of renal cell carcinomas, including clear cell carcinomas, papillary lesions, chromophobe tumors and renal oncocytomas. Spot pattern was compared to the corresponding normal kidney from the same patients and distinctive, differentially expressed proteins were characterized by mass spectrometry. Twenty-one protein spots were found differentially expressed between clear cell RCC and normal tissue and 38 spots were found expressed in chromophobe tumors. Eleven proteins were identified, with most differentially expressed -by fold change- between clear cell tumors and the corresponding normal tissue. Two of the identified proteins, Triosephosphate isomerase 1 (TPI-1 and Heat Shock protein 27 (Hsp27, were further validated in a separate set of tumors by immunohistochemistry and expression levels were correlated with clinicopathologic features of the patients. Hsp27 was highly expressed in 82% of the tumors used for validation, and all cases showed strong immunoreactivity for TPI-1. In both Hsp27 and TPI-1, protein expression positively correlated with histologic features of the disease. Our results suggest that the subjacent cytogenetic abnormalities seen in different histological types of RCC are followed by specific changes in protein expression. From these changes, Hsp27 and TPI-1 emerged as potential candidates for the differentiation and prognosis in RCC.

  7. Prolactin-inducible proteins in human breast cancer cells

    International Nuclear Information System (INIS)

    The mechanism of action of prolactin in target cells and the role of prolactin in human breast cancer are poorly understood phenomena. The present study examines the effect of human prolactin (hPRL) on the synthesis of unique proteins by a human breast cancer cell line, T-47D, in serum-free medium containing bovine serum albumin. [35S]Methionine-labeled proteins were analysed by sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis and fluorography. Treatment of cells with hPRL (1-1000 ng/ml) and hydrocortisone (1 microgram/ml) for 36 h or longer resulted in the synthesis and secretion of three proteins having molecular weights of 11,000, 14,000, and 16,000. Neither hPRL nor hydrocortisone alone induced these proteins. Of several other peptide hormones tested, only human growth hormone, a hormone structurally and functionally similar to hPRL, could replace hPRL in causing protein induction. These three proteins were, therefore, referred to as prolactin-inducible proteins (PIP). Each of the three PIPs was purified to homogeneity by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and specific antibodies were generated to them in rabbits. By immunoprecipitation and immunoblotting (Western blot) of proteins secreted by T-47D cells, it was demonstrated that the three PIPs were immunologically identical to one another. In addition, the 16-kDa and 14-kDa proteins (PIP-16 and PIP-14), and not the 11-kDa protein (PIP-11), incorporated [3H]glycosamine. Furthermore, 2-deoxyglucose (2 mM) and tunicamycin (0.5 micrograms/ml), two compounds known to inhibit glycosylation, blocked the production of PIP-16 and PIP-14, with a concomitant increase in the accumulation of PIP-11

  8. Alterations of proteins in MDCK cells during acute potassium deficiency.

    Science.gov (United States)

    Peerapen, Paleerath; Ausakunpipat, Nardtaya; Chanchaem, Prangwalai; Thongboonkerd, Visith

    2016-06-01

    Chronic K(+) deficiency can cause hypokalemic nephropathy associated with metabolic alkalosis, polyuria, tubular dilatation, and tubulointerstitial injury. However, effects of acute K(+) deficiency on the kidney remained unclear. This study aimed to explore such effects by evaluating changes in levels of proteins in renal tubular cells during acute K(+) deficiency. MDCK cells were cultivated in normal K(+) (NK) (K(+)=5.3mM), low K(+) (LK) (K(+)=2.5mM), or K(+) depleted (KD) (K(+)=0mM) medium for 24h and then harvested. Cellular proteins were resolved by two-dimensional gel electrophoresis (2-DE) and visualized by SYPRO Ruby staining (5 gels per group). Spot matching and quantitative intensity analysis revealed a total 48 protein spots that had significantly differential levels among the three groups. Among these, 46 and 30 protein spots had differential levels in KD group compared to NK and LK groups, respectively. Comparison between LK and NK groups revealed only 10 protein spots that were differentially expressed. All of these differentially expressed proteins were successfully identified by Q-TOF MS and/or MS/MS analyses. The altered levels of heat shock protein 90 (HSP90), ezrin, lamin A/C, tubulin, chaperonin-containing TCP1 (CCT1), and calpain 1 were confirmed by Western blot analysis. Global protein network analysis showed three main functional networks, including 1) cell growth and proliferation, 2) cell morphology, cellular assembly and organization, and 3) protein folding in which the altered proteins were involved. Further investigations on these networks may lead to better understanding of pathogenic mechanisms of low K(+)-induced renal injury. PMID:26976750

  9. Surface immobilized protein multilayers for cell seeding

    Czech Academy of Sciences Publication Activity Database

    Brynda, Eduard; Pacherník, J.; Houska, Milan; Pientka, Zbyněk; Dvořák, P.

    2005-01-01

    Roč. 21, č. 17 (2005), s. 7877. ISSN 0743-7463 R&D Projects: GA ČR(CZ) GA203/02/1326; GA ČR GA102/03/0633; GA MŠk(CZ) LN00A065 Keywords : surface modification * layer-by-layer deposition * protein multilayers Subject RIV: CE - Biochemistry Impact factor: 3.705, year: 2005

  10. Adiponectin Protein Exists in Aortic Endothelial Cells

    OpenAIRE

    Noriyuki Komura; Norikazu Maeda; Takuya Mori; Shinji Kihara; Hideaki Nakatsuji; Ayumu Hirata; Yoshihiro Tochino; Tohru Funahashi; Iichiro Shimomura

    2013-01-01

    AIMS: Inflammation is closely associated with the development of atherosclerosis and metabolic syndrome. Adiponectin, an adipose-derived secretory protein, possesses an anti-atherosclerotic property. The present study was undertaken to elucidate the presence and significance of adiponectin in vasculature. METHODS AND RESULTS: Immunofluorescence staining was performed in aorta of wild-type (WT) mice and demonstrated that adiponectin was co-stained with CD31. Thoracic aorta was cut through and ...

  11. Unstimulated secretion of protein from rat exocrine pancreas cells.

    Science.gov (United States)

    Kramer, M F; Poort, C

    1972-01-01

    Our earlier work demonstrated that the rate of protein synthesis in the exocrine cells of the rat pancreas is constant in different physiological states, including prolonged fasting. In this study we have followed the fate of the protein in the pancreatic cells of the fasting animal in vivo as well as in vitro. The data were obtained by quantitative radioautography and by biochemical determinations. In nonanesthesized, fasting rats, without cannulated pancreatic duct, some 80% of the proteins synthesized at a given time leaves the cell within 12 hr by way of secretion, intracellular breakdown not being important. Two mechanisms of fasting secretion exist. The first, starting at a slow rate after 20 min, is inferred to result from fortuitous contacts of young secretory granules with the apical cell membrane. The rate of secretion is the same in vivo as in vitro, at least during the first 4 hr after pulse labeling. Within 7 hr about 20% of the total amount of newly synthesized protein has left the cell. The second mechanism consists of an orderly movement of the mass of secretory granules towards the apical cell membrane as caused by the continuous assembly of new granules. The granules that come into contact with the cell membrane are discharged. It takes about 7-12 hr for secretory protein transported in this way to reach the cell membrane. The addition of new secretory granules to those present is essential for the second mechanism, for the blockade of protein synthesis by cycloheximide decreases the rate of this phase of secretion without interfering with the secretory process proper. Atropin does not inhibit the fasting secretion in vitro, nor does extensive washing of the tissue slices, excluding possible secretagogues as important factors in fasting secretion. PMID:4331296

  12. Specific Protein Markers for Stem Cell Cross-Talk with Neighboring Cells in the Environment

    OpenAIRE

    Park, Kyung Soo; Shin, Seung Won; Choi, Jeong-Woo; Um, Soong Ho

    2013-01-01

    A stem cell interacts with the neighboring cells in its environment. To maintain a living organism’s metabolism, either cell-cell or cell-environment interactions may be significant. Usually, these cells communicate with each other through biological signaling by interactive behaviors of primary proteins or complementary chemicals. The signaling intermediates offer the stem cell’s functionality on its metabolism. With the rapid advent of omics technologies, various specific markers by which s...

  13. Stem-cell-abundant proteins Nanog, Nucleostemin and Musashi1 are highly expressed in malignant cervical epithelial cells

    International Nuclear Information System (INIS)

    Nanog, nucleostemin (NS) and musashi1 (Msi1) are proteins that are highly expressed in undifferentiated embryonic stem (ES) cells and have been shown to be essential in maintaining the pluripotency and regulating the proliferation and asymmetric division of ES cells and several nervous system tumor cells. The roles of Nanog, NS and Msi1 in development and progression of cervical carcinoma have, until now, not been well documented. In this study, expression of Nanog, NS and Msi1 was detected by immunohistochemistry analysis in 235 patients with various degrees of cervical epithelial lesions, including 49 with normal cervical epithelia, 31 with mild dysplasia (CIN I), 77 with moderate-severe dysplasia (CIN II-III) and 78 with squamous cervical carcinomas (SCCs). Associations with various clinical pathological prognostic variables were analyzed in 50 early-stage SCC patients. Nanog, NS and Msi1 expression levels were significantly higher in SCC patients compared with CIN patients, and were higher in CIN patients compared with those with normal cervical epithelia. Nanog expression levels showed significantly differences according to different tumor sizes (P < 0.05), whereas there were no differences in NS and Msi1 expression levels according to different clinical pathological parameters. Our findings indicate that Nanog, NS and Msi1 may be involved in carcinogenesis of the cervix and progression of cervical carcinoma

  14. Combining in Vitro Folding with Cell Free Protein Synthesis for Membrane Protein Expression.

    Science.gov (United States)

    Focke, Paul J; Hein, Christopher; Hoffmann, Beate; Matulef, Kimberly; Bernhard, Frank; Dötsch, Volker; Valiyaveetil, Francis I

    2016-08-01

    Cell free protein synthesis (CFPS) has emerged as a promising methodology for protein expression. While polypeptide production is very reliable and efficient using CFPS, the correct cotranslational folding of membrane proteins during CFPS is still a challenge. In this contribution, we describe a two-step protocol in which the integral membrane protein is initially expressed by CFPS as a precipitate followed by an in vitro folding procedure using lipid vesicles for converting the protein precipitate to the correctly folded protein. We demonstrate the feasibility of using this approach for the K(+) channels KcsA and MVP and the amino acid transporter LeuT. We determine the crystal structure of the KcsA channel obtained by CFPS and in vitro folding to show the structural similarity to the cellular expressed KcsA channel and to establish the feasibility of using this two-step approach for membrane protein production for structural studies. Our studies show that the correct folding of these membrane proteins with complex topologies can take place in vitro without the involvement of the cellular machinery for membrane protein biogenesis. This indicates that the folding instructions for these complex membrane proteins are contained entirely within the protein sequence. PMID:27384110

