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Sample records for pombe cell wall

  1. Chromatin and Cell Wall Staining of Schizosaccharomyces pombe.

    PubMed

    Hagan, Iain M

    2016-01-01

    Fission yeasts grow by tip extension, maintaining a constant width until they reach a critical size threshold and divide. Division by medial fission-which gives these yeast their name-generates a new end that arises from the site of cytokinesis. The old end, which was produced during the previous cell cycle, initiates progression of the new cell cycle, and in G2, the new end is activated in a process termed new-end takeoff (NETO). In this protocol, the fluorescent stains calcofluor and 4',6-diamidino-2-phenylindole (DAPI) are used to give a rapid and informative assessment of morphogenesis and cell-cycle progression in the fission yeast Schizosaccharomyces pombe Calcofluor reveals the timing of NETO because it stains the birth scars that are generated at new ends by cytokinesis less efficiently than the rest of the cell wall. Intense calcofluor staining of the septum and measurement of cell length are also widely used to identify dividing cells and to gauge the timing of mitotic commitment. Staining nuclei with DAPI identifies mono- and binucleated cells and complements the calcofluor staining procedure to evaluate the stages of the cell cycle and identify mitotic errors. Equally simple DAPI staining procedures reveal chromatin structure in higher resolution, facilitating more accurate staging of mitotic progression and characterization of mitotic errors. PMID:27250942

  2. Fusion and erosion of cell walls during confugation in the fussion yeast (Schizosaccharomyces pombe).

    PubMed

    Calleja, G B; Yoo, B Y; Johnson, B F

    1977-06-01

    Conjugation in Schizosaccharomyces pombe was studied by transmission electron microscopy. Mural and nuclear events were scored from induction, the initial event, to meiosis I, the start of sporulation. These morphogenic markers were separately identifiable as flocculation, copulation, conjugation-tube formation, cross-wall formation, cross-wall erosion, conjugation-tube expansion, cytoplasmic fusion, de-differentiation of site of union, nuclear migration and karyogamy. The following were identified as new structural elements: sex hairs, which presumably mediate hydrogen bonding between cells during flocculation; crimp at the site of union; dark patch, which presumably serves as a leak-proof seal at the time of cross-wall erosion; suture, an electron-dense seam formed by the union of a copulant pair; and small electron-dense particles close to the site of wall erosion. No special structures on the cell wall could be identified as indicative of specific sites for potential copulatory activity. The discontinuity of the 2 cell walls at the site of union became so de-differentiated after fusion and erosion that it was no longer possible to pinpoint the site of union.

  3. Regulation of cell wall synthesis by the clathrin light chain is essential for viability in Schizosaccharomyces pombe.

    PubMed

    de León, Nagore; Sharifmoghadam, Mohammad Reza; Hoya, Marta; Curto, M-Ángeles; Doncel, Cristina; Valdivieso, M-Henar

    2013-01-01

    The regulation of cell wall synthesis by the clathrin light chain has been addressed. Schizosaccharomyces pombe clc1Δ mutant was inviable in the absence of osmotic stabilization; when grown in sorbitol-supplemented medium clc1Δ cells grew slowly, formed aggregates, and had strong defects in morphology. Additionally, clc1Δ cells exhibited an altered cell wall composition. A mutant that allowed modulating the amount of Clc1p was created to analyze in more detail the dependence of cell wall synthesis on clathrin. A 40% reduction in the amount of Clc1p did not affect acid phosphatase secretion and bulk lipid internalization. Under these conditions, β(1,3)glucan synthase activity and cell wall synthesis were reduced. Also, the delivery of glucan synthases to the cell surface, and the secretion of the Eng1p glucanase were defective. These results suggest that the defects in the cell wall observed in the conditional mutant were due to a defective secretion of enzymes involved in the synthesis/remodelling of this structure, rather than to their endocytosis. Our results show that a reduction in the amount of clathrin that has minor effects on general vesicle trafficking has a strong impact on cell wall synthesis, and suggest that this is the reason for the lethality of clc1Δ cells in the absence of osmotic stabilization. PMID:23977061

  4. Discovery of genes involved in mitosis, cell division, cell wall integrity and chromosome segregation through construction of Schizosaccharomyces pombe deletion strains.

    PubMed

    Chen, Jun-Song; Beckley, Janel R; Ren, Liping; Feoktistova, Anna; Jensen, Michael A; Rhind, Nicholas; Gould, Kathleen L

    2016-09-01

    The fission yeast model system Schizosaccharomyces pombe is used to study fundamental biological processes. To continue to fill gaps in the Sz. pombe gene deletion collection, we constructed a set of 90 haploid gene deletion strains covering many previously uncharacterized genes. To begin to understand the function of these genes, we exposed this collection of strains to a battery of stress conditions. Using this information in combination with microscopy, proteomics and mini-chromosome loss assays, we identified genes involved in cell wall integrity, cytokinesis, chromosome segregation and DNA metabolism. This subset of non-essential gene deletions will add to the toolkits available for the study of biological processes in Sz. pombe. Copyright © 2016 John Wiley & Sons, Ltd.

  5. Analysis of the Schizosaccharomyces pombe Cell Cycle.

    PubMed

    Hagan, Iain M; Grallert, Agnes; Simanis, Viesturs

    2016-01-01

    Schizosaccharomyces pombe cells are rod shaped, and they grow by tip elongation. Growth ceases during mitosis and cell division; therefore, the length of a septated cell is a direct measure of the timing of mitotic commitment, and the length of a wild-type cell is an indicator of its position in the cell cycle. A large number of documented stage-specific changes can be used as landmarks to characterize cell cycle progression under specific experimental conditions. Conditional mutations can permanently or transiently block the cell cycle at almost any stage. Large, synchronously dividing cell populations, essential for the biochemical analysis of cell cycle events, can be generated by induction synchrony (arrest-release of a cell cycle mutant) or selection synchrony (centrifugal elutriation or lactose-gradient centrifugation). Schizosaccharomyces pombe cell cycle studies routinely combine particular markers, mutants, and synchronization procedures to manipulate the cycle. We describe these techniques and list key landmarks in the fission yeast mitotic cell division cycle.

  6. Analysis of the Schizosaccharomyces pombe Cell Cycle.

    PubMed

    Hagan, Iain M; Grallert, Agnes; Simanis, Viesturs

    2016-01-01

    Schizosaccharomyces pombe cells are rod shaped, and they grow by tip elongation. Growth ceases during mitosis and cell division; therefore, the length of a septated cell is a direct measure of the timing of mitotic commitment, and the length of a wild-type cell is an indicator of its position in the cell cycle. A large number of documented stage-specific changes can be used as landmarks to characterize cell cycle progression under specific experimental conditions. Conditional mutations can permanently or transiently block the cell cycle at almost any stage. Large, synchronously dividing cell populations, essential for the biochemical analysis of cell cycle events, can be generated by induction synchrony (arrest-release of a cell cycle mutant) or selection synchrony (centrifugal elutriation or lactose-gradient centrifugation). Schizosaccharomyces pombe cell cycle studies routinely combine particular markers, mutants, and synchronization procedures to manipulate the cycle. We describe these techniques and list key landmarks in the fission yeast mitotic cell division cycle. PMID:27587785

  7. Buoyant density constancy of Schizosaccharomyces pombe cells

    SciTech Connect

    Kubitschek, H.E.; Ward, R.A.

    1985-06-01

    Buoyant densities of cells from exponentially growing cultures of the fission yeast Schizosaccharomyces pombe 972h/sup -/ with division rates from 0.14 to 0.5 per h were determined by equilibrium centrifugation in Percoll gradients. Buoyant densities were independent of growth rate, with an average value (+/- standard error) of 1.0945 (+/- 0.00037) g/ml. When cells from these cultures were separated by size, mean cell volumes were independent of buoyant density, indicating that buoyant densities also were independent of cell age during the division cycle. These results support the suggestion that most or all kinds of cells that divide by equatorial fission may have similar, evolutionarily conserved mechanisms for regulation of buoyant density.

  8. Bgs3p, a Putative 1,3-β-Glucan Synthase Subunit, Is Required for Cell Wall Assembly in Schizosaccharomyces pombe

    PubMed Central

    Martín, Victoria; García, Blanca; Carnero, Elena; Durán, Angel; Sánchez, Yolanda

    2003-01-01

    β-Glucans are the main components of the fungal cell wall. Fission yeast possesses a family of β-glucan synthase-related genes. We describe here the cloning and characterization of bgs3+, a new member of this family. bgs3+ was cloned as a suppressor of a mutant hypersensitive to Echinocandin and Calcofluor White, drugs that interfere with cell wall biosynthesis. Disruption of the gene is lethal, and a decrease in Bgs3p levels leads to rounded cells with thicker walls, slightly reduces the amount of the β-glucan, and raises the amount of α-glucan polymer. These cells finally died. bgs3+ is expressed in vegetative cells grown in different conditions and during mating and germination and is not enhanced by stress situations. Consistent with the observed expression pattern, Bgs3-green fluorescence protein (GFP-Bgs3p) was found at the growing tips during interphase and at the septum prior to cytokinesis, always localized to growth areas. We also found GFP-Bgs3p in mating projections, during the early stages of zygote formation, and at the growing pole during ascospore germination. We conclude that Bgs3p localization is restricted to growth areas and that Bgs3p is a glucan synthase homologue required for cell wall biosynthesis and cell elongation in the fission yeast life cycle. PMID:12582133

  9. POMB/ACE chemotherapy for mediastinal germ cell tumours.

    PubMed

    Bower, M; Brock, C; Holden, L; Nelstrop, A; Makey, A R; Rustin, G J; Newlands, E S

    1997-05-01

    Mediastinal germ cell tumours (MGCT) are rare and most published series reflect the experiences of individual institutions over many years. Since 1979, we have treated 16 men (12 non-seminomatous germ cell tumours and 4 seminomas) with newly diagnosed primary MGCT with POMB/ACE chemotherapy and elective surgical resection of residual masses. This approach yielded complete remissions in 15/16 (94%) patients. The median follow-up was 6.0 years and no relapses occurred more than 2 years after treatment. The 5 year overall survival in the non-seminomatous germ cell tumours (NSGCT) is 73% (95% confidence interval 43-90%). One patient with NSGCT developed drug-resistant disease and died without achieving remission and 2 patients died of relapsed disease. In addition, 4 patients with bulky and/or metastatic seminoma were treated with POMB/ACE. One died of treatment-related neutropenic sepsis in complete remission and one died of relapsed disease. Finally, 4 patients (2 NSGCT and 2 seminomas) referred at relapse were treated with POMB/ACE and one was successfully salvaged. The combination of POMB/ACE chemotherapy and surgery is effective management for MGCT producing high long-term survival rates.

  10. Fixed-Cell Imaging of Schizosaccharomyces pombe.

    PubMed

    Hagan, Iain M; Bagley, Steven

    2016-01-01

    The acknowledged genetic malleability of fission yeast has been matched by impressive cytology to drive major advances in our understanding of basic molecular cell biological processes. In many of the more recent studies, traditional approaches of fixation followed by processing to accommodate classical staining procedures have been superseded by live-cell imaging approaches that monitor the distribution of fusion proteins between a molecule of interest and a fluorescent protein. Although such live-cell imaging is uniquely informative for many questions, fixed-cell imaging remains the better option for others and is an important-sometimes critical-complement to the analysis of fluorescent fusion proteins by live-cell imaging. Here, we discuss the merits of fixed- and live-cell imaging as well as specific issues for fluorescence microscopy imaging of fission yeast. PMID:27371603

  11. Time-lapse electrical impedance spectroscopy for monitoring the cell cycle of single immobilized S. pombe cells

    NASA Astrophysics Data System (ADS)

    Zhu, Zhen; Frey, Olivier; Haandbaek, Niels; Franke, Felix; Rudolf, Fabian; Hierlemann, Andreas

    2015-11-01

    As a complement and alternative to optical methods, wide-band electrical impedance spectroscopy (EIS) enables multi-parameter, label-free and real-time detection of cellular and subcellular features. We report on a microfluidics-based system designed to reliably capture single rod-shaped Schizosaccharomyces pombe cells by applying suction through orifices in a channel wall. The system enables subsequent culturing of immobilized cells in an upright position, while dynamic changes in cell-cycle state and morphology were continuously monitored through EIS over a broad frequency range. Besides measuring cell growth, clear impedance signals for nuclear division have been obtained. The EIS system has been characterized with respect to sensitivity and detection limits. The spatial resolution in measuring cell length was 0.25 μm, which corresponds to approximately a 5-min interval of cell growth under standard conditions. The comprehensive impedance data sets were also used to determine the occurrence of nuclear division and cytokinesis. The obtained results have been validated through concurrent confocal imaging and plausibilized through comparison with finite-element modeling data. The possibility to monitor cellular and intracellular features of single S. pombe cells during the cell cycle at high spatiotemporal resolution renders the presented microfluidics-based EIS system a suitable tool for dynamic single-cell investigations.

  12. Time-lapse electrical impedance spectroscopy for monitoring the cell cycle of single immobilized S. pombe cells.

    PubMed

    Zhu, Zhen; Frey, Olivier; Haandbaek, Niels; Franke, Felix; Rudolf, Fabian; Hierlemann, Andreas

    2015-11-26

    As a complement and alternative to optical methods, wide-band electrical impedance spectroscopy (EIS) enables multi-parameter, label-free and real-time detection of cellular and subcellular features. We report on a microfluidics-based system designed to reliably capture single rod-shaped Schizosaccharomyces pombe cells by applying suction through orifices in a channel wall. The system enables subsequent culturing of immobilized cells in an upright position, while dynamic changes in cell-cycle state and morphology were continuously monitored through EIS over a broad frequency range. Besides measuring cell growth, clear impedance signals for nuclear division have been obtained. The EIS system has been characterized with respect to sensitivity and detection limits. The spatial resolution in measuring cell length was 0.25 μm, which corresponds to approximately a 5-min interval of cell growth under standard conditions. The comprehensive impedance data sets were also used to determine the occurrence of nuclear division and cytokinesis. The obtained results have been validated through concurrent confocal imaging and plausibilized through comparison with finite-element modeling data. The possibility to monitor cellular and intracellular features of single S. pombe cells during the cell cycle at high spatiotemporal resolution renders the presented microfluidics-based EIS system a suitable tool for dynamic single-cell investigations.

  13. Time-lapse electrical impedance spectroscopy for monitoring the cell cycle of single immobilized S. pombe cells

    PubMed Central

    Zhu, Zhen; Frey, Olivier; Haandbaek, Niels; Franke, Felix; Rudolf, Fabian; Hierlemann, Andreas

    2015-01-01

    As a complement and alternative to optical methods, wide-band electrical impedance spectroscopy (EIS) enables multi-parameter, label-free and real-time detection of cellular and subcellular features. We report on a microfluidics-based system designed to reliably capture single rod-shaped Schizosaccharomyces pombe cells by applying suction through orifices in a channel wall. The system enables subsequent culturing of immobilized cells in an upright position, while dynamic changes in cell-cycle state and morphology were continuously monitored through EIS over a broad frequency range. Besides measuring cell growth, clear impedance signals for nuclear division have been obtained. The EIS system has been characterized with respect to sensitivity and detection limits. The spatial resolution in measuring cell length was 0.25 μm, which corresponds to approximately a 5-min interval of cell growth under standard conditions. The comprehensive impedance data sets were also used to determine the occurrence of nuclear division and cytokinesis. The obtained results have been validated through concurrent confocal imaging and plausibilized through comparison with finite-element modeling data. The possibility to monitor cellular and intracellular features of single S. pombe cells during the cell cycle at high spatiotemporal resolution renders the presented microfluidics-based EIS system a suitable tool for dynamic single-cell investigations. PMID:26608589

  14. Interplanetary Migration of Eucaryotic Cell, Spore of Schizosaccharomyces Pombe

    NASA Astrophysics Data System (ADS)

    Hayashi, N.; Nosaka, J.; Ando, R.; Hashimoto, H.; Yokobori, S.; Narumi, I.; Nakagawa, K.; Yamagishi, A.; Tohda, H.

    2013-11-01

    The Tanpopo mission to examine possible interplanetary migration of microbes is progressing. Spore of Schizosaccharomyces pombe are considered as the exposed samples. In this paper, results of preliminary experiments for the exposure are shown.

  15. Schizosaccharomyces pombe glycosylation mutant with altered cell surface properties.

    PubMed Central

    Ballou, C E; Ballou, L; Ball, G

    1994-01-01

    Mutagenesis of Schizosaccharomyces pombe cells yielded a strain that made reduced amounts of invertase. A comparison of the O- and N-linked carbohydrate chains of the wild-type and mutant glycoproteins revealed that a single type of alpha 1-->2-linked mannose was missing in the mutant. Analysis of the wild-type galactomannoprotein showed that it contained a heterogeneous small "core" oligosaccharide fraction linked to asparagine with sugar compositions that ranged from Man9(GlcNAc)2- to Gal4Man10(GlcNAc)2-. The galactose units are in terminal positions of a Man10(GlcNAc)2- unit that is similar to the mannoprotein core of Saccharomyces cerevisiae. Attached to this core in a larger oligosaccharide fraction is an alpha 1-->6-linked polymannose chain that is substituted at position 2 with alpha-linked mannose and galactose. The O-linked sugars consist of mannose, alpha 1-->2-linked mannosylmannose and alpha 1-->2-linked galactosylmannose, along with small amounts of tri- and tetrasaccharides. The glycosylation mutant lacks alpha 1-->2-linked mannose on both the O-linked chains and the outer chain of the large N-linked chains, suggesting that it may be defective in regulation of an alpha 1,2-mannosyltransferase that adds mannose to glycoproteins in the Golgi. PMID:7937765

  16. Regulation of contractile ring formation and septation in Schizosaccharomyces pombe.

    PubMed

    Willet, Alaina H; McDonald, Nathan A; Gould, Kathleen L

    2015-12-01

    The fission yeast Schizosaccharomyces pombe has become a powerful model organism for cytokinesis studies, propelled by pioneering genetic screens in the 1980s and 1990s. S. pombe cells are rod-shaped and divide similarly to mammalian cells, utilizing a medially-placed actin-and myosin-based contractile ring. A cell wall division septum is deposited behind the constricting ring, forming the new ends of each daughter cell. Here we discuss recent advances in our understanding of the regulation of contractile ring formation through formin proteins and the role of the division septum in S. pombe cell division.

  17. Schizosaccharomyces pombe: a novel transport vehicle of functional DNA and mRNA into mammalian antigen-presenting cells.

    PubMed

    Walch-Rückheim, Barbara; Schmitt, Manfred J; Breinig, Frank

    2014-10-21

    Vaccine vehicles based on recombinant yeasts have become promising candidates for the induction of cellular immune responses. In this study, we investigated the capacity of the fission yeast Sz. pombe for the delivery of functional nucleic acids into murine and human antigen-presenting cells. We demonstrate that Sz. pombe cells effectively induce maturation of human dendritic cells (DC), an important prerequisite for T-cell activation. Further, recombinant fission yeast efficiently delivers functional DNA and mRNA into murine macrophages and human DC resulting in the expression of the model antigen eGFP in these cells. Thus, Sz. pombe suggests itself as a promising candidate for a novel live vaccine.

  18. [Comparing Cell Toxicity of Schizosaccharomyces pombe Exposure to Airborne PM2.5 from Beijing and Inert Particle SiO2].

    PubMed

    Liu, Meng-jiao; Huang, Yi; Wen, Hang; Qiu, Guo-yu

    2015-11-01

    To figure out the main factor of PM2.5 toxicity to cell, this study compared the cell toxicity of Schizosaccharomyces pombe (S. pombe), a model organism, exposed to inert ultrafine SiO2 particles, a model particle, and airborne PM2.5 collected from campus of Peking University Beijing China. Using ultraviolet spectrophotometry to measure cell proliferation ratio, and environmental scanning microscope to observe the particle adhesion on the cell surface, and detecting cellular ROS generation with DHE fluorescent dye chromogenic method, and using single cell gel electrophoresis to test cell DNA damage, the experiment results indicated that the ultrafine SiO2 particles (< 60 nm) could inhibit the cell proliferation of S. pombe, mainly through adsorbing onto the cell surface to change the permeability of the cell wall; but it could not induce cells to generate ROS to cause the oxidative damage. PM2.5, the average particle size of which was larger than that of SiO2 particles, could cause oxidative damages to cells mainly by inducing cells to generate ROS, and damage DNA simultaneously. It might illustrate that there was no direct relationship between the toxicity of PM2.5 and its physical properties such as the particle size.

  19. [Comparing Cell Toxicity of Schizosaccharomyces pombe Exposure to Airborne PM2.5 from Beijing and Inert Particle SiO2].

    PubMed

    Liu, Meng-jiao; Huang, Yi; Wen, Hang; Qiu, Guo-yu

    2015-11-01

    To figure out the main factor of PM2.5 toxicity to cell, this study compared the cell toxicity of Schizosaccharomyces pombe (S. pombe), a model organism, exposed to inert ultrafine SiO2 particles, a model particle, and airborne PM2.5 collected from campus of Peking University Beijing China. Using ultraviolet spectrophotometry to measure cell proliferation ratio, and environmental scanning microscope to observe the particle adhesion on the cell surface, and detecting cellular ROS generation with DHE fluorescent dye chromogenic method, and using single cell gel electrophoresis to test cell DNA damage, the experiment results indicated that the ultrafine SiO2 particles (< 60 nm) could inhibit the cell proliferation of S. pombe, mainly through adsorbing onto the cell surface to change the permeability of the cell wall; but it could not induce cells to generate ROS to cause the oxidative damage. PM2.5, the average particle size of which was larger than that of SiO2 particles, could cause oxidative damages to cells mainly by inducing cells to generate ROS, and damage DNA simultaneously. It might illustrate that there was no direct relationship between the toxicity of PM2.5 and its physical properties such as the particle size. PMID:26910977

  20. Morphogenesis of the Fission Yeast Cell through Cell Wall Expansion.

    PubMed

    Atilgan, Erdinc; Magidson, Valentin; Khodjakov, Alexey; Chang, Fred

    2015-08-17

    The shape of walled cells such as fungi, bacteria, and plants are determined by the cell wall. Models for cell morphogenesis postulate that the effects of turgor pressure and mechanical properties of the cell wall can explain the shapes of these diverse cell types. However, in general, these models await validation through quantitative experiments. Fission yeast Schizosaccharomyces pombe are rod-shaped cells that grow by tip extension and then divide medially through formation of a cell wall septum. Upon cell separation after cytokinesis, the new cell ends adopt a rounded morphology. Here, we show that this shape is generated by a very simple mechanical-based mechanism in which turgor pressure inflates the elastic cell wall in the absence of cell growth. This process is independent of actin and new cell wall synthesis. To model this morphological change, we first estimate the mechanical properties of the cell wall using several approaches. The lateral cell wall behaves as an isotropic elastic material with a Young's modulus of 50 ± 10 MPa inflated by a turgor pressure estimated to be 1.5 ± 0.2 MPa. Based upon these parameters, we develop a quantitative mechanical-based model for new end formation that reveals that the cell wall at the new end expands into its characteristic rounded shape in part because it is softer than the mature lateral wall. These studies provide a simple example of how turgor pressure expands the elastic cell wall to generate a particular cell shape.

  1. Long-term single cell analysis of S. pombe on a microfluidic microchemostat array.

    PubMed

    Nobs, Jean-Bernard; Maerkl, Sebastian J

    2014-01-01

    Although Schyzosaccharomyces pombe is one of the principal model organisms for studying the cell cycle, surprisingly few methods have characterized S. pombe growth on the single cell level, and no methods exist capable of analyzing thousands of cells and tens of thousands of cell division events. We developed an automated microfluidic platform permitting S. pombe to be grown on-chip for several days under defined and changeable conditions. We developed an image processing pipeline to extract and quantitate several physiological parameters including cell length, time to division, and elongation rate without requiring synchronization of the culture. Over a period of 50 hours our platform analyzed over 100000 cell division events and reconstructed single cell lineages up to 10 generations in length. We characterized cell lengths and division times in a temperature shift experiment in which cells were initially grown at 30°C and transitioned to 25°C. Although cell length was identical at both temperatures at steady-state, we observed transient changes in cell length if the temperature shift took place during a critical phase of the cell cycle. We further show that cells born with normal length do divide over a wide range of cell lengths and that cell length appears to be controlled in the second generation, were large newly born cells have a tendency to divide more rapidly and thus at a normalized cell size. The platform is thus applicable to measure fine-details in cell cycle dynamics, should be a useful tool to decipher the molecular mechanism underlying size homeostasis, and will be generally applicable to study processes on the single cell level that require large numbers of precision measurements and single cell lineages.

  2. Aging and Cell Death in the Other Yeasts, Schizosaccharomyces pombe and Candida albicans

    PubMed Central

    Lin, Su-Ju; Austriaco, Nicanor

    2013-01-01

    How do cells age and die? For the past twenty years, the budding yeast, Saccharomyces cerevisiae, has been used as a model organism to uncover the genes that regulate lifespan and cell death. More recently, investigators have begun to interrogate the other yeasts, the fission yeast, Schizosaccharomyces pombe, and the human fungal pathogen, Candida albicans, to determine if similar longevity and cell death pathways exist in these organisms. After summarizing the longevity and cell death phenotypes in S. cerevisiae, this mini-review surveys the progress made in the study of both aging and programmed cell death (PCD) in the yeast models, with a focus on the biology of S. pombe and C. albicans. Particular emphasis is placed on the similarities and differences between the two types of aging, replicative aging and chronological aging, and between the three types of cell death, intrinsic apoptosis, autophagic cell death, and regulated necrosis, found in these yeasts. The development of the additional microbial models for aging and PCD in the other yeasts may help further elucidate the mechanisms of longevity and cell death regulation in eukaryotes. PMID:24205865

  3. A New Schizosaccharomyces pombe Chronological Lifespan Assay Reveals that Caloric Restriction Promotes Efficient Cell Cycle Exit and Extends Longevity

    PubMed Central

    Chen, Bo-Ruei; Runge, Kurt W.

    2009-01-01

    We describe a new chronological lifespan (CLS) assay for the yeast Schizosaccharomyces pombe. Yeast CLS assays monitor the loss of cell viability in a culture over time, and this new assay shows a continuous decline in viability without detectable regrowth until all cells in the culture are dead. Thus, the survival curve is not altered by the generation of mutants that can grow during the experiments, and one can monitor the entire lifespan of a strain until the number of viable cells has decreased over 106-fold. This CLS assay recapitulates the evolutionarily conserved features of lifespan shortening by over nutrition, lifespan extension by caloric restriction, increased stress resistance of calorically restricted cells and lifespan control by the AKT kinases. Both S. pombe AKT kinase orthologs regulate CLS: loss of sck1+ extended lifespan in over nutrition conditions, loss of sck2+ extended lifespan under both normal and over nutrition conditions, and loss of both genes showed that sck1+ and sck2+ control different longevity pathways. The longest-lived S. pombe cells showed the most efficient cell cycle exit, demonstrating that caloric restriction links these two processes. This new S. pombe CLS assay will provide a valuable tool for aging research. PMID:19409973

  4. Shu1 is a cell-surface protein involved in iron acquisition from heme in Schizosaccharomyces pombe.

    PubMed

    Mourer, Thierry; Jacques, Jean-François; Brault, Ariane; Bisaillon, Martin; Labbé, Simon

    2015-04-17

    Iron is an essential metal cofactor that is required for many biological processes. Eukaryotic cells have consequently developed different strategies for its acquisition. Until now, Schizosaccharomyces pombe was known to use reductive iron uptake and siderophore-bound iron transport to scavenge iron from the environment. Here, we report the identification of a gene designated shu1(+) that encodes a protein that enables S. pombe to take up extracellular heme for cell growth. When iron levels are low, the transcription of shu1(+) is induced, although its expression is repressed when iron levels rise. The iron-dependent down-regulation of shu1(+) requires the GATA-type transcriptional repressor Fep1, which strongly associates with a proximal promoter region of shu1(+) in vivo in response to iron repletion. HA4-tagged Shu1 localizes to the plasma membrane in cells expressing a functional shu1(+)-HA4 allele. When heme biosynthesis is selectively blocked in mutated S. pombe cells, their ability to acquire exogenous hemin or the fluorescent heme analog zinc mesoporphyrin IX is dependent on the expression of Shu1. Further analysis by absorbance spectroscopy and hemin-agarose pulldown assays showed that Shu1 interacts with hemin, with a KD of ∼2.2 μm. Taken together, results reported here revealed that S. pombe possesses an unexpected pathway for heme assimilation, which may also serve as a source of iron for cell growth.