  15. Nanoparticles-cell association predicted by protein corona fingerprints.

    Science.gov (United States)

    Palchetti, S; Digiacomo, L; Pozzi, D; Peruzzi, G; Micarelli, E; Mahmoudi, M; Caracciolo, G

    2016-07-01

    In a physiological environment (e.g., blood and interstitial fluids) nanoparticles (NPs) will bind proteins shaping a "protein corona" layer. The long-lived protein layer tightly bound to the NP surface is referred to as the hard corona (HC) and encodes information that controls NP bioactivity (e.g. cellular association, cellular signaling pathways, biodistribution, and toxicity). Decrypting this complex code has become a priority to predict the NP biological outcomes. Here, we use a library of 16 lipid NPs of varying size (Ø≈ 100-250 nm) and surface chemistry (unmodified and PEGylated) to investigate the relationships between NP physicochemical properties (nanoparticle size, aggregation state and surface charge), protein corona fingerprints (PCFs), and NP-cell association. We found out that none of the NPs' physicochemical properties alone was exclusively able to account for association with human cervical cancer cell line (HeLa). For the entire library of NPs, a total of 436 distinct serum proteins were detected. We developed a predictive-validation modeling that provides a means of assessing the relative significance of the identified corona proteins. Interestingly, a minor fraction of the HC, which consists of only 8 PCFs were identified as main promoters of NP association with HeLa cells. Remarkably, identified PCFs have several receptors with high level of expression on the plasma membrane of HeLa cells. PMID:27279572

  16. Cell-free system for synthesizing membrane proteins cell free method for synthesizing membrane proteins

    Science.gov (United States)

    Laible, Philip D; Hanson, Deborah K

    2013-06-04

    The invention provides an in vitro method for producing proteins, membrane proteins, membrane-associated proteins, and soluble proteins that interact with membrane-associated proteins for assembly into an oligomeric complex or that require association with a membrane for proper folding. The method comprises, supplying intracytoplasmic membranes from organisms; modifying protein composition of intracytoplasmic membranes from organism by modifying DNA to delete genes encoding functions of the organism not associated with the formation of the intracytoplasmic membranes; generating appropriate DNA or RNA templates that encode the target protein; and mixing the intracytoplasmic membranes with the template and a transcription/translation-competent cellular extract to cause simultaneous production of the membrane proteins and encapsulation of the membrane proteins within the intracytoplasmic membranes.

  17. Cell ``vision'': complementary factor of protein corona in nanotoxicology

    Science.gov (United States)

    Mahmoudi, Morteza; Saeedi-Eslami, Seyyed N.; Shokrgozar, Mohammad A.; Azadmanesh, Kayhan; Hassanlou, Maryam; Kalhor, Hamid R.; Burtea, Carmen; Rothen-Rutishauser, Barbara; Laurent, Sophie; Sheibani, Sara; Vali, Hojatollah

    2012-08-01

    Engineered nanoparticles are increasingly being considered for use as biosensors, imaging agents and drug delivery vehicles. Their versatility in design and applications make them an attractive proposition for new biological and biomedical approaches. Despite the remarkable speed of development in nanoscience, relatively little is known about the interaction of nanoscale objects with living systems. In a biological fluid, proteins associate with nanoparticles, and the amount and the presentation of the proteins on their surface could lead to a different in vivo response than an uncoated particle. Here, in addition to protein adsorption, we are going to introduce concept of cell ``vision'', which would be recognized as another crucial factor that should be considered for the safe design of any type of nanoparticles that will be used in specific biomedical applications. The impact of exactly the same nanoparticles on various cells is significantly different and could not be assumed for other cells; the possible mechanisms that justify this cellular response relate to the numerous detoxification strategies that any particular cell can utilize in response to nanoparticles. The uptake and defence mechanism could be considerably different according to the cell type. Thus, what the cell ``sees'', when it is faced with nanoparticles, is most likely dependent on the cell type.Engineered nanoparticles are increasingly being considered for use as biosensors, imaging agents and drug delivery vehicles. Their versatility in design and applications make them an attractive proposition for new biological and biomedical approaches. Despite the remarkable speed of development in nanoscience, relatively little is known about the interaction of nanoscale objects with living systems. In a biological fluid, proteins associate with nanoparticles, and the amount and the presentation of the proteins on their surface could lead to a different in vivo response than an uncoated particle. Here

  18. Cell-cycle protein modifications in the kidney of Mus spretus from Doñana National Park.

    Science.gov (United States)

    Leoni, Silvia; Torrice, Alessia; Chiuchiarelli, Giorgia; Bruscalupi, Giovannella; Cristaldi, Mauro

    2008-02-01

    On April 1998 a tailing dam of the Aznalcollar pyrite mine partially collapsed and released to the Guadiamar river acidic water and mud containing toxic metals threatening the Doñana National Park, a Spanish wildlife reserve located near the estuary of Guadalquivir river. To assess the possible biological effects on terrestrial ecosystems, biochemical markers were assayed in the kidneys of Algerian mice (Mus spretus) collected in several areas of Doñana and Guadiamar river. Biomarkers assayed are proteins involved in cell cycle regulation, in particular cyclins and their associated kinases, and some cell cycle inhibitors. Moreover Mitogen Activated Protein Kinases (MAPK), a signal transduction system involved in cell division, p53, a protein involved in growth arrest after DNA damage, and HSP70, an early stress-induced protein, were assayed. The kidneys of animals collected one year after the ecological disaster had increased levels of PCNA (proliferating cell nuclear antigen), indicating an increased number of cells in the S phase of cell cycle. This shift of cells from G0 to S phase is due to increased levels of cyclins D1, E, and A, to decreased levels of p21 and p27 cdk inhibitors, and to activation of MAPK cascade. On the other hand, p53 and HSP70 levels are not changed. These data demonstrate that the presence of toxic metals after ecological disaster provoked the induction of kidney cell proliferation interpretable as a compensatory cell growth after tissue damage and apoptosis, and that could lead to the genomic instability characteristic of cancer cell. PMID:18214926

  19. Protein carbonylation, protein aggregation and neuronal cell death in a murine model of multiple sclerosis

    Science.gov (United States)

    Dasgupta, Anushka

    Many studies have suggested that oxidative stress plays an important role in the pathophysiology of both multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Yet, the mechanism by which oxidative stress leads to tissue damage in these disorders is unclear. Recent work from our laboratory has revealed that protein carbonylation, a major oxidative modification caused by severe and/or chronic oxidative stress conditions, is elevated in MS and EAE. Furthermore, protein carbonylation has been shown to alter protein structure leading to misfolding/aggregation. These findings prompted me to hypothesize that carbonylated proteins, formed as a consequence of oxidative stress and/or decreased proteasomal activity, promote protein aggregation to mediate neuronal apoptosis in vitro and in EAE. To test this novel hypothesis, I first characterized protein carbonylation, protein aggregation and apoptosis along the spinal cord during the course of myelin-oligodendrocyte glycoprotein (MOG)35-55 peptide-induced EAE in C57BL/6 mice [Chapter 2]. The results show that carbonylated proteins accumulate throughout the course of the disease, albeit by different mechanisms: increased oxidative stress in acute EAE and decreased proteasomal activity in chronic EAE. I discovered not only that there is a temporal correlation between protein carbonylation and apoptosis but also that carbonyl levels are significantly higher in apoptotic cells. A high number of juxta-nuclear and cytoplasmic protein aggregates containing the majority of the oxidized proteins are also present during the course of EAE, which seems to be due to reduced autophagy. In chapter 3, I show that when gluthathione levels are reduced to those in EAE spinal cord, both neuron-like PC12 (nPC12) cells and primary neuronal cultures accumulate carbonylated proteins and undergo cell death (both by necrosis and apoptosis). Immunocytochemical and biochemical studies also revealed a temporal

  20. Identification of chikungunya virus interacting proteins in mammalian cells

    Indian Academy of Sciences (India)

    Mandar S Paingankar; Vidya A Arankalle

    2014-06-01

    Identification and characterization of virus host interactions is an essential step for the development of novel antiviral strategies. Very few studies have been targeted towards identification of chikungunya virus (CHIKV) interacting host proteins. In current study, virus overlay protein binding assay (VOPBA) and matrix-assisted laser desorption/ionization time of flight analysis (MALDI TOF/TOF) were employed for the identification of CHIKV binding proteins in mammalian cells. HSP70 and actin were identified as virus binding proteins in HEK-293T and Vero-E6 cells, whereas STAT-2 was identified as an additional protein in Vero-E6 cells. Pre-incubation with anti-HSP70 antibody and miRNA silencing of HSP70 significantly reduced the CHIKV production in HEK-293T and Vero-E6 cells at early time points. These results suggest that CHIKV exploits the housekeeping molecules such as actin, HSP70 and STAT-2 to establish infection in the mammalian cells.

  1. Cdon, a cell surface protein, mediates oligodendrocyte differentiation and myelination.

    Science.gov (United States)

    Wang, Li-Chun; Almazan, Guillermina

    2016-06-01

    During central nervous system development, oligodendrocyte progenitors (OLPs) establish multiple branched processes and axonal contacts to initiate myelination. A complete understanding of the molecular signals implicated in cell surface interaction to initiate myelination/remyelination is currently lacking. The objective of our study was to assess whether Cdon, a cell surface protein that was shown to participate in muscle and neuron cell development, is involved in oligodendrocyte (OLG) differentiation and myelination. Here, we demonstrate that endogenous Cdon protein is expressed in OLPs, increasing in the early differentiation stages and decreasing in mature OLGs. Immunocytochemistry of endogenous Cdon showed localization on both OLG cell membranes and cellular processes exhibiting puncta- or varicosity-like structures. Cdon knockdown with siRNA decreased protein levels by 62% as well as two myelin-specific proteins, MBP and MAG. Conversely, overexpression of full-length rat Cdon increased myelin proteins in OLGs. The complexity of OLGs branching and contact point numbers with axons were also increased in Cdon overexpressing cells growing alone or in coculture with dorsal root ganglion neurons (DRGNs). Furthermore, myelination of DRGNs was decreased when OLPs were transfected with Cdon siRNA. Altogether, our results suggest that Cdon participates in OLG differentiation and myelination, most likely in the initial stages of development. GLIA 2016;64:1021-1033. PMID:26988125

  2. Yeast spore germination: a requirement for Ras protein activity during re-entry into the cell cycle.

    OpenAIRE

    Herman, P K; Rine, J.