  5. Cell wall integrity

    PubMed Central

    Pogorelko, Gennady; Lionetti, Vincenzo; Bellincampi, Daniela; Zabotina, Olga

    2013-01-01

    The plant cell wall, a dynamic network of polysaccharides and glycoproteins of significant compositional and structural complexity, functions in plant growth, development and stress responses. In recent years, the existence of plant cell wall integrity (CWI) maintenance mechanisms has been demonstrated, but little is known about the signaling pathways involved, or their components. Examination of key mutants has shed light on the relationships between cell wall remodeling and plant cell responses, indicating a central role for the regulatory network that monitors and controls cell wall performance and integrity. In this review, we present a short overview of cell wall composition and discuss post-synthetic cell wall modification as a valuable approach for studying CWI perception and signaling pathways. PMID:23857352

  6. The Lamportian cell wall

    SciTech Connect

    Keiliszewski, M.; Lamport, D. )

    1991-05-01

    The Lamportian Warp-Weft hypothesis suggests a cellulose-extensin interpenetrating network where extensin mechanically couples the load-bearing cellulose microfibrils in a wall matrix that is best described as a microcomposite. This model is based on data gathered from the extensin-rich walls of tomato and sycamore cell suspension culture, wherein extensin precursors are insolubilized into the wall by undefined crosslinks. The authors recent work with cell walls isolated from intact tissue as well as walls from suspension cultured cells of the graminaceous monocots maize and rice, the non-graminaceous monocot asparagus, the primitive herbaceous dicot sugar beet, and the gymnosperm Douglas Fir indicate that although extensins are ubiquitous to all plant species examined, they are not the major structural protein component of most walls examined. Amino acid analyses of intact and HF-treated walls shows a major component neither an HRGP, nor directly comparable to the glycine-rich wall proteins such as those associated with seed coat walls or the 67 mole% glycine-rich proteins cloned from petunia and soybean. Clearly, structural wall protein alternatives to extensin exist and any cell wall model must take that into account. If we assume that extracellular matrices are a priori network structures, then new Hypless' structural proteins in the maize cell wall raise questions about the sort of network these proteins create: the kinds of crosslinks involved; how they are formed; and the roles played by the small amounts of HRGPs.

  7. Modulating the level of the Rpb7 subunit of RNA polymerase II affects cell separation in Schizosaccharomyces pombe.

    PubMed

    Kumar, Deepak; Sharma, Nimisha

    2015-01-01

    The rpb7(+) gene encodes the seventh largest subunit of RNA polymerase II and is essential for survival of yeast cells. To gain insight into its functions, we expressed rpb7(+) under the control of the nmt1 promoter and investigated its role in regulating multiple phenotypes in Schizosaccharomyces pombe. We observed that low rpb7(+) levels resulted in slow growth of cells under optimum growth conditions. However, no growth defect was observed under different stress conditions tested in this study. Our results also showed that the most prominent phenotype of cells expressing reduced rpb7(+) is a defect in cell separation. Quantitative real-time PCR analysis further revealed that the transcription of specific cell septation genes was significantly reduced in these cells. Collectively, results presented in this study highlight the distinct role of Rpb7p in regulating cell separation in S. pombe.

  8. Linear increase in cell volume during the growth cycle of Schizosaccharomyces pombe

    SciTech Connect

    Kubitschek, H.E.; Clay, K.

    1985-01-01

    Classical observations on the growth of cells of the fission yeast Schizosaccharomyces pombe indicate that cell length and volume increase to a maximum value, which is reached about three-quarters of the way through the growth cycle, followed by a constant volume plateau. The authors have reexamined the growth of these cells by phase microscopy. When growth conditions were perturbed by inoculating cells at high density, in the presence of contaminant, or upon agar slips containing the growth medium in 4% agar, all cells grew in the classical pattern. But, at smaller agar concentrations and lower cell densities, many cells grew at a constant rate throughout the entire cell cycle. Also, the frequency of this linear pattern of cell growth increased as growth perturbations were reduced. They interpret these results as evidence for a sensitivity of this microorganism to perturbations of steady-state growth. The constant rate of volume increase throughout the cell cycle in unperturbed cells, when considered along with Mitchison's earlier results for constant dry mass increase, suggests that the buoyant densities of these cells remain constant during the entire cell cycle.

  9. Preparation and performance of immobilized yeast cells in columns containing no inert carrier. [Schizosaccharomyces pombe

    SciTech Connect

    Hsiao, H.Y.; Chiang, L.C.; Yang, C.M.; Chen, L.F.; Tsao, G.T.

    1983-02-01

    Schizosaccharomyes pombe was cultivated in a medium of glucose (10 g/l), malt extract (3 g/l), yeast extract (3 g/l), and bactopeptone (5 g/l) to form flocs. More than 95% of the cell population were flocculated. Variation in glucose concentration (from 10 to 11 g/l) did not affect flocculation. Yeast extract helped induce flocculation. Application of the immobilized yeast for the continuous production of ethanol was tested in a column reactor. Soft yeast flocs (50-200 mesh) underwent morphological changes to heavy particles (0.1-9.3 cm diameter) after continuously being fed with fresh substrates in the column. Productivity as high as 87 g EtOH/l/hour was obtained when a 150 g/l glucose medium was fed. The performance of this yeast reactor was stable over a two-month period. The ethanol yield was 97% of the theoretical maximum based upon glucose consumed. (Refs. 16).

  10. Cross talk between NDR kinase pathways coordinates cytokinesis with cell separation in Schizosaccharomyces pombe.

    PubMed

    Gupta, Sneha; Govindaraghavan, Meera; McCollum, Dannel

    2014-08-01

    NDR (nuclear Dbf-2-related) kinases constitute key regulatory nodes in signaling networks that control multiple biological processes such as growth, proliferation, mitotic exit, morphogenesis, and apoptosis. Two NDR pathways called the septation initiation network (SIN) and the morphogenesis Orb6 network (MOR) exist in the fission yeast Schizosaccharomyces pombe. The SIN promotes cytokinesis, and the MOR drives cell separation at the end of cytokinesis and polarized growth during interphase. We showed previously that cross talk exists between these two pathways, with the SIN inhibiting the MOR during cytokinesis through phosphorylation of the MOR component Nak1 by the SIN Sid2 kinase. The reason for this inhibition remained uncertain. We show here that failure to inhibit MOR signaling during cytokinesis results in cell lysis at the site of septum formation. Time-lapse analysis revealed that MOR signaling during cytokinesis causes cells to prematurely initiate septum degradation/cell separation. The cell lysis phenotype is due to premature initiation of cell separation because it can be rescued by mutations in genes required for cell separation/septum degradation. We also shed further light on how the SIN inhibits the MOR. Sid2 phosphorylation of the MOR proteins Sog2 and Nak1 is required to prevent cell lysis during cytokinesis. Together, these results show that SIN inhibition of the MOR enforces proper temporal ordering of cytokinetic events.

  11. Lipid droplets form from distinct regions of the cell in the fission yeast Schizosaccharomyces pombe

    DOE PAGES

    Meyers, Alex; del Rio, Zuania P.; Beaver, Rachael A.; Morris, Ryan M.; Weiskittel, Taylor M.; Alshibli, Amany K.; Mannik, Jaana; Morrell-Falvey, Jennifer; Dalhaimer, Paul

    2016-04-29

    Eukaryotic cells store cholesterol/sterol esters (SEs) and triacylglycerols (TAGs) in lipid droplets, which form from the contiguous endoplasmic reticulum (ER) network. However, it is not known if droplets preferentially form from certain regions of the ER over others. Here, we used fission yeast Schizosaccharomyces pombe cells where the nuclear and cortical/peripheral ER domains are distinguishable by light microscopy to show that SE-enriched lipid droplets form away from the nucleus at the cell tips, whereas TAG-enriched lipid droplets form around the nucleus. Sterols localize to the regions of the cells where droplets enriched in SEs are observed. TAG droplet formation aroundmore » the nucleus appears to be a strong function of diacylglycerol (DAG) homeostasis with Cpt1p, which coverts DAG into phosphatidylcholine and phosphatidylethanolamine localized exclusively to the nuclear ER. Also, Dgk1p, which converts DAG into phosphatidic acid localized strongly to the nuclear ER over the cortical/peripheral ER. We also show that TAG more readily translocates from the ER to lipid droplets than do SEs. Lastly, the results augment the standard lipid droplet formation model, which has SEs and TAGs flowing into the same nascent lipid droplet regardless of its biogenesis point in the cell.« less

  12. Lipid Droplets Form from Distinct Regions of the Cell in the Fission Yeast Schizosaccharomyces pombe.

    PubMed

    Meyers, Alex; Del Rio, Zuania P; Beaver, Rachael A; Morris, Ryan M; Weiskittel, Taylor M; Alshibli, Amany K; Mannik, Jaana; Morrell-Falvey, Jennifer; Dalhaimer, Paul

    2016-06-01

    Eukaryotic cells store cholesterol/sterol esters (SEs) and triacylglycerols (TAGs) in lipid droplets, which form from the contiguous endoplasmic reticulum (ER) network. However, it is not known if droplets preferentially form from certain regions of the ER over others. Here, we used fission yeast Schizosaccharomyces pombe cells where the nuclear and cortical/peripheral ER domains are distinguishable by light microscopy to show that SE-enriched lipid droplets form away from the nucleus at the cell tips, whereas TAG-enriched lipid droplets form around the nucleus. Sterols localize to the regions of the cells where droplets enriched in SEs are observed. TAG droplet formation around the nucleus appears to be a strong function of diacylglycerol (DAG) homeostasis with Cpt1p, which coverts DAG into phosphatidylcholine and phosphatidylethanolamine localized exclusively to the nuclear ER. Also, Dgk1p, which converts DAG into phosphatidic acid localized strongly to the nuclear ER over the cortical/peripheral ER. We also show that TAG more readily translocates from the ER to lipid droplets than do SEs. The results augment the standard lipid droplet formation model, which has SEs and TAGs flowing into the same nascent lipid droplet regardless of its biogenesis point in the cell.

  13. A microPIXE investigation of the interaction of cells of Schizosaccharomyces pombe with the culture medium

    NASA Astrophysics Data System (ADS)

    Rombouts, P. M. M.; Gomez-Morilla, I.; Grime, G. W.; Webb, R. P.; Cuenca, L.; Rodriguez, R.; Browton, M.; Wardell, N.; Underwood, B.; Kirkby, N. F.; Kirkby, K. J.

    2007-07-01

    Schizosaccharomyces pombe ( S. pombe) is a eucaryotic cell type similar to mammalian cells but much more simple. As it also executes its cell cycle rapidly it is very useful for investigating basic processes in cells. In this paper we report a feasibility study of the applicability of microPIXE to investigate the interaction between S. pombe cells and the surrounding culture medium. Cells were cultured in various growth medium prior to preparation for analysis. 1 μl drops of medium and cells were spotted onto polypropylene foils held in contact with a polished copper block previously cooled in liquid nitrogen. The samples were dehydrated by freeze-drying. Micro PIXE analysis was carried out with the IBC microbeam facility using a beam of 2.5 MeV protons focused to 1-2 μm diameter. Initially no elemental contrast was observed between the cells and the medium, but by modifying the dilution of the cell suspension, the cells could be distinguished from the surrounding medium through an increased concentration of P and reduced concentration of Cl. The distribution of Na in the medium around the cells showed evidence of the action of the Na pump. Sporulation appears to be induced in the cells by adding Cu to the growth medium and the uptake of Cu by the cells could be clearly observed. This study shows that it is possible to analyse the mass transport of elements in and out of cells In the future this will enable concentration gradients to be analysed and allow the rate of production or consumption of individual cells to be calculated. By observing these patterns for individual cells (not populations) at various known points in the cell cycle, fundamental data can be derived.

  14. Characterization of Schizosaccharomyces pombe copper transporter proteins in meiotic and sporulating cells.

    PubMed

    Plante, Samuel; Ioannoni, Raphaël; Beaudoin, Jude; Labbé, Simon

    2014-04-01

    Meiosis requires copper to undertake its program in which haploid gametes are produced from diploid precursor cells. In Schizosaccharomyces pombe, copper is transported by three members of the copper transporter (Ctr) family, namely Ctr4, Ctr5, and Ctr6. Although central for sexual differentiation, very little is known about the expression profile, cellular localization, and physiological contribution of the Ctr proteins during meiosis. Analysis of gene expression of ctr4(+) and ctr5(+) revealed that they are primarily expressed in early meiosis under low copper conditions. In the case of ctr6(+), its expression is broader, being detected throughout the entire meiotic process with an increase during middle- and late-phase meiosis. Whereas the expression of ctr4(+) and ctr5(+) is exclusively dependent on the presence of Cuf1, ctr6(+) gene expression relies on two distinct regulators, Cuf1 and Mei4. Ctr4 and Ctr5 proteins co-localize at the plasma membrane shortly after meiotic induction, whereas Ctr6 is located on the membrane of vacuoles. After meiotic divisions, Ctr4 and Ctr5 disappear from the cell surface, whereas Ctr6 undergoes an intracellular re-location to co-localize with the forespore membrane. Under copper-limiting conditions, disruption of ctr4(+) and ctr6(+) results in altered SOD1 activity, whereas these mutant cells exhibit substantially decreased levels of CAO activity mostly in early- and middle-phase meiosis. Collectively, these results emphasize the notion that Ctr proteins exhibit differential expression, localization, and contribution in delivering copper to SOD1 and Cao1 proteins during meiosis.

  15. Bacterial Cell Wall Components

    NASA Astrophysics Data System (ADS)

    Ginsberg, Cynthia; Brown, Stephanie; Walker, Suzanne

    Bacterial cell-surface polysaccharides cells are surrounded by a variety of cell-surface structures that allow them to thrive in extreme environments. Components of the cell envelope and extracellular matrix are responsible for providing the cells with structural support, mediating intercellular communication, allowing the cells to move or to adhere to surfaces, protecting the cells from attack by antibiotics or the immune system, and facilitating the uptake of nutrients. Some of the most important cell wall components are polysaccharide structures. This review discusses the occurrence, structure, function, and biosynthesis of the most prevalent bacterial cell surface polysaccharides: peptidoglycan, lipopolysaccharide, arabinogalactan, and lipoarabinomannan, and capsular and extracellular polysaccharides. The roles of these polysaccharides in medicine, both as drug targets and as therapeutic agents, are also described.

  16. Posttranscriptional Regulation of Cell-Cell Interaction Protein-Encoding Transcripts by Zfs1p in Schizosaccharomyces pombe

    PubMed Central

    Wells, Melissa L.; Huang, Weichun; Li, Leping; Gerrish, Kevin E.; Fargo, David C.; Ozsolak, Fatih

    2012-01-01

    Members of the tristetraprolin (TTP) family of CCCH tandem zinc finger proteins can bind directly to AU-rich elements in mRNAs and promote transcript deadenylation and decay. The yeast Schizosaccharomyces pombe expresses a single TTP family member, Zfs1p. In this study, we identified probable Zfs1p target mRNAs by comparing transcript levels in wild-type yeast and zfs1Δ mutants, using deep sequencing and microarray approaches. We also used direct RNA sequencing to determine polyadenylation site locations and to confirm the presence of potential Zfs1p target sequences within the target mRNA. These studies identified a set of transcripts containing potential Zfs1p binding sites that accumulated significantly in the zfs1Δ mutants; a subset of these transcripts decayed more slowly in the zfs1Δ mutants and bound directly to Zfs1p in coimmunoprecipitation assays. One apparent direct target encodes the transcription factor Cbf12p, which is known to increase cell-cell adhesion and flocculation when overexpressed. Studies of zfs1Δ cbf12Δ double mutants demonstrated that the increased flocculation seen in zfs1Δ mutants is due, at least in part, to a direct effect on the turnover of cbf12 mRNA. These data suggest that Zfs1p can both directly and indirectly regulate the levels of transcripts involved in cell-cell adhesion in this species. PMID:22907753

  17. Dual functions for the Schizosaccharomyces pombe inositol kinase Ipk1 in nuclear mRNA export and polarized cell growth.

    PubMed

    Sarmah, Bhaskarjyoti; Wente, Susan R

    2009-02-01

    The inositol 1,3,4,5,6-pentakisphosphate (IP(5)) 2-kinase (Ipk1) catalyzes the production of inositol hexakisphosphate (IP(6)) in eukaryotic cells. Previous studies have shown that IP(6) is required for efficient nuclear mRNA export in the budding yeast Saccharomyces cerevisiae. Here, we report the first functional analysis of ipk1(+) in Schizosaccharomyces pombe. S. pombe Ipk1 (SpIpk1) is unique among Ipk1 orthologues in that it harbors a novel amino (N)-terminal domain with coiled-coil structural motifs similar to those of BAR (Bin-amphiphysin-Rvs) domain proteins. Mutants with ipk1(+) deleted (ipk1Delta) had mRNA export defects as well as pleiotropic defects in polarized growth, cell morphology, endocytosis, and cell separation. The SpIpk1 catalytic carboxy-terminal domain was required to rescue these defects, and the mRNA export block was genetically linked to SpDbp5 function and, likely, IP(6) production. However, the overexpression of the N-terminal domain alone also inhibited these functions in wild-type cells. This revealed a distinct noncatalytic function for the N-terminal domain. To test for connections with other inositol polyphosphates, we also analyzed whether the loss of asp1(+) function, encoding an IP(6) kinase downstream of Ipk1, had an effect on ipk1Delta cells. The asp1Delta mutant alone did not block mRNA export, and its cell morphology, polarized growth, and endocytosis defects were less severe than those of ipk1Delta cells. Moreover, ipk1Delta asp1Delta double mutants had altered inositol polyphosphate levels distinct from those of the ipk1Delta mutant. This suggested novel roles for asp1(+) upstream of ipk1(+). We propose that IP(6) production is a key signaling linchpin for regulating multiple essential cellular processes. PMID:19047361

  18. Extraction of Chromosomal DNA from Schizosaccharomyces pombe.

    PubMed

    Murray, Johanne M; Watson, Adam T; Carr, Antony M

    2016-05-02

    Extraction of DNA from Schizosaccharomyces pombe cells is required for various uses, including templating polymerase chain reactions (PCRs), Southern blotting, library construction, and high-throughput sequencing. To purify high-quality DNA, the cell wall is removed by digestion with Zymolyase or Lyticase and the resulting spheroplasts lysed using sodium dodecyl sulfate (SDS). Cell debris, SDS, and SDS-protein complexes are subsequently precipitated by the addition of potassium acetate and removed by centrifugation. Finally, DNA is precipitated using isopropanol. At this stage, purity is usually sufficient for PCR. However, for more sensitive procedures, such as restriction enzyme digestion, additional purification steps, including proteinase K digestion and phenol-chloroform extraction, are recommended. All of these steps are described in detail here.

  19. Colony Polymerase Chain Reaction with Schizosaccharomyces pombe.

    PubMed

    Murray, Johanne M; Watson, Adam T; Carr, Antony M

    2016-01-01

    When screening a large number of individual Schizosaccharomyces pombe strains by polymerase chain reaction (PCR), a rapid "colony PCR" approach may be used. Numerous colony PCR protocols are available, and fundamental to them all is that the colony must be fresh (grown overnight) and that as few cells as possible are used. In this protocol, we present three reliable methods for preparing S. pombe cells for colony PCR.

  20. Featured Organism: Schizosaccharomyces pombe, The Fission Yeast

    PubMed Central

    2002-01-01

    Schizosaccharomyces pombe, the fission yeast, has long been a crucial model for the study of the eukaryote cell cycle. We take a look at this important yeast, whose genome has recently been completed, featuring comments from Valerie Wood, Jürg Bähler, Ramsay McFarlane, Susan Forsburg, Iain Hagan and Paul Nurse on the implications of having the complete sequence and future prospects for pombe genomics. PMID:18628834

  1. Plant cell walls to ethanol.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Conversion of plant cell walls to ethanol constitutes generation 2 bioethanol production. The process consists of several steps: biomass selection/genetic modification, physiochemical pretreatment, enzymatic saccharification, fermentation, and separation. Ultimately, it is desired to combine as man...

  2. Regulation of the antioxidant system in cells of the fission yeast Schizosaccharomyces pombe after combined treatment with patulin and citrinin.

    PubMed

    Papp, Gábor; Máté, Gábor; Mike, Nóra; Gazdag, Zoltán; Pesti, Miklós

    2016-03-01

    The effects of combined treatment with patulin (PAT) and citrinin (CTN) on Schizosaccharomyces pombe cells were investigated in acute toxicity tests. In comparison with the controls the exposure of fission yeast cells (10(7) cells ml(-1)) to PAT + CTN (250 μM each) for 1 h at a survival rate of 66.6% significantly elevated the concentration of total reactive oxygen species (ROS) via increased levels of peroxides without affecting the concentrations of superoxides or the hydroxyl radical. This treatment induced a 3.08-fold increase in the specific concentration of glutathione and elevated specific activities of catalase and glutathione S-transferase, while at the same time the activity of glutathione reductase decreased. The pattern of the ROS was the same as that induced by CTN (Máté et al., 2014), while the presence of PAT in the PAT + CTN combination treatment modified the activities of the antioxidant system (Papp et al., 2012) in comparison with the individual PAT or CTN treatment, suggesting toxin-specific regulation of glutathione and the enzymes of the antioxidant system and the possibility that the transcription factor (pap1 and atf1) -regulated processes might be influenced directly by ROS. PMID:26752674

  3. Rho 1 GTPase activates the (1-3)beta-D-glucan synthase and is involved in Schizosaccharomyces pombe morphogenesis.

    PubMed Central

    Arellano, M; Durán, A; Pérez, P

    1996-01-01

    The Schizosaccharomyces pombe Cdc42 and Rho1 GTPases were tested for their ability to complement the cwg2-1 mutant phenotype of a decrease in (1-3)beta-D-glucan synthase activity when grown at the non-permissive temperature. Only Rho1 is able to partly complement the defect in glucan synthase associated with the cwg2-1 mutation. Moreover, overexpression of the rho1 gene in wild-type S.pombe cells causes aberrant morphology with loss of polarity and cells with several septa. Under this condition (1-3)beta-D-glucan synthase activity is increased four times, but is still dependent on GTP. When S.pombe is transformed with constitutively active rho1 mutant alleles (rho1-G15V or rho1-Q64L), cells stop growing and show a very thick cell wall with hardly any septum. Under this condition the level of (1-3)beta-D-glucan synthase activity is at least 20 times higher than wild-type and is independent of GTP. Neither cdc42+ nor the cdc42-V12G or cdc42-Q61L constitutively active mutant alleles affect (1-3)beta-D-glucan synthase activity when overexpressed in S.pombe. Cells overproducing Rho1 are hypersensitive to inhibitors of cell wall biosynthesis or to cell wall degrading enzymes. We conclude that Rho1 GTPase directly activates (1-3)beta-D-glucan synthase and regulates S.pombe morphogenesis. Images PMID:8887550

  4. Cell Lysis in S. pombe ura4 Mutants Is Suppressed by Loss of Functional Pub1, Which Regulates the Uracil Transporter Fur4.

    PubMed

    Nishino, Kohei; Kushima, Misaki; Matsuo, Yuzy; Matsuo, Yasuhiro; Kawamukai, Makoto

    2015-01-01

    Schizosaccharomyces pombe Δura4 cells lyse when grown on YPD medium. A S. pombe non-essential gene deletion library was screened to determine suppressors of the lysis phenotype. Deletion of the pub1 gene, which encoded E3 ubiquitin ligase, strongly suppressed cell lysis in Δura4 cells. The Δpub1 cells displayed high sensitivity to 5-fluorouracil, a toxic analog of uracil, and this sensitivity was suppressed by deletion of fur4, which encoded a uracil transporter. Fur4 localized primarily to the Golgi apparatus and vacuoles in wild-type cells, but localization was predominantly at the plasma membrane in Δpub1 cells. Fur4 was necessary for the utilization of extracellular uracil, cytosine, or UMP. Uracil uptake activity increased in the Δpub1 strain in a Fur4-dependent manner. In addition, uracil starvation was critical for induction of cell lysis of Δura4 strains and uracil supplementation suppressed lysis. In summary, the increased uracil uptake ability of Δpub1 cells, where Fur4 was predominantly localized to the plasma membrane, resulted in suppression of cell lysis in the Δura4 background.

  5. Cell Lysis in S. pombe ura4 Mutants Is Suppressed by Loss of Functional Pub1, Which Regulates the Uracil Transporter Fur4

    PubMed Central

    Nishino, Kohei; Kushima, Misaki; Matsuo, Yuzy; Matsuo, Yasuhiro; Kawamukai, Makoto

    2015-01-01

    Schizosaccharomyces pombe Δura4 cells lyse when grown on YPD medium. A S. pombe non-essential gene deletion library was screened to determine suppressors of the lysis phenotype. Deletion of the pub1 gene, which encoded E3 ubiquitin ligase, strongly suppressed cell lysis in Δura4 cells. The Δpub1 cells displayed high sensitivity to 5-fluorouracil, a toxic analog of uracil, and this sensitivity was suppressed by deletion of fur4, which encoded a uracil transporter. Fur4 localized primarily to the Golgi apparatus and vacuoles in wild-type cells, but localization was predominantly at the plasma membrane in Δpub1 cells. Fur4 was necessary for the utilization of extracellular uracil, cytosine, or UMP. Uracil uptake activity increased in the Δpub1 strain in a Fur4-dependent manner. In addition, uracil starvation was critical for induction of cell lysis of Δura4 strains and uracil supplementation suppressed lysis. In summary, the increased uracil uptake ability of Δpub1 cells, where Fur4 was predominantly localized to the plasma membrane, resulted in suppression of cell lysis in the Δura4 background. PMID:26536126

  6. [Characterization of Schizosaccharomyces pombe secreted proteins].

    PubMed

    Liu, Yu-Ling; Liu, Yun-Fan; Xie, Jian-Ping

    2007-02-01

    Secreted proteins play a vital physiological role. Schizosaccharomyces pombe is an important model organism for cell cycle study and a potential useful drug screen model. Secreted proteins also initiate the mating. However, few global studies concerning the secreted proteins of S. pombe was reported. To address this issue, bioinformatics were used to reveal the global secreted proteins of S. pombe. The 4997 proteins deduced from the S. pombe genome were analyzed by combined several programs. One hundred and sixty proteins were identified carrying an NH2-terminal secretory signal peptide by signalP3.0. Among them, 117 proteins are integral membrane proteins (TMpred), 13 proteins are lipoproteins (PrositeS-can), and 66 proteins are secreted proteins. The location of the secreted proteins was also predicted by Target P. Some of the secreted proteins are involved in the nutrition, reproduction, as well as the communication between cells and environment. The global information of the secreted proteins of S. pombe will benefit further studies in drug screening model and host searching for heterologous gene expression.

  7. Back wall solar cell

    NASA Technical Reports Server (NTRS)

    Brandhorst, H. W., Jr. (Inventor)

    1978-01-01

    A solar cell is disclosed which comprises a first semiconductor material of one conductivity type with one face having the same conductivity type but more heavily doped to form a field region arranged to receive the radiant energy to be converted to electrical energy, and a layer of a second semiconductor material, preferably highly doped, of opposite conductivity type on the first semiconductor material adjacent the first semiconductor material at an interface remote from the heavily doped field region. Instead of the opposite conductivity layer, a metallic Schottky diode layer may be used, in which case no additional back contact is needed. A contact such as a gridded contact, previous to the radiant energy may be applied to the heavily doped field region of the more heavily doped, same conductivity material for its contact.

  8. Catalysts of plant cell wall loosening

    PubMed Central

    Cosgrove, Daniel J.

    2016-01-01

    The growing cell wall in plants has conflicting requirements to be strong enough to withstand the high tensile forces generated by cell turgor pressure while selectively yielding to those forces to induce wall stress relaxation, leading to water uptake and polymer movements underlying cell wall expansion. In this article, I review emerging concepts of plant primary cell wall structure, the nature of wall extensibility and the action of expansins, family-9 and -12 endoglucanases, family-16 xyloglucan endotransglycosylase/hydrolase (XTH), and pectin methylesterases, and offer a critical assessment of their wall-loosening activity PMID:26918182

  9. The Schizosaccharomyces pombe rhp3+ gene required for DNA repair and cell viability is functionally interchangeable with the RAD3 gene of Saccharomyces cerevisiae.

    PubMed Central

    Reynolds, P R; Biggar, S; Prakash, L; Prakash, S

    1992-01-01

    The RAD3 gene of Saccharomyces cerevisiae is required for excision repair and is essential for cell viability. RAD3 encoded protein possesses a single stranded DNA-dependent ATPase and DNA and DNA.RNA helicase activities. Mutational studies have indicated a requirement for the RAD3 helicase activities in excision repair. To examine the extent of conservation of structure and function of RAD3 during eukaryotic evolution, we have cloned the RAD3 homolog, rhp3+, from the distantly related yeast Schizosaccharomyces pombe. RAD3 and rhp3+ encoded proteins are highly similar, sharing 67% identical amino acids. We show that like RAD3, rhp3+ is indispensable for excision repair and cell viability, and our studies indicate a requirement of the putative rhp3+ DNA helicase activity in DNA repair. We find that the RAD3 and rhp3+ genes can functionally substitute for one another. The level of complementation provided by the rhp3+ gene in S.cerevisiae rad3 mutants or by the RAD3 gene in S.pombe rhp3 mutants is remarkable in that both the excision repair and viability defects in both yeasts are restored to wild type levels. These observations suggest a parallel evolutionary conservation of other protein components with which RAD3 interacts in mediating its DNA repair and viability functions. Images PMID:1534406

  10. Moss cell walls: structure and biosynthesis

    PubMed Central

    Roberts, Alison W.; Roberts, Eric M.; Haigler, Candace H.

    2012-01-01

    The genome sequence of the moss Physcomitrella patens has stimulated new research examining the cell wall polysaccharides of mosses and the glycosyl transferases that synthesize them as a means to understand fundamental processes of cell wall biosynthesis and plant cell wall evolution. The cell walls of mosses and vascular plants are composed of the same classes of polysaccharides, but with differences in side chain composition and structure. Similarly, the genomes of P. patens and angiosperms encode the same families of cell wall glycosyl transferases, yet, in many cases these families have diversified independently in each lineage. Our understanding of land plant evolution could be enhanced by more complete knowledge of the relationships among glycosyl transferase functional diversification, cell wall structural and biochemical specialization, and the roles of cell walls in plant adaptation. As a foundation for these studies, we review the features of P. patens as an experimental system, analyses of cell wall composition in various moss species, recent studies that elucidate the structure and biosynthesis of cell wall polysaccharides in P. patens, and phylogenetic analysis of P. patens genes potentially involved in cell wall biosynthesis. PMID:22833752

  11. Secondary cell walls: biosynthesis and manipulation.

    PubMed

    Kumar, Manoj; Campbell, Liam; Turner, Simon

    2016-01-01

    Secondary cell walls (SCWs) are produced by specialized plant cell types, and are particularly important in those cells providing mechanical support or involved in water transport. As the main constituent of plant biomass, secondary cell walls are central to attempts to generate second-generation biofuels. Partly as a consequence of this renewed economic importance, excellent progress has been made in understanding how cell wall components are synthesized. SCWs are largely composed of three main polymers: cellulose, hemicellulose, and lignin. In this review, we will attempt to highlight the most recent progress in understanding the biosynthetic pathways for secondary cell wall components, how these pathways are regulated, and how this knowledge may be exploited to improve cell wall properties that facilitate breakdown without compromising plant growth and productivity. While knowledge of individual components in the pathway has improved dramatically, how they function together to make the final polymers and how these individual polymers are incorporated into the wall remain less well understood.

  12. Polyamines in cell walls of chlorococcalean microalgae.

    PubMed

    Burczyk, Jan; Zych, Maria; Ioannidis, Nikolaos E; Kotzabasis, Kiriakos

    2014-01-01

    Biotechnology of microalgae represents a very attractive alternative as a source of energy and substances of high value when compared with plant cultivation. Cell walls of green microalgae have an extraordinary chemical and mechanical resistance and may impede some steps in the biotechnological/industrial exploitation of algae. The aim of the present contribution was to check the presence of polyamines in the cell walls of chlorococcalean green microalgae. Polyamines are nitrogenous compounds synthesized normally in cells and may affect the properties of the cell wall. Our work included strains either forming or not forming the polymer algaenan, allowing us to conclude that algaenan is not a prerequisite for the presence of polyamines in the cell walls. Polyamines were detected in isolated cell walls of Scenedesmus obliquus, Chlorella fusca, Chlorella saccharophila, and Chlorella vulgaris. Their concentration in isolated cell walls ranged between 0.4 and 8.4 nmol/mg dry weight. PMID:24772826

  13. Architecture of dermatophyte cell Walls: Electron microscopic and biochemical analysis

    NASA Technical Reports Server (NTRS)

    Nozawa, Y.; Kitajima, Y.

    1984-01-01

    A review with 83 references on the cell wall structure of dermatophytes is presented. Topics discussed include separation and preparation of cell walls; microstructure of cell walls by electron microscopy; chemical composition of cell walls; structural model of cell walls; and morphological structure of cell walls.

  14. Plant cell walls to ethanol.

    PubMed

    Jordan, Douglas B; Bowman, Michael J; Braker, Jay D; Dien, Bruce S; Hector, Ronald E; Lee, Charles C; Mertens, Jeffrey A; Wagschal, Kurt

    2012-03-01

    Conversion of plant cell walls to ethanol constitutes second generation bioethanol production. The process consists of several steps: biomass selection/genetic modification, physiochemical pretreatment, enzymatic saccharification, fermentation and separation. Ultimately, it is desirable to combine as many of the biochemical steps as possible in a single organism to achieve CBP (consolidated bioprocessing). A commercially ready CBP organism is currently unreported. Production of second generation bioethanol is hindered by economics, particularly in the cost of pretreatment (including waste management and solvent recovery), the cost of saccharification enzymes (particularly exocellulases and endocellulases displaying kcat ~1 s-1 on crystalline cellulose), and the inefficiency of co-fermentation of 5- and 6-carbon monosaccharides (owing in part to redox cofactor imbalances in Saccharomyces cerevisiae). PMID:22329798

  15. Plant cell walls to ethanol.

    PubMed

    Jordan, Douglas B; Bowman, Michael J; Braker, Jay D; Dien, Bruce S; Hector, Ronald E; Lee, Charles C; Mertens, Jeffrey A; Wagschal, Kurt

    2012-03-01

    Conversion of plant cell walls to ethanol constitutes second generation bioethanol production. The process consists of several steps: biomass selection/genetic modification, physiochemical pretreatment, enzymatic saccharification, fermentation and separation. Ultimately, it is desirable to combine as many of the biochemical steps as possible in a single organism to achieve CBP (consolidated bioprocessing). A commercially ready CBP organism is currently unreported. Production of second generation bioethanol is hindered by economics, particularly in the cost of pretreatment (including waste management and solvent recovery), the cost of saccharification enzymes (particularly exocellulases and endocellulases displaying kcat ~1 s-1 on crystalline cellulose), and the inefficiency of co-fermentation of 5- and 6-carbon monosaccharides (owing in part to redox cofactor imbalances in Saccharomyces cerevisiae).

  16. Natural Paradigms of Plant Cell Wall Degradation

    SciTech Connect

    Wei, H.; Xu, Q.; Taylor, L. E.; Baker, J. O.; Tucker, M. P.; Ding, S. Y.

    2009-01-01

    Natural processes of recycling carbon from plant cell walls are slow but very efficient, generally involving microbial communities and their secreted enzymes. Efficient combinations of microbial communities and enzymes act in a sequential and synergistic manner to degrade plant cell walls. Recent understanding of plant cell wall ultra-structure, as well as the carbon metabolism, ATP production, and ecology of participating microbial communities, and the biochemical properties of their cellulolytic enzymes have led to new perspectives on saccharification of biomass. Microbial communities are dynamic functions of the chemical and structural compositions of plant cell wall components. The primitive 'multicellularity' exhibited by certain cellulolytic microorganisms may play a role in facilitating cell-cell communication and cell-plant cell wall-substrate interaction.

  17. Structure of Plant Cell Walls

    PubMed Central

    Weinstein, Larry; Albersheim, Peter

    1979-01-01

    Wild type Bacillus subtilis, when grown on beet araban, secretes into its culture medium an endo-arabanase and two arabinosidases. An alternate procedure to one previously described (Kaji A, T Saheki 1975 Biochim Biophys Acta 410: 354-360) has been developed for the purification of the endo-arabanase. The purified endo-arabanase is shown to be homogeneous by sodium dodecyl sulfate-urea disc gel electrophoresis (molecular weight ≃ 32,000) and by isoelectric focusing (pI = 9.3). The endo-arabanase, acting on a branched araban substrate, has maximal activity at pH 6.0 and preferentially cleaves 5-linked arabinosyl residues. One of the arabinosidases (molecular weight ≃ 65,000, pI = 5.3) has been purified to the point that it contains only one quantitatively minor contaminant, as shown by sodium dodecyl sulfate-urea disc gel electrophoresis and isoelectric focusing. The purified arabinosidase, acting on p-nitrophenyl-α-l-arabinofuranoside, has maximal activity at pH 6.5, and, when acting on a branched araban substrate, preferentially attacks nonreducing terminal arabinosyl residues linked to the 2 or 3 position of other arabinosyl residues. Neither of the two purified enzymes is capable of hydrolyzing a variety of carbohydrate substrates which lack arabinosidic linkages. The purified endo-arabinase is shown to be capable of releasing arabinosyl oligomers from the walls of suspension-cultured sycamore cells, thereby suggesting its usefulness as a probe in studying the structure of the araban component of primary cell walls. PMID:16660741

  18. How do plant cell walls extend?

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1993-01-01

    This article briefly summarizes recent work that identifies the biophysical and biochemical processes that give rise to the extension of plant cell walls. I begin with the biophysical notion of stress relaxation of the wall and follow with recent studies of wall enzymes thought to catalyze wall extension and relaxation. Readers should refer to detailed reviews for more comprehensive discussion of earlier literature (Taiz, 1984; Carpita and Gibeaut, 1993; Cosgrove, 1993).

  19. Transformation of Schizosaccharomyces pombe: Protoplast Procedure.

    PubMed

    Murray, Johanne M; Watson, Adam T; Carr, Antony M

    2016-04-01

    Transformation of Schizosaccharomyces pombe with DNA requires the conditioning of cells to promote DNA uptake followed by cell growth under conditions that select and maintain the plasmid or integration event. The three main methodologies are electroporation, treatment with lithium cations, and transformation of protoplasts. The protocol for protoplast transformation, which is described here, is more complicated than those for electroporation or lithium acetate and thus less often used. However, for some strains, it remains the only reliable protocol. PMID:27037076

  20. Do plant cell walls have a code?

    PubMed

    Tavares, Eveline Q P; Buckeridge, Marcos S

    2015-12-01

    A code is a set of rules that establish correspondence between two worlds, signs (consisting of encrypted information) and meaning (of the decrypted message). A third element, the adaptor, connects both worlds, assigning meaning to a code. We propose that a Glycomic Code exists in plant cell walls where signs are represented by monosaccharides and phenylpropanoids and meaning is cell wall architecture with its highly complex association of polymers. Cell wall biosynthetic mechanisms, structure, architecture and properties are addressed according to Code Biology perspective, focusing on how they oppose to cell wall deconstruction. Cell wall hydrolysis is mainly focused as a mechanism of decryption of the Glycomic Code. Evidence for encoded information in cell wall polymers fine structure is highlighted and the implications of the existence of the Glycomic Code are discussed. Aspects related to fine structure are responsible for polysaccharide packing and polymer-polymer interactions, affecting the final cell wall architecture. The question whether polymers assembly within a wall display similar properties as other biological macromolecules (i.e. proteins, DNA, histones) is addressed, i.e. do they display a code?

  1. The cell walls of Chara aspera Willd. (Charophyta) vegetative cells.

    PubMed

    Nyberg, H; Saranpää, P

    1989-01-01

    The ultrastructure of the vegetative cell walls of the charophyte Chara aspera Willd was studied with TEM. Thallus cells, rhizoid bulbil and rhizoidal node cells were investigated. The internodal cells transverse walls contained plasmodesmata. The longitudinal walls of the internodal cells were uniform, fibrillar, with two thin structurally distinct layers with different structure facing the cytoplasm. The outermost layers of internodal, cortical and rhizoid bulbil cells were composed of randomly orientated fibrils. The longitudinal walls of the cortical cells were helicoidal in structure. In the rhizoid bulbil cell walls, six different layers could be distinguished, but their occurrence seemed to depend on the fixation, staining and cutting procedures. A middle lamella and osmophilic deposits were found in the wall between rhizoidal node cells. The cytoplasmic structure of the internodal and cortical cells was not found to differ from other species of Chara. Charasomes were observed only in cortical cells.

  2. Polyphosphorylated fungal cell wall glycopeptides

    SciTech Connect

    Bonetti, S.J.; Black, B.; Gander, J.E.

    1987-05-01

    Penicillium charlesii secretes a 65 kDa peptidophosphogalactomannan (pPGM) containing 10 phosphodiester residues and 10 galactofuranosyl-containing galactin chains attached to a linear mannan; the polysaccharides is attached to a 3 kDa seryl- and threonyl-rich peptide. The authors have now isolated and partially characterized a form of pPGM released from mycelia of P. charlesii treated at 50/sup 0/C for 15, 30, 60 or 120 min. Two- to 3-fold more pPGM was released by heat treatment than is secreted. Crude pPGM, released by heat, was fractionated on DE-52 and was fractionated into two major fractions on the basis of its difference in negative charge. /sup 1/H-decoupled /sup 13/C NMR spectroscopy of these two fractions provided spectra very similar to that of secreted pPGM previously reported from this laboratory. /sup 1/H-decoupled /sup 31/P NMR showed major signals at 1.47, and 0.22 ppm and minor signals at 1.32, 1.15, 1.00, 0.91 and 0.76 ppm. These signals are upfield from phosphomonoesters and are in the region observed for (6-O-phosphorylcholine)- and (6-O-phosphorylethanolamine)-..cap alpha..-D-mannopyranosyl residues which are 0.22 and 0.90 ppm, respectively. These polymers contain 30 phosphodiester residues per molecule of 70 kDa mass compared with 10 phosphodiesters in secreted pPGM. Acid phosphatase and alkaline protease were the only lytic enzymes released by heat treatment. The evidence suggests that much of the pPGM is derived from cell walls; and that the polysaccharide is highly phosphorylated.

  3. Cell wall, cytoskeleton, and cell expansion in higher plants.

    PubMed

    Bashline, Logan; Lei, Lei; Li, Shundai; Gu, Ying

    2014-04-01

    To accommodate two seemingly contradictory biological roles in plant physiology, providing both the rigid structural support of plant cells and the adjustable elasticity needed for cell expansion, the composition of the plant cell wall has evolved to become an intricate network of cellulosic, hemicellulosic, and pectic polysaccharides and protein. Due to its complexity, many aspects of the cell wall influence plant cell expansion, and many new and insightful observations and technologies are forthcoming. The biosynthesis of cell wall polymers and the roles of the variety of proteins involved in polysaccharide synthesis continue to be characterized. The interactions within the cell wall polymer network and the modification of these interactions provide insight into how the plant cell wall provides its dual function. The complex cell wall architecture is controlled and organized in part by the dynamic intracellular cytoskeleton and by diverse trafficking pathways of the cell wall polymers and cell wall-related machinery. Meanwhile, the cell wall is continually influenced by hormonal and integrity sensing stimuli that are perceived by the cell. These many processes cooperate to construct, maintain, and manipulate the intricate plant cell wall--an essential structure for the sustaining of the plant stature, growth, and life.

  4. Molecular regulation of plant cell wall extensibility

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1998-01-01

    Gravity responses in plants often involve spatial and temporal changes in cell growth, which is regulated primarily by controlling the ability of the cell wall to extend. The wall is thought to be a cellulose-hemicellulose network embedded in a hydrated matrix of complex polysaccharides and a small amount of structural protein. The wall extends by a form of polymer creep, which is mediated by expansins, a novel group of wall-loosening proteins. Expansins were discovered during a molecular dissection of the "acid growth" behavior of cell walls. Expansin alters the rheology of plant walls in profound ways, yet its molecular mechanism of action is still uncertain. It lacks detectable hydrolytic activity against the major components of the wall, but it is able to disrupt noncovalent adhesion between wall polysaccharides. The discovery of a second family of expansins (beta-expansins) sheds light on the biological role of a major group of pollen allergens and implies that expansins have evolved for diverse developmental functions. Finally, the contribution of other processes to wall extensibility is briefly summarized.

  5. Molecular regulation of plant cell wall extensibility.

    PubMed

    Cosgrove, D J

    1998-05-01

    Gravity responses in plants often involve spatial and temporal changes in cell growth, which is regulated primarily by controlling the ability of the cell wall to extend. The wall is thought to be a cellulose-hemicellulose network embedded in a hydrated matrix of complex polysaccharides and a small amount of structural protein. The wall extends by a form of polymer creep, which is mediated by expansins, a novel group of wall-loosening proteins. Expansins were discovered during a molecular dissection of the "acid growth" behavior of cell walls. Expansin alters the rheology of plant walls in profound ways, yet its molecular mechanism of action is still uncertain. It lacks detectable hydrolytic activity against the major components of the wall, but it is able to disrupt noncovalent adhesion between wall polysaccharides. The discovery of a second family of expansins (beta-expansins) sheds light on the biological role of a major group of pollen allergens and implies that expansins have evolved for diverse developmental functions. Finally, the contribution of other processes to wall extensibility is briefly summarized. PMID:11540640

  6. 2003 Plant Cell Walls Gordon Conference

    SciTech Connect

    Daniel J. Cosgrove

    2004-09-21

    This conference will address recent progress in many aspects of cell wall biology. Molecular, genetic, and genomic approaches are yielding major advances in our understanding of the composition, synthesis, and architecture of plant cell walls and their dynamics during growth, and are identifying the genes that encode the machinery needed to make their biogenesis possible. This meeting will bring together international scientists from academia, industry and government labs to share the latest breakthroughs and perspectives on polysaccharide biosynthesis, wood formation, wall modification, expansion and interaction with other organisms, and genomic & evolutionary analyses of wall-related genes, as well as to discuss recent ''nanotechnological'' advances that take wall analysis to the level of a single cell.

  7. Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe.

    PubMed

    Caspari, Thomas; Hilditch, Victoria

    2015-01-01

    The activity of Cdc2 (CDK1) kinase, which coordinates cell cycle progression and DNA break repair, is blocked upon its phosphorylation at tyrosine 15 (Y15) by Wee1 kinase in the presence of DNA damage. How Cdc2 can support DNA repair whilst being inactivated by the DNA damage checkpoint remains to be explained. Human CDK1 is phosphorylated by Myt1 kinase at threonine 14 (T14) close to its ATP binding site before being modified at threonine 161 (T167Sp) in its T-loop by the CDK-activating kinase (CAK). While modification of T161 promotes association with the cyclin partner, phosphorylation of T14 inhibits the CDK1-cyclin complex. This inhibition is further enforced by the modification of Y15 by Wee1 in the presence of DNA lesions. In S.pombe, the dominant inhibition of Cdc2 is provided by the phosphorylation of Y15 and only a small amount of Cdc2 is modified at T14 when cells are in S phase. Unlike human cells, both inhibitory modifications are executed by Wee1. Using the novel IEFPT technology, which combines isoelectric focusing (IEF) with Phos-tag SDS electrophoresis (PT), we report here that S.pombe Cdc2 kinase exists in seven forms. While five forms are phosphorylated, two species are not. Four phospho-forms associate with cyclin B (Cdc13) of which only two are modified at Y15 by Wee1. Interestingly, only one Y15-modified species carries also the T14 modification. The fifth phospho-form has a low affinity for cyclin B and is neither Y15 nor T14 modified. The two unphosphorylated forms may contribute directly to the DNA damage response as only they associate with the DNA damage checkpoint kinase Chk1. Interestingly, cyclin B is also present in the unphosphorylated pool. We also show that the G146D mutation in Cdc2.1w, which renders Cdc2 insensitive to Wee1 inhibition, is aberrantly modified in a Wee1-dependent manner. In conclusion, our work adds support to the idea that two distinct Cdc2 pools regulate cell cycle progression and the response to DNA damage.

  8. Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe.

    PubMed

    Caspari, Thomas; Hilditch, Victoria

    2015-01-01

    The activity of Cdc2 (CDK1) kinase, which coordinates cell cycle progression and DNA break repair, is blocked upon its phosphorylation at tyrosine 15 (Y15) by Wee1 kinase in the presence of DNA damage. How Cdc2 can support DNA repair whilst being inactivated by the DNA damage checkpoint remains to be explained. Human CDK1 is phosphorylated by Myt1 kinase at threonine 14 (T14) close to its ATP binding site before being modified at threonine 161 (T167Sp) in its T-loop by the CDK-activating kinase (CAK). While modification of T161 promotes association with the cyclin partner, phosphorylation of T14 inhibits the CDK1-cyclin complex. This inhibition is further enforced by the modification of Y15 by Wee1 in the presence of DNA lesions. In S.pombe, the dominant inhibition of Cdc2 is provided by the phosphorylation of Y15 and only a small amount of Cdc2 is modified at T14 when cells are in S phase. Unlike human cells, both inhibitory modifications are executed by Wee1. Using the novel IEFPT technology, which combines isoelectric focusing (IEF) with Phos-tag SDS electrophoresis (PT), we report here that S.pombe Cdc2 kinase exists in seven forms. While five forms are phosphorylated, two species are not. Four phospho-forms associate with cyclin B (Cdc13) of which only two are modified at Y15 by Wee1. Interestingly, only one Y15-modified species carries also the T14 modification. The fifth phospho-form has a low affinity for cyclin B and is neither Y15 nor T14 modified. The two unphosphorylated forms may contribute directly to the DNA damage response as only they associate with the DNA damage checkpoint kinase Chk1. Interestingly, cyclin B is also present in the unphosphorylated pool. We also show that the G146D mutation in Cdc2.1w, which renders Cdc2 insensitive to Wee1 inhibition, is aberrantly modified in a Wee1-dependent manner. In conclusion, our work adds support to the idea that two distinct Cdc2 pools regulate cell cycle progression and the response to DNA damage. PMID

  9. Refractive index of plant cell walls

    NASA Technical Reports Server (NTRS)

    Gausman, H. W.; Allen, W. A.; Escobar, D. E.

    1974-01-01

    Air was replaced with media of higher refractive indices by vacuum infiltration in leaves of cucumber, blackeye pea, tomato, and string bean plants, and reflectance of noninfiltrated and infiltrated leaves was spectrophotometrically measured. Infiltrated leaves reflected less light than noninfiltrated leaves over the 500-2500-nm wavelength interval because cell wall-air interfaces were partly eliminated. Minimal reflectance should occur when the average refractive index of plant cell walls was matched by the infiltrating fluid. Although refractive indices that resulted in minimal reflectance differed among the four plant genera, an average value of 1.425 approximates the refractive index of plant cell walls for the four plant genera.

  10. Differential scanning calorimetry of plant cell walls

    SciTech Connect

    Lin, Liangshiou; Varner, J.E. ); Yuen, H.K. )

    1991-03-15

    High-sensitivity differential scanning calorimetry has been used to study the phase transition of cell wall preparations of the elongating and mature regions of soybean hypocotyls and of celery epidermis and collenchyma strands. A step-like transition believed to be glass transition was observed in walls isolated from the elongating region of soybean hypocotyls at 52.9C. Addition of 1 mM CaCl{sub 2} to the cell wall preparation increased the transition temperature to 60.8C and greatly reduced the transition magnitude. In walls from the mature region, the transition was small and occurred at a higher temperature (60.1C). Addition of calcium to the mature region cell wall had little effect on the transition. Based on the known interactions between calcium and pectin, the authors propose that calcium affects the glass transition by binding to the polygalacturonate backbone of wall pectin, resulting in a more rigid wall with a smaller transition at a higher temperature. The mature region either has more calcium in the wall or has more methyl-esterified pectin, making it less responsive to added calcium.

  11. Cell wall remodeling under abiotic stress

    PubMed Central

    Tenhaken, Raimund

    2015-01-01

    Plants exposed to abiotic stress respond to unfavorable conditions on multiple levels. One challenge under drought stress is to reduce shoot growth while maintaining root growth, a process requiring differential cell wall synthesis and remodeling. Key players in this process are the formation of reactive oxygen species (ROS) and peroxidases, which initially cross-link phenolic compounds and glycoproteins of the cell walls causing stiffening. The function of ROS shifts after having converted all the peroxidase substrates in the cell wall. If ROS-levels remain high during prolonged stress, OH°-radicals are formed which lead to polymer cleavage. In concert with xyloglucan modifying enzymes and expansins, the resulting cell wall loosening allows further growth of stressed organs. PMID:25709610

  12. Identification of Schizosaccharomyces pombe prenol as dolichol-16,17.

    PubMed

    Quellhorst, G J; Piotrowski, J S; Steffen, S E; Krag, S S

    1998-03-17

    The identity of the prenol involved in N-linked glycosylation in the fission yeast Schizosaccharomyces pombe was unknown. In order to determine the identity of the prenol, S. pombe cells were incubated with a metabolic precursor of prenol, tritiated mevalonolactone. The cells incorporated only a modest amount of label, about 1000 dpm per million cells, into base-stable lipid and only 1% of that radioactivity was incorporated into prenol. We found by normal phase silica HPLC and more directly by the lack of reactivity with MnO2 that the labeled lipid was predominantly dolichol, not polyprenol. Reverse phase HPLC demonstrated that in S. pombe dolichol ranged between 14 and 18 isoprene units with dolichol-16,17 being the most abundant prenol. This dolichol is of an intermediate length, between the dolichol of S. cerevisiae and that of mammalian cells.

  13. Role of cell wall deconstructing enzymes in the proanthocyanidin-cell wall adsorption-desorption phenomena.

    PubMed

    Castro-López, Liliana del Rocío; Gómez-Plaza, Encarna; Ortega-Regules, Ana; Lozada, Daniel; Bautista-Ortín, Ana Belén

    2016-04-01

    The transference of proanthocyanidins from grapes to wine is quite low. This could be due, among other causes, to proanthocyanidins being bound to grape cell wall polysaccharides, which are present in high concentrations in the must. Therefore, the effective extraction of proanthocyanidins from grapes will depend on the ability to disrupt these associations, and, in this respect, enzymes that degrade these polysaccharides could play an important role. The main objective of this work was to test the behavior of proanthocyanidin-cell wall interactions when commercial maceration enzymes are present in the solution. The results showed that cell wall polysaccharides adsorbed a high amount of proanthocyanidins and only a limited quantity of proanthocyanidins could be desorbed from the cell walls after washing with a model solution. The presence of enzymes in the solution reduced the proanthocyanidin-cell wall interaction, probably through the elimination of pectins from the cell wall network.

  14. Structure of Plant Cell Walls 1

    PubMed Central

    Ishii, Tadashi; Thomas, Jerry; Darvill, Alan; Albersheim, Peter

    1989-01-01

    Considerable information has been obtained about the primary structures of suspension-cultured sycamore (Acer pseudoplatanus) cell-wall pectic polysaccharides, i.e. rhamnogalacturonan I, rhamnogalacturonan II, and homogalacturonan. However, these polysaccharides, which are solubilized from the walls by endo-α-1,4-polygalacturonase, account for only about half of the pectic polysaccharides known to be present in sycamore cell walls. We now report that, after exhaustive treatment with endo-α-1,4-polygalacturonase, additional pectic polysaccharides were extracted from sycamore cell walls by treatment with Na2CO3 at 1 and 22°C. These previously uncharacterized polysaccharides accounted for ∼4% of the cell wall. Based on the glycosyl and glycosyl-linkage compositions and the nature of the products obtained by treating the quantitatively predominant NaCO3-extracted polysaccharides with lithium metal dissolved in ethylenediamine, the polysaccharides were found to strongly resemble rhamnogalacturonan I. However, unlike rhamnogalacturonan I that characteristically had equal amounts of 2- and 2,4-linked rhamnosyl residues in its backbone, the polysaccharides extracted in Na2CO3 at 1°C had markedly disparate ratios of 2- to 2,4-linked rhamnosyl residues. We concluded that polysaccharides similar to rhamnogalacturonan I but with different degrees of branching are present in the walls of suspension-cultured sycamore cells. PMID:16666559

  15. Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

    PubMed Central

    Levin, David E.

    2011-01-01

    The yeast cell wall is a strong, but elastic, structure that is essential not only for the maintenance of cell shape and integrity, but also for progression through the cell cycle. During growth and morphogenesis, and in response to environmental challenges, the cell wall is remodeled in a highly regulated and polarized manner, a process that is principally under the control of the cell wall integrity (CWI) signaling pathway. This pathway transmits wall stress signals from the cell surface to the Rho1 GTPase, which mobilizes a physiologic response through a variety of effectors. Activation of CWI signaling regulates the production of various carbohydrate polymers of the cell wall, as well as their polarized delivery to the site of cell wall remodeling. This review article centers on CWI signaling in Saccharomyces cerevisiae through the cell cycle and in response to cell wall stress. The interface of this signaling pathway with other pathways that contribute to the maintenance of cell wall integrity is also discussed. PMID:22174182

  16. Identification of Novel Cell Wall Components

    SciTech Connect

    Michelle Momany

    2009-10-26

    Our DOE Biosciences-funded work focused on the fungal cell wall and morphogenesis. We are especially interested in how new cell wall material is targeted to appropriate areas for polar (asymmetric) growth. Polar growth is the only way that filamentous fungi explore the environment to find suitable substrates to degrade. Work funded by this grant has resulted in a total of twenty peer-reviewed publications. In work funded by this grant, we identified nine Aspergillus nidulans temperature-sensitive (ts) mutants that fail to send out a germ tube and show a swollen cell phenotype at restrictive temperature, the swo mutants. In other organisms, a swollen cell phenotype is often associated with misdirected growth or weakened cell walls. Our work shows that several of the A. nidulans swo mutants have defects in the establishment and maintenance of polarity. Cloning of several swo genes by complementation also showed that secondary modification of proteins seems is important in polarity. We also investigated cell wall biosynthesis and branching based on leads in literature from other organisms and found that branching and nuclear division are tied and that the cell wall reorganizes during development. In our most recent work we have focused on gene expression during the shift from isotropic to polar growth. Surprisingly we found that genes previously thought to be involved only in spore formation are important in early vegetative growth as well.

  17. Modes of deformation of walled cells.

    PubMed

    Dumais, Jacques

    2013-11-01

    The bewildering morphological diversity found in cells is one of the starkest illustrations of life's ability to self-organize. Yet the morphogenetic mechanisms that produce the multifarious shapes of cells are still poorly understood. The shared similarities between the walled cells of prokaryotes, many protists, fungi, and plants make these groups particularly appealing to begin investigating how morphological diversity is generated at the cell level. In this review, I attempt a first classification of the different modes of surface deformation used by walled cells. Five modes of deformation were identified: inextensional bending, equi-area shear, elastic stretching, processive intussusception, and chemorheological growth. The two most restrictive modes-inextensional and equi-area deformations-are embodied in the exine of pollen grains and the wall-like pellicle of euglenoids, respectively. For these modes, it is possible to express the deformed geometry of the cell explicitly in terms of the undeformed geometry and other easily observable geometrical parameters. The greatest morphogenetic power is reached with the processive intussusception and chemorheological growth mechanisms that underlie the expansive growth of walled cells. A comparison of these two growth mechanisms suggests a possible way to tackle the complexity behind wall growth.