    1997-01-01

    Saccharomyces cerevisiae spore germination is a process in which quiescent, non-dividing spores become competent for mitotic cell division. Using a novel assay for spore uncoating, we found that spore germination was a multi-step process whose nutritional requirements differed from those for mitotic division. Although both processes were controlled by nutrient availability, efficient spore germination occurred in conditions that did not support cell division. In addition, germination did not ...

  3. Surface Analyses and Immune Reactivities of Major Cell Wall-Associated Proteins of Group A Streptococcus

    OpenAIRE

    Cole, Jason N; Ramirez, Ruben D.; Currie, Bart J.; Cordwell, Stuart J.; Djordjevic, Steven P.; Mark J Walker

    2005-01-01

    A proteomic analysis was undertaken to identify cell wall-associated proteins of Streptococcus pyogenes. Seventy-four distinct cell wall-associated proteins were identified, 66 of which were novel. Thirty-three proteins were immunoreactive with pooled S. pyogenes-reactive human antisera. Biotinylation of the GAS cell surface identified 23 cell wall-associated proteins that are surface exposed.

  4. Synthesis of the cell surface during the division cycle of rod-shaped, gram-negative bacteria.

    OpenAIRE

    Cooper, S

    1991-01-01

    When the growth of the gram-negative bacterial cell wall is considered in relation to the synthesis of the other components of the cell, a new understanding of the pattern of wall synthesis emerges. Rather than a switch in synthesis between the side wall and pole, there is a partitioning of synthesis such that the volume of the cell increases exponentially and thus perfectly encloses the exponentially increasing cytoplasm. This allows the density of the cell to remain constant during the divi...

  5. Femtosecond UV-laser pulses to unveil protein-protein interactions in living cells.

    Science.gov (United States)

    Itri, Francesco; Monti, Daria M; Della Ventura, Bartolomeo; Vinciguerra, Roberto; Chino, Marco; Gesuele, Felice; Lombardi, Angelina; Velotta, Raffaele; Altucci, Carlo; Birolo, Leila; Piccoli, Renata; Arciello, Angela

    2016-02-01

    A hallmark to decipher bioprocesses is to characterize protein-protein interactions in living cells. To do this, the development of innovative methodologies, which do not alter proteins and their natural environment, is particularly needed. Here, we report a method (LUCK, Laser UV Cross-linKing) to in vivo cross-link proteins by UV-laser irradiation of living cells. Upon irradiation of HeLa cells under controlled conditions, cross-linked products of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were detected, whose yield was found to be a linear function of the total irradiation energy. We demonstrated that stable dimers of GAPDH were formed through intersubunit cross-linking, as also observed when the pure protein was irradiated by UV-laser in vitro. We proposed a defined patch of aromatic residues located at the enzyme subunit interface as the cross-linking sites involved in dimer formation. Hence, by this technique, UV-laser is able to photofix protein surfaces that come in direct contact. Due to the ultra-short time scale of UV-laser-induced cross-linking, this technique could be extended to weld even transient protein interactions in their native context. PMID:26265182

  6. Comparative proteomics reveals highly and differentially expressed proteins in field-collected and laboratory-cultured blooming cells of the diatom Skeletonema costatum.

    Science.gov (United States)

    Zhang, Hao; Wang, Da-Zhi; Xie, Zhang-Xian; Zhang, Shu-Fei; Wang, Ming-Hua; Lin, Lin

    2015-10-01

    Diatoms are a major phytoplankton group causing extensive blooms in the ocean. However, little is known about the intracellular biological processes occurring during the blooming period. This study compared the protein profiles of field-collected and laboratory-cultured blooming cells of Skeletonema costatum, and identified highly and differentially expressed proteins using the shotgun proteomic approach. A total of 1372 proteins were confidently identified with two or more peptides. Among them, 222 and 311 proteins were unique to the laboratory and field samples respectively. Proteins involved in photosynthesis, translation, nucleosome assembly, carbohydrate and energy metabolism dominated the protein profiles in both samples. However, different features of specific proteins were also found: proteins participated in light harvesting, photosynthetic pigment biosynthesis, photoprotection, cell division and redox homeostasis were highly detected in the field sample, whereas proteins involved in translation, amino acid and protein metabolic processes, and nitrogen and carbon assimilation presented high detection rates in the laboratory sample. ATP synthase cf1 subunit beta and light harvest complex protein were the most abundant protein in the laboratory and field samples respectively. These results indicated that S. costatum had evolved adaptive mechanisms to the changing environment, and integrating field and laboratory proteomic data should provide comprehensive understanding of bloom mechanisms. PMID:26014042

  7. Lineage-specific interface proteins match up the cell cycle and differentiation in embryo stem cells

    DEFF Research Database (Denmark)

    Re, Angela; Workman, Christopher; Waldron, Levi;

    2014-01-01

    The shortage of molecular information on cell cycle changes along embryonic stem cell (ESC) differentiation prompts an in silico approach, which may provide a novel way to identify candidate genes or mechanisms acting in coordinating the two programs. We analyzed germ layer specific gene expression...... changes during the cell cycle and ESC differentiation by combining four human cell cycle transcriptome profiles with thirteen in vitro human ESC differentiation studies. To detect cross-talk mechanisms we then integrated the transcriptome data that displayed differential regulation with protein...... interaction data. A new class of non-transcriptionally regulated genes was identified, encoding proteins which interact systematically with proteins corresponding to genes regulated during the cell cycle or cell differentiation, and which therefore can be seen as interface proteins coordinating the two...

  8. Isoelectric focusing technology quantifies protein signaling in 25 cells

    Science.gov (United States)

    O'Neill, Roger A.; Bhamidipati, Arunashree; Bi, Xiahui; Deb-Basu, Debabrita; Cahill, Linda; Ferrante, Jason; Gentalen, Erik; Glazer, Marc; Gossett, John; Hacker, Kevin; Kirby, Celeste; Knittle, James; Loder, Robert; Mastroieni, Catherine; MacLaren, Michael; Mills, Thomas; Nguyen, Uyen; Parker, Nineveh; Rice, Audie; Roach, David; Suich, Daniel; Voehringer, David; Voss, Karl; Yang, Jade; Yang, Tom; Vander Horn, Peter B.

    2006-01-01

    A previously undescribed isoelectric focusing technology allows cell signaling to be quantitatively assessed in <25 cells. High-resolution capillary isoelectric focusing allows isoforms and individual phosphorylation forms to be resolved, often to baseline, in a 400-nl capillary. Key to the method is photochemical capture of the resolved protein forms. Once immobilized, the proteins can be probed with specific antibodies flowed through the capillary. Antibodies bound to their targets are detected by chemiluminescence. Because chemiluminescent substrates are flowed through the capillary during detection, localized substrate depletion is overcome, giving excellent linearity of response across several orders of magnitude. By analyzing pan-specific antibody signals from individual resolved forms of a protein, each of these can be quantified, without the problems associated with using multiple antibodies with different binding avidities to detect individual protein forms. PMID:17053065

  9. In vivo protein synthesis determinations in human immune cells

    OpenAIRE

    Januszkiewicz, Anna

    2005-01-01

    Intact immune responses are essential for defeating severe infections in individual patients. Insufficient function of the immune system contributes to a poor prognosis in these patients, in particular the ICU patients. Nevertheless, the immune system function is not easily monitored and evaluated. The ongoing metabolic activity of immune competent cells is reflected by their in vivo protein synthesis rate. The aim of this thesis was to apply in vivo protein synthesis measur...

  10. Actinomycetes as host cells for production of recombinant proteins

    OpenAIRE

    Tamura Tomohiro; Mitani Yasuo; Nakashima Nobutaka

    2005-01-01

    Abstract Actinomycetes (Actinobacteria) are highly attractive as cell factories or bioreactors for applications in industrial, agricultural, environmental, and pharmaceutical fields. Genome sequencing of several species of actinomycetes has paved the way for biochemical and structural analysis of important proteins and the production of such proteins as recombinants on a commercial scale. In this regard, there is a need for improved expression vectors that will be applicable to actinomycetes....

  11. Cell-free protein expression based on extracts from CHO cells.

    Science.gov (United States)

    Brödel, Andreas K; Sonnabend, Andrei; Kubick, Stefan

    2014-01-01

    Protein expression systems are widely used in biotechnology and medicine for the efficient and economic production of therapeutic proteins. Today, cultivated Chinese hamster ovary (CHO) cells are the market dominating mammalian cell-line for the production of complex therapeutic proteins. Despite this outstanding potential of CHO cells, no high-yield cell-free system based on translationally active lysates from these cells has been reported so far. To date, CHO cell extracts have only been used as a foundational research tool for understanding mRNA translation (Lodish et al., 1974; McDowell et al., 1972). In the present study, we address this fact by establishing a novel cell-free protein expression system based on extracts from cultured CHO cells. Lysate preparation, adaptation of in vitro reaction conditions and the construction of particular expression vectors are considered for high-yield protein production. A specific in vitro expression vector, which includes an internal ribosome entry site (IRES) from the intergenic region (IGR) of the Cricket paralysis virus (CrPV), has been constructed in order to obtain optimal performance. The IGR IRES is supposed to bind directly to the eukaryotic 40S ribosomal subunit thereby bypassing the process of translation initiation, which is often a major bottleneck in cell-free systems. The combination of expression vector and optimized CHO cell extracts enables the production of approximately 50 µg/mL active firefly luciferase within 4 h. The batch-type cell-free coupled transcription-translation system has the potential to perform post-translational modifications, as shown by the glycosylation of erythropoietin. Accordingly, the system contains translocationally active endogenous microsomes, enabling the co-translational incorporation of membrane proteins into biological membranes. Hence, the presented in vitro translation system is a powerful tool for the fast and convenient optimization of expression constructs, the

  12. Determinants of Cell-to-Cell Variability in Protein Kinase Signaling

    OpenAIRE

    Matthias Jeschke; Stephan Baumgärtner; Stefan Legewie

    2013-01-01

    Cells reliably sense environmental changes despite internal and external fluctuations, but the mechanisms underlying robustness remain unclear. We analyzed how fluctuations in signaling protein concentrations give rise to cell-to-cell variability in protein kinase signaling using analytical theory and numerical simulations. We characterized the dose-response behavior of signaling cascades by calculating the stimulus level at which a pathway responds ('pathway sensitivity') and the maximal act...