  18. Planctomycetes do possess a peptidoglycan cell wall

    PubMed Central

    Jeske, Olga; Schüler, Margarete; Schumann, Peter; Schneider, Alexander; Boedeker, Christian; Jogler, Mareike; Bollschweiler, Daniel; Rohde, Manfred; Mayer, Christoph; Engelhardt, Harald; Spring, Stefan; Jogler, Christian

    2015-01-01

    Most bacteria contain a peptidoglycan (PG) cell wall, which is critical for maintenance of shape and important for cell division. In contrast, Planctomycetes have been proposed to produce a proteinaceous cell wall devoid of PG. The apparent absence of PG has been used as an argument for the putative planctomycetal ancestry of all bacterial lineages. Here we show, employing multiple bioinformatic methods, that planctomycetal genomes encode proteins required for PG synthesis. Furthermore, we biochemically demonstrate the presence of the sugar and the peptide components of PG in Planctomycetes. In addition, light and electron microscopic experiments reveal planctomycetal PG sacculi that are susceptible to lysozyme treatment. Finally, cryo-electron tomography demonstrates that Planctomycetes possess a typical PG cell wall and that their cellular architecture is thus more similar to that of other Gram-negative bacteria. Our findings shed new light on the cellular architecture and cell division of the maverick Planctomycetes. PMID:25964217

  19. The Structure of Plant Cell Walls

    PubMed Central

    Bauer, Wolfgang D.; Talmadge, Kenneth W.; Keegstra, Kenneth; Albersheim, Peter

    1973-01-01

    The molecular structure, chemical properties, and biological function of the xyloglucan polysaccharide isolated from cell walls of suspension-cultured sycamore (Acer pseudoplatanus) cells are described. The sycamore wall xyloglucan is compared to the extracellular xyloglucan secreted by suspension-cultured sycamore cells into their culture medium and is also compared to the seed “amyloid” xyloglucans. Xyloglucan—or fragments of xyloglucan—and acidic fragments of the pectic polysaccharides are released from endopolygalacturonase-pretreated sycamore walls by treatment of these walls with 8 m urea, endoglucanase, or 0.5 n NaOH. Some of the xyloglucan thus released is found to cochromatograph with the acidic pectic fragments on diethylaminoethyl Sephadex. The chemical or enzymic treatments required for the release of xyloglucan from the walls and the cochromatography of xyloglucan with the acidic pectic fragments indicate that xyloglucan is covalently linked to the pectic polysaccharides and is noncovalently bound to the cellulose fibrils of the sycamore cell wall. The molecular structure of sycamore xyloglucan was characterized by methylation analysis of the oligosaccharides obtained by endoglucanase treatment of the polymer. The structure of the polymer is based on a repeating heptasaccharide unit which consists of 4 residues of β-1-4-linked glucose and 3 residues of terminal xylose. A single xylose residue is glycosidically linked to carbon 6 of 3 of the glucosyl residues. PMID:16658281

  20. Fission yeast Schizosaccharomyces pombe in continuous culture

    SciTech Connect

    Vrana, D.

    1983-08-01

    The fission yeast Schizosaccharomyces pombe was cultivated in a chemostat at dilution rates of D = 0.03, 0.05, 0.10, and 0.20/h. After steady state has been reached, the amount of dry matter, number of cells, concentration of residual sugar, yield coefficient (Y), and some morphological properties of the cells were estimated. Curves reflecting the dry mass, number of cells, and cell mean volume show a changing coordination between the growth rate and the rate of cell division, with respect of D. In addition, it could be concluded that in dividing cells the cell septum is localized asymmetrically; two nonidentical cells differing both in length and volume result. The degree of asymmetry is a function of the dilution rate. (25 Refs.)

  1. Saccharomyces cerevisiae structural cell wall mannoprotein.

    PubMed

    Frevert, J; Ballou, C E

    1985-01-29

    A novel mannoprotein fraction with an average molecular weight of 180 000 has been isolated from Saccharomyces cerevisiae mnn9 mutant cell wall that was solubilized by beta-glucanase digestion. The same material could be extracted from purified wall fragments with 1% sodium dodecyl sulfate. The protein component, 12% by weight, is rich in proline, whereas the carbohydrate, mainly mannose, is about evenly distributed between asparagine and hydroxyamino acids. Endoglucosaminidase H digestion of the isolated mannoprotein reduced its average molecular weight to 150 000, but the mannoprotein, while still embedded in the cell wall, was inaccessible to the enzyme. Biosynthesis and translocation of the mannoprotein were investigated by following incorporation of [3H]proline into this fraction. In the presence of tunicamycin, both mnn9 and wild-type X2180 cells made a mannoprotein fraction with an average molecular weight of 140 000, whereas in the absence of the glycosylation inhibitor, the mnn9 mutant made material with a molecular weight of 180 000 and the mannoprotein made by wild-type cells was too large to penetrate the polyacrylamide gel. Although the cell wall mannoprotein was resistant to heat and proteolytic enzymes, attempts to isolate the carbohydrate-free component failed to yield any characteristic peptide material. PMID:3888262

  2. Roles of membrane trafficking in plant cell wall dynamics

    PubMed Central

    Ebine, Kazuo; Ueda, Takashi

    2015-01-01

    The cell wall is one of the characteristic components of plant cells. The cell wall composition differs among cell types and is modified in response to various environmental conditions. To properly generate and modify the cell wall, many proteins are transported to the plasma membrane or extracellular space through membrane trafficking, which is one of the key protein transport mechanisms in eukaryotic cells. Given the diverse composition and functions of the cell wall in plants, the transport of the cell wall components and proteins that are involved in cell wall-related events could be specialized for each cell type, i.e., the machinery for cell wall biogenesis, modification, and maintenance could be transported via different trafficking pathways. In this review, we summarize the recent progress in the current understanding of the roles and mechanisms of membrane trafficking in plant cells and focus on the biogenesis and regulation of the cell wall. PMID:26539200

  3. Reconstitution of a Secondary Cell Wall in a Secondary Cell Wall-Deficient Arabidopsis Mutant

    PubMed Central

    Sakamoto, Shingo; Mitsuda, Nobutaka

    2015-01-01

    The secondary cell wall constitutes a rigid frame of cells in plant tissues where rigidity is required. Deposition of the secondary cell wall in fiber cells contributes to the production of wood in woody plants. The secondary cell wall is assembled through co-operative activities of many enzymes, and their gene expression is precisely regulated by a pyramidal cascade of transcription factors. Deposition of a transmuted secondary cell wall in empty fiber cells by expressing selected gene(s) in this cascade has not been attempted previously. In this proof-of-concept study, we expressed chimeric activators of 24 transcription factors that are preferentially expressed in the stem, in empty fiber cells of the Arabidopsis nst1-1 nst3-1 double mutant, which lacks a secondary cell wall in fiber cells, under the control of the NST3 promoter. The chimeric activators of MYB46, SND2 and ANAC075, as well as NST3, reconstituted a secondary cell wall with different characteristics from those of the wild type in terms of its composition. The transgenic lines expressing the SND2 or ANAC075 chimeric activator showed increased glucose and xylose, and lower lignin content, whereas the transgenic line expressing the MYB46 chimeric activator showed increased mannose content. The expression profile of downstream genes in each transgenic line was also different from that of the wild type. This study proposed a new screening strategy to identify factors of secondary wall formation and also suggested the potential of the artificially reconstituted secondary cell walls as a novel raw material for production of bioethanol and other chemicals. PMID:25535195

  4. Reconstitution of a secondary cell wall in a secondary cell wall-deficient Arabidopsis mutant.

    PubMed

    Sakamoto, Shingo; Mitsuda, Nobutaka

    2015-02-01

    The secondary cell wall constitutes a rigid frame of cells in plant tissues where rigidity is required. Deposition of the secondary cell wall in fiber cells contributes to the production of wood in woody plants. The secondary cell wall is assembled through co-operative activities of many enzymes, and their gene expression is precisely regulated by a pyramidal cascade of transcription factors. Deposition of a transmuted secondary cell wall in empty fiber cells by expressing selected gene(s) in this cascade has not been attempted previously. In this proof-of-concept study, we expressed chimeric activators of 24 transcription factors that are preferentially expressed in the stem, in empty fiber cells of the Arabidopsis nst1-1 nst3-1 double mutant, which lacks a secondary cell wall in fiber cells, under the control of the NST3 promoter. The chimeric activators of MYB46, SND2 and ANAC075, as well as NST3, reconstituted a secondary cell wall with different characteristics from those of the wild type in terms of its composition. The transgenic lines expressing the SND2 or ANAC075 chimeric activator showed increased glucose and xylose, and lower lignin content, whereas the transgenic line expressing the MYB46 chimeric activator showed increased mannose content. The expression profile of downstream genes in each transgenic line was also different from that of the wild type. This study proposed a new screening strategy to identify factors of secondary wall formation and also suggested the potential of the artificially reconstituted secondary cell walls as a novel raw material for production of bioethanol and other chemicals.

  5. Examination and Disruption of the Yeast Cell Wall.

    PubMed

    Okada, Hiroki; Kono, Keiko; Neiman, Aaron M; Ohya, Yoshikazu

    2016-01-01

    The cell wall of Saccharomyces cerevisiae is a complicated extracellular organelle. Although the barrier may seem like a technical nuisance for researchers studying intracellular biomolecules or conditions, the rigid wall is an essential aspect of the yeast cell. Without it, yeast cells are unable to proliferate or carry out their life cycle. The chemical composition of the cell wall and the biosynthetic pathways and signal transduction mechanisms involved in cell wall remodeling have been studied extensively, but many unanswered questions remain. This introduction describes techniques for investigating abnormalities in the cell and spore walls and performing cell wall disruption. PMID:27480724

  6. The sensitivity of yeast and yeast-like cells to new lysosomotropic agents.

    PubMed

    Krasowska, Anna; Chmielewska, Lucyna; Adamski, Ryszard; Luczyński, Jacek; Witek, Stanisław; Sigler, Karel

    2004-01-01

    The lysosomotropic action of the compounds DM-11 and DMAL-12s against Saccharomyces cerevisiae, Schizosaccharomyces pombe and Candida albicans is species- and pH-dependent. At pH 6.0, DMAL-12s is less effective against S. cerevisiae and S. pombe but more effective against C. albicans than DM-11. At pH 8.0, DMAL-12s strongly inhibits the growth of S. cerevisiae but has only a marginal effect on the resistant C. albicans. S. pombe did not grow at pH 8.0. As shown by quinacrine accumulation, DM-11 causes a general intracellular acidification in all three species, while with DMAL-12s, the acidification is marginal. Morphological changes caused by DMAL-12s in S. cerevisiae affect the cell interior but not surface structures, while S. pombe cells exhibit a thickened and wrinkled cell wall, shrunken protoplast and "grainy" plasma membrane. A large number of blisters resembling lipid droplets were observed inside S. cerevisiae and S. pombe vacuoles. The high susceptibility of S. pombe cells to the action of DM-11 and DMAL-12s contrasts with the low sensitivity of S. pombe H+-ATPase to the agents. In our C. albicans isolate, DMAL 12s did not have an effect on cell morphology and appeared to be unable to penetrate the cells, especially at pH 8.0.

  7. Characterization of the Sclerotinia sclerotiorum cell wall proteome.

    PubMed

    Liu, Longzhou; Free, Stephen J

    2016-08-01

    We used a proteomic analysis to identify cell wall proteins released from Sclerotinia sclerotiorum hyphal and sclerotial cell walls via a trifluoromethanesulfonic acid (TFMS) digestion. Cell walls from hyphae grown in Vogel's glucose medium (a synthetic medium lacking plant materials), from hyphae grown in potato dextrose broth and from sclerotia produced on potato dextrose agar were used in the analysis. Under the conditions used, TFMS digests the glycosidic linkages in the cell walls to release intact cell wall proteins. The analysis identified 24 glycosylphosphatidylinositol (GPI)-anchored cell wall proteins and 30 non-GPI-anchored cell wall proteins. We found that the cell walls contained an array of cell wall biosynthetic enzymes similar to those found in the cell walls of other fungi. When comparing the proteins in hyphal cell walls grown in potato dextrose broth with those in hyphal cell walls grown in the absence of plant material, it was found that a core group of cell wall biosynthetic proteins and some proteins associated with pathogenicity (secreted cellulases, pectin lyases, glucosidases and proteases) were expressed in both types of hyphae. The hyphae grown in potato dextrose broth contained a number of additional proteins (laccases, oxalate decarboxylase, peroxidase, polysaccharide deacetylase and several proteins unique to Sclerotinia and Botrytis) that might facilitate growth on a plant host. A comparison of the proteins in the sclerotial cell wall with the proteins in the hyphal cell wall demonstrated that sclerotia formation is not marked by a major shift in the composition of cell wall protein. We found that the S. sclerotiorum cell walls contained 11 cell wall proteins that were encoded only in Sclerotinia and Botrytis genomes.

  8. Characterization of the Sclerotinia sclerotiorum cell wall proteome.

    PubMed

    Liu, Longzhou; Free, Stephen J

    2016-08-01

    We used a proteomic analysis to identify cell wall proteins released from Sclerotinia sclerotiorum hyphal and sclerotial cell walls via a trifluoromethanesulfonic acid (TFMS) digestion. Cell walls from hyphae grown in Vogel's glucose medium (a synthetic medium lacking plant materials), from hyphae grown in potato dextrose broth and from sclerotia produced on potato dextrose agar were used in the analysis. Under the conditions used, TFMS digests the glycosidic linkages in the cell walls to release intact cell wall proteins. The analysis identified 24 glycosylphosphatidylinositol (GPI)-anchored cell wall proteins and 30 non-GPI-anchored cell wall proteins. We found that the cell walls contained an array of cell wall biosynthetic enzymes similar to those found in the cell walls of other fungi. When comparing the proteins in hyphal cell walls grown in potato dextrose broth with those in hyphal cell walls grown in the absence of plant material, it was found that a core group of cell wall biosynthetic proteins and some proteins associated with pathogenicity (secreted cellulases, pectin lyases, glucosidases and proteases) were expressed in both types of hyphae. The hyphae grown in potato dextrose broth contained a number of additional proteins (laccases, oxalate decarboxylase, peroxidase, polysaccharide deacetylase and several proteins unique to Sclerotinia and Botrytis) that might facilitate growth on a plant host. A comparison of the proteins in the sclerotial cell wall with the proteins in the hyphal cell wall demonstrated that sclerotia formation is not marked by a major shift in the composition of cell wall protein. We found that the S. sclerotiorum cell walls contained 11 cell wall proteins that were encoded only in Sclerotinia and Botrytis genomes. PMID:26661933

  9. Cell Wall Heterogeneity in Root Development of Arabidopsis

    PubMed Central

    Somssich, Marc; Khan, Ghazanfar Abbas; Persson, Staffan

    2016-01-01

    Plant cell walls provide stability and protection to plant cells. During growth and development the composition of cell walls changes, but provides enough strength to withstand the turgor of the cells. Hence, cell walls are highly flexible and diverse in nature. These characteristics are important during root growth, as plant roots consist of radial patterns of cells that have diverse functions and that are at different developmental stages along the growth axis. Young stem cell daughters undergo a series of rapid cell divisions, during which new cell walls are formed that are highly dynamic, and that support rapid anisotropic cell expansion. Once the cells have differentiated, the walls of specific cell types need to comply with and support different cell functions. For example, a newly formed root hair needs to be able to break through the surrounding soil, while endodermal cells modify their walls at distinct positions to form Casparian strips between them. Hence, the cell walls are modified and rebuilt while cells transit through different developmental stages. In addition, the cell walls of roots readjust to their environment to support growth and to maximize nutrient uptake. Many of these modifications are likely driven by different developmental and stress signaling pathways. However, our understanding of how such pathways affect cell wall modifications and what enzymes are involved remain largely unknown. In this review we aim to compile data linking cell wall content and re-modeling to developmental stages of root cells, and dissect how root cell walls respond to certain environmental changes. PMID:27582757

  10. Cell Wall Heterogeneity in Root Development of Arabidopsis.

    PubMed

    Somssich, Marc; Khan, Ghazanfar Abbas; Persson, Staffan

    2016-01-01

    Plant cell walls provide stability and protection to plant cells. During growth and development the composition of cell walls changes, but provides enough strength to withstand the turgor of the cells. Hence, cell walls are highly flexible and diverse in nature. These characteristics are important during root growth, as plant roots consist of radial patterns of cells that have diverse functions and that are at different developmental stages along the growth axis. Young stem cell daughters undergo a series of rapid cell divisions, during which new cell walls are formed that are highly dynamic, and that support rapid anisotropic cell expansion. Once the cells have differentiated, the walls of specific cell types need to comply with and support different cell functions. For example, a newly formed root hair needs to be able to break through the surrounding soil, while endodermal cells modify their walls at distinct positions to form Casparian strips between them. Hence, the cell walls are modified and rebuilt while cells transit through different developmental stages. In addition, the cell walls of roots readjust to their environment to support growth and to maximize nutrient uptake. Many of these modifications are likely driven by different developmental and stress signaling pathways. However, our understanding of how such pathways affect cell wall modifications and what enzymes are involved remain largely unknown. In this review we aim to compile data linking cell wall content and re-modeling to developmental stages of root cells, and dissect how root cell walls respond to certain environmental changes. PMID:27582757

  11. At the border: the plasma membrane-cell wall continuum.

    PubMed

    Liu, Zengyu; Persson, Staffan; Sánchez-Rodríguez, Clara

    2015-03-01

    Plant cells rely on their cell walls for directed growth and environmental adaptation. Synthesis and remodelling of the cell walls are membrane-related processes. During cell growth and exposure to external stimuli, there is a constant exchange of lipids, proteins, and other cell wall components between the cytosol and the plasma membrane/apoplast. This exchange of material and the localization of cell wall proteins at certain spots in the plasma membrane seem to rely on a particular membrane composition. In addition, sensors at the plasma membrane detect changes in the cell wall architecture, and activate cytoplasmic signalling schemes and ultimately cell wall remodelling. The apoplastic polysaccharide matrix is, on the other hand, crucial for preventing proteins diffusing uncontrollably in the membrane. Therefore, the cell wall-plasma membrane link is essential for plant development and responses to external stimuli. This review focuses on the relationship between the cell wall and plasma membrane, and its importance for plant tissue organization.

  12. Measuring in vitro extensibility of growing plant cell walls.

    PubMed

    Cosgrove, Daniel J

    2011-01-01

    This article summarizes the theory and practical aspects of measuring cell wall properties by four different extensometer techniques and how the results of these methods relate to the concept and ideal measurement of cell wall extensibility in the context of cell growth. These in vivo techniques are particularly useful for studies of the molecular basis of cell wall extension. Measurements of breaking strength, elastic compliance, and plastic compliance may be informative about changes in cell wall structure, whereas measurements of wall stress relaxation and creep are sensitive to both changes in wall structure and wall-loosening processes, such as those mediated by expansins and some lytic enzymes. A combination of methods is needed to obtain a broader view of cell wall behavior and properties connected with the concept of cell wall extensibility.

  13. Measuring in-vitro extensibility of growth plant cell walls

    SciTech Connect

    Cosgrove, Daniel

    2011-01-01

    This article summarizes the theory and practical aspects of measuring cell wall properties by four different extensometer techniques and how the results of these methods relate to the concept and ideal measurement of cell wall extensibility in the context of cell growth. These in vivo techniques are particularly useful for studies of the molecular basis of cell wall extension. Measurements of breaking strength, elastic compliance, and plastic compliance may be informative about changes in cell wall structure, whereas measurements of wall stress relaxation and creep are sensitive to both changes in wall structure and wall-loosening processes, such as those mediated by expansins and some lytic enzymes. A combination of methods is needed to obtain a broader view of cell wall behavior and properties connected with the concept of cell wall extensibility.

  14. Interconnections between cell wall polymers, wall mechanics, and cortical microtubules: Teasing out causes and consequences.

    PubMed

    Xiao, Chaowen; Anderson, Charles T

    2016-09-01

    In plants, cell wall components including cellulose, hemicelluloses, and pectins interact with each other to form complex extracellular network structures that control cell growth and maintain cell shape. However, it is still not clear exactly how different wall polymers interact, how the conformations and interactions of cell wall polymers relate to wall mechanics, and how these factors impinge on intracellular structures such as the cortical microtubule cytoskeleton. Here, based on studies of Arabidopsis thaliana xxt1 xxt2 mutants, which lack detectable xyloglucan in their walls and display aberrant wall mechanics, altered cellulose patterning and biosynthesis, and reduced cortical microtubule stability, we discuss the potential relationships between cell wall biosynthesis, wall mechanics, and cytoskeletal dynamics in an effort to better understand their roles in controlling plant growth and morphogenesis. PMID:27611066

  15. Interconnections between cell wall polymers, wall mechanics, and cortical microtubules: Teasing out causes and consequences.

    PubMed

    Xiao, Chaowen; Anderson, Charles T

    2016-09-01

    In plants, cell wall components including cellulose, hemicelluloses, and pectins interact with each other to form complex extracellular network structures that control cell growth and maintain cell shape. However, it is still not clear exactly how different wall polymers interact, how the conformations and interactions of cell wall polymers relate to wall mechanics, and how these factors impinge on intracellular structures such as the cortical microtubule cytoskeleton. Here, based on studies of Arabidopsis thaliana xxt1 xxt2 mutants, which lack detectable xyloglucan in their walls and display aberrant wall mechanics, altered cellulose patterning and biosynthesis, and reduced cortical microtubule stability, we discuss the potential relationships between cell wall biosynthesis, wall mechanics, and cytoskeletal dynamics in an effort to better understand their roles in controlling plant growth and morphogenesis.

  16. Protective roles of methionine-R-sulfoxide reductase against stresses in Schizosaccharomyces pombe.

    PubMed

    Jo, Hannah; Cho, Young-Wook; Ji, Sun-Young; Kang, Ga-Young; Lim, Chang-Jin

    2014-01-01

    The Schizosaccharomyces pombe msrB(+) gene encoding methionine-R-sulfoxide reductase (MsrB) was cloned into the shuttle vector pRS316 to generate the recombinant plasmid pFMetSO. The msrB(+) mRNA level was significantly increased in the S. pombe cells harboring pFMetSO, indicating that the cloned msrB(+) gene is functioning. In the presence of 0.1 mM L-methionine-(R,S)-sulfoxide, the S. pombe cells harboring pFMetSO could grow normally but the growth of the vector control cells was almost arrested. The S. pombe cells harboring pFMetSO exhibited the enhanced growth on the minimal medium plates with stress-inducing agents, such as hydrogen peroxide, superoxide radical-generating menadione (MD), nitric oxide (NO)-generating sodium nitroprusside (SNP), and cadmium (Cd), when compared with the vector control cells. They also gave rise to the enhanced growth at the high incubation temperature of 37 °C than the vector control cells. The S. pombe cells harboring pFMetSO contained lower reactive oxygen species (ROS) and higher total glutathione (GSH) levels than the vector control cells. In brief, the S. pombe MsrB plays a protective role against oxidative, nitrosative, and thermal stresses, and is involved in diminishing intracellular ROS level.

  17. Celery (Apium graveolens) parenchyma cell walls: cell walls with minimal xyloglucan.

    PubMed

    Thimm, Julian C.; Burritt, David J.; Sims, Ian M.; Newman, Roger H.; Ducker, William A.; Melton, Laurence D.

    2002-10-01

    The primary walls of celery (Apium graveolens L.) parenchyma cells were isolated and their polysaccharide components characterized by glycosyl linkage analysis, cross-polarization magic-angle spinning solid-state 13C nuclear magnetic resonance (CP/MAS 13C NMR) and X-ray diffraction. Glycosyl linkage analysis showed that the cell walls consisted of mainly cellulose (43 mol%) and pectic polysaccharides (51 mol%), comprising rhamnogalacturonan (28 mol%), arabinan (12 mol%) and galactan (11 mol%). The amounts of xyloglucan (2 mol%) and xylan (2 mol%) detected in the cell walls were strikingly low. The small amount of xyloglucan present means that it cannot coat the cellulose microfibrils. Solid-state 13C NMR signals were consistent with the constituents identified by glycosyl linkage analysis and allowed the walls to be divided into three domains, based on the rigidity of the polymers. Cellulose (rigid) and rhamnogalacturonan (semi-mobile) polymers responded to the CP/MAS 13C NMR pulse sequence and were distinguished by differences in proton spin relaxation time constants. The arabinans, the most mobile polymers, responded to single-pulse excitation (SPE), but not CP/MAS 13C NMR. From solid-state 13C NMR of the cell walls the diameter of the crystalline cellulose microfibrils was determined to be approximately 3 nm while X-ray diffraction of the cell walls gave a value for the diameter of approximately 2 nm.

  18. Food applications of bacterial cell wall hydrolases.

    PubMed

    Callewaert, Lien; Walmagh, Maarten; Michiels, Chris W; Lavigne, Rob

    2011-04-01

    Bacterial cell wall hydrolases (BCWHs) display a remarkable structural and functional diversity that offers perspectives for novel food applications, reaching beyond those of the archetype BCWH and established biopreservative hen egg white lysozyme. Insights in BCWHs from bacteriophages to animals have provided concepts for tailoring BCWHs to target specific pathogens or spoilage bacteria, or, conversely, to expand their working range to Gram-negative bacteria. Genetically modified foods expressing BCWHs in situ showed successful, but face regulatory and ethical concerns. An interesting spin-off development is the use of cell wall binding domains of bacteriophage BCWHs for detection and removal of foodborne pathogens. Besides for improving food safety or stability, BCWHs may also find use as functional food ingredients with specific health effects.

  19. Metabolism of the phospholipid precursor inositol and its relationship to growth and viability in the natural auxotroph Schizosaccharomyces pombe.

    PubMed Central

    Fernandez, S; Homann, M J; Henry, S A; Carman, G M

    1986-01-01

    Phospholipid metabolism in the fission yeast Schizosaccharomyces pombe was examined. Three enzymes of phospholipid biosynthesis, cytidine diphosphate diacylglycerol synthase (CDP-DG), phosphatidylinositol (PI) synthase, and phosphatidylserine (PS) synthase, were characterized in extracts of S. pombe cells. Contrary to an earlier report, we were able to demonstrate that CDP-DG served as a precursor for PI and PS biosynthesis in S. pombe. S. pombe is naturally auxotrophic for the phospholipid precursor inositol. We found that S. pombe was much more resistant to loss of viability during inositol starvation than artificially generated inositol auxotrophs of Saccharomyces cerevisiae. The phospholipid composition of S. pombe cells grown in inositol-rich medium (50 microM) was similar to that of S. cerevisiae cells grown under similar conditions. However, growth of S. pombe at low inositol concentrations (below 30 microM) affected the ratio of the anionic phospholipids PI and PS, while the relative proportions of other glycerophospholipids remained unchanged. During inositol starvation, the rate of PI synthesis decreased rapidly, and there was a concomitant increase in the rate of PS synthesis. Phosphatidic acid and CDP-DG, which are precursors to these phospholipids, also increased when PI synthesis was blocked by lack of exogenous inositol. The major product of turnover of inositol-containing phospholipids in S. pombe was found to be free inositol, which accumulated in the medium and could be reused by the cell. Images PMID:3011744

  20. Revealing the structural and functional diversity of plant cell walls.

    PubMed

    Knox, J Paul

    2008-06-01

    The extensive knowledge of the chemistry of isolated cell wall polymers, and that relating to the identification and partial annotation of gene families involved in their synthesis and modification, is not yet matched by a sophisticated understanding of the occurrence of the polymers within cell walls of the diverse cell types within a growing organ. Currently, the main sets of tools that are used to determine cell-type-specific configurations of cell wall polymers and aspects of cell wall microstructures are antibodies, carbohydrate-binding modules (CBMs) and microspectroscopies. As these tools are applied we see that cell wall polymers are extensively developmentally regulated and that there is a range of structurally distinct primary and secondary cell walls within organs and across species. The challenge now is to document cell wall structures in relation to diverse cell biological events and to integrate this knowledge with the emerging understanding of polymer functions.

  1. Wall relaxation and the driving forces for cell expansive growth

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1987-01-01

    When water uptake by growing cells is prevented, the turgor pressure and the tensile stress in the cell wall are reduced by continued wall loosening. This process, termed in vivo stress relaxation, provides a new way to study the dynamics of wall loosening and to measure the wall yield threshold and the physiological wall extensibility. Stress relaxation experiments indicate that wall stress supplies the mechanical driving force for wall yielding. Cell expansion also requires water absorption. The driving force for water uptake during growth is created by wall relaxation, which lowers the water potential of the expanding cells. New techniques for measuring this driving force show that it is smaller than believed previously; in elongating stems it is only 0.3 to 0.5 bar. This means that the hydraulic resistance of the water transport pathway is small and that rate of cell expansion is controlled primarily by wall loosening and yielding.

  2. Cell wall of Fusarium sulphureum; I. Chemical composition of the hyphal wall.

    PubMed

    Barran, L R; Schneider, E F; Wood, P J; Madhosingh, C; Miller, R W

    1975-05-01

    The hyphae wall of Fusarium sulphureum Schlect. (Isolate 1) was isolated and purified. Electron microscopy studies showed that the isolated cell wall consisted of two distinct layers, an outer electron dense layer and a broader electron transparent inner layer. Chemical analysis revealed that the cell wall contained 66% carbohydrate, 7.3% protein, 5.5% lipid and 1.8% ash. The major cell wall component N-acetylglucosamine (39%) was shown by X-ray diffraction analysis to be present as chitin. Glucose constituted 14% of the cell wall, while mannose, galactose, and glucuronic acid, accounted for 15% of the cell wall. Glucuronic acid appears to be predominantly linked to galactose in the intact wall.

  3. The Structure of Plant Cell Walls

    PubMed Central

    Talmadge, Kenneth W.; Keegstra, Kenneth; Bauer, Wolfgang D.; Albersheim, Peter

    1973-01-01

    This is the first in a series of papers dealing with the structure of cell walls isolated from suspension-cultured sycamore cells (Acer pseudoplatanus). These studies have been made possible by the availability of purified hydrolytic enzymes and by recent improvements in the techniques of methylation analysis. These techniques have permitted us to identify and quantitate the macromolecular components of sycamore cell walls. These walls are composed of 10% arabinan, 2% 3,6-linked arabinogalactan, 23% cellulose, 9% oligo-arabinosides (attached to hydroxyproline), 8% 4-linked galactan, 10% hydroxyproline-rich protein, 16% rhamnogalacturonan, and 21% xyloglucan. The structures of the pectic polymers (the neutral arabinan, the neutral galactan, and the acidic rhamnogalacturonan) were obtained, in part, by methylation analysis of fragments of these polymers which were released from the sycamore walls by the action of a highly purified endopolygalacturonase. The data suggest a branched arabinan and a linear 4-linked galactan occurring as side chains on the rhamnogalacturonan. Small amounts or pieces of a xyloglucan, the wall hemicellulose, appear to be covalently linked to some of the galactan chains. Thus, the galactan appears to serve as a bridge between the xyloglucan and rhamnogalacturonan components of the wall. The rhamnogalacturonan consists of an α-(1 → 4)-linked galacturonan chain which is interspersed with 2-linked rhamnosyl residues. The rhamnosyl residues are not randomly distributed in the chain but probably occur in units of rhamnosyl- (1 → 4)-galacturonosyl- (1 → 2)-rhamnosyl. This sequence appears to alternate with a homogalacturonan sequence containing approximately 8 residues of 4-linked galacturonic acid. About half of the rhamnosyl residues are branched, having a substituent attached to carbon 4. This is likely to be the site of attachment of the 4-linked galactan. The hydroxyprolyl oligo-arabinosides of the hydroxyproline-rich glycoprotein

  4. Beyond growth: novel functions for bacterial cell wall hydrolases.

    PubMed

    Wyckoff, Timna J; Taylor, Jennifer A; Salama, Nina R

    2012-11-01

    The peptidoglycan cell wall maintains turgor pressure and cell shape of most bacteria. Cell wall hydrolases are essential, together with synthases, for growth and daughter cell separation. Recent work in diverse organisms has uncovered new cell wall hydrolases that act autonomously or on neighboring cells to modulate invasion of prey cells, cell shape, innate immune detection, intercellular communication, and competitor lysis. The hydrolases involved in these processes catalyze the cleavage of bonds throughout the sugar and peptide moities of peptidoglycan. Phenotypes associated with these diverse hydrolases reveal new functions of the bacterial cell wall beyond growth and division.

  5. Monoclonal antibodies against plant cell wall polysaccharides

    SciTech Connect

    Hahn, M.G.; Bucheli, E.; Darvill, A.; Albersheim, P. )

    1989-04-01

    Monoclonal antibodies (McAbs) are useful tools to probe the structure of plant cell wall polysaccharides and to localize these polysaccharides in plant cells and tissues. Murine McAbs were generated against the pectic polysaccharide, rhamnogalacturonan I (RG-I), isolated from suspension-cultured sycamore cells. The McAbs that were obtained were grouped into three classes based upon their reactivities with a variety of plant polysaccharides and membrane glycoproteins. Eleven McAbs (Class I) recognize epitope(s) that appear to be immunodominant and are found in RG-I from sycamore and maize, citrus pectin, polygalacturonic acid, and membrane glycoproteins from suspension-cultured cells of sycamore, maize, tobacco, parsley, and soybean. A second group of five McAbs (Class II) recognize epitope(s) present in sycamore RG-I, but do not bind to any of the other polysaccharides or glycoproteins recognized by Class I. Lastly, one McAb (Class III) reacts with sycamore RG-I, sycamore and tamarind xyloglucan, and sycamore and rice glucuronoarabinoxylan, but does not bind to maize RG-I, polygalacturonic acid or the plant membrane glycoproteins recognized by Class I. McAbs in Classes II and III are likely to be useful in studies of the structure, biosynthesis and localization of plant cell wall polysaccharides.

  6. Plant cell wall proteomics: the leadership of Arabidopsis thaliana

    PubMed Central

    Albenne, Cécile; Canut, Hervé; Jamet, Elisabeth

    2013-01-01

    Plant cell wall proteins (CWPs) progressively emerged as crucial components of cell walls although present in minor amounts. Cell wall polysaccharides such as pectins, hemicelluloses, and cellulose represent more than 90% of primary cell wall mass, whereas hemicelluloses, cellulose, and lignins are the main components of lignified secondary walls. All these polymers provide mechanical properties to cell walls, participate in cell shape and prevent water loss in aerial organs. However, cell walls need to be modified and customized during plant development and in response to environmental cues, thus contributing to plant adaptation. CWPs play essential roles in all these physiological processes and particularly in the dynamics of cell walls, which requires organization and rearrangements of polysaccharides as well as cell-to-cell communication. In the last 10 years, plant cell wall proteomics has greatly contributed to a wider knowledge of CWPs. This update will deal with (i) a survey of plant cell wall proteomics studies with a focus on Arabidopsis thaliana; (ii) the main protein families identified and the still missing peptides; (iii) the persistent issue of the non-canonical CWPs; (iv) the present challenges to overcome technological bottlenecks; and (v) the perspectives beyond cell wall proteomics to understand CWP functions. PMID:23641247

  7. Global transcriptomic profiling of Schizosaccharomyces pombe in response to nitrosative stress.

    PubMed

    Biswas, Pranjal; Ghosh, Sanjay

    2015-03-10

    Excess production of nitric oxide (NO) and reactive nitrogen intermediates (RNIs) cause nitrosative stress on cells. Schizosaccharomyces pombe was used as a model to study nitrosative stress response. This is the first report on the global gene expression profile in response to NO in S. pombe using microarray. Among the 4824 genes reported for S. pombe, 818 were differentially expressed by at least 2-fold upon NO donor treatment. We previously showed that Pap1, the Activator Protein 1 transcription factor is required to combat nitrosative stress. In this study, the transcriptional response to NO in a null mutant for pap1 identified 45 genes that seem to be controlled by Pap1. Surprisingly, Pap1 regulated genes in S. pombe were distinctly different under nitrosative stress than those reported under oxidative stress. Genes of the pathway meiosis, cell cycle, spliceosome and oxidative phosphorylation were mostly affected under nitrosative stress in the fission yeast.

  8. Roles and regulation of plant cell walls surrounding plasmodesmata.

    PubMed

    Knox, J Paul; Benitez-Alfonso, Yoselin

    2014-12-01

    In plants, the intercellular transport of simple and complex molecules can occur symplastically through plasmodesmata. These are membranous channels embedded in cell walls that connect neighbouring cells. The properties of the cell walls surrounding plasmodesmata determine their transport capacity and permeability. These cell wall micro-domains are enriched in callose and have a characteristic pectin distribution. Cell wall modifications, leading to changes in plasmodesmata structure, have been reported to occur during development and in response to environmental signals. Cell wall remodelling enzymes target plasmodesmata to rapidly control intercellular communication in situ. Here we describe current knowledge on the composition of cell walls at plasmodesmata sites and on the proteins and signals that modify cell walls to regulate plasmodesmata aperture.

  9. Cell wall sorting of lipoproteins in Staphylococcus aureus.

    PubMed Central

    Navarre, W W; Daefler, S; Schneewind, O

    1996-01-01

    Many surface proteins are thought to be anchored to the cell wall of gram-positive organisms via their C termini, while the N-terminal domains of these molecules are displayed on the bacterial surface. Cell wall anchoring of surface proteins in Staphylococcus aureus requires both an N-terminal leader peptide and a C-terminal cell wall sorting signal. By fusing the cell wall sorting of protein A to the C terminus of staphylococcal beta-lactamase, we demonstrate here that lipoproteins can also be anchored to the cell wall of S. aureus. The topology of cell wall-anchored beta-lactamase is reminiscent of that described for Braun's murein lipoprotein in that the N terminus of the polypeptide chain is membrane anchored whereas the C-terminal end is tethered to the bacterial cell wall. PMID:8550464

  10. Cell Wall Loosening in the Fungus, Phycomyces blakesleeanus

    PubMed Central

    Ortega, Joseph K. E.; Truong, Jason T.; Munoz, Cindy M.; Ramirez, David G.

    2015-01-01

    A considerable amount of research has been conducted to determine how cell walls are loosened to produce irreversible wall deformation and expansive growth in plant and algal cells. The same cannot be said about fungal cells. Almost nothing is known about how fungal cells loosen their walls to produce irreversible wall deformation and expansive growth. In this study, anoxia is used to chemically isolate the wall from the protoplasm of the sporangiophores of Phycomyces blakesleeanus. The experimental results provide direct evidence of the existence of chemistry within the fungal wall that is responsible for wall loosening, irreversible wall deformation and elongation growth. In addition, constant-tension extension experiments are conducted on frozen-thawed sporangiophore walls to obtain insight into the wall chemistry and wall loosening mechanism. It is found that a decrease in pH to 4.6 produces creep extension in the frozen-thawed sporangiophore wall that is similar, but not identical, to that found in frozen-thawed higher plant cell walls. Experimental results from frozen-thawed and boiled sporangiophore walls suggest that protein activity may be involved in the creep extension. PMID:27135318

  11. Cell wall-associated kinases and pectin perception.

    PubMed

    Kohorn, Bruce D

    2016-01-01

    The pectin matrix of the angiosperm cell wall is regulated in both synthesis and modification and greatly influences the direction and extent of cell growth. Pathogens, herbivory and mechanical stresses all influence this pectin matrix and consequently plant form and function. The cell wall-associated kinases (WAKs) bind to pectin and regulate cell expansion or stress responses depending upon the state of the pectin. This review explores the WAKs in the context of cell wall biology and signal transduction pathways.

  12. Cortical microtubule rearrangements and cell wall patterning

    PubMed Central

    Oda, Yoshihisa

    2015-01-01

    Plant cortical microtubules, which form a highly ordered array beneath the plasma membrane, play essential roles in determining cell shape and function by directing the arrangement of cellulosic and non-cellulosic compounds on the cell surface. Interphase transverse arrays of cortical microtubules self-organize through their dynamic instability and inter-microtubule interactions, and by branch-form microtubule nucleation and severing. Recent studies revealed that distinct spatial signals including ROP GTPase, cellular geometry, and mechanical stress regulate the behavior of cortical microtubules at the subcellular and supercellular levels, giving rise to dramatic rearrangements in the cortical microtubule array in response to internal and external cues. Increasing evidence indicates that negative regulators of microtubules also contribute to the rearrangement of the cortical microtubule array. In this review, I summarize recent insights into how the rearrangement of the cortical microtubule array leads to proper, flexible cell wall patterning. PMID:25904930

  13. Cell Wall Invertase in Tobacco Crown Gall Cells 1

    PubMed Central

    Weil, Marion; Rausch, Thomas

    1990-01-01

    The cell wall invertase from an Agrobacterium tumefaciens-transformed Nicotiana tabacum cell line (SR1-C58) was purified. The heterogeneously glycosylated enzyme has the following properties: Mr 63,000, pH optimum at 4.7, Km sucrose 0.6 millimolar (at pH 4.7), pl 9.5. Enzyme activity is inhibited by micromolar concentrations of HgCl2 but is insensitive to H2O2, N-ethylmaleimide and dithiothreitol. Upon transfer of transformed cells from the stationary phase to fresh medium, a cycloheximide- and tunicamycin-sensitive de novo formation of cell wall invertase is demonstrated in the absence or presence of sucrose. While in an auxin mutant (lacking gene 1;SR1-3845) 1 micromolar 1-naphthaleneacetic acid led to a further increased activity, the wild-type transformed cell line (SR1-C58) responded with a decreased activity compared to the control. An analysis of cell wall invertase in and around tumors initiated with Agrobacterium tumefaciens (strain C58) on Nicotiana tabacum stem and Kalanchoë daigremontiana leaves revealed gradients of activity. The results indicate that the auxin-stimulated cell wall invertase is essential for the establishment of the tumor sink. Images Figure 1 PMID:16667892

  14. (Hydroxyproline-rich glycoproteins of the plant cell wall)

    SciTech Connect

    Varner, J.E.

    1990-01-01

    We are studying the chemistry and architecture of plant cells walls, the extracellular matrices that taken together shape the plant and provide mechanical support for the plant. Cell walls are dynamic structures that regulate, or are the site of, many physiological processes, in addition to being the cells' first line of defense against invading pathogens. In the past year we have examined the role of the cell wall enzyme ascorbic acid oxidase as related to the structure of the wall and its possible interactions with hydroxyproline-rich glycoproteins of the wall.

  15. Plant and algal cell walls: diversity and functionality

    PubMed Central

    Popper, Zoë A.; Ralet, Marie-Christine; Domozych, David S.

    2014-01-01

    Background Although plants and many algae (e.g. the Phaeophyceae, brown, and Rhodophyceae, red) are only very distantly related they are united in their possession of carbohydrate-rich cell walls, which are of integral importance being involved in many physiological processes. Furthermore, wall components have applications within food, fuel, pharmaceuticals, fibres (e.g. for textiles and paper) and building materials and have long been an active topic of research. As shown in the 27 papers in this Special Issue, as the major deposit of photosynthetically fixed carbon, and therefore energy investment, cell walls are of undisputed importance to the organisms that possess them, the photosynthetic eukaryotes (plants and algae). The complexities of cell wall components along with their interactions with the biotic and abiotic environment are becoming increasingly revealed. Scope The importance of plant and algal cell walls and their individual components to the function and survival of the organism, and for a number of industrial applications, are illustrated by the breadth of topics covered in this issue, which includes papers concentrating on various plants and algae, developmental stages, organs, cell wall components, and techniques. Although we acknowledge that there are many alternative ways in which the papers could be categorized (and many would fit within several topics), we have organized them as follows: (1) cell wall biosynthesis and remodelling, (2) cell wall diversity, and (3) application of new technologies to cell walls. Finally, we will consider future directions within plant cell wall research. Expansion of the industrial uses of cell walls and potentially novel uses of cell wall components are both avenues likely to direct future research activities. Fundamentally, it is the continued progression from characterization (structure, metabolism, properties and localization) of individual cell wall components through to defining their roles in almost every

  16. Enzymes and other agents that enhance cell wall extensibility

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1999-01-01

    Polysaccharides and proteins are secreted to the inner surface of the growing cell wall, where they assemble into a network that is mechanically strong, yet remains extensible until the cells cease growth. This review focuses on the agents that directly or indirectly enhance the extensibility properties of growing walls. The properties of expansins, endoglucanases, and xyloglucan transglycosylases are reviewed and their postulated roles in modulating wall extensibility are evaluated. A summary model for wall extension is presented, in which expansin is a primary agent of wall extension, whereas endoglucanases, xyloglucan endotransglycosylase, and other enzymes that alter wall structure act secondarily to modulate expansin action.

  17. Nedd8 processing enzymes in Schizosaccharomyces pombe

    PubMed Central

    2013-01-01

    Background Conjugation of the ubiquitin-like modifier Nedd8 to cullins is critical for the function of SCF-type ubiquitin ligases and thus facilitates ubiquitin conjugation and ultimately degradation of SCF substrates, including several cell cycle regulators. Like ubiquitin, Nedd8 is produced as a precursor that must first be processed before it becomes active. In Saccharomyces cerevisiae this is carried out exclusively by the enzyme Yuh1. Results Here we show that in the fission yeast, Schizosaccharomyces pombe, the Yuh1 orthologue, Uch1, is not the sole Nedd8 processing enzyme. Instead it appears that deubiquitylating enzymes can efficiently process the Nedd8 precursor in vivo. Conclusions Several enzymes contribute to Nedd8 precursor processing including a number of deubiquitylating enzymes. PMID:23496905

  18. Plant cell wall dynamics and wall-related susceptibility in plant-pathogen interactions.

    PubMed

    Bellincampi, Daniela; Cervone, Felice; Lionetti, Vincenzo

    2014-01-01

    The cell wall is a dynamic structure that often determines the outcome of the interactions between plants and pathogens. It is a barrier that pathogens need to breach to colonize the plant tissue. While fungal necrotrophs extensively destroy the integrity of the cell wall through the combined action of degrading enzymes, biotrophic fungi require a more localized and controlled degradation of the cell wall in order to keep the host cells alive and utilize their feeding structures. Also bacteria and nematodes need to degrade the plant cell wall at a certain stage of their infection process, to obtain nutrients for their growth. Plants have developed a system for sensing pathogens and monitoring the cell wall integrity, upon which they activate defense responses that lead to a dynamic cell wall remodeling required to prevent the disease. Pathogens, on the other hand, may exploit the host cell wall metabolism to support the infection. We review here the strategies utilized by both plants and pathogens to prevail in the cell wall battleground.

  19. Plant cell wall dynamics and wall-related susceptibility in plant–pathogen interactions

    PubMed Central

    Bellincampi, Daniela; Cervone, Felice; Lionetti, Vincenzo

    2014-01-01

    The cell wall is a dynamic structure that often determines the outcome of the interactions between plants and pathogens. It is a barrier that pathogens need to breach to colonize the plant tissue. While fungal necrotrophs extensively destroy the integrity of the cell wall through the combined action of degrading enzymes, biotrophic fungi require a more localized and controlled degradation of the cell wall in order to keep the host cells alive and utilize their feeding structures. Also bacteria and nematodes need to degrade the plant cell wall at a certain stage of their infection process, to obtain nutrients for their growth. Plants have developed a system for sensing pathogens and monitoring the cell wall integrity, upon which they activate defense responses that lead to a dynamic cell wall remodeling required to prevent the disease. Pathogens, on the other hand, may exploit the host cell wall metabolism to support the infection. We review here the strategies utilized by both plants and pathogens to prevail in the cell wall battleground. PMID:24904623

  20. Disruption of cell walls for enhanced lipid recovery

    DOEpatents

    Knoshaug, Eric P; Donohoe, Bryon S; Gerken, Henri; Laurens, Lieve; Van Wychen, Stefanie Rose

    2015-03-24

    Presented herein are methods of using cell wall degrading enzymes for recovery of internal lipid bodies from biomass sources such as algae. Also provided are algal cells that express at least one exogenous gene encoding a cell wall degrading enzyme and methods for recovering lipids from the cells.

  1. An Ancient Yeast for Young Geneticists: A Primer on the Schizosaccharomyces pombe Model System.

    PubMed

    Hoffman, Charles S; Wood, Valerie; Fantes, Peter A

    2015-10-01

    The fission yeast Schizosaccharomyces pombe is an important model organism for the study of eukaryotic molecular and cellular biology. Studies of S. pombe, together with studies of its distant cousin, Saccharomyces cerevisiae, have led to the discovery of genes involved in fundamental mechanisms of transcription, translation, DNA replication, cell cycle control, and signal transduction, to name but a few processes. However, since the divergence of the two species approximately 350 million years ago, S. pombe appears to have evolved less rapidly than S. cerevisiae so that it retains more characteristics of the common ancient yeast ancestor, causing it to share more features with metazoan cells. This Primer introduces S. pombe by describing the yeast itself, providing a brief description of the origins of fission yeast research, and illustrating some genetic and bioinformatics tools used to study protein function in fission yeast. In addition, a section on some key differences between S. pombe and S. cerevisiae is included for readers with some familiarity with budding yeast research but who may have an interest in developing research projects using S. pombe.

  2. An Ancient Yeast for Young Geneticists: A Primer on the Schizosaccharomyces pombe Model System

    PubMed Central

    Hoffman, Charles S.; Wood, Valerie; Fantes, Peter A.

    2015-01-01

    The fission yeast Schizosaccharomyces pombe is an important model organism for the study of eukaryotic molecular and cellular biology. Studies of S. pombe, together with studies of its distant cousin, Saccharomyces cerevisiae, have led to the discovery of genes involved in fundamental mechanisms of transcription, translation, DNA replication, cell cycle control, and signal transduction, to name but a few processes. However, since the divergence of the two species approximately 350 million years ago, S. pombe appears to have evolved less rapidly than S. cerevisiae so that it retains more characteristics of the common ancient yeast ancestor, causing it to share more features with metazoan cells. This Primer introduces S. pombe by describing the yeast itself, providing a brief description of the origins of fission yeast research, and illustrating some genetic and bioinformatics tools used to study protein function in fission yeast. In addition, a section on some key differences between S. pombe and S. cerevisiae is included for readers with some familiarity with budding yeast research but who may have an interest in developing research projects using S. pombe. PMID:26447128

  3. Calnexin is essential for survival under nitrogen starvation and stationary phase in Schizosaccharomyces pombe.

    PubMed

    Núñez, Andrés; Dulude, Dominic; Jbel, Mehdi; Rokeach, Luis A

    2015-01-01

    Cell fate is determined by the balance of conserved molecular mechanisms regulating death (apoptosis) and survival (autophagy). Autophagy is a process by which cells recycle their organelles and macromolecules through degradation within the vacuole in yeast and plants, and lysosome in metazoa. In the yeast Schizosaccharomyces pombe, autophagy is strongly induced under nitrogen starvation and in aging cells. Previously, we demonstrated that calnexin (Cnx1p), a highly conserved transmembrane chaperone of the endoplasmic reticulum (ER), regulates apoptosis under ER stress or inositol starvation. Moreover, we showed that in stationary phase, Cnx1p is cleaved into two moieties, L_Cnx1p and S_Cnx1p. Here, we show that the processing of Cnx1p is regulated by autophagy, induced by nitrogen starvation or cell aging. The cleavage of Cnx1p involves two vacuolar proteases: Isp6, which is essential for autophagy, and its paralogue Psp3. Blocking autophagy through the knockout of autophagy-related genes (atg) results in inhibition of both, the cleavage and the trafficking of Cnx1p from the ER to the vacuole. We demonstrate that Cnx1p is required for cell survival under nitrogen-starvation and in chronological aging cultures. The death of the mini_cnx1 mutant (overlapping S_cnx1p) cells is accompanied by accumulation of high levels of reactive-oxygen species (ROS), a slowdown in endocytosis and severe cell-wall defects. Moreover, mutant cells expressing only S_Cnx1p showed cell wall defects. Co-expressing mutant overlapping the L_Cnx1p and S_Cnx1p cleavage products reverses the death, ROS phenotype and cell wall defect to wild-type levels. As it is involved in both apoptosis and autophagy, Cnx1p could be a nexus for the crosstalk between these pro-death and pro-survival mechanisms. Ours, and observations in mammalian systems, suggest that the multiple roles of calnexin depend on its sub-cellular localization and on its cleavage. The use of S. pombe should assist in further

  4. Plant cell wall extensibility: connecting plant cell growth with cell wall structure, mechanics, and the action of wall-modifying enzymes.

    PubMed

    Cosgrove, Daniel J

    2016-01-01

    The advent of user-friendly instruments for measuring force/deflection curves of plant surfaces at high spatial resolution has resulted in a recent outpouring of reports of the 'Young's modulus' of plant cell walls. The stimulus for these mechanical measurements comes from biomechanical models of morphogenesis of meristems and other tissues, as well as single cells, in which cell wall stress feeds back to regulate microtubule organization, auxin transport, cellulose deposition, and future growth directionality. In this article I review the differences between elastic modulus and wall extensibility in the context of cell growth. Some of the inherent complexities, assumptions, and potential pitfalls in the interpretation of indentation force/deflection curves are discussed. Reported values of elastic moduli from surface indentation measurements appear to be 10- to >1000-fold smaller than realistic tensile elastic moduli in the plane of plant cell walls. Potential reasons for this disparity are discussed, but further work is needed to make sense of the huge range in reported values. The significance of wall stress relaxation for growth is reviewed and connected to recent advances and remaining enigmas in our concepts of how cellulose, hemicellulose, and pectins are assembled to make an extensible cell wall. A comparison of the loosening action of α-expansin and Cel12A endoglucanase is used to illustrate two different ways in which cell walls may be made more extensible and the divergent effects on wall mechanics.

  5. Evolution and diversity of green plant cell walls.

    PubMed

    Popper, Zoë A

    2008-06-01

    Plant cells are surrounded by a dynamic cell wall that performs many essential biological roles, including regulation of cell expansion, the control of tissue cohesion, ion-exchange and defence against microbes. Recent evidence shows that the suite of polysaccharides and wall proteins from which the plant cell wall is composed shows variation between monophyletic plant taxa. This is likely to have been generated during the evolution of plant groups in response to environmental stress. Understanding the natural variation and diversity that exists between cell walls from different taxa is key to facilitating their future exploitation and manipulation, for example by increasing lignocellulosic content or reducing its recalcitrance for use in biofuel generation.

  6. Two endogenous proteins that induce cell wall extension in plants

    NASA Technical Reports Server (NTRS)

    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.

  7. Deubiquitinating activity of Sdu1, a putative member of the PPPDE peptidase family, in Schizosaccharomyces pombe.

    PubMed

    Kim, Yunsik; Jo, Hannah; Lim, Chang-Jin

    2013-12-01

    The Schizosaccharomyces pombe sdu⁺ gene encoding a putative member of the PPPDE (Permuted Papain fold Peptidases of DsRNA viruses and Eukaryotes) superfamily was cloned into an Escherichia coli - yeast shuttle vector pRS316, resulting in the recombinant plasmid pYSTP. The determined nucleotide sequence carries 1207 bp, which would encode a protein of 201 amino acid residues. The S. pombe cells harboring pYSTP contained higher sdu1⁺ mRNA and deubiquitinating activity levels than the vector control cells, indicating that the sdu1⁺ gene is functioning. They exhibited a better growth in normal rich medium than the vector control cells. When shifted into the fresh medium containing hydrogen peroxide, menadione, or sodium nitroprusside, the S. pombe cells harboring pYSTP were able to grow reasonably well, while the growth of the vector control cells was arrested. The reactive oxygen species and total glutathione levels of the S. pombe cells harboring pYSTP were lower and higher than those of the vector control cells under the same stressful conditions, respectively. They exhibited a lower nitric oxide level than the vector control cells when subjected to sodium nitroprusside. Taken together, the sdu1⁺ gene encodes an actual protein having deubiquitinating activity and is involved in the response against oxidative and nitrosative stresses in S. pombe.

  8. Multidimensional solid-state NMR spectroscopy of plant cell walls.

    PubMed

    Wang, Tuo; Phyo, Pyae; Hong, Mei

    2016-09-01

    Plant biomass has become an important source of bio-renewable energy in modern society. The molecular structure of plant cell walls is difficult to characterize by most atomic-resolution techniques due to the insoluble and disordered nature of the cell wall. Solid-state NMR (SSNMR) spectroscopy is uniquely suited for studying native hydrated plant cell walls at the molecular level with chemical resolution. Significant progress has been made in the last five years to elucidate the molecular structures and interactions of cellulose and matrix polysaccharides in plant cell walls. These studies have focused on primary cell walls of growing plants in both the dicotyledonous and grass families, as represented by the model plants Arabidopsis thaliana, Brachypodium distachyon, and Zea mays. To date, these SSNMR results have shown that 1) cellulose, hemicellulose, and pectins form a single network in the primary cell wall; 2) in dicot cell walls, the protein expansin targets the hemicellulose-enriched region of the cellulose microfibril for its wall-loosening function; and 3) primary wall cellulose has polymorphic structures that are distinct from the microbial cellulose structures. This article summarizes these key findings, and points out future directions of investigation to advance our fundamental understanding of plant cell wall structure and function.

  9. Multidimensional solid-state NMR spectroscopy of plant cell walls.

    PubMed

    Wang, Tuo; Phyo, Pyae; Hong, Mei

    2016-09-01

    Plant biomass has become an important source of bio-renewable energy in modern society. The molecular structure of plant cell walls is difficult to characterize by most atomic-resolution techniques due to the insoluble and disordered nature of the cell wall. Solid-state NMR (SSNMR) spectroscopy is uniquely suited for studying native hydrated plant cell walls at the molecular level with chemical resolution. Significant progress has been made in the last five years to elucidate the molecular structures and interactions of cellulose and matrix polysaccharides in plant cell walls. These studies have focused on primary cell walls of growing plants in both the dicotyledonous and grass families, as represented by the model plants Arabidopsis thaliana, Brachypodium distachyon, and Zea mays. To date, these SSNMR results have shown that 1) cellulose, hemicellulose, and pectins form a single network in the primary cell wall; 2) in dicot cell walls, the protein expansin targets the hemicellulose-enriched region of the cellulose microfibril for its wall-loosening function; and 3) primary wall cellulose has polymorphic structures that are distinct from the microbial cellulose structures. This article summarizes these key findings, and points out future directions of investigation to advance our fundamental understanding of plant cell wall structure and function. PMID:27552739

  10. Impregnation of softwood cell walls with melamine-formaldehyde resin.

    PubMed

    Gindl, W; Zargar-Yaghubi, F; Wimmer, R

    2003-05-01

    Melamine-formaldehyde (MF) resin impregnation has shown considerable potential to improve a number of wood properties, such as surface hardness and weathering resistance. In this study, selected factors influencing the uptake of MF resin into the cell wall of softwood were studied. Using UV-microspectroscopy, it could be shown that water soluble MF diffused well into the secondary cell wall and the middle lamella. Concentrations as high as 24% (v/v) were achieved after an impregnation of 20 h. High cell wall moisture content, high water content of the resin used for impregnation, and low extractive content are factors which are favourable for MF resin uptake into the cell wall. For dry cell walls, solvent exchange drying improved resin uptake to a similar extent, as was the case when cell walls were soaked in water.