  13. MicroRNA-194 restrains the cell progression of non-small cell lung cancer by targeting human nuclear distribution protein C.

    Science.gov (United States)

    Zhou, Lirong; Di, Qingguo; Sun, Baohua; Wang, Xiaosheng; Li, Min; Shi, Jian

    2016-06-01

    NSCLC accounts for over 80% of all lung cancers and is associated with poor prognosis. Human nuclear distribution C (hNUDC) was predicted to be the target gene of microRNA-194 (miR-194). The present study was designed to demonstrate the mechanism of miR-194 in the regulation of non-small cell lung cancer (NSCLC) via targeting the hNUDC. The hNUDC expression was found to strongly be increased while the miR-194 decreased significantly in the NSCLC cell lines when compared with the healthy controls. Moreover, the luciferase report confirmed the targeting reaction between miR-194 and hNUDC. After transfection with miR-194 mimic into NSCLC cells, we found that the miR-194 overexpression resulted in abnormal nuclear division, decreased cell proliferation and inhibited the expression of hNUDC and Mpl/ERK pathway proteins. Furthermore, the hNUDC overexpression affected the suppression effect of miR-194 in 95D cells, indicating that miR-194 suppresses tumor cell process by inhibiting the hNUDC expression. In brief, the present study suggests that the upregulation of miR-194 affects the hNUDC expression, leading to a downregulated expression of Mpl/ERK pathway proteins, and suppresses the mitosis and proliferation of NSCLC cells. These results offer a potential therapeutic strategy for the treatment of lung cancer. PMID:27035759

  14. MICROORGANISMS: A MARVELOUS SOURCE OF SINGLE CELL PROTEINS

    Directory of Open Access Journals (Sweden)

    Agam Nangul

    2013-08-01

    Full Text Available The increasing global population living below the poverty line is driving the scientific community to search for non-conventional protein sources that can replace conventional expensive ones. Microbial proteins, or single-cell protein (SCP, represent a potential future nutrient source for human food and animal feed. These microbial proteins can be grown rapidly on substrates with minimum dependence on soil, water and climate conditions. They can be produced from algae, fungi and bacteria the chief sources of SCP. It is convenient to use microorganisms for production of SCP as they grow rapidly and have high protein content. Industrially, they can be produced from algal biomass, yeast, fungi. There are several other ways of getting SCP as well. Despite numerous advantages of SCP, they have disadvantages and toxic effects too, especially related to mycotoxins and bacterial toxins.

  15. Effect of space flight on the frequency of micronuclei and expression of stress-responsive proteins in cultured mammalian cells

    Energy Technology Data Exchange (ETDEWEB)

    Ikenaga, Mituo; Hirayama, Jun; Kato, Tomohisa [Kyoto Univ. (Japan). Radiation Biology Center] [and others

    2002-12-01

    Results of past space experiments suggest that the biological effect of space radiation could been hanced under microgravity in some cases, especially ininsects. To examine if such a synergistic effect of radiation and microgravity also exists in human cells, frequencies of chromosome instability and cellular levels of several stress-responsive proteins were analyzed incultured human and rodent cells afterspace flight. Human (MCF7 and ataxia telangiectasia(AT)2KY), mouse (m5S) and hamster (Syrian hamster embryo (SHE)) cell lines were loaded on the Space Shuttle Discovery (STS-95 mission) and grown during a 9-daymission. After landing, the micronuclei resulting from abnormal nuclear division and accumulationof stress-responsive proteins such as p53 and mitogen-activated protein kinases (MAPKs), which are involved in radiation-induced signal transduction cascades, were analyzed. The frequencies of micronucleiin all the four mammalian cell strains tested were not significantly different between flight and ground control samples. Also, the cellular amounts of p53, p21 (WAF1/SDI1/CIP1) and activated (phosphorylated) forms of three distinct MAPKs in MCF7 and m5S cells of flight samples were similar to those of ground control samples. These results indicated that anyeffect of space radiation, microgravity, or combination of both were not detectable, at least under thepresent experimental conditions. (author)

  16. Effect of space flight on the frequency of micronuclei and expression of stress-responsive proteins in cultured mammalian cells

    International Nuclear Information System (INIS)

    Results of past space experiments suggest that the biological effect of space radiation could been hanced under microgravity in some cases, especially ininsects. To examine if such a synergistic effect of radiation and microgravity also exists in human cells, frequencies of chromosome instability and cellular levels of several stress-responsive proteins were analyzed incultured human and rodent cells afterspace flight. Human (MCF7 and ataxia telangiectasia(AT)2KY), mouse (m5S) and hamster (Syrian hamster embryo (SHE)) cell lines were loaded on the Space Shuttle Discovery (STS-95 mission) and grown during a 9-daymission. After landing, the micronuclei resulting from abnormal nuclear division and accumulationof stress-responsive proteins such as p53 and mitogen-activated protein kinases (MAPKs), which are involved in radiation-induced signal transduction cascades, were analyzed. The frequencies of micronucleiin all the four mammalian cell strains tested were not significantly different between flight and ground control samples. Also, the cellular amounts of p53, p21 (WAF1/SDI1/CIP1) and activated (phosphorylated) forms of three distinct MAPKs in MCF7 and m5S cells of flight samples were similar to those of ground control samples. These results indicated that anyeffect of space radiation, microgravity, or combination of both were not detectable, at least under thepresent experimental conditions. (author)

  17. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins

    DEFF Research Database (Denmark)

    Weber, Daniela; Davies, Michael J.; Grune, Tilman

    2015-01-01

    the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the...... different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in...... spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples....

  18. Proteins present in tannin cenocyte mother cells in Sambucus racemosa L.

    OpenAIRE

    Zobel, A M

    2015-01-01

    It has been demonstrated that protein content and concentration are higher in mononucleate tannin-cells than in the parenchyma cells of Sambucus racemosa. Cytoplasmic and nuclear free acid proteins markedly prevail here. It is believed that they may be enzymatic proteins. Increase of acid proteins content within the nucleus of tannin cells causes an increase of the nucleus size. The content of nuclear bound basic proteins in tannin cells, may be lower than in the neighbouring parenchymal cells.

  19. Histochemical approaches to assess cell-to-cell transmission of misfolded proteins in neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    G. Natale

    2013-03-01

    Full Text Available Formation, aggregation and transmission of abnormal proteins are common features in neurodegenerative disorders including Parkinson’s disease, Alzheimer’s disease, amyotrophic lateral sclerosis, and Huntington’s disease. The mechanisms underlying protein alterations in neurodegenerative diseases remain controversial. Novel findings highlighted altered protein clearing systems as common biochemical pathways which generate protein misfolding, which in turn causes protein aggregation and protein spreading. In fact, proteinaceous aggregates are prone to cell-to-cell propagation. This is reminiscent of what happens in prion disorders, where the prion protein misfolds thus forming aggregates which spread to neighbouring cells. For this reason, the term prionoids is currently used to emphasize how several misfolded proteins are transmitted in neurodegenerative diseases following this prion-like pattern. Histochemical techniques including the use of specific antibodies covering both light and electron microscopy offer a powerful tool to describe these phenomena and investigate specific molecular steps. These include: prion like protein alterations; glycation of prion-like altered proteins to form advanced glycation end-products (AGEs; mechanisms of extracellular secretion; interaction of AGEs with specific receptors placed on neighbouring cells (RAGEs. The present manuscript comments on these phenomena aimed to provide a consistent scenario of the available histochemical approaches to dissect each specific step.

  20. Cell kinetic effects of incorporated 3H-thymidine on proliferating human lymphocytes: flow cytometric analysis using the DNA/nuclear protein method

    International Nuclear Information System (INIS)

    Phytohemagglutinin-stimulated human peripheral blood lymphocytes incorporating high concentrations of 3H-thymidine accumulate in G2 and show a consequent reduction in the number of cells entering M (division delay). The simultaneous flow cytometric analysis of DNA content (propidium iodide fluorescence) and nuclear protein content (fluorescein isothiocyanate fluorescence) allows for the accurate quantitation of these events; G2 and M are separated in the bivariate distributions. A good correlation was observed between mitotic indices, quantitated by manually counting mitotic cells, and integration of the M area in DNA/nuclear protein histograms. Moreover, significant differences in G2 nuclear protein levels were found between untreated and 3H-thymidine-treated lymphocytes. In order to characterize this effect, G2 was empirically divided into low nuclear protein (G2A) and high nuclear protein (G2B) compartments. 3H-thymidine caused an initial accumulation of lymphocytes in G2A, followed within 3-6 h by a gradual movement of some cells into G2B, with a subsequent accumulation of cells in G2B. The results suggest that the distribution of cells in G2 (G2A and G2B), the average nuclear protein content of G2B cells, and the proportion of cells in M are parameters that when used in combination provide a unique description of radiobiological effects

  1. Two endogenous proteins that induce cell wall extension in plants

    Science.gov (United States)

    McQueen-Mason, S.; Durachko, D. M.; Cosgrove, D. J.

    1992-01-01

    Plant cell enlargement is regulated by wall relaxation and yielding, which is thought to be catalyzed by elusive "wall-loosening" enzymes. By employing a reconstitution approach, we found that a crude protein extract from the cell walls of growing cucumber seedlings possessed the ability to induce the extension of isolated cell walls. This activity was restricted to the growing region of the stem and could induce the extension of isolated cell walls from various dicot stems and the leaves of amaryllidaceous monocots, but was less effective on grass coleoptile walls. Endogenous and reconstituted wall extension activities showed similar sensitivities to pH, metal ions, thiol reducing agents, proteases, and boiling in methanol or water. Sequential HPLC fractionation of the active wall extract revealed two proteins with molecular masses of 29 and 30 kD associated with the activity. Each protein, by itself, could induce wall extension without detectable hydrolytic breakdown of the wall. These proteins appear to mediate "acid growth" responses of isolated walls and may catalyze plant cell wall extension by a novel biochemical mechanism.