  11. Impregnation of softwood cell walls with melamine-formaldehyde resin.

    PubMed

    Gindl, W; Zargar-Yaghubi, F; Wimmer, R

    2003-05-01

    Melamine-formaldehyde (MF) resin impregnation has shown considerable potential to improve a number of wood properties, such as surface hardness and weathering resistance. In this study, selected factors influencing the uptake of MF resin into the cell wall of softwood were studied. Using UV-microspectroscopy, it could be shown that water soluble MF diffused well into the secondary cell wall and the middle lamella. Concentrations as high as 24% (v/v) were achieved after an impregnation of 20 h. High cell wall moisture content, high water content of the resin used for impregnation, and low extractive content are factors which are favourable for MF resin uptake into the cell wall. For dry cell walls, solvent exchange drying improved resin uptake to a similar extent, as was the case when cell walls were soaked in water. PMID:12507874

  12. A protective role of methionine-R-sulfoxide reductase against cadmium in Schizosaccharomyces pombe.

    PubMed

    Lim, Chang-Jin; Jo, Hannah; Kim, Kyunghoon

    2014-11-01

    The Schizosaccharomyces pombe cells harboring the methionine- R-sulfoxide reductase (MsrB)-overexpressing recombinant plasmid pFMetSO exhibited better growth than vector control cells, when shifted into fresh medium containing cadmium chloride (abbreviated as Cd). Although both groups of cells contained enhanced reactive oxygen species (ROS) and nitric oxide (NO) levels in the presence of Cd, ROS and NO levels were significantly lower in the S. pombe cells harboring pFMetSO than in vector control cells. Conversely, the S. pombe cells harboring pFMetSO possessed higher total glutathione (GSH) levels and a greater reduced/oxidized GSH ratio than vector control cells under the same conditions.

  13. Conjugation in S. pombe: identification of a microtubule-organising centre, a requirement for microtubules and a role for Mad2.

    PubMed

    Petersen, J; Heitz, M J; Hagan, I M

    1998-08-27

    During the G1 phase of the cell cycle, cells of the fission yeast Schizosaccharomyces pombe can be induced to mate by nitrogen starvation and the presence of mating pheromones. Polarised growth towards cells of the opposite mating type (P or M) leads to the formation of a projection tip and, upon contact, localised cell wall degradation results in conjugation and cell fusion [1]. Here, we have investigated the role of microtubules in this process. We describe a previously unidentified microtubule-organising centre (MTOC) that forms at projection tips upon cell-to-cell contact, before cells fuse. Treatment of mating cells with the microtubule-destabilising drug thiabendazole (TBZ) showed that microtubule integrity was required for mating at two distinct stages: during projection tip formation and cell fusion. Projection tip formation requires filamentous (F) actin function [2] and microtubules are required for the localisation of F actin to the projection tip. We also identify a role during mating for Mad2--a mitotic checkpoint protein that is required in all eukaryotes to maintain the mitotic state in response to microtubule depolymerisation [3]. S. pombe mad2 mutant cells were compromised in their ability to mate upon removal of TBZ, indicating that in fission yeast, in the absence of microtubules, Mad2 is also required to maintain mating competence.

  14. Bacterial cell wall composition and the influence of antibiotics by cell-wall and whole-cell NMR.

    PubMed

    Romaniuk, Joseph A H; Cegelski, Lynette

    2015-10-01

    The ability to characterize bacterial cell-wall composition and structure is crucial to understanding the function of the bacterial cell wall, determining drug modes of action and developing new-generation therapeutics. Solid-state NMR has emerged as a powerful tool to quantify chemical composition and to map cell-wall architecture in bacteria and plants, even in the context of unperturbed intact whole cells. In this review, we discuss solid-state NMR approaches to define peptidoglycan composition and to characterize the modes of action of old and new antibiotics, focusing on examples in Staphylococcus aureus. We provide perspectives regarding the selected NMR strategies as we describe the exciting and still-developing cell-wall and whole-cell NMR toolkit. We also discuss specific discoveries regarding the modes of action of vancomycin analogues, including oritavancin, and briefly address the reconsideration of the killing action of β-lactam antibiotics. In such chemical genetics approaches, there is still much to be learned from perturbations enacted by cell-wall assembly inhibitors, and solid-state NMR approaches are poised to address questions of cell-wall composition and assembly in S. aureus and other organisms.

  15. Bacterial cell wall composition and the influence of antibiotics by cell-wall and whole-cell NMR

    PubMed Central

    Romaniuk, Joseph A. H.; Cegelski, Lynette

    2015-01-01

    The ability to characterize bacterial cell-wall composition and structure is crucial to understanding the function of the bacterial cell wall, determining drug modes of action and developing new-generation therapeutics. Solid-state NMR has emerged as a powerful tool to quantify chemical composition and to map cell-wall architecture in bacteria and plants, even in the context of unperturbed intact whole cells. In this review, we discuss solid-state NMR approaches to define peptidoglycan composition and to characterize the modes of action of old and new antibiotics, focusing on examples in Staphylococcus aureus. We provide perspectives regarding the selected NMR strategies as we describe the exciting and still-developing cell-wall and whole-cell NMR toolkit. We also discuss specific discoveries regarding the modes of action of vancomycin analogues, including oritavancin, and briefly address the reconsideration of the killing action of β-lactam antibiotics. In such chemical genetics approaches, there is still much to be learned from perturbations enacted by cell-wall assembly inhibitors, and solid-state NMR approaches are poised to address questions of cell-wall composition and assembly in S. aureus and other organisms. PMID:26370936

  16. (The structure of pectins from cotton suspension culture cell walls)

    SciTech Connect

    Mort, A.

    1990-01-01

    We have made progress on several projects to do with determining the structure of pectins. These include: (1) Devising a new sensitive method to determine the degree of methyl esterification (DOM) of pectins; (2) solubilization of all of RGI from cotton cell walls; (3) solubilization of RGII from cotton cell walls; (4) characterization of xyloglucan from cotton cell walls; and (5) investigation giving an indication of a cross-link between extension and pectin.

  17. An arabidopsis gene regulatory network for secondary cell wall synthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The plant cell wall is an important factor for determining cell shape, function and response to the environment. Secondary cell walls, such as those found in xylem, are composed of cellulose, hemicelluloses and lignin and account for the bulk of plant biomass. The coordination between transcriptiona...

  18. Cell wall degradation in the autolysis of filamentous fungi.

    PubMed

    Perez-Leblic, M I; Reyes, F; Martinez, M J; Lahoz, R

    1982-12-27

    A systematic study on autolysis of the cell walls of fungi has been made on Neurospora crassa, Botrytis cinerea, Polystictus versicolor, Aspergillus nidulans, Schizophyllum commune, Aspergillus niger, and Mucor mucedo. During autolysis each fungus produces the necessary lytic enzymes for its autodegradation. From autolyzed cultures of each fungus enzymatic precipitates were obtained. The degree of lysis of the cell walls, obtained from non-autolyzed mycelia, was studied by incubating these cell walls with and without a supply of their own lytic enzymes. The degree of lysis increased with the incubation time and generally was higher with a supply of lytic enzymes. Cell walls from mycelia of different ages were obtained. A higher degree of lysis was always found, in young cell walls than in older cell walls, when exogenous lytic enzymes were present. In all the fungi studied, there is lysis of the cell walls during autolysis. This is confirmed by the change of the cell wall structure as well as by the degree of lysis reached by the cell wall and the release of substances, principally glucose and N-acetylglucosamine in the medium.

  19. Secondary cell walls: biosynthesis, patterned deposition and transcriptional regulation.

    PubMed

    Zhong, Ruiqin; Ye, Zheng-Hua

    2015-02-01

    Secondary walls are mainly composed of cellulose, hemicelluloses (xylan and glucomannan) and lignin, and are deposited in some specialized cells, such as tracheary elements, fibers and other sclerenchymatous cells. Secondary walls provide strength to these cells, which lend mechanical support and protection to the plant body and, in the case of tracheary elements, enable them to function as conduits for transporting water. Formation of secondary walls is a complex process that requires the co-ordinated expression of secondary wall biosynthetic genes, biosynthesis and targeted secretion of secondary wall components, and patterned deposition and assembly of secondary walls. Here, we provide a comprehensive review of genes involved in secondary wall biosynthesis and deposition. Most of the genes involved in the biosynthesis of secondary wall components, including cellulose, xylan, glucomannan and lignin, have been identified and their co-ordinated activation has been shown to be mediated by a transcriptional network encompassing the secondary wall NAC and MYB master switches and their downstream transcription factors. It has been demonstrated that cortical microtubules and microtubule-associated proteins play important roles in the targeted secretion of cellulose synthase complexes, the oriented deposition of cellulose microfibrils and the patterned deposition of secondary walls. Further investigation of many secondary wall-associated genes with unknown functions will provide new insights into the mechanisms controlling the formation of secondary walls that constitute the bulk of plant biomass.

  20. Visualization of cellulose synthases in Arabidopsis secondary cell walls.

    PubMed

    Watanabe, Y; Meents, M J; McDonnell, L M; Barkwill, S; Sampathkumar, A; Cartwright, H N; Demura, T; Ehrhardt, D W; Samuels, A L; Mansfield, S D

    2015-10-01

    Cellulose biosynthesis in plant secondary cell walls forms the basis of vascular development in land plants, with xylem tissues constituting the vast majority of terrestrial biomass. We used plant lines that contained an inducible master transcription factor controlling xylem cell fate to quantitatively image fluorescently tagged cellulose synthase enzymes during cellulose deposition in living protoxylem cells. The formation of secondary cell wall thickenings was associated with a redistribution and enrichment of CESA7-containing cellulose synthase complexes (CSCs) into narrow membrane domains. The velocities of secondary cell wall-specific CSCs were faster than those of primary cell wall CSCs during abundant cellulose production. Dynamic intracellular of endomembranes, in combination with increased velocity and high density of CSCs, enables cellulose to be synthesized rapidly in secondary cell walls. PMID:26450210

  1. Visualization of cellulose synthases in Arabidopsis secondary cell walls.

    PubMed

    Watanabe, Y; Meents, M J; McDonnell, L M; Barkwill, S; Sampathkumar, A; Cartwright, H N; Demura, T; Ehrhardt, D W; Samuels, A L; Mansfield, S D

    2015-10-01

    Cellulose biosynthesis in plant secondary cell walls forms the basis of vascular development in land plants, with xylem tissues constituting the vast majority of terrestrial biomass. We used plant lines that contained an inducible master transcription factor controlling xylem cell fate to quantitatively image fluorescently tagged cellulose synthase enzymes during cellulose deposition in living protoxylem cells. The formation of secondary cell wall thickenings was associated with a redistribution and enrichment of CESA7-containing cellulose synthase complexes (CSCs) into narrow membrane domains. The velocities of secondary cell wall-specific CSCs were faster than those of primary cell wall CSCs during abundant cellulose production. Dynamic intracellular of endomembranes, in combination with increased velocity and high density of CSCs, enables cellulose to be synthesized rapidly in secondary cell walls.

  2. Cell wall structure and biogenesis in Aspergillus species.

    PubMed

    Yoshimi, Akira; Miyazawa, Ken; Abe, Keietsu

    2016-09-01

    Aspergillus species are among the most important filamentous fungi from the viewpoints of industry, pathogenesis, and mycotoxin production. Fungal cells are exposed to a variety of environmental stimuli, including changes in osmolality, temperature, and pH, which create stresses that primarily act on fungal cell walls. In addition, fungal cell walls are the first interactions with host cells in either human or plants. Thus, understanding cell wall structure and the mechanism of their biogenesis is important for the industrial, medical, and agricultural fields. Here, we provide a systematic review of fungal cell wall structure and recent findings regarding the cell wall integrity signaling pathways in aspergilli. This accumulated knowledge will be useful for understanding and improving the use of industrial aspergilli fermentation processes as well as treatments for some fungal infections.

  3. Unicellular Algal Growth: A Biomechanical Approach to Cell Wall Dynamics

    NASA Astrophysics Data System (ADS)

    Kam, Royce; Levine, Herbert

    1997-11-01

    We model a growing cell in a calcium solution as an elastic shell on short time scales. The turgor pressure and elastic properties (Young's modulus, thickness) of the cell wall determine a stressed cell shape. Enzyme-mediated relaxation of the unstressed toward the stressed configuration results in a slow (plastic) deformation of the cell. The cell wall thickness is then modulated by calcium-mediated fusion of material and elongation. We analyze small perturbations to a circular cell and find an instability related to modulations of the wall thickness, leading to growth rates which peak at a finite wave number.

  4. Preparation of Cell Wall Antigens of Staphylococcus aureus

    PubMed Central

    Kowalski, J. J.; Tipper, Donald J.; Berman, David T.

    1970-01-01

    Cell walls were prepared from Staphylococcus aureus strains Copenhagen and 263 by high-speed mixing in the presence of glass beads followed by differential centrifugation. Insoluble peptidoglycan complexes were derived from cell walls by extraction of teichoic acid with 10% trichloroacetic acid. Intact teichoic acid was prepared from each strain by digestion of cell walls with lysostaphin and isolated by column chromatography. Soluble glycopeptide (peptidoglycan in which only the glycan has been fragmented) and the stable complex of teichoic acid with glycopeptide were prepared by digestion of cell walls with Chalaropsis B endo-N-acetylmuramidase and were separated by column chromatography. Amino acid and amino sugar contents of walls and subunits of walls were comparable to those reported by others. Images PMID:16557799

  5. Screening and characterization of plant cell walls using carbohydrate microarrays.

    PubMed

    Sørensen, Iben; Willats, William G T

    2011-01-01

    Plant cells are surrounded by cell walls built largely from complex carbohydrates. The primary walls of growing plant cells consist of interdependent networks of three polysaccharide classes: cellulose, cross-linking glycans (also known as hemicelluloses), and pectins. Cellulose microfibrils are tethered together by cross-linking glycans, and this assembly forms the major load-bearing component of primary walls, which is infiltrated with pectic polymers. In the secondary walls of woody tissues, pectins are much reduced and walls are reinforced with the phenolic polymer lignin. Plant cell walls are essential for plant life and also have numerous industrial applications, ranging from wood to nutraceuticals. Enhancing our knowledge of cell wall biology and the effective use of cell wall materials is dependent to a large extent on being able to analyse their fine structures. We have developed a suite of techniques based on microarrays probed with monoclonal antibodies with specificity for cell wall components, and here we present practical protocols for this type of analysis.

  6. Ultrastructural localization of capsules, cell wall polysaccharide, cell wall proteins, and F antigen in pneumococci.

    PubMed Central

    Skov Sørensen, U B; Blom, J; Birch-Andersen, A; Henrichsen, J

    1988-01-01

    The localization of pneumococcal capsular and cell wall antigens was examined by immunoelectron microscopy. C polysaccharide (C-Ps), a common component of all pneumococci, was uniformly distributed on both the inside and outside of the cell walls. The thickness of the C-Ps varied with the strain. Encapsulated strains were covered by varied amounts of capsular polysaccharide concealing the C-Ps of the bacteria so as to render it inaccessible to anti-C-Ps antibodies. In addition to C-Ps, protein antigens were demonstrable on the surface of nonencapsulated pneumococci. The proteins were not masked by the C-Ps layer. An extra layer on the cell walls was conspicuous on electron micrographs of both rough and encapsulated pneumococci. The nature of this extra layer has not been disclosed. F antigen, another common antigen of pneumococci, was uniformly distributed on the surface of the plasma membranes. During the course of the experimental work a reproducible method of gold labeling immunoglobulins was developed. Images PMID:3397179

  7. Genes and plant cell walls: a difficult relationship.

    PubMed

    Wojtaszek, P

    2000-08-01

    Chemical information, carried by genes, is one of several types of information important for the functioning of cells and organisms. While genes govern the two-dimensional flow of information, the cell walls are at the basis of a structural, three-dimensional framework of plant form and growth. Recent data show the walls to be a cellular 'organelle' undergoing dynamic changes in response to a plethora of stimuli. In this review, an integrated approach, rooted in the organismal perspective, is taken to consider the role of cell walls in the biology of plants. First, the complexity of molecular and biochemical events leading to the biosynthesis of wall components is described within the framework of its spatial cellular organisation, and the major regulatory check-points are characterised. Second, cell walls form a structural and functional continuum within the whole plant and thus could be defined in relation to the protoplasts that produce them and in relation to the plant itself. Model systems of suspension-cultured cells are used to reveal the existence of a bidirectional exchange of information between the protoplast and its walls. The 'plasticity' of plant cell reactions, seen in defence responses or in changes in wall composition, to e.g. stress, plant growth regulators or chemical agents as well as the role of cell walls and/or wall components in somatic embryogenesis are also discussed. Third, being a continuum within the plant body, the walls fulfil vital functions in plant growth and development. The examples characterised include the determination of cellular polarity and the plane of cell division, cytokinesis, and the role of plasmodesmata in cell-to-cell communication and the formation of functional symplastic domains. Fourth, the exocellular control of morphogenetic processes is described and the potential of cell walls as determinants or reservoirs of positional information is indicated. Particular emphasis is put on the (bio)chemical signals coming

  8. Assembly and enlargement of the primary cell wall in plants

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1997-01-01

    Growing plant cells are shaped by an extensible wall that is a complex amalgam of cellulose microfibrils bonded noncovalently to a matrix of hemicelluloses, pectins, and structural proteins. Cellulose is synthesized by complexes in the plasma membrane and is extruded as a self-assembling microfibril, whereas the matrix polymers are secreted by the Golgi apparatus and become integrated into the wall network by poorly understood mechanisms. The growing wall is under high tensile stress from cell turgor and is able to enlarge by a combination of stress relaxation and polymer creep. A pH-dependent mechanism of wall loosening, known as acid growth, is characteristic of growing walls and is mediated by a group of unusual wall proteins called expansins. Expansins appear to disrupt the noncovalent bonding of matrix hemicelluloses to the microfibril, thereby allowing the wall to yield to the mechanical forces generated by cell turgor. Other wall enzymes, such as (1-->4) beta-glucanases and pectinases, may make the wall more responsive to expansin-mediated wall creep whereas pectin methylesterases and peroxidases may alter the wall so as to make it resistant to expansin-mediated creep.

  9. Structural Studies of Complex Carbohydrates of Plant Cell Walls

    SciTech Connect

    Darvill, Alan; Hahn, Michael G.; O'Neill, Malcolm A.; York, William S.

    2015-02-17

    Most of the solar energy captured by land plants is converted into the polysaccharides (cellulose, hemicellulose, and pectin) that are the predominant components of the cell wall. These walls, which account for the bulk of plant biomass, have numerous roles in the growth and development of plants. Moreover, these walls have a major impact on human life as they are a renewable source of biomass, a source of diverse commercially useful polymers, a major component of wood, and a source of nutrition for humans and livestock. Thus, understanding the molecular mechanisms that lead to wall assembly and how cell walls and their component polysaccharides contribute to plant growth and development is essential to improve and extend the productivity and value of plant materials. The proposed research will develop and apply advanced analytical and immunological techniques to study specific changes in the structures and interactions of the hemicellulosic and pectic polysaccharides that occur during differentiation and in response to genetic modification and chemical treatments that affect wall biosynthesis. These new techniques will make it possible to accurately characterize minute amounts of cell wall polysaccharides so that subtle changes in structure that occur in individual cell types can be identified and correlated to the physiological or developmental state of the plant. Successful implementation of this research will reveal fundamental relationships between polysaccharide structure, cell wall architecture, and cell wall functions.

  10. Engineering the Oryza sativa cell wall with rice NAC transcription factors regulating secondary wall formation

    PubMed Central

    Yoshida, Kouki; Sakamoto, Shingo; Kawai, Tetsushi; Kobayashi, Yoshinori; Sato, Kazuhito; Ichinose, Yasunori; Yaoi, Katsuro; Akiyoshi-Endo, Miho; Sato, Hiroko; Takamizo, Tadashi; Ohme-Takagi, Masaru; Mitsuda, Nobutaka

    2013-01-01

    Plant tissues that require structural rigidity synthesize a thick, strong secondary cell wall of lignin, cellulose and hemicelluloses in a complicated bridged structure. Master regulators of secondary wall synthesis were identified in dicots, and orthologs of these regulators have been identified in monocots, but regulation of secondary cell wall formation in monocots has not been extensively studied. Here we demonstrate that the rice transcription factors SECONDARY WALL NAC DOMAIN PROTEINs (SWNs) can regulate secondary wall formation in rice (Oryza sativa) and are potentially useful for engineering the monocot cell wall. The OsSWN1 promoter is highly active in sclerenchymatous cells of the leaf blade and less active in xylem cells. By contrast, the OsSWN2 promoter is highly active in xylem cells and less active in sclerenchymatous cells. OsSWN2 splicing variants encode two proteins; the shorter protein (OsSWN2S) has very low transcriptional activation ability, but the longer protein (OsSWN2L) and OsSWN1 have strong transcriptional activation ability. In rice, expression of an OsSWN2S chimeric repressor, driven by the OsSWN2 promoter, resulted in stunted growth and para-wilting (leaf rolling and browning under normal water conditions) due to impaired vascular vessels. The same OsSWN2S chimeric repressor, driven by the OsSWN1 promoter, caused a reduction of cell wall thickening in sclerenchymatous cells, a drooping leaf phenotype, reduced lignin and xylose contents and increased digestibility as forage. These data suggest that OsSWNs regulate secondary wall formation in rice and manipulation of OsSWNs may enable improvements in monocotyledonous crops for forage or biofuel applications. PMID:24098302

  11. Engineering the Oryza sativa cell wall with rice NAC transcription factors regulating secondary wall formation.

    PubMed

    Yoshida, Kouki; Sakamoto, Shingo; Kawai, Tetsushi; Kobayashi, Yoshinori; Sato, Kazuhito; Ichinose, Yasunori; Yaoi, Katsuro; Akiyoshi-Endo, Miho; Sato, Hiroko; Takamizo, Tadashi; Ohme-Takagi, Masaru; Mitsuda, Nobutaka

    2013-01-01

    Plant tissues that require structural rigidity synthesize a thick, strong secondary cell wall of lignin, cellulose and hemicelluloses in a complicated bridged structure. Master regulators of secondary wall synthesis were identified in dicots, and orthologs of these regulators have been identified in monocots, but regulation of secondary cell wall formation in monocots has not been extensively studied. Here we demonstrate that the rice transcription factors SECONDARY WALL NAC DOMAIN PROTEINs (SWNs) can regulate secondary wall formation in rice (Oryza sativa) and are potentially useful for engineering the monocot cell wall. The OsSWN1 promoter is highly active in sclerenchymatous cells of the leaf blade and less active in xylem cells. By contrast, the OsSWN2 promoter is highly active in xylem cells and less active in sclerenchymatous cells. OsSWN2 splicing variants encode two proteins; the shorter protein (OsSWN2S) has very low transcriptional activation ability, but the longer protein (OsSWN2L) and OsSWN1 have strong transcriptional activation ability. In rice, expression of an OsSWN2S chimeric repressor, driven by the OsSWN2 promoter, resulted in stunted growth and para-wilting (leaf rolling and browning under normal water conditions) due to impaired vascular vessels. The same OsSWN2S chimeric repressor, driven by the OsSWN1 promoter, caused a reduction of cell wall thickening in sclerenchymatous cells, a drooping leaf phenotype, reduced lignin and xylose contents and increased digestibility as forage. These data suggest that OsSWNs regulate secondary wall formation in rice and manipulation of OsSWNs may enable improvements in monocotyledonous crops for forage or biofuel applications. PMID:24098302

  12. 7. ENGINE TEST CELL BUILDING INTERIOR. WALL MAP IN CENTRAL ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    7. ENGINE TEST CELL BUILDING INTERIOR. WALL MAP IN CENTRAL BASEMENT OFFICE AREA. LOOKING SOUTHWEST. - Fairchild Air Force Base, Engine Test Cell Building, Near intersection of Arnold Street & George Avenue, Spokane, Spokane County, WA

  13. The role of wall calcium in the extension of cell walls of soybean hypocotyls

    NASA Technical Reports Server (NTRS)

    Virk, S. S.; Cleland, R. E.

    1990-01-01

    Calcium crosslinks are load-bearing bonds in soybean (Glycine max (L.) Merr.) hypocotyl cell walls, but they are not the same load-bearing bonds that are broken during acid-mediated cell elongation. This conclusion is reached by studying the relationship between wall calcium, pH and the facilitated creep of frozen-thawed soybean hypocotyl sections. Supporting data include the following observations: 1) 2-[(2-bis-[carboxymethyl]amino-5-methylphenoxy)methyl]-6-methoxy-8-bis[car boxymethyl]aminoquinoline (Quin 2) and ethylene glycol-bis(2-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) caused only limited facilitated creep as compared with acid, despite removal of comparable or larger amounts of wall calcium; 2) the pH-response curves for calcium removal and acid-facilitated creep were different; 3) reversible acid-extension occurred even after removal of almost all wall calcium with Quin 2; and 4) growth of abraded sections did not involve a proportional loss of wall calcium. Removal of wall calcium, however, increased the capacity of the walls to undergo acid-facilitated creep. These data indicate that breakage of calcium crosslinks is not a major mechanism of cell-wall loosening in soybean hypocotyl tissues.

  14. Epigenetic Regulation of Chromatin States in Schizosaccharomyces pombe.

    PubMed

    Allshire, Robin C; Ekwall, Karl

    2015-07-01

    This article discusses the advances made in epigenetic research using the model organism fission yeast Schizosaccharomyces pombe. S. pombe has been used for epigenetic research since the discovery of position effect variegation (PEV). This is a phenomenon in which a transgene inserted within heterochromatin is variably expressed, but can be stably inherited in subsequent cell generations. PEV occurs at centromeres, telomeres, ribosomal DNA (rDNA) loci, and mating-type regions of S. pombe chromosomes. Heterochromatin assembly in these regions requires enzymes that modify histones and the RNA interference (RNAi) machinery. One of the key histone-modifying enzymes is the lysine methyltransferase Clr4, which methylates histone H3 on lysine 9 (H3K9), a classic hallmark of heterochromatin. The kinetochore is assembled on specialized chromatin in which histone H3 is replaced by the variant CENP-A. Studies in fission yeast have contributed to our understanding of the establishment and maintenance of CENP-A chromatin and the epigenetic activation and inactivation of centromeres.

  15. On-off switches for secondary cell wall biosynthesis.

    PubMed

    Wang, Huan-Zhong; Dixon, Richard A

    2012-03-01

    Secondary cell walls provide plants with rigidity and strength to support their body weight and ensure water and nutrient transport. They also provide textiles, timber, and potentially second-generation biofuels for human use. Genes responsible for synthesis of the different cell wall components, namely cellulose, hemicelluloses, and lignin, are coordinately expressed and under transcriptional regulation. In the past several years, cell wall-related NAC and MYB transcription factors have been intensively investigated in different species and shown to be master switches of secondary cell wall biosynthesis. Positive and negative regulators, which function upstream of NAC master switches, have also been identified in different plant tissues. Further elucidation of the regulatory mechanisms of cell wall synthesis will facilitate the engineering of plant feedstocks suitable for biofuel production. PMID:22138968

  16. Signaling role of oligogalacturonides derived during cell wall degradation

    PubMed Central

    Vallarino, José G.; Osorio, Sonia

    2012-01-01

    In addition to the role of the cell wall as a physical barrier against pathogens, some of its constituents, such as pectin-derived oligogalacturonides (OGAs) are essential components to trigger signaling pathways that induce rapid defense responses. Many pathogens directly penetrate the cell wall to access water and nutrients of the plant protoplast, and a rigid cell wall can fend off pathogen attack by forming an impenetrable physical barrier. Thus, cell wall integrity sensing is one mechanism by which plants may detect pathogen attack. Moreover, when the plant-pathogen interaction occurred, OGAs released during cell wall modification can trigger plant defense (e.g., production of reactive oxygen species, production of anti-microbial metabolites and synthesis of pathogenesis-related proteins). This review documents and discusses studies suggesting that OGAs play a dual signaling role during pathogen attack by inducing defense responses and plant architecture adjustment. PMID:22918501

  17. Methods for degrading or converting plant cell wall polysaccharides

    DOEpatents

    Berka, Randy; Cherry, Joel

    2008-08-19

    The present invention relates to methods for converting plant cell wall polysaccharides into one or more products, comprising: treating the plant cell wall polysaccharides with an effective amount of a spent whole fermentation broth of a recombinant microorganism, wherein the recombinant microorganism expresses one or more heterologous genes encoding enzymes which degrade or convert the plant cell wall polysaccharides into the one or more products. The present invention also relates to methods for producing an organic substance, comprising: (a) saccharifying plant cell wall polysaccharides with an effective amount of a spent whole fermentation broth of a recombinant microorganism, wherein the recombinant microorganism expresses one or more heterologous genes encoding enzymes which degrade or convert the plant cell wall polysaccharides into saccharified material; (b) fermenting the saccharified material of step (a) with one or more fermenting microoganisms; and (c) recovering the organic substance from the fermentation.