  2. Genetically Encoded Molecular Tension Probe for Tracing Protein-Protein Interactions in Mammalian Cells.

    Science.gov (United States)

    Kim, Sung Bae; Nishihara, Ryo; Citterio, Daniel; Suzuki, Koji

    2016-02-17

    Optical imaging of protein-protein interactions (PPIs) facilitates comprehensive elucidation of intracellular molecular events. We demonstrate an optical measure for visualizing molecular tension triggered by any PPI in mammalian cells. Twenty-three kinds of candidate designs were fabricated, in which a full-length artificial luciferase (ALuc) was sandwiched between two model proteins of interest, e.g., FKBP and FRB. One of the designs greatly enhanced the bioluminescence in response to varying concentrations of rapamycin. It is confirmed with negative controls that the elevated bioluminescence is solely motivated from the molecular tension. The probe design was further modified toward eliminating the C-terminal end of ALuc and was found to improve signal-to-background ratios, named "a combinational probe". The utilities were elucidated with detailed substrate selectivity, bioluminescence imaging of live cells, and different PPI models. This study expands capabilities of luciferases as a tool for analyses of molecular dynamics and cell signaling in living subjects. PMID:26322739

  3. Cell-Free Expression of Protein Kinase A for Rapid Activity Assays

    OpenAIRE

    Leippe, Donna M.; Kate Qin Zhao; Kevin Hsiao; Slater, Michael R.

    2010-01-01

    Functional protein analysis often calls for lengthy, laborious in vivo protein expression and purification, and can be complicated by the lack of stability of the purified protein. In this study, we demonstrate the feasibility of a simplified procedure for functional protein analysis on magnetic particles using cell-free protein synthesis of the catalytic subunit of human cAMP-dependent protein kinase as a HaloTag® fusion protein. The cell-free protein synthesis systems provide quick access t...

  4. Molecular evolution in bacteria: cell division Evolução molecular em bactérias: divisão celular

    Directory of Open Access Journals (Sweden)

    J.T. Trevors

    1998-10-01

    Full Text Available Molecular evolution in bacteria is examined with an emphasis on the self-assembly of cells capable of primitive division and growth during early molecular evolution. Also, the possibility that some type of encapsulation structure preceeded biochemical pathways and the assembly of genetic material is examined. These aspects will be considered from an evolutionary perspective.A evolução molecular em bactérias é examinada com ênfase na auto-organização de uma célula capaz de divisão primitiva e multiplicação durante o princípio da evolução molecular. Também se discute a possibilidade de que algum tipo de estrutura de encapsulação tenha antecedido as vias bioquímicas e o agrupamento de material genético. Esses aspectos são considerados sob uma perspectiva evolutiva.

  5. Heterotrimeric G proteins precouple with G protein-coupled receptors in living cells

    OpenAIRE

    Nobles, M.; Benians, A.; Tinker, A

    2005-01-01

    Using fluorescence resonance energy transfer (FRET) microscopy, we investigate how heterotrimeric G proteins interact with G protein-coupled receptors (GPCRs). In the absence of receptor activation, the alpha 2A adrenergic and muscarinic M4 receptors are present on the cell membrane as dimers. Furthermore, there is an interaction between the G protein subunits alpha o, beta 1, and gamma 2 and a number of GPCRs including M4, a2A, the adenosine All receptor, and the dopamine D2 receptor under r...

  6. Identification of a cell membrane protein that binds alveolar surfactant.

    OpenAIRE

    Strayer, D. S.

    1991-01-01

    Alveolar surfactants are complex mixtures of proteins and phospholipids produced by type II alveolar cells and responsible for lowering pulmonary surface tension. The process by which surfactant is produced and exported and by which its production by pulmonary cells is regulated are not well understood. This study was designed to identify a cellular receptor for surfactant constituents. To do so, monoclonal anti-idiotypic antibodies directed against antibodies to porcine and rabbit surfactant...

  7. Mechanism Suggests How HIV Protein Disrupts Immune Cell Migration

    OpenAIRE

    Janardhan Ajit; Swigut Tomek; Hill Brian; Myers Michael P; Skowronski Jacek

    2004-01-01

    The infectious cycle of primate lentiviruses is intimately linked to interactions between cells of the immune system. Nef, a potent virulence factor, alters cellular environments to increase lentiviral replication in the host, yet the mechanisms underlying these effects have remained elusive. Since Nef likely functions as an adaptor protein, we exploited a proteomic approach to directly identify molecules that Nef targets to subvert the signaling machinery in T cells. We purified to near homo...

  8. Multistage Magnetic Separator of Cells and Proteins

    Science.gov (United States)

    Barton, Ken; Ainsworth, Mark; Daily, Bruce; Dunn, Scott; Metz, Bill; Vellinger, John; Taylor, Brock; Meador, Bruce

    2005-01-01

    The multistage electromagnetic separator for purifying cells and magnetic particles (MAGSEP) is a laboratory apparatus for separating and/or purifying particles (especially biological cells) on the basis of their magnetic susceptibility and magnetophoretic mobility. Whereas a typical prior apparatus based on similar principles offers only a single stage of separation, the MAGSEP, as its full name indicates, offers multiple stages of separation; this makes it possible to refine a sample population of particles to a higher level of purity or to categorize multiple portions of the sample on the basis of magnetic susceptibility and/or magnetophoretic mobility. The MAGSEP includes a processing unit and an electronic unit coupled to a personal computer. The processing unit includes upper and lower plates, a plate-rotation system, an electromagnet, an electromagnet-translation system, and a capture-magnet assembly. The plates are bolted together through a roller bearing that allows the plates to rotate with respect to each other. An interface between the plates acts as a seal for separating fluids. A lower cuvette can be aligned with as many as 15 upper cuvette stations for fraction collection during processing. A two-phase stepping motor drives the rotation system, causing the upper plate to rotate for the collection of each fraction of the sample material. The electromagnet generates a magnetic field across the lower cuvette, while the translation system translates the electromagnet upward along the lower cuvette. The current supplied to the electromagnet, and thus the magnetic flux density at the pole face of the electromagnet, can be set at a programmed value between 0 and 1,400 gauss (0.14 T). The rate of translation can be programmed between 5 and 2,000 m/s so as to align all sample particles in the same position in the cuvette. The capture magnet can be a permanent magnet. It is mounted on an arm connected to a stepping motor. The stepping motor rotates the arm to

  9. Intercellular Protein Transfer from Thymocytes to Thymic Epithelial Cells

    Science.gov (United States)

    Wang, Hong-Xia; Qiu, Yu-Rong; Zhong, Xiao-Ping

    2016-01-01

    Promiscuous expression of tissue restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs) is crucial for negative selection of self-reactive T cells to establish central tolerance. Intercellular transfer of self-peptide-MHC complexes from mTECs to thymic dendritic cells (DCs) allows DCs to acquire TRAs, which in turn contributes to negative selection and regulatory T cell generation. However, mTECs are unlikely to express all TRAs, such as immunoglobulins generated only in B cells after somatic recombination, hyper-mutation, or class-switches. We report here that both mTECs and cortical TECs can efficiently acquire not only cell surface but also intracellular proteins from thymocytes. This reveals a previously unappreciated intercellular sharing of molecules from thymocytes to TECs, which may broaden the TRA inventory in mTECs for establishing a full spectrum of central tolerance. PMID:27022746

  10. Comparison of cell wall proteins of Saccharomyces cerevisiae as anchors for cell surface expression of heterologous proteins.

    Science.gov (United States)

    Van der Vaart, J M; te Biesebeke, R; Chapman, J W; Toschka, H Y; Klis, F M; Verrips, C T

    1997-01-01

    The carboxyl-terminal regions of five cell wall proteins (Cwp1p, Cwp2p, Ag alpha 1p, Tip1p, and Flo1p) and three potential cell wall proteins (Sed1p, YCR89w, and Tir1p) all proved capable of immobilizing alpha-galactosidase in the cell wall of Saccharomyces cerevisiae. The fraction of the total amount of fusion protein that was localized to the cell wall varied depending on the anchor domain used. The highest proportion of cell wall incorporation was achieved with Cwp2p, Ag alpha 1p, or Sed1p as an anchor. Although 80% of these fusion proteins were incorporated in the cell wall, the total production of alpha-galactosidase-Ag alpha 1p was sixfold lower than that of alpha-galactosidase-Cwp2p and eightfold lower than that of alpha-galactosidase-Sed1p. Differences in mRNA levels were not responsible for this discrepancy, nor was an intracellular accumulation of alpha-galactosidase-Ag alpha 1p detectable. A lower translation efficiency of the alpha-galactosidase-AG alpha 1 fusion construct is most likely to be responsible for the low level of protein production. alpha-Galactosidase immobilized by the carboxyl-terminal 67 amino acids of Cwp2p was most effective in the hydrolysis of the high-molecular-weight substrate guar gum from Cyamopsis tetragonoloba. This indicates that the use of a large anchoring domain does not necessarily result in a better exposure of the immobilized enzyme to the exterior of the yeast cell. PMID:9023939

  11. Muscle Stem Cell Fate Is Controlled by the Cell-Polarity Protein Scrib

    Directory of Open Access Journals (Sweden)

    Yusuke Ono

    2015-02-01

    Full Text Available Satellite cells are resident skeletal muscle stem cells that supply myonuclei for homeostasis, hypertrophy, and repair in adult muscle. Scrib is one of the major cell-polarity proteins, acting as a potent tumor suppressor in epithelial cells. Here, we show that Scrib also controls satellite-cell-fate decisions in adult mice. Scrib is undetectable in quiescent cells but becomes expressed during activation. Scrib is asymmetrically distributed in dividing daughter cells, with robust accumulation in cells committed to myogenic differentiation. Low Scrib expression is associated with the proliferative state and preventing self-renewal, whereas high Scrib levels reduce satellite cell proliferation. Satellite-cell-specific knockout of Scrib in mice causes a drastic and insurmountable defect in muscle regeneration. Thus, Scrib is a regulator of tissue stem cells, controlling population expansion and self-renewal with Scrib expression dynamics directing satellite cell fate.

  12. Geminin deploys multiple mechanisms to regulate Cdt1 before cell division thus ensuring the proper execution of DNA replication

    DEFF Research Database (Denmark)

    Ballabeni, Andrea; Zamponi, Raffaella; Moore, Jodene K;

    2013-01-01

    accumulation of Cdt1 in mitosis, because decreasing the Geminin levels prevents Cdt1 accumulation and impairs DNA replication. Geminin is known to inhibit Cdt1 function; its depletion during G2 leads to DNA rereplication and checkpoint activation. Here we show that, despite rapid Cdt1 protein turnover in G2...

  13. The Seckel syndrome and centrosomal protein Ninein localizes asymmetrically to stem cell centrosomes but is not required for normal development, behavior, or DNA damage response in Drosophila.

    Science.gov (United States)

    Zheng, Yiming; Mennella, Vito; Marks, Steven; Wildonger, Jill; Elnagdi, Esraa; Agard, David; Megraw, Timothy L

    2016-06-01

    Ninein (Nin) is a centrosomal protein whose gene is mutated in Seckel syndrome (SCKL, MIM 210600), an inherited recessive disease that results in primordial dwarfism, cognitive deficiencies, and increased sensitivity to genotoxic stress. Nin regulates neural stem cell self-renewal, interkinetic nuclear migration, and microtubule assembly in mammals. Nin is evolutionarily conserved, yet its role in cell division and development has not been investigated in a model organism. Here we characterize the single Nin orthologue in Drosophila Drosophila Nin localizes to the periphery of the centrosome but not at centriolar structures as in mammals. However, Nin shares the property of its mammalian orthologue of promoting microtubule assembly. In neural and germline stem cells, Nin localizes asymmetrically to the younger (daughter) centrosome, yet it is not required for the asymmetric division of stem cells. In wing epithelia and muscle, Nin localizes to noncentrosomal microtubule-organizing centers. Surprisingly, loss of nin expression from a nin mutant does not significantly affect embryonic and brain development, fertility, or locomotor performance of mutant flies or their survival upon exposure to DNA-damaging agents. Although it is not essential, our data suggest that Nin plays a supportive role in centrosomal and extracentrosomal microtubule organization and asymmetric stem cell division. PMID:27053665

  14. In vivo Biotinylation Based Method for the Study of Protein-Protein Proximity in Eukaryotic Cells

    Directory of Open Access Journals (Sweden)

    Arman Kulyyassov

    2014-01-01

    Full Text Available Introduction: The spatiotemporal order plays an important role in cell functioning and is affected in many pathologies such as cancer and neurodegenerative diseases. One of the ultimate goals of molecular biology is reconstruction of the spatiotemporal structure of a living cell at the molecular level. This task includes determination of proximities between different molecular components in the cell and monitoring their time- and physiological state-dependent changes. In many cases, proximity between macromolecules arises due to their interactions; however, the contribution of dynamic self-organization in generation of spatiotemporal order is emerging as another viable possibility. Specifically, in proteomics, this implies that the detection of protein-protein proximity is a more general task than gaining information about physical interactions between proteins, as it could detail aspects of spatial order in vivo that are challenging to reconstitute in binding experiments in vitro. Methods: In this work, we have developed a method of monitoring protein-protein proximity in vivo. For this purpose, the BirA was fused to one of the interaction partners, whereas the BAP was modified to make the detection of its biotinylation possible by mass spectrometry. Results: Using several experimental systems, we showed that the biotinylation is interaction dependent. In addition, we demonstrated that BAP domains with different primary amino acid structures and thus with different molecular weights can be used in the same experiment, providing the possibility of multiplexing. Alternatively to the changes in primary amino acid structure, the stable isotope format can also be used, providing another way to perform multiplexing experiments. Finally, we also demonstrated that our system could help to overcome another limitation of current methodologies to detect protein-protein proximity. For example, one can follow the state of a protein of interest at a defined

  15. Detection of cytoskeletal proteins in small cell lung carcinoma

    Czech Academy of Sciences Publication Activity Database

    Hložánková, M.; Lukáš, Z.; Viklický, Vladimír

    1999-01-01

    Roč. 18, - (1999), s. 47-49. ISSN 0231-5882 Grant ostatní: MŠk1(CZ) OE10a/EU1450 Keywords : cytoskeletal proteins * small cell lung carcinoma Subject RIV: EI - Biotechnology ; Bionics Impact factor: 0.400, year: 1999

  16. Protein Phosphorylation and Redox Modification in Stomatal Guard Cells.

    Science.gov (United States)

    Balmant, Kelly M; Zhang, Tong; Chen, Sixue

    2016-01-01

    Post-translational modification (PTM) is recognized as a major process accounting for protein structural variation, functional diversity, and the dynamics and complexity of the proteome. Since PTMs can change the structure and function of proteins, they are essential to coordinate signaling networks and to regulate important physiological processes in eukaryotes. Plants are constantly challenged by both biotic and abiotic stresses that reduce productivity, causing economic losses in crops. The plant responses involve complex physiological, cellular, and molecular processes, with stomatal movement as one of the earliest responses. In order to activate such a rapid response, stomatal guard cells employ cellular PTMs of key protein players in the signaling pathways to regulate the opening and closure of the stomatal pores. Here we discuss two major types of PTMs, protein phosphorylation and redox modification that play essential roles in stomatal movement under stress conditions. We present an overview of PTMs that occur in stomatal guard cells, especially the methods and technologies, and their applications in PTM identification and quantification. Our focus is on PTMs that modify molecular components in guard cell signaling at the stages of signal perception, second messenger production, as well as downstream signaling events and output. Improved understanding of guard cell signaling will enable generation of crops with enhanced stress tolerance, and increased yield and bioenergy through biotechnology and molecular breeding. PMID:26903877

  17. Effects of Static Magnetic Field on Growth of Leptospire, Leptospira interrogans serovar canicola: Immunoreactivity and Cell Division

    CERN Document Server

    Triampo, W; Triampo, D; Wong-Ekkabut, J; Tang, I M; Triampo, Wannapong; Doungchawee, Galayanee; Triampo, Darapond; Wong-Ekkabut, Jirasak

    2004-01-01

    The effects of the exposure of the bacterium, Leptospira interrogans serovar canicola to a constant magnetic field with magnetic flux density from a permanent ferrite magnet = 140 mT were studied. Changes in Leptospira cells after their exposure to the field were determined on the basis of changes in their growth behavior and agglutination immunoreactivity with a homologous antiserum using darkfield microscopy together with visual imaging. The data showed that the exposed Leptospira cells have lower densities and lower agglutination immunoreactivity than the unexposed control group. Interestingly, some of the exposed Leptospira cells showed abnormal morphologies such as large lengths. We discussed some of the possible reasons for these observations.

  18. Germ cells may survive clipping and division of the spermatic vessels in surgery for intra-abdominal testes

    DEFF Research Database (Denmark)

    Thorup, J M; Cortes, Dina; Visfeldt, J

    1999-01-01

    Laparoscopy is a well described modality that provides an accurate visual diagnosis upon which further management of intra-abdominal testes may be based. Laparoscopic ligation of spermatic vessels as stage 1 of the procedure is a natural extension of laparoscopy. A staged approach provides adequate...... viability of the intra-abdominal testis. However, it is uncertain whether the more sensitive germ cells survive this procedure in addition to the Sertoli and interstitial cells of the human testis. Survival of germ cells is a prerequisite of later fertility potential....

  19. Histochemical approaches to assess cell-to-cell transmission of misfolded proteins in neurodegenerative diseases

    Science.gov (United States)

    Natale, G.; Pompili, E.; Biagioni, F.; Paparelli, S.; Lenzi, P.; Fornai, F.

    2013-01-01

    Formation, aggregation and transmission of abnormal proteins are common features in neurodegenerative disorders including Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, and Huntington's disease. The mechanisms underlying protein alterations in neurodegenerative diseases remain controversial. Novel findings highlighted altered protein clearing systems as common biochemical pathways which generate protein misfolding, which in turn causes protein aggregation and protein spreading. In fact, proteinaceous aggregates are prone to cell-tocell propagation. This is reminiscent of what happens in prion disorders, where the prion protein misfolds thus forming aggregates which spread to neighbouring cells. For this reason, the term prionoids is currently used to emphasize how several misfolded proteins are transmitted in neurodegenerative diseases following this prion-like pattern. Histochemical techniques including the use of specific antibodies covering both light and electron microscopy offer a powerful tool to describe these phenomena and investigate specific molecular steps. These include: prion like protein alterations; glycation of prion-like altered proteins to form advanced glycation end-products (AGEs); mechanisms of extracellular secretion; interaction of AGEs with specific receptors placed on neighbouring cells (RAGEs). The present manuscript comments on these phenomena aimed to provide a consistent scenario of the available histochemical approaches to dissect each specific step. PMID:23549464

  20. Buffalo milk: proteins electrophoretic profile and somatic cell count

    Directory of Open Access Journals (Sweden)

    S. Mattii

    2011-03-01

    Full Text Available Water buffalo milk differs from the cow’s milk for greater fat and protein content, very important features in cheese making. Proteins, casein and whey-proteins in particular, are the most important factors determining cheese yield. Several previous research discussed the rule of SCC in cow milk production (Varisco, 1999 and the close relationship existing between cow’s milk cheese yield and somatic cell count (Barbano, 2000. In particular the inverse correlation between cheese yields and somatic cells’content have been demonstrated. In Italy the regulation in force DPR 54/97 acknowledges what expressed in EEC 46/92 Directive (Tripodi, 1999 without fixing the limit threshold of somatic cells for buffalo’s milk....

  1. Simultaneous detection of mRNA and protein stem cell markers in live cells

    Directory of Open Access Journals (Sweden)

    Bao Gang

    2009-04-01

    Full Text Available Abstract Background Biological studies and medical application of stem cells often require the isolation of stem cells from a mixed cell population, including the detection of cancer stem cells in tumor tissue, and isolation of induced pluripotent stem cells after eliciting the expression of specific genes in adult cells. Here we report the detection of Oct-4 mRNA and SSEA-1 protein in live carcinoma stem cells using respectively molecular beacon and dye-labeled antibody, aiming to establish a new method for stem cells detection and isolation. Results Quantification of Oct-4 mRNA and protein in P19 mouse carcinoma stem cells using respectively RT-PCR and immunocytochemistry confirmed that their levels drastically decreased after differentiation. To visualize Oct-4 mRNA in live stem cells, molecular beacons were designed, synthesized and validated, and the detection specificity was confirmed using control studies. We found that the fluorescence signal from Oct-4-targeting molecular beacons provides a clear discrimination between undifferentiated and retinoic acid-induced differentiated cells. Using deconvolution fluorescence microscopy, Oct-4 mRNAs were found to reside on one side of the cytosol. We demonstrated that, using a combination of Oct-4 mRNA-targeting molecular beacon with SSEA-1 antibody in flow cytometric analysis, undifferentiated stem cells can be clearly distinguished from differentiated cells. We revealed that Oct-4 targeting molecular beacons do not seem to affect stem cell biology. Conclusion Molecular beacons have the potential to provide a powerful tool for highly specific detection and isolation of stem cells, including cancer stem cells and induced pluripotent stem (iPS cells without disturbing cell physiology. It is advantageous to perform simultaneous detection of intracellular (mRNA and cell-surface (protein stem cell markers in flow cytometric analysis, which may lead to high detection sensitivity and efficiency.