  18. Collenchyma: a versatile mechanical tissue with dynamic cell walls

    PubMed Central

    Leroux, Olivier

    2012-01-01

    Background Collenchyma has remained in the shadow of commercially exploited mechanical tissues such as wood and fibres, and therefore has received little attention since it was first described. However, collenchyma is highly dynamic, especially compared with sclerenchyma. It is the main supporting tissue of growing organs with walls thickening during and after elongation. In older organs, collenchyma may become more rigid due to changes in cell wall composition or may undergo sclerification through lignification of newly deposited cell wall material. While much is known about the systematic and organographic distribution of collenchyma, there is rather less information regarding the molecular architecture and properties of its cell walls. Scope and conclusions This review summarizes several aspects that have not previously been extensively discussed including the origin of the term ‘collenchyma’ and the history of its typology. As the cell walls of collenchyma largely determine the dynamic characteristics of this tissue, I summarize the current state of knowledge regarding their structure and molecular composition. Unfortunately, to date, detailed studies specifically focusing on collenchyma cell walls have not been undertaken. However, generating a more detailed understanding of the structural and compositional modifications associated with the transition from plastic to elastic collenchyma cell wall properties is likely to provide significant insights into how specific configurations of cell wall polymers result in specific functional properties. This approach, focusing on architecture and functional properties, is likely to provide improved clarity on the controversial definition of collenchyma. PMID:22933416

  19. Structure of plant cell walls: XIX. Isolation and characterization of wall polysaccharides from suspension-cultured Douglas fir cells

    SciTech Connect

    Thomas, J.R.; McNeil, M.; Darvill, A.G.; Albersheim, P.

    1987-03-01

    The partial purification and characterization of cell wall polysaccharides isolated from suspension-cultured Douglas fir (Pseudotsuga menziesii) cells are described. Extraction of isolated cell walls from 1.0 M LiCl solubilized pectic polysaccharides with glycosyl-linkage compositions similar to those of rhamnogalacturonans I and II, pectic polysaccharides isolated from walls of suspension-cultured sycamore cells. Treatment of LiCl-extracted Douglas fir walls with an endo-..cap alpha..-1,4-polygalacturonase released only small, additional amounts of pectic polysaccharide, which had a glycosyl-linkage composition similar to that of rhamnogalacturonan I. Xyloglucan oligosaccharides were released from the endo-..cap alpha..-1,4-polygalacturonase-treated walls by treatment with an endo-..beta..-1,4-glucanase. These oligosaccharides included hepta- and nonasaccharides similar or identical to those released from sycamore cell walls by the same enzyme, and structurally related octa- and decasaccharides similar to those isolated from various angiosperms. Finally, additional xyloglucan and small amounts of xylan were extracted from the endo-..beta..-1,4-glucanase-treated walls by 0.5 N NaOH. The xylan resembled that extracted by NaOH from dicot cell walls in that it contained 2,4- but not 3,4-linked xylosyl residues. In this study, a total of 15% of the cell wall was isolated as pectic material, 10% as xyloglucan, and less than 1% as xylan. The noncellulosic polysaccharides accounted for 25% of the cell walls, cellulose for 23%, protein for 34%, and ash for 5%, for a total of 88% of the cell wall.

  20. Multinet growth in the cell wall of Nitella.

    PubMed

    GREEN, P B

    1960-04-01

    Plant cell walls typically consist of crystalline microfibrils embedded in a non-crystalline matrix. The growing cylindrical Nitella cell wall contains microfibrils predominantly oriented in the transverse direction. The present study has shown that the transversely oriented microfibrils are primarily located toward the inner surface of the wall and that, proceeding outward from the inner surface, the wall contains microfibrils of ever poorer transverse orientation, the fibrils being randomly or axially arranged in the outermost regions of the wall. Because cell expansion is primarily in the axial direction, the texture of the fibrillar elements of the wall can be explained by assuming that new microfibrils of transverse orientation are added only at the inner surface of the wall and that they become passively reoriented to the axial direction during cell elongation. The described structure corresponds to that proposed by Roelofsen and Houwink for cells showing "multi-net growth." The demonstration of a continuous gradient of microfibrillar arrangement and its partial quantitative description was accomplished by the analysis, with the polarized light and interference microscopes, of wedge-like torn edges of developing cell walls which were 1 micron or less in optical thickness.

  1. Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

    PubMed Central

    Orlean, Peter

    2012-01-01

    The wall gives a Saccharomyces cerevisiae cell its osmotic integrity; defines cell shape during budding growth, mating, sporulation, and pseudohypha formation; and presents adhesive glycoproteins to other yeast cells. The wall consists of β1,3- and β1,6-glucans, a small amount of chitin, and many different proteins that may bear N- and O-linked glycans and a glycolipid anchor. These components become cross-linked in various ways to form higher-order complexes. Wall composition and degree of cross-linking vary during growth and development and change in response to cell wall stress. This article reviews wall biogenesis in vegetative cells, covering the structure of wall components and how they are cross-linked; the biosynthesis of N- and O-linked glycans, glycosylphosphatidylinositol membrane anchors, β1,3- and β1,6-linked glucans, and chitin; the reactions that cross-link wall components; and the possible functions of enzymatic and nonenzymatic cell wall proteins. PMID:23135325

  2. The plant cell wall: a dynamic barrier against pathogen invasion.

    PubMed

    Underwood, William

    2012-01-01

    Prospective plant pathogens must overcome the physical barrier presented by the plant cell wall. In addition to being a preformed, passive barrier limiting access of pathogens to plant cells, the cell wall is actively remodeled and reinforced specifically at discrete sites of interaction with potentially pathogenic microbes. Active reinforcement of the cell wall through the deposition of cell wall appositions, referred to as papillae, is an early response to perception of numerous categories of pathogens including fungi and bacteria. Rapid deposition of papillae is generally correlated with resistance to fungal pathogens that attempt to penetrate plant cell walls for the establishment of feeding structures. Despite the ubiquity and apparent importance of this early defense response, relatively little is known about the underlying molecular mechanisms and cellular processes involved in the targeting and assembly of papillae. This review summarizes recent advances in our understanding of cell wall-associated defenses induced by pathogen perception as well as the impact of changes in cell wall polymers on interactions with pathogens and highlights significant unanswered questions driving future research in the area.

  3. cps1+, a Schizosaccharomyces pombe gene homolog of Saccharomyces cerevisiae FKS genes whose mutation confers hypersensitivity to cyclosporin A and papulacandin B.

    PubMed Central

    Ishiguro, J; Saitou, A; Durán, A; Ribas, J C

    1997-01-01

    The Schizosaccharomyces pombe cps1-12 (for chlorpropham supersensitive) mutant strain was originally isolated as hypersensitive to the spindle poison isopropyl N-3-chlorophenyl carbamate (chlorpropham) (J. Ishiguro and Y. Uhara, Jpn. J. Genet. 67:97-109, 1992). We have found that the cps1-12 mutation also confers (i) hypersensitivity to the immunosuppressant cyclosporin A (CsA), (ii) hypersensitivity to the drug papulacandin B, which specifically inhibits 1,3-beta-D-glucan synthesis both in vivo and in vitro, and (iii) thermosensitive growth at 37 degrees C. Under any of these restrictive treatments, cells swell up and finally lyse. With an osmotic stabilizer, cells do not lyse, but at 37 degrees C they become multiseptated and multibranched. The cps1-12 mutant, grown at a restrictive temperature, showed an increase in sensitivity to lysis by enzymatic cell wall degradation, in in vitro 1,3-beta-D-glucan synthase activity (173% in the absence of GTP in the reaction), and in cell wall biosynthesis (130% of the wild-type amount). Addition of Ca2+ suppresses hypersensitivity to papulacandin B and septation and branching phenotypes. All of these data suggest a relationship between the cps1+ gene and cell wall synthesis. A DNA fragment containing the cps1+ gene was cloned, and sequence analysis indicated that it encodes a predicted membrane protein of 1,729 amino acids with 15 to 16 transmembrane domains. S. pombe cps1p has overall 55% sequence identity with Fks1p or Fks2p, proposed to be catalytic or associated subunits of Saccharomyces cerevisiae 1,3-beta-D-glucan synthase. Thus, the cps1+ product might be a catalytic or an associated copurifying subunit of the fission yeast 1,3-beta-D-glucan synthase that plays an essential role in cell wall synthesis. PMID:9401022

  4. A Fungal Endoglucanase with Plant Cell Wall Extension Activity1

    PubMed Central

    Yuan, Sheng; Wu, Yajun; Cosgrove, Daniel J.

    2001-01-01

    We have identified a wall hydrolytic enzyme from Trichoderma reesei with potent ability to induce extension of heat-inactivated type I cell walls. It is a small (23-kD) endo-1,4-β-glucanase (Cel12A) belonging to glycoside hydrolase family 12. Extension of heat-inactivated walls from cucumber (Cucumis sativus cv Burpee Pickler) hypocotyls was induced by Cel12A after a distinct lag time and was accompanied by a large increase in wall plasticity and elasticity. Cel12A also increased the rate of stress relaxation of isolated walls at very short times (<200 ms; equivalent to reducing t0, a parameter that estimates the minimum relaxation time). Similar changes in wall plasticity and elasticity were observed in wheat (Triticum aestivum cv Pennmore Winter) coleoptile (type II) walls, which showed only a negligible extension in response to Cel12A treatment. Thus, Cel12A modifies both type I and II walls, but substantial extension is found only in type I walls. Cel12A has strong endo-glucanase activity against xyloglucan and (1→3,1→4)-β-glucan, but did not exhibit endo-xylanase, endo-mannase, or endo-galactanase activities. In terms of kinetics of action and effects on wall rheology, wall loosening by Cel12A differs qualitatively from the action by expansins, which induce wall extension by a non-hydrolytic polymer creep mechanism. The action by Cel12A mimics some of the changes in wall rheology found after auxin-induced growth. The strategy used here to identify Cel12A could be used to identify analogous plant enzymes that cause auxin-induced changes in cell wall rheology. PMID:11553760

  5. Fungal Cell Wall Septation and Cytokinesis Are Inhibited by Bleomycins

    PubMed Central

    Moore, Carol W.; McKoy, Judith; Del Valle, Robert; Armstrong, Donald; Bernard, Edward M.; Katz, Norman; Gordon, Ronald E.

    2003-01-01

    When the essential and distinctive cell walls of either pathogenic or nonpathogenic fungi break, cytoplasmic membranes rupture and fungi die. This fungicidal activity was discovered previously on nonproliferating Saccharomyces cerevisiae cells treated briefly with the oxidative tool and anticancer drug family of bleomycins. The present studies investigated effects of bleomycin on growing fungal organisms. These included the medically important Aspergillus fumigatus and Cryptococcus neoformans, as well as the emerging human pathogen and fungal model, S. cerevisiae. Bleomycin had its highest potency against A. fumigatus. Scanning electron microscopy and thin-section transmission electron microscopy were used to study morphological growth characteristics. Killing and growth inhibition were also measured. Long, thin, and segmented hyphae were observed when A. fumigatus was grown without bleomycin but were never observed when the mold was grown with the drug. Bleomycin arrested conidial germination, hyphal development, and the progression and completion of cell wall septation. Similarly, the drug inhibited the construction of yeast cell wall septa, preventing cytokinesis and progression in the cell division cycle of S. cerevisiae. Even when cytoplasms of mother and daughter cells separated, septation and cell division did not necessarily occur. Bizarre cell configurations, abnormally thickened cell walls at mother-daughter necks, abnormal polarized growth, large undivided cells, fragmented cells, and empty cell ghosts were also produced. This is the first report of a fungicidal agent that arrests fungal growth and development, septum formation, and cytokinesis and that also preferentially localizes to cell walls and alters isolated cell walls as well as intact cell walls on nongrowing cells. PMID:14506042

  6. A proteomic and genetic analysis of the Neurospora crassa conidia cell wall proteins identifies two glycosyl hydrolases involved in cell wall remodeling.

    PubMed

    Ao, Jie; Aldabbous, Mash'el; Notaro, Marysa J; Lojacono, Mark; Free, Stephen J

    2016-09-01

    A proteomic analysis of the conidial cell wall identified 35 cell wall proteins. A comparison with the proteome of the vegetative hyphae showed that 16 cell wall proteins were shared, and that these shared cell wall proteins were cell wall biosynthetic proteins or cell wall structural proteins. Deletion mutants for 34 of the genes were analyzed for phenotypes indicative of conidial cell wall defects. Mutants for two cell wall glycosyl hydrolases, the CGL-1 β-1,3-glucanase (NCU07523) and the NAG-1 exochitinase (NCU10852), were found to have a conidial separation phenotype. These two enzymes function in remodeling the cell wall between adjacent conidia to facilitate conidia formation and dissemination. Using promoter::RFP and promoter::GFP constructs, we demonstrated that the promoters for 15 of the conidia-specific cell wall genes, including cgl-1 and nag-1, provided for conidia-specific gene expression or for a significant increase in their expression during conidiation.

  7. Dynamic microtubules and the texture of plant cell walls.

    PubMed

    Lloyd, Clive

    2011-01-01

    The relationship between microtubules and cell-wall texture has had a fitful history in which progress in one area has not been matched by progress in the other. For example, the idea that wall texture arises entirely from self-assembly, independently of microtubules, originated with electron microscopic analyses of fixed cells that gave no clue to the ability of microtubules to reorganize. Since then, live-cell studies have established the surprising dynamicity of plant microtubules involving collisions, changes in angle, parallelization, and rotation of microtubule tracks. Combined with proof that cellulose synthases do track along shifting microtubules, this offers more realistic models for the dynamic influence of microtubules on wall texture than could have been imagined in the electron microscopic era-the era from which most ideas on wall texture originate. This review revisits the classical literature on wall organization from the vantage point of current knowledge of microtubule dynamics.

  8. Plant expansins: diversity and interactions with plant cell walls.

    PubMed

    Cosgrove, Daniel J

    2015-06-01

    Expansins were discovered two decades ago as cell wall proteins that mediate acid-induced growth by catalyzing loosening of plant cell walls without lysis of wall polymers. In the interim our understanding of expansins has gotten more complex through bioinformatic analysis of expansin distribution and evolution, as well as through expression analysis, dissection of the upstream transcription factors regulating expression, and identification of additional classes of expansin by sequence and structural similarities. Molecular analyses of expansins from bacteria have identified residues essential for wall loosening activity and clarified the bifunctional nature of expansin binding to complex cell walls. Transgenic modulation of expansin expression modifies growth and stress physiology of plants, but not always in predictable or even understandable ways.

  9. Plant expansins: diversity and interactions with plant cell walls

    PubMed Central

    Cosgrove, Daniel J.

    2015-01-01

    Expansins were discovered two decades ago as cell wall proteins that mediate acid-induced growth by catalyzing loosening of plant cell walls without lysis of wall polymers. In the interim our understanding of expansins has gotten more complex through bioinformatic analysis of expansin distribution and evolution, as well as through expression analysis, dissection of the upstream transcription factors regulating expression, and identification of additional classes of expansin by sequence and structural similarities. Molecular analyses of expansins from bacteria have identified residues essential for wall loosening activity and clarified the bifunctional nature of expansin binding to complex cell walls. Transgenic modulation of expansin expression modifies growth and stress physiology of plants, but not always in predictable and even understandable ways. PMID:26057089

  10. Uncleavable Nup98-Nup96 is functional in the fission yeast Schizosaccharomyces pombe.

    PubMed

    Asakawa, Haruhiko; Mori, Chie; Ohtsuki, Chizuru; Iwamoto, Masaaki; Hiraoka, Yasushi; Haraguchi, Tokuko

    2015-01-01

    Essential nucleoporins Nup98 and Nup96 are coded by a single open reading frame, and produced by autopeptidase cleavage. The autocleavage site of Nup98-Nup96 is highly conserved in a wide range of organisms. To understand the importance of autocleavage, we examined a mutant that produces the Nup98-Nup96 joint molecule as a sole protein product of the nup189 (+) gene in the fission yeast Schizosaccharomyces pombe. Cells expressing only the joint molecule were found to be viable. This result indicates that autocleavage of Nup98-Nup96 is dispensable for cell growth, at least under normal culture conditions in S. pombe.

  11. Influence of polyethylene glycol on the size of Schizosaccharomyces pombe electropores

    SciTech Connect

    Hood, M.T.; Stachow, C. )

    1992-04-01

    The role of polyethylene glycol (PEG) in the transformation of Schizosaccharomyces pombe by electroporation is investigated by fluorescein isothiocyanate-dextran uptake and transformation studies. It is shown that when S. pombe cells are electroporated in the presence of PEG, the permeability state created is sustained until removal of PEG. In addition, the permeability of electroporated S. pombe envelopes is further increased with longer incubation times in PEG. The increased permeability is apparently a result of enlarged pores (electropores) due to the presence of PEG. Comparison of a heat pulse transformation protocol with electroporation suggests a second role for PEG in the uptake of macromolecules. Since pores are not thought to be created during a heat pulse, the PEG may be facilitating the uptake of plasmid DNA. This facilitation of uptake would also be expected to affect DNA uptake by electroporated cells.

  12. Measurement of pectin methylation in plant cell walls

    SciTech Connect

    McFeeters, R.F.; Armstrong, S.A.

    1984-01-01

    A procedure was developed to measure the degree of pectin methylation in small samples of isolated cell walls from nonlignified plant tissues or pectin solutions. Galacturonic acid was determined colorimetrically with the 3,5-dimethylphenol reagent. Methylation was measured by base hydrolysis of galacturonic acid methyl esters, followed by gas chromatographic determination of released methanol. Estimates of the precision of analysis of pectin and cell wall samples were made. The coefficient of variation for estimates of the pectin esterification in cell walls isolated from 10-g samples of cucumber tissue ranged from 7.7 to 13.2%.

  13. Cell Wall Metabolism in Response to Abiotic Stress.

    PubMed

    Le Gall, Hyacinthe; Philippe, Florian; Domon, Jean-Marc; Gillet, Françoise; Pelloux, Jérôme; Rayon, Catherine

    2015-01-01

    This review focuses on the responses of the plant cell wall to several abiotic stresses including drought, flooding, heat, cold, salt, heavy metals, light, and air pollutants. The effects of stress on cell wall metabolism are discussed at the physiological (morphogenic), transcriptomic, proteomic and biochemical levels. The analysis of a large set of data shows that the plant response is highly complex. The overall effects of most abiotic stress are often dependent on the plant species, the genotype, the age of the plant, the timing of the stress application, and the intensity of this stress. This shows the difficulty of identifying a common pattern of stress response in cell wall architecture that could enable adaptation and/or resistance to abiotic stress. However, in most cases, two main mechanisms can be highlighted: (i) an increased level in xyloglucan endotransglucosylase/hydrolase (XTH) and expansin proteins, associated with an increase in the degree of rhamnogalacturonan I branching that maintains cell wall plasticity and (ii) an increased cell wall thickening by reinforcement of the secondary wall with hemicellulose and lignin deposition. Taken together, these results show the need to undertake large-scale analyses, using multidisciplinary approaches, to unravel the consequences of stress on the cell wall. This will help identify the key components that could be targeted to improve biomass production under stress conditions. PMID:27135320

  14. Cell Wall Metabolism in Response to Abiotic Stress

    PubMed Central

    Gall, Hyacinthe Le; Philippe, Florian; Domon, Jean-Marc; Gillet, Françoise; Pelloux, Jérôme; Rayon, Catherine

    2015-01-01

    This review focuses on the responses of the plant cell wall to several abiotic stresses including drought, flooding, heat, cold, salt, heavy metals, light, and air pollutants. The effects of stress on cell wall metabolism are discussed at the physiological (morphogenic), transcriptomic, proteomic and biochemical levels. The analysis of a large set of data shows that the plant response is highly complex. The overall effects of most abiotic stress are often dependent on the plant species, the genotype, the age of the plant, the timing of the stress application, and the intensity of this stress. This shows the difficulty of identifying a common pattern of stress response in cell wall architecture that could enable adaptation and/or resistance to abiotic stress. However, in most cases, two main mechanisms can be highlighted: (i) an increased level in xyloglucan endotransglucosylase/hydrolase (XTH) and expansin proteins, associated with an increase in the degree of rhamnogalacturonan I branching that maintains cell wall plasticity and (ii) an increased cell wall thickening by reinforcement of the secondary wall with hemicellulose and lignin deposition. Taken together, these results show the need to undertake large-scale analyses, using multidisciplinary approaches, to unravel the consequences of stress on the cell wall. This will help identify the key components that could be targeted to improve biomass production under stress conditions. PMID:27135320

  15. Cell Wall Metabolism in Response to Abiotic Stress.

    PubMed

    Le Gall, Hyacinthe; Philippe, Florian; Domon, Jean-Marc; Gillet, Françoise; Pelloux, Jérôme; Rayon, Catherine

    2015-02-16

    This review focuses on the responses of the plant cell wall to several abiotic stresses including drought, flooding, heat, cold, salt, heavy metals, light, and air pollutants. The effects of stress on cell wall metabolism are discussed at the physiological (morphogenic), transcriptomic, proteomic and biochemical levels. The analysis of a large set of data shows that the plant response is highly complex. The overall effects of most abiotic stress are often dependent on the plant species, the genotype, the age of the plant, the timing of the stress application, and the intensity of this stress. This shows the difficulty of identifying a common pattern of stress response in cell wall architecture that could enable adaptation and/or resistance to abiotic stress. However, in most cases, two main mechanisms can be highlighted: (i) an increased level in xyloglucan endotransglucosylase/hydrolase (XTH) and expansin proteins, associated with an increase in the degree of rhamnogalacturonan I branching that maintains cell wall plasticity and (ii) an increased cell wall thickening by reinforcement of the secondary wall with hemicellulose and lignin deposition. Taken together, these results show the need to undertake large-scale analyses, using multidisciplinary approaches, to unravel the consequences of stress on the cell wall. This will help identify the key components that could be targeted to improve biomass production under stress conditions.

  16. An Arabidopsis Gene Regulatory Network for Secondary Cell Wall Synthesis

    PubMed Central

    Taylor-Teeples, M; Lin, L; de Lucas, M; Turco, G; Toal, TW; Gaudinier, A; Young, NF; Trabucco, GM; Veling, MT; Lamothe, R; Handakumbura, PP; Xiong, G; Wang, C; Corwin, J; Tsoukalas, A; Zhang, L; Ware, D; Pauly, M; Kliebenstein, DJ; Dehesh, K; Tagkopoulos, I; Breton, G; Pruneda-Paz, JL; Ahnert, SE; Kay, SA; Hazen, SP; Brady, SM

    2014-01-01

    Summary The plant cell wall is an important factor for determining cell shape, function and response to the environment. Secondary cell walls, such as those found in xylem, are composed of cellulose, hemicelluloses and lignin and account for the bulk of plant biomass. The coordination between transcriptional regulation of synthesis for each polymer is complex and vital to cell function. A regulatory hierarchy of developmental switches has been proposed, although the full complement of regulators remains unknown. Here, we present a protein-DNA network between Arabidopsis transcription factors and secondary cell wall metabolic genes with gene expression regulated by a series of feed-forward loops. This model allowed us to develop and validate new hypotheses about secondary wall gene regulation under abiotic stress. Distinct stresses are able to perturb targeted genes to potentially promote functional adaptation. These interactions will serve as a foundation for understanding the regulation of a complex, integral plant component. PMID:25533953

  17. On the growth of walled cells: From shells to vesicles.

    NASA Astrophysics Data System (ADS)

    Boudaoud, Arezki

    2003-03-01

    The growth of isolated walled cells is investigated. Examples of such cells range from bacteria to giant algae, and include cochlear hair, plant root hair, fungi and yeast cells. They are modeled as elastic shells inflated by a liquid. Cell growth is driven by fluid pressure and is similar to a plastic deformation of the wall. The requirement of mechanical equilibrium leads to two new scaling laws for cell size that are in quantitative agreement with the compiled biological data. Given these results, possible shapes for growing cells are computed by analogy with those of vesicle membranes.

  18. Growth of Walled Cells: From Shells to Vesicles

    NASA Astrophysics Data System (ADS)

    Boudaoud, Arezki

    2003-07-01

    The growth of isolated walled cells is investigated. Examples of such cells range from bacteria to giant algae, and include cochlear hair, plant root hair, fungi, and yeast cells. They are modeled as elastic shells containing a liquid. Cell growth is driven by fluid pressure and is is similar to a plastic deformation of the wall. The requirement of mechanical equilibrium leads to two new scaling laws for cell size that are in quantitative agreement with the compiled biological data. Given these results, possible shapes for growing cells are computed by analogy with those of vesicle membranes.

  19. Structural characteristics of developing Nitella internodal cell walls.

    PubMed

    GREEN, P B

    1958-09-25

    The Nilella intermodal cell is formed by a division of the segment cell, the latter being a direct derivative of the shoot apical cell. The internodal cell is remarkable in that it elongates from an initial length of about 20 microns to a mature length of about 60 millimeters. The structures of the apical and segment cells, and the internodal cells in all stages of development were examined with the techniques of interference, polarization, and electron microscopy. The apical and segment cells were found to be isotropic. The upper part of the segment cell, destined to form a node, shows a curious pitted structure that was characteristic of certain node structures. The lower part of the segment cell, destined to become an internodal cell, shows a vague transverse arrangement of fibrils at the inner wall surface. The internodal cells, from the time they are first formed, show negative birefringence and a transverse arrangement of microfibrils at the inner wall surface. The elongation of the internodal cell is characterized by a rise, dip, and rise in both the optical thickness and retardation of the cell wall. The dip in both these variables coincides with the attainment of the maximum relative elongation rate. After the cessation of elongation, wall deposition continues, but the fibrils at .the inner surface of the wall are now seen to occur in fields of nearly parallel microfibrils. These fields, with varying fibrillar directions, may partly overlap each other or may merge with one another. Unlike the growing wall, this wall which is deposited after the end of elongation is isotropic.

  20. Up against the wall: is yeast cell wall integrity ensured by mechanosensing in plasma membrane microdomains?

    PubMed

    Kock, Christian; Dufrêne, Yves F; Heinisch, Jürgen J

    2015-02-01

    Yeast cell wall integrity (CWI) signaling serves as a model of the regulation of fungal cell wall synthesis and provides the basis for the development of antifungal drugs. A set of five membrane-spanning sensors (Wsc1 to Wsc3, Mid2, and Mtl1) detect cell surface stress and commence the signaling pathway upon perturbations of either the cell wall structure or the plasma membrane. We here summarize the latest advances in the structure/function relationship primarily of the Wsc1 sensor and critically review the evidence that it acts as a mechanosensor. The relevance and physiological significance of the information obtained for the function of the other CWI sensors, as well as expected future developments, are discussed.

  1. Cell Wall Composition, Biosynthesis and Remodeling during Pollen Tube Growth

    PubMed Central

    Mollet, Jean-Claude; Leroux, Christelle; Dardelle, Flavien; Lehner, Arnaud

    2013-01-01

    The pollen tube is a fast tip-growing cell carrying the two sperm cells to the ovule allowing the double fertilization process and seed setting. To succeed in this process, the spatial and temporal controls of pollen tube growth within the female organ are critical. It requires a massive cell wall deposition to promote fast pollen tube elongation and a tight control of the cell wall remodeling to modify the mechanical properties. In addition, during its journey, the pollen tube interacts with the pistil, which plays key roles in pollen tube nutrition, guidance and in the rejection of the self-incompatible pollen. This review focuses on our current knowledge in the biochemistry and localization of the main cell wall polymers including pectin, hemicellulose, cellulose and callose from several pollen tube species. Moreover, based on transcriptomic data and functional genomic studies, the possible enzymes involved in the cell wall remodeling during pollen tube growth and their impact on the cell wall mechanics are also described. Finally, mutant analyses have permitted to gain insight in the function of several genes involved in the pollen tube cell wall biosynthesis and their roles in pollen tube growth are further discussed. PMID:27137369

  2. Identification and refinement of two strong constitutive promoters for gene expression system of Schizosaccharomyces pombe.

    PubMed

    Wang, Hongcheng; Wang, Haiyang; Wang, Meng; Zhang, Lei; Wang, Ren; Mei, Yanzhen; Shao, Weilan

    2014-06-01

    Fission yeast Schizosaccharomyces pombe shares various important properties with higher eukaryotes and is now considered a useful host for elevated production of mammalian proteins for medicinal applications. The full-length nmt1 promoter has been widely used as a strong promoter in S. pombe expression system. In the present study, the promoters of the eno101 and gpd3 genes in S. pombe were identified as strong constitutive promoters. For convenient applications in the plasmids of S. pombe, these promoters were refined to 276-bp eno and 273-bp gpd promoters by deleting undesired sequences and examining the expression of reporter genes including lacZ and xynA. Both the refined eno and gpd promoters provided approximately 1.5-fold higher expression of LacZ than nmt1 promoter. Furthermore, gene expression under the control of the eno or gpd promoter was not repressed by the components of YES medium while nmt1 promoter was inhibited by thiamine in yeast extract. Therefore, both eno and gpd promoters offer opportunities for efficient production of recombinant proteins by S. pombe in high cell-density fermentation.