  2. Translational control in Drosophila female germline stem cell asymmetric division%mRNA翻译水平调控参与果蝇生殖干细胞不对称分裂的机制研究

    Institute of Scientific and Technical Information of China (English)

    夏来新; 陈大华

    2013-01-01

    Germline stem cells have the sole character of transmitting their genetic information to the next generation. Drosophila female germline stem cells are a very powerful and sophisticated system to study stem cell asymmetric division behavior, which is controlled by the niche signal. These stem cells are regulated at many different levels including transcriptional level, post-transcriptional level, translational level and protein level. The study from the last two decades shows that many RNA binding proteins mutants in Drosophila have either germline stem cell loss or over-proliferation phenotypes. Here, this review tried to summarize the relationship between translational control and germline stem cell fate determination, especially focused on recent progresses in the field. And we also discuss the probable molecular detailed mechanisms involved in the process.%生殖干细胞是唯一能够将遗传物质传给下一代的成体干细胞.果蝇的雌性生殖干细胞是一个非常成熟的用来研究干细胞自身不对称分裂的系统,来自微环境的信号控制其增殖和分化.果蝇的生殖干细胞受到转录水平、转录后水平、mRNA翻译水平和蛋白质水平的精确调控,多年来的研究表明很多翻译相关的RNA结合蛋白的突变都会导致果蝇的生殖干细胞提前分化或者不分化.综述了mRNA翻译水平调控与果蝇生殖干细胞命运决定的关系,特别是该领域近年来的一些重大进展,并讨论了翻译水平调控参与果蝇生殖干细胞命运决定的可能具体分子机制.

  3. Effects of Static Magnetic Field on Growth of Leptospire, Leptospira interrogans serovar canicola: Immunoreactivity and Cell Division

    OpenAIRE

    Triampo, Wannapong; Doungchawee, Galayanee; Triampo, Darapond; Wong-Ekkabut, Jirasak; Tang, I-Ming

    2004-01-01

    The effects of the exposure of the bacterium, Leptospira interrogans serovar canicola to a constant magnetic field with magnetic flux density from a permanent ferrite magnet = 140 mT were studied. Changes in Leptospira cells after their exposure to the field were determined on the basis of changes in their growth behavior and agglutination immunoreactivity with a homologous antiserum using darkfield microscopy together with visual imaging. The data showed that the exposed Leptospira cells hav...

  4. 31P NMR studies of intracellular pH and phosphate metabolism during cell division cycle of Saccharomyces cerevisiae.

    OpenAIRE

    Gillies, R.J.; Ugurbil, K; den Hollander, J A; Shulman, R G

    1981-01-01

    We have analyzed changes in intracellular pH and phosphate metabolism during the cell cycle of Saccharomyces cerevisiae (NCYC 239) by using high-resolution 31P NMR spectroscopy. High-density yeast cultures (2 x 10(8) cells per ml) were arrested prior to "start" by sequential glucose deprivation, after which they synchronously replicated DNA and divided after a final glucose feeding. Oxygenation of arrested cultures in the absence of glucose led to increased levels of sugar phosphates and ATP ...

  5. Study of protein metabolism and cell proliferation using 15N

    International Nuclear Information System (INIS)

    Investigations of nitrogen and protein metabolism with the stable isotope 15N were carried out in 11 patients with arteriosclerosis and 7 healthy controls. After oral application of 3 g 15NH4Cl (95 At% 15N) per 70 kg body weight the incorporation of the isotope 15N in plasma proteins and blood cells and the 15N elimination in urine were followed up. Retardations of 15N elimination, an accelerated incorporation of 15N in fibrin and a retarded 15N incorporation in platelet protein were observed in patients with arteriosclerosis. The described method enables complex assertions about protein metabolism of the whole body and so represents a possibility to evaluate objectively the influence of an intervention on metabolism. (author)

  6. Enhanced cell-permeant Cre protein for site-specific recombination in cultured cells

    Directory of Open Access Journals (Sweden)

    Ruley H Earl

    2004-10-01

    Full Text Available Abstract Background Cell-permeant Cre DNA site-specific recombinases provide an easily controlled means to regulate gene structure and function in living cells. Since recombination provides a stable and unambiguous record of protein uptake, the enzyme may also be used for quantitative studies of cis- and trans-acting factors that influence the delivery of proteins into cells. Results In the present study, 11 recombinant fusion proteins were analyzed to characterize sequences and conditions that affect protein uptake and/or activity and to develop more active cell-permeant enzymes. We report that the native enzyme has a low, but intrinsic ability to enter cells. The most active Cre proteins tested contained either an N-terminal 6xHis tag and a nuclear localization sequence from SV40 large T antigen (HNC or the HIV Tat transduction sequence and a C-terminal 6xHis tag (TCH6. The NLS and 6xHis elements separately enhanced the delivery of the HNC protein into cells; moreover, transduction sequences from fibroblast growth factor 4, HIV Tat or consisting of the (KFF3K sequence were not required for efficient protein transduction and adversely affected enzyme solubility. Transduction of the HNC protein required 10 to 15 min for half-maximum uptake, was greatly decreased at 4°C and was inhibited by serum. Efficient recombination was observed in all cell types tested (a T-cell line, NIH3T3, Cos7, murine ES cells, and primary splenocytes, and did not require localization of the enzyme to the nucleus. Conclusions The effects of different sequences on the delivery and/or activity of Cre in cultured cells could not be predicted in advance. Consequently, the process of developing more active cell-permeant recombinases was largely empirical. The HNC protein, with an excellent combination of activity, solubility and yield, will enhance the use of cell-permeant Cre proteins to regulate gene structure and function in living cells.

  7. Synthesis of protein in intestinal cells exposed to cholera toxin

    International Nuclear Information System (INIS)

    The mechanism by which cyclic adenosine monophosphate (AMP), formed by intestinal epithelial cells in response to cholera toxin, ultimately results in alterations in water and electrolyte transport is poorly understood. Several studies have indicated that inhibitors of transcription or translation block much of the transport of ions and water in the intestine and edema formation in tissue elicited by cholera toxin. Data presented in this study confirmed the inhibitory effects of cycloheximide on cholera toxin-induced fluid accumulation in the rabbit intestinal loop model. Neither cycloheximide nor actinomycin D altered the amount of cyclic AMP that accumulated in intestinal cells and Chinese hamster ovary cells exposed to cholera toxin. An increase in [3H] leucine incorporation was readily demonstrable in intestinal epithelial cells from rabbits challenged with Vibrio cholerae. Similarly, intestinal epithelial cells incubated with cholera toxin for 4 hr synthesized substantially more protein than controls as determined by relative incorporation of [35S] methionine. Most of the new protein synthesized in response to cholera toxin was membrane associated and of high molecular weight. The possible significance of the toxin-induced protein relative to cholera pathogenesis was discussed

  8. G Protein-Coupled Receptor Signaling in Stem Cells and Cancer

    OpenAIRE

    Jennifer R. Lynch; Jenny Yingzi Wang

    2016-01-01

    G protein-coupled receptors (GPCRs) are a large superfamily of cell-surface signaling proteins that bind extracellular ligands and transduce signals into cells via heterotrimeric G proteins. GPCRs are highly tractable drug targets. Aberrant expression of GPCRs and G proteins has been observed in various cancers and their importance in cancer stem cells has begun to be appreciated. We have recently reported essential roles for G protein-coupled receptor 84 (GPR84) and G protein subunit Gαq in ...

  9. A Novel Protein Is Lower Expressed in Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Ruili Guan

    2014-04-01

    Full Text Available Engrailed-2 (EN2 has been identified as a candidate oncogene in breast cancer and prostate cancer. It is usually recognized as a mainly nuclear staining in the cells. However, recent studies showed a cytoplasmic staining occurred in prostate cancer, bladder cancer and clear cell renal cell carcinoma. The inconsistency makes us confused. To clarify the localization and expression of EN2 in renal cell carcinoma, anti-EN2 antibody (ab28731 and anti-EN2 antibody (MAB2600 were used for immunohistochemistry (IHC respectively. Interestingly, we found that EN2 detected by ab28731 was mainly presented in cytoplasm while EN2 detected by MAB2600 was mainly presented in nucleus. To further investigate the different patterns observed above, lysates from full-length EN2 over expression in HEK293T cells were used to identify which antibody the EN2 molecule bound by western blot. Results showed ab28731 did not react with the lysates. For this reason, the novel specific protein detected by ab28731 was not the EN2 molecule and was named nonEN2. Then using the renal carcinoma tissue microarray and renal tissues, we found that the protein expression levels of nonEN2 in kidney tumor tissues was significantly lower than that in kidney normal tissues (p < 0.05, so was in renal cell lines. Taken together, nonEN2 is lower expressed and may play an important role in renal cell carcinoma.

  10. Stable persistence of the yeast plasmid by hitchhiking on chromosomes during vegetative and germ-line divisions of host cells

    OpenAIRE

    Sau, Soumitra; Liu, Yen-Ting; Ma, Chien-Hui; Jayaram, Makkuni

    2015-01-01

    The chromosome-like stability of the Saccharomyces cerevisiae plasmid 2 micron circle likely stems from its ability to tether to chromosomes and segregate by a hitchhiking mechanism. The plasmid partitioning system, responsible for chromosome-coupled segregation, is comprised of 2 plasmid coded proteins Rep1 and Rep2 and a partitioning locus STB. The evidence for the hitchhiking model for mitotic plasmid segregation, although compelling, is almost entirely circumstantial. Direct tests for pla...

  11. YEAST A SINGLE CELL PROTEIN: CHARACTERISTICS and METABOLISM

    Directory of Open Access Journals (Sweden)

    AMATA, I.A

    2013-02-01

    Full Text Available Most of the developing countries of the world are facing a major problem of malnutrition. Due to rapid growth in the population, food and feed scarcity are prevalent leading to a deficiency of protein and essential nutrients amongst human beings and livestock. It is therefore important to take necessary measures to stem this trend by increasing protein production and making it available and more affordable to the population by utilizing methods available for the production of alternative sources of nutrients. The increased world demand for food and in particular protein has engineered the search for non-conventional protein sources to supplement the available protein sources. Since the early fifties, intense efforts have been made to explore these alternate and non-conventional protein sources. In 1996, new sources mainly yeast, fungi, bacteria and algae have been used to ferment biomass in the form of biological waste to produce single cell proteins. Microbial biomass has been considered an alternative to conventional sources of food and feed. Large scale production processes for SCP production reveal interesting features.

  12. Detecting protein association at the T cell plasma membrane.

    Science.gov (United States)

    Baumgart, Florian; Schütz, Gerhard J

    2015-04-01

    At the moment, many models on T cell signaling rely on results obtained via rather indirect methodologies, which makes direct comparison and conclusions to the in vivo situation difficult. Recently, a variety of new imaging methods were developed, which have the potential to directly shed light onto the mysteries of protein association at the T cell membrane. While the new modalities are extremely promising, for a broad readership it may be difficult to judge the results, since technological shortcomings are not always obvious. In this review article, we put key questions on the mechanism of protein interactions in the T cell plasma membrane into relation with techniques that allow to address such questions. We discuss applicability of the techniques, their strengths and weaknesses. This article is part of a Special Issue entitled: Nanoscale membrane organisation and signalling. PMID:25300585

  13. Molecular Cloning and Functional Analysis of ESGP, an Embryonic Stem Cell and Germ Cell Specific Protein

    Institute of Scientific and Technical Information of China (English)

    Yan-Mei CHEN; Zhong-Wei DU; Zhen YAO

    2005-01-01

    Several putative Oct-4 downstream genes from mouse embryonic stem (ES) cells have been identified using the suppression-subtractive hybridization method. In this study, one of the novel genes encoding an ES cell and germ cell specific protein (ESGP) was cloned by rapid amplification of cDNA ends.ESGP contains 801 bp encoding an 84 amino acid small protein and has no significant homology to any known genes. There is a signal peptide at the N-terminal of ESGP protein as predicted by SeqWeb (GCG)(SeqWeb version 2.0.2, http://gcg.biosino.org:8080/). The result of immunofluorescence assay suggested that ESGP might encode a secretory protein. The expression pattern of ESGP is consistent with the expression of Oct-4 during embryonic development. ESGP protein was detected in fertilized oocyte, from 3.5 day postcoital (dpc) blastocyst to 17.5 dpc embryo, and was only detected in testis and ovary tissues in adult. In vitro, ESGP was only expressed in pluripotent cell lines, such as embryonic stem cells, embryonic carcinoma cells and embryonic germ cells, but not in their differentiated progenies. Despite its specific expression,forced expression of ESGP is not indispensable for the effect of Oct-4 on ES cell self-renewal, and does not affect the differentiation to three germ layers.

  14. Estimating the Energetic State of Malignant Cells from RNA Transcription and Protein Interaction Network Data

    OpenAIRE

    Rietman, Edward A.; Sachs, Rainer; Hahnfeldt, Philip; Hlatky, Lynn

    2014-01-01

    Gene expression data, or transcription data, are surrogates for actual protein concentrations in the cells. In addition protein-protein interactions are static diagrams of all the protein-protein interactions in the cell. These interactions may consist of covalent bonding or maybe just secondary bonding such as hydrogen bonding. Given these two surrogate data types we show a technique to compute the Gibbs free energy of a cell. We apply this to yeast cell cycle and to cancer.

  15. Discovering Protein-Protein Interactions within the Programmed Cell Death Network Using a Protein-Fragment Complementation Screen

    Directory of Open Access Journals (Sweden)

    Yuval Gilad

    2014-08-01

    Full Text Available Apoptosis and autophagy are distinct biological processes, each driven by a different set of protein-protein interactions, with significant crosstalk via direct interactions among apoptotic and autophagic proteins. To measure the global profile of these interactions, we adapted the Gaussia luciferase protein-fragment complementation assay (GLuc PCA, which monitors binding between proteins fused to complementary fragments of a luciferase reporter. A library encompassing 63 apoptotic and autophagic proteins was constructed for the analysis of ∼3,600 protein-pair combinations. This generated a detailed landscape of the apoptotic and autophagic modules and points of interface between them, identifying 46 previously unknown interactions. One of these interactions, between DAPK2, a Ser/Thr kinase that promotes autophagy, and 14-3-3τ, was further investigated. We mapped the region responsible for 14-3-3τ binding and proved that this interaction inhibits DAPK2 dimerization and activity. This proof of concept underscores the power of the GLuc PCA platform for the discovery of biochemical pathways within the cell death network.

  16. Cell-free Protein Synthesis in an Autoinduction System for NMR Studies of Protein-Protein Interactions

    International Nuclear Information System (INIS)

    Cell-free protein synthesis systems provide facile access to proteins in a nascent state that enables formation of soluble, native protein-protein complexes even if one of the protein components is prone to self-aggregation and precipitation. Combined with selective isotope-labeling, this allows the rapid analysis of protein-protein interactions with few 15N-HSQC spectra. The concept is demonstrated with binary and ternary complexes between the χ, ψ and γ subunits of Escherichia coli DNA polymerase III: nascent, selectively 15N-labeled ψ produced in the presence of χ resulted in a soluble, correctly folded χ-ψ complex, whereas ψ alone precipitated irrespective of whether γ was present or not. The 15N-HSQC spectra showed that the N-terminal segment of ψ is mobile in the χ-ψ complex, yet important for its binding to γ. The sample preparation was greatly enhanced by an autoinduction strategy, where the T7 RNA polymerase needed for transcription of a gene in a T7-promoter vector was produced in situ

  17. Silver nanoparticle protein corona composition in cell culture media.

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    Jonathan H Shannahan

    Full Text Available The potential applications of nanomaterials as drug delivery systems and in other products continue to expand. Upon introduction into physiological environments and driven by energetics, nanomaterials readily associate proteins forming a protein corona (PC on their surface. This PC influences the nanomaterial's surface characteristics and may impact their interaction with cells. To determine the biological impact of nanomaterial exposure as well as nanotherapeutic applications, it is necessary to understand PC formation. Utilizing a label-free mass spectrometry-based proteomics approach, we examined the composition of the PC for a set of four silver nanoparticles (AgNPs including citrate-stabilized and polyvinlypyrrolidone-stabilized (PVP colloidal silver (20 or 110 nm diameter. To simulate cell culture conditions, AgNPs were incubated for 1 h in Dulbecco's Modified Eagle Medium supplemented with 10% fetal bovine serum, washed, coronal proteins solubilized, and proteins identified and quantified by label-free LC-MS/MS. To determine which attributes influence PC formation, the AgNPs were characterized in both water and cell culture media with 10% FBS. All AgNPs associated a common subset of 11 proteins including albumin, apolipoproteins, keratins, and other serum proteins. 110 nm citrate- and PVP-stabilized AgNPs were found to bind the greatest number of proteins (79 and 85 respectively compared to 20 nm citrate- and PVP-stabilized AgNPs (45 and 48 respectively, suggesting a difference in PC formation based on surface curvature. While no relationships were found for other protein parameters (isoelectric point or aliphatic index, the PC on 20 nm AgNPs (PVP and citrate consisted of more hydrophobic proteins compared to 110 nm AgNPs implying that this class of proteins are more receptive to curvature-induced folding and crowding in exchange for an increased hydration in the aqueous environment. These observations demonstrate the significance of

  18. [Comparative analysis of total cell protein electrophoregram of pathogenic Burkholderia].

    Science.gov (United States)

    Budchenko, A A; Iliukhin, V I; Viktorov, D V

    2005-01-01

    Whole-cell proteins of 22 strain of Burkhoderia pseudomallei, including 13 B. mallei, 5 B. cepacia strains and 14 strains of opportunistically pathogenic Pseudomonas defined by 1D SDC-PAAG electrophoresis. Electrophoregrams contained 35 to 45 protein fractions sized 19 to 130 kDa, which were highly reproductive. On the basis of computer-aided comparative analysis of protein patterns the interspecies and intraspecies grouping of studied microorganisms was made. The cluster analysis of the similarity matrix of protein spectra made it possible to allocate two groups of strains at the level of similarity of 78%. Group I was formed by Burkholderia species that previously belonged to the II RNA-DNA homology group of Pseudomonas: B. pseudomallei, B. mallei, B. cepacia. All Pseudomonas species were added to the 2nd Group: P. aeruginosa, P. stutzeri, P. testosterone, P. fluorescens, P. putida, P. mendocina. Four phenons were isolated among the strains of B. pseudomallei and 2 phenons--among the strains of B. mallei at the threshold similarity level (89%). The authors conclude that the comparative analysis of electrophoregrams of whole-cell proteins can be useful in the identification and typing of pathogenic Burkholderia. PMID:15954473

  19. Interfacial Polymerization for Colorimetric Labeling of Protein Expression in Cells

    Science.gov (United States)

    Lilly, Jacob L.; Sheldon, Phillip R.; Hoversten, Liv J.; Romero, Gabriela; Balasubramaniam, Vivek; Berron, Brad J.

    2014-01-01

    Determining the location of rare proteins in cells typically requires the use of on-sample amplification. Antibody based recognition and enzymatic amplification is used to produce large amounts of visible label at the site of protein expression, but these techniques suffer from the presence of nonspecific reactivity in the biological sample and from poor spatial control over the label. Polymerization based amplification is a recently developed alternative means of creating an on-sample amplification for fluorescence applications, while not suffering from endogenous labels or loss of signal localization. This manuscript builds upon polymerization based amplification by developing a stable, archivable, and colorimetric mode of amplification termed Polymer Dye Labeling. The basic concept involves an interfacial polymer grown at the site of protein expression and subsequent staining of this polymer with an appropriate dye. The dyes Evans Blue and eosin were initially investigated for colorimetric response in a microarray setting, where both specifically stained polymer films on glass. The process was translated to the staining of protein expression in human dermal fibroblast cells, and Polymer Dye Labeling was specific to regions consistent with desired protein expression. The labeling is stable for over 200 days in ambient conditions and is also compatible with modern mounting medium. PMID:25536421

  20. The anti-cell death FNK protein protects cells from death induced by freezing and thawing

    International Nuclear Information System (INIS)

    The FNK protein, constructed from anti-apoptotic Bcl-xL with enhanced activity, was fused with the protein transduction domain (PTD) of the HIV/Tat protein to mediate the delivery of FNK into cells. The fusion protein PTD-FNK was introduced into chondrocytes in isolated articular cartilage-bone sections, cultured neurons, and isolated bone marrow mononuclear cells to evaluate its ability to prevent cell death induced by freezing and thawing. PTD-FNK protected the cells from freeze-thaw damage in a concentration-dependent manner. Addition of PTD-FNK with conventional cryoprotectants (dimethyl sulfoxide and hydroxyethyl starch) increased surviving cell numbers around 2-fold compared with controls treated only with the cryoprotectants. Notably, PTD-FNK allowed CD34+ cells among bone marrow mononuclear cells to survive more efficiently (12-fold more than the control cells) from two successive freeze-thaw cycles. Thus, PTD-FNK prevented cell death induced by freezing and thawing, suggesting that it provides for the successful cryopreservation of biological materials