  3. Quantification and characterization of cell wall polysaccharides released by non-Saccharomyces yeast strains during alcoholic fermentation.

    PubMed

    Giovani, Giovanna; Rosi, Iolanda; Bertuccioli, Mario

    2012-11-15

    In order to improve knowledge about the oenological characteristics of non-Saccharomyces yeast strains, and to reconsider their contribution to wine quality, we studied the release of polysaccharides by 13 non-Saccharomyces strains of different species (three wine yeasts, six grape yeasts, and three spoilage yeasts) during alcoholic fermentation in synthetic must. Three Saccharomyces cerevisiae strains were included for comparison. All of the non-Saccharomyces strains released polysaccharides into fermentation medium; the amount released depended on the yeast species, the number of cells formed and their physiological conditions. Normalizing the quantity of macromolecules released to the cell biomass revealed that most non-Saccharomyces strains produced a greater quantity of polysaccharides compared to S. cerevisiae strains after 7 and 14days of fermentation. This capacity was particularly expressed in the studied wine spoilage yeasts (Saccharomycodes ludwigii, Zygosaccharomyces bailii, and Brettanomyces bruxellensis). Chemical characterization of exocellular polysaccharides produced by non-Saccharomyces yeasts revealed them to essentially be mannoproteins with high mannose contents, ranging from 93% for S'codes. ludwigii to 73-74% for Pichia anomala and Starmerella bombicola. Protein contents varied from 9% for P. anomala to 29% for Z. bailii. These compositions were very similar to those of the S. cerevisiae strains, and to the chemical composition of the cell wall mannoproteins of different yeast species. The presence of galactose, in addition to mannose and glucose, in the exocellular polysaccharides released by Schizosaccharomyces pombe, confirmed the parietal nature of the polysaccharides released by non-Saccharomyces yeasts; only this species has a galactomannan located in the outer layer of the cell wall.

  4. A formin-nucleated actin aster concentrates cell wall hydrolases for cell fusion in fission yeast

    PubMed Central

    Dudin, Omaya; Bendezú, Felipe O.; Groux, Raphael; Laroche, Thierry; Seitz, Arne

    2015-01-01

    Cell–cell fusion is essential for fertilization. For fusion of walled cells, the cell wall must be degraded at a precise location but maintained in surrounding regions to protect against lysis. In fission yeast cells, the formin Fus1, which nucleates linear actin filaments, is essential for this process. In this paper, we show that this formin organizes a specific actin structure—the actin fusion focus. Structured illumination microscopy and live-cell imaging of Fus1, actin, and type V myosins revealed an aster of actin filaments whose barbed ends are focalized near the plasma membrane. Focalization requires Fus1 and type V myosins and happens asynchronously always in the M cell first. Type V myosins are essential for fusion and concentrate cell wall hydrolases, but not cell wall synthases, at the fusion focus. Thus, the fusion focus focalizes cell wall dissolution within a broader cell wall synthesis zone to shift from cell growth to cell fusion. PMID:25825517

  5. Characterisation of Schizosaccharomyces pombe α-actinin.

    PubMed

    Addario, Barbara; Sandblad, Linda; Persson, Karina; Backman, Lars

    2016-01-01

    The actin cytoskeleton plays a fundamental role in eukaryotic cells. Its reorganization is regulated by a plethora of actin-modulating proteins, such as a-actinin. In higher organisms, α-actinin is characterized by the presence of three distinct structural domains: an N-terminal actin-binding domain and a C-terminal region with EF-hand motif separated by a central rod domain with four spectrin repeats. Sequence analysis has revealed that the central rod domain of α-actinin from the fission yeast Schizosaccharomyces pombe consists of only two spectrin repeats. To obtain a firmer understanding of the structure and function of this unconventional α-actinin, we have cloned and characterized each structural domain. Our results show that this a-actinin isoform is capable of forming dimers and that the rod domain is required for this. However, its actin-binding and cross-linking activity appears less efficient compared to conventional α-actinins. The solved crystal structure of the actin-binding domain indicates that the closed state is stabilised by hydrogen bonds and a salt bridge not present in other α-actinins, which may reduce the affinity for actin.

  6. Characterisation of Schizosaccharomyces pombe α-actinin

    PubMed Central

    Addario, Barbara; Sandblad, Linda; Persson, Karina

    2016-01-01

    The actin cytoskeleton plays a fundamental role in eukaryotic cells. Its reorganization is regulated by a plethora of actin-modulating proteins, such as a-actinin. In higher organisms, α-actinin is characterized by the presence of three distinct structural domains: an N-terminal actin-binding domain and a C-terminal region with EF-hand motif separated by a central rod domain with four spectrin repeats. Sequence analysis has revealed that the central rod domain of α-actinin from the fission yeast Schizosaccharomyces pombe consists of only two spectrin repeats. To obtain a firmer understanding of the structure and function of this unconventional α-actinin, we have cloned and characterized each structural domain. Our results show that this a-actinin isoform is capable of forming dimers and that the rod domain is required for this. However, its actin-binding and cross-linking activity appears less efficient compared to conventional α-actinins. The solved crystal structure of the actin-binding domain indicates that the closed state is stabilised by hydrogen bonds and a salt bridge not present in other α-actinins, which may reduce the affinity for actin. PMID:27069798

  7. Modification of cell wall polysaccharides during retting of cassava roots.

    PubMed

    Ngolong Ngea, Guillaume Legrand; Guillon, Fabienne; Essia Ngang, Jean Justin; Bonnin, Estelle; Bouchet, Brigitte; Saulnier, Luc

    2016-12-15

    Retting is an important step in traditional cassava processing that involves tissue softening of the roots to transform the cassava into flour and various food products. The tissue softening that occurs during retting was attributed to the degradation of cell wall pectins through the action of pectin-methylesterase and pectate-lyase that possibly originated from a microbial source or the cassava plant itself. Changes in cell wall composition were investigated during retting using chemical analysis, specific glycanase degradation and immuno-labelling of cell wall polysaccharides. Pectic 1,4-β-d-galactan was the main cell wall polysaccharide affected during the retting of cassava roots. This result suggested that better control of pectic galactan degradation and a better understanding of the degradation mechanism by endogenous endo-galactanase and/or exogenous microbial enzymes might contribute to improve the texture properties of cassava products. PMID:27451197

  8. Plant cell wall characterization using scanning probe microscopy techniques

    PubMed Central

    Yarbrough, John M; Himmel, Michael E; Ding, Shi-You

    2009-01-01

    Lignocellulosic biomass is today considered a promising renewable resource for bioenergy production. A combined chemical and biological process is currently under consideration for the conversion of polysaccharides from plant cell wall materials, mainly cellulose and hemicelluloses, to simple sugars that can be fermented to biofuels. Native plant cellulose forms nanometer-scale microfibrils that are embedded in a polymeric network of hemicelluloses, pectins, and lignins; this explains, in part, the recalcitrance of biomass to deconstruction. The chemical and structural characteristics of these plant cell wall constituents remain largely unknown today. Scanning probe microscopy techniques, particularly atomic force microscopy and its application in characterizing plant cell wall structure, are reviewed here. We also further discuss future developments based on scanning probe microscopy techniques that combine linear and nonlinear optical techniques to characterize plant cell wall nanometer-scale structures, specifically apertureless near-field scanning optical microscopy and coherent anti-Stokes Raman scattering microscopy. PMID:19703302

  9. Modification of cell wall polysaccharides during retting of cassava roots.

    PubMed

    Ngolong Ngea, Guillaume Legrand; Guillon, Fabienne; Essia Ngang, Jean Justin; Bonnin, Estelle; Bouchet, Brigitte; Saulnier, Luc

    2016-12-15

    Retting is an important step in traditional cassava processing that involves tissue softening of the roots to transform the cassava into flour and various food products. The tissue softening that occurs during retting was attributed to the degradation of cell wall pectins through the action of pectin-methylesterase and pectate-lyase that possibly originated from a microbial source or the cassava plant itself. Changes in cell wall composition were investigated during retting using chemical analysis, specific glycanase degradation and immuno-labelling of cell wall polysaccharides. Pectic 1,4-β-d-galactan was the main cell wall polysaccharide affected during the retting of cassava roots. This result suggested that better control of pectic galactan degradation and a better understanding of the degradation mechanism by endogenous endo-galactanase and/or exogenous microbial enzymes might contribute to improve the texture properties of cassava products.

  10. A versatile strategy for grafting polymers to wood cell walls.

    PubMed

    Keplinger, T; Cabane, E; Chanana, M; Hass, P; Merk, V; Gierlinger, N; Burgert, I

    2015-01-01

    The hierarchical structure of wood is composed of a cellulose skeleton of high structural order at various length scales. At the nanoscale and microscale the specific structural features of the cells and cell walls result in a lightweight structure with an anisotropic material profile of excellent mechanical performance. By being able to specifically functionalize wood at the level of cell and cell walls one can insert new properties and inevitably upscale them along the intrinsic hierarchical structure, to a level of large-scale engineering materials applications. For this purpose, however, precise control of the spatial distribution of the modifying substances in the complex wood structure is needed. Here we demonstrate a method to insert methacryl groups into wood cell walls using two different chemistry routes. By using these methacryl groups as the anchor points for grafting, various polymers can be inserted into the wood structure. Strikingly, depending on the methacryl precursor, the spatial distribution of the polymer differs strongly. As a proof of concept we grafted polystyrene as a model compound in the second modification step. In the case of methacryloyl chloride the polymer was located mainly at the interface between the cell lumina and the cell wall covering the inner surface of the cells and being traceable up to 2-3 μm in the cell wall, whereas in the case of methacrylic anhydride the polymer was located inside the whole cell wall. Scanning electron microscopy, Fourier transform infrared spectroscopy and especially Raman spectroscopy were used for an in-depth analysis of the modified wood at the cell wall level.

  11. Vascular wall progenitor cells in health and disease.

    PubMed

    Psaltis, Peter J; Simari, Robert D

    2015-04-10

    The vasculature plays an indispensible role in organ development and maintenance of tissue homeostasis, such that disturbances to it impact greatly on developmental and postnatal health. Although cell turnover in healthy blood vessels is low, it increases considerably under pathological conditions. The principle sources for this phenomenon have long been considered to be the recruitment of cells from the peripheral circulation and the re-entry of mature cells in the vessel wall back into cell cycle. However, recent discoveries have also uncovered the presence of a range of multipotent and lineage-restricted progenitor cells in the mural layers of postnatal blood vessels, possessing high proliferative capacity and potential to generate endothelial, smooth muscle, hematopoietic or mesenchymal cell progeny. In particular, the tunica adventitia has emerged as a progenitor-rich compartment with niche-like characteristics that support and regulate vascular wall progenitor cells. Preliminary data indicate the involvement of some of these vascular wall progenitor cells in vascular disease states, adding weight to the notion that the adventitia is integral to vascular wall pathogenesis, and raising potential implications for clinical therapies. This review discusses the current body of evidence for the existence of vascular wall progenitor cell subpopulations from development to adulthood and addresses the gains made and significant challenges that lie ahead in trying to accurately delineate their identities, origins, regulatory pathways, and relevance to normal vascular structure and function, as well as disease.

  12. Mechanics and morphogenesis of fission yeast cells.

    PubMed

    Davì, Valeria; Minc, Nicolas

    2015-12-01

    The integration of biochemical and biomechanical elements is at the heart of morphogenesis. While animal cells are relatively soft objects which shape and mechanics is mostly regulated by cytoskeletal networks, walled cells including those of plants, fungi and bacteria are encased in a rigid cell wall which resist high internal turgor pressure. How these particular mechanical properties may influence basic cellular processes, such as growth, shape and division remains poorly understood. Recent work using the model fungal cell fission yeast, Schizosaccharomyces pombe, highlights important contribution of cell mechanics to various morphogenesis processes. We envision this genetically tractable system to serve as a novel standard for the mechanobiology of walled cell.

  13. The Permeability of Plant Cell Walls as Measured by Gel Filtration Chromatography

    NASA Astrophysics Data System (ADS)

    Tepeer, Mark; Taylor, Iain E. P.

    1981-08-01

    The permeability of plant cell walls to macromolecules may limit the ability of enzymes to alter the biochemical and physical properties of the wall. Proteins of molecular weight up to 60,000 can permeate a substantial portion of the cell wall. Measurements of wall permeability in which cells are exposed to hypertonic solutions of macromolecules may seriously underestimate wall permeability.

  14. Ultrastructure and Composition of the Nannochloropsis gaditana Cell Wall

    PubMed Central

    Scholz, Matthew J.; Weiss, Taylor L.; Jinkerson, Robert E.; Jing, Jia; Roth, Robyn; Goodenough, Ursula; Posewitz, Matthew C.

    2014-01-01

    Marine algae of the genus Nannochloropsis are promising producers of biofuel precursors and nutraceuticals and are also harvested commercially for aquaculture feed. We have used quick-freeze, deep-etch electron microscopy, Fourier transform infrared spectroscopy, and carbohydrate analyses to characterize the architecture of the Nannochloropsis gaditana (strain CCMP 526) cell wall, whose recalcitrance presents a significant barrier to biocommodity extraction. The data indicate a bilayer structure consisting of a cellulosic inner wall (∼75% of the mass balance) protected by an outer hydrophobic algaenan layer. Cellulase treatment of walls purified after cell lysis generates highly enriched algaenan preparations without using the harsh chemical treatments typically used in algaenan isolation and characterization. Nannochloropsis algaenan was determined to comprise long, straight-chain, saturated aliphatics with ether cross-links, which closely resembles the cutan of vascular plants. Chemical identification of >85% of the isolated cell wall mass is detailed, and genome analysis is used to identify candidate biosynthetic enzymes. PMID:25239976

  15. Role of the plant cell wall in gravity resistance.

    PubMed

    Hoson, Takayuki; Wakabayashi, Kazuyuki

    2015-04-01

    Gravity resistance, mechanical resistance to the gravitational force, is a principal graviresponse in plants, comparable to gravitropism. The cell wall is responsible for the final step of gravity resistance. The gravity signal increases the rigidity of the cell wall via the accumulation of its constituents, polymerization of certain matrix polysaccharides due to the suppression of breakdown, stimulation of cross-link formation, and modifications to the wall environment, in a wide range of situations from microgravity in space to hypergravity. Plants thus develop a tough body to resist the gravitational force via an increase in cell wall rigidity and the modification of growth anisotropy. The development of gravity resistance mechanisms has played an important role in the acquisition of responses to various mechanical stresses and the evolution of land plants.

  16. Ferulic acid is esterified to glucuronoarabinoxylans in pineapple cell walls.

    PubMed

    Smith, B G; Harris, P J

    2001-03-01

    The ester-linkage of ferulic acid (mainly E) to polysaccharides in primary cell walls of pineapple fruit (Ananas comosus) (Bromeliaceae) was investigated by treating a cell-wall preparation with 'Driselase' which contains a mixture of endo- and exo-glycanases, but no hydroxycinnamoyl esterase activity. The most abundant feruloyl oligosaccharide released was O-[5-O-(E-feruloyl)-alpha-L-arabinofuranosyl](1-->3)-O-beta-D-xylopyranosyl-(1-->4)-D-xylopyranose (FAXX). This indicated that the ferulic acid is ester-linked to glucuronoarabinoxylans in the same way as in the primary walls of grasses and cereals (Poaceae). Glucuronoarabinoxylans are the major non-cellulosic polysaccharides in the pineapple cell walls.

  17. Live cell imaging of the cytoskeleton and cell wall enzymes in plant cells.

    PubMed

    Sampathkumar, Arun; Wightman, Raymond

    2015-01-01

    The use of live imaging techniques to visualize the dynamic changes and interactions within plant cells has given us detailed information on the function and organization of the cytoskeleton and cell wall associated proteins. This information has grown with the constant improvement in imaging hardware and molecular tools. In this chapter, we describe the procedure for the preparation and live visualization of fluorescent protein fusions associated with the cytoskeleton and the cell wall in Arabidopsis. PMID:25408450

  18. A model for cell wall dissolution in mating yeast cells: polarized secretion and restricted diffusion of cell wall remodeling enzymes induces local dissolution.

    PubMed

    Huberman, Lori B; Murray, Andrew W

    2014-01-01

    Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells.

  19. A model for cell wall dissolution in mating yeast cells: polarized secretion and restricted diffusion of cell wall remodeling enzymes induces local dissolution.

    PubMed

    Huberman, Lori B; Murray, Andrew W

    2014-01-01

    Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells. PMID:25329559

  20. A Model for Cell Wall Dissolution in Mating Yeast Cells: Polarized Secretion and Restricted Diffusion of Cell Wall Remodeling Enzymes Induces Local Dissolution

    PubMed Central

    Huberman, Lori B.; Murray, Andrew W.

    2014-01-01

    Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells. PMID:25329559

  1. Determining the polysaccharide composition of plant cell walls.

    PubMed

    Pettolino, Filomena A; Walsh, Cherie; Fincher, Geoffrey B; Bacic, Antony

    2012-09-01

    The plant cell wall is a chemically complex structure composed mostly of polysaccharides. Detailed analyses of these cell wall polysaccharides are essential for our understanding of plant development and for our use of plant biomass (largely wall material) in the food, agriculture, fabric, timber, biofuel and biocomposite industries. We present analytical techniques not only to define the fine chemical structures of individual cell wall polysaccharides but also to estimate the overall polysaccharide composition of cell wall preparations. The procedure covers the preparation of cell walls, together with gas chromatography-mass spectrometry (GC-MS)-based methods, for both the analysis of monosaccharides as their volatile alditol acetate derivatives and for methylation analysis to determine linkage positions between monosaccharide residues as their volatile partially methylated alditol acetate derivatives. Analysis time will vary depending on both the method used and the tissue type, and ranges from 2 d for a simple neutral sugar composition to 2 weeks for a carboxyl reduction/methylation linkage analysis. PMID:22864200

  2. Determining the polysaccharide composition of plant cell walls.

    PubMed

    Pettolino, Filomena A; Walsh, Cherie; Fincher, Geoffrey B; Bacic, Antony

    2012-09-01

    The plant cell wall is a chemically complex structure composed mostly of polysaccharides. Detailed analyses of these cell wall polysaccharides are essential for our understanding of plant development and for our use of plant biomass (largely wall material) in the food, agriculture, fabric, timber, biofuel and biocomposite industries. We present analytical techniques not only to define the fine chemical structures of individual cell wall polysaccharides but also to estimate the overall polysaccharide composition of cell wall preparations. The procedure covers the preparation of cell walls, together with gas chromatography-mass spectrometry (GC-MS)-based methods, for both the analysis of monosaccharides as their volatile alditol acetate derivatives and for methylation analysis to determine linkage positions between monosaccharide residues as their volatile partially methylated alditol acetate derivatives. Analysis time will vary depending on both the method used and the tissue type, and ranges from 2 d for a simple neutral sugar composition to 2 weeks for a carboxyl reduction/methylation linkage analysis.

  3. Production Model Press for the Preparation of Bacterial Cell Walls

    PubMed Central

    Perrine, T. D.; Ribi, E.; Maki, W.; Miller, B.; Oertli, E.

    1962-01-01

    A modification of the apparatus previously described permits the preparation of cell walls in quantity. This consists of a heavy duty, double-acting hydraulic press with motor-driven pump, and a superstrength alloy steel pressure cell which is corrosion resistant. Liquid cooling of the jet is substituted for the previously used gas cooling to minimize aerosol formation and to facilitate subsequent treatment of the products. The device produces cell walls of excellent quality in good yield. The pressure cell has been used satisfactorily up to about 60,000 psi. Design details are given. Images FIG. 1 FIG. 2 FIG. 6 PMID:14485524

  4. Electron microscopy of Staphylococcus aureus cell wall lysis.

    PubMed

    Virgilio, R; González, C; Muñoz, N; Mendoza, S

    1966-05-01

    Virgilio, Rafael (Escuela de Química y Farmacia, Universidad de Chile, Santiago, Chile), C. González, Nubia Muñoz, and Silvia Mendoza. Electron microscopy of Staphylococcus aureus cell wall lysis. J. Bacteriol. 91:2018-2024. 1966.-A crude suspension of Staphylococcus aureus cell walls (strain Cowan III) in buffer solution was shown by electron microscopy to lyse slightly after 16 hr, probably owing to the action of autolysin. The lysis was considerably faster and more intense after the addition of lysozyme. A remarkable reduction in thickness and rigidity of the cell walls, together with the appearance of many irregular protrusions in their outlines, was observed after 2 hr; after 16 hr, there remained only a few recognizable cell wall fragments but many residual particulate remnants. When autolysin was previously inactivated by trypsin, there was a complete inhibition of the lytic action of lysozyme; on the other hand, when autolysin was inactivated by heat and lysozyme was added, a distinct decrease in the thickness of the cell walls was observed, but there was no destruction of the walls. The lytic action of lysozyme, after treatment with hot 5% trichloroacetic acid, gave rise to a marked dissolution of the structure of the cell walls, which became lost against the background, without, however, showing ostensible alteration of wall outlines. From a morphological point of view, the lytic action of autolysin plus lysozyme was quite different from that of trichloroacetic acid plus lysozyme, as shown by electron micrographs, but in both cases it was very intense. This would suggest different mechanisms of action for these agents.

  5. Another brick in the cell wall: biosynthesis dependent growth model.

    PubMed

    Barbacci, Adelin; Lahaye, Marc; Magnenet, Vincent

    2013-01-01

    Expansive growth of plant cell is conditioned by the cell wall ability to extend irreversibly. This process is possible if (i) a tensile stress is developed in the cell wall due to the coupling effect between turgor pressure and the modulation of its mechanical properties through enzymatic and physicochemical reactions and if (ii) new cell wall elements can be synthesized and assembled to the existing wall. In other words, expansive growth is the result of coupling effects between mechanical, thermal and chemical energy. To have a better understanding of this process, models must describe the interplay between physical or mechanical variable with biological events. In this paper we propose a general unified and theoretical framework to model growth in function of energy forms and their coupling. This framework is based on irreversible thermodynamics. It is then applied to model growth of the internodal cell of Chara corallina modulated by changes in pressure and temperature. The results describe accurately cell growth in term of length increment but also in term of cell pectate biosynthesis and incorporation to the expanding wall. Moreover, the classical growth model based on Lockhart's equation such as the one proposed by Ortega, appears as a particular and restrictive case of the more general growth equation developed in this paper.

  6. Another brick in the cell wall: biosynthesis dependent growth model.

    PubMed

    Barbacci, Adelin; Lahaye, Marc; Magnenet, Vincent

    2013-01-01

    Expansive growth of plant cell is conditioned by the cell wall ability to extend irreversibly. This process is possible if (i) a tensile stress is developed in the cell wall due to the coupling effect between turgor pressure and the modulation of its mechanical properties through enzymatic and physicochemical reactions and if (ii) new cell wall elements can be synthesized and assembled to the existing wall. In other words, expansive growth is the result of coupling effects between mechanical, thermal and chemical energy. To have a better understanding of this process, models must describe the interplay between physical or mechanical variable with biological events. In this paper we propose a general unified and theoretical framework to model growth in function of energy forms and their coupling. This framework is based on irreversible thermodynamics. It is then applied to model growth of the internodal cell of Chara corallina modulated by changes in pressure and temperature. The results describe accurately cell growth in term of length increment but also in term of cell pectate biosynthesis and incorporation to the expanding wall. Moreover, the classical growth model based on Lockhart's equation such as the one proposed by Ortega, appears as a particular and restrictive case of the more general growth equation developed in this paper. PMID:24066142

  7. A widespread family of bacterial cell wall assembly proteins

    PubMed Central

    Kawai, Yoshikazu; Marles-Wright, Jon; Cleverley, Robert M; Emmins, Robyn; Ishikawa, Shu; Kuwano, Masayoshi; Heinz, Nadja; Bui, Nhat Khai; Hoyland, Christopher N; Ogasawara, Naotake; Lewis, Richard J; Vollmer, Waldemar; Daniel, Richard A; Errington, Jeff

    2011-01-01

    Teichoic acids and acidic capsular polysaccharides are major anionic cell wall polymers (APs) in many bacteria, with various critical cell functions, including maintenance of cell shape and structural integrity, charge and cation homeostasis, and multiple aspects of pathogenesis. We have identified the widespread LytR–Cps2A–Psr (LCP) protein family, of previously unknown function, as novel enzymes required for AP synthesis. Structural and biochemical analysis of several LCP proteins suggest that they carry out the final step of transferring APs from their lipid-linked precursor to cell wall peptidoglycan (PG). In Bacillus subtilis, LCP proteins are found in association with the MreB cytoskeleton, suggesting that MreB proteins coordinate the insertion of the major polymers, PG and AP, into the cell wall. PMID:21964069

  8. Serine 26 in the PomB Subunit of the Flagellar Motor Is Essential for Hypermotility of Vibrio cholerae

    PubMed Central

    Halang, Petra; Vorburger, Thomas; Steuber, Julia

    2015-01-01

    Vibrio cholerae is motile by means of its single polar flagellum which is driven by the sodium-motive force. In the motor driving rotation of the flagellar filament, a stator complex consisting of subunits PomA and PomB converts the electrochemical sodium ion gradient into torque. Charged or polar residues within the membrane part of PomB could act as ligands for Na+, or stabilize a hydrogen bond network by interacting with water within the putative channel between PomA and PomB. By analyzing a large data set of individual tracks of swimming cells, we show that S26 located within the transmembrane helix of PomB is required to promote very fast swimming of V. cholerae. Loss of hypermotility was observed with the S26T variant of PomB at pH 7.0, but fast swimming was restored by decreasing the H+ concentration of the external medium. Our study identifies S26 as a second important residue besides D23 in the PomB channel. It is proposed that S26, together with D23 located in close proximity, is important to perturb the hydration shell of Na+ before its passage through a constriction within the stator channel. PMID:25874792

  9. Microfabricated alkali vapor cell with anti-relaxation wall coating

    SciTech Connect

    Straessle, R.; Pétremand, Y.; Briand, D.; Rooij, N. F. de; Pellaton, M.; Affolderbach, C.; Mileti, G.

    2014-07-28

    We present a microfabricated alkali vapor cell equipped with an anti-relaxation wall coating. The anti-relaxation coating used is octadecyltrichlorosilane and the cell was sealed by thin-film indium-bonding at a low temperature of 140 °C. The cell body is made of silicon and Pyrex and features a double-chamber design. Depolarizing properties due to liquid Rb droplets are avoided by confining the Rb droplets to one chamber only. Optical and microwave spectroscopy performed on this wall-coated cell are used to evaluate the cell's relaxation properties and a potential gas contamination. Double-resonance signals obtained from the cell show an intrinsic linewidth that is significantly lower than the linewidth that would be expected in case the cell had no wall coating but only contained a buffer-gas contamination on the level measured by optical spectroscopy. Combined with further experimental evidence this proves the presence of a working anti-relaxation wall coating in the cell. Such cells are of interest for applications in miniature atomic clocks, magnetometers, and other quantum sensors.

  10. Induction of arginase and ornithine transaminase in the fission yeast Schizosaccharomyces pombe.

    PubMed Central

    Benítez, T; Farrar, L

    1980-01-01

    The induction of arginase and ornithine transaminase in the fission yeast Schizosaccharomyces pombe requires the absence of ammonia and the presence of the inducer arginine. It seems that immediate arginase degradation is initiated by starved cells or ones from which arginine has been removed. PMID:7430074

  11. Characterization of rhamnogalacturonan I from cotton suspension culture cell walls

    SciTech Connect

    Not Available

    1991-01-01

    Progress has been made on the project of determining the structure of pectins. From recent progress, a covalent crosslink between rhamnogalacturonan I (RGI) and xyloglucan was hypothesized and a structure for RGI was proposed. The development of a method to determine the distribution of methyl esterification with pectins also progressed. The degree of methyl esterification of cotton cotyledon cell walls was compared to that of cotton suspension cultures. Cotyledon wall were found to have {approximately}55% of the galacturonic acid esterified whereas suspension culture wall were only about 14% methyl esterified. 10 refs. (SM)

  12. Co-delivery of cell-wall-forming enzymes in the same vesicle for coordinated fungal cell wall formation.

    PubMed

    Schuster, Martin; Martin-Urdiroz, Magdalena; Higuchi, Yujiro; Hacker, Christian; Kilaru, Sreedhar; Gurr, Sarah J; Steinberg, Gero

    2016-01-01

    Fungal cells are surrounded by an extracellular cell wall. This complex matrix of proteins and polysaccharides protects against adverse stresses and determines the shape of fungal cells. The polysaccharides of the fungal wall include 1,3-β-glucan and chitin, which are synthesized by membrane-bound synthases at the growing cell tip. A hallmark of filamentous fungi is the class V chitin synthase, which carries a myosin-motor domain. In the corn smut fungus Ustilago maydis, the myosin-chitin synthase Mcs1 moves to the plasma membrane in secretory vesicles, being delivered by kinesin-1 and myosin-5. The myosin domain of Mcs1 enhances polar secretion by tethering vesicles at the site of exocytosis. It remains elusive, however, how other cell-wall-forming enzymes are delivered and how their activity is coordinated post secretion. Here, we show that the U. maydis class VII chitin synthase and 1,3-β-glucan synthase travel in Mcs1-containing vesicles, and that their apical secretion depends on Mcs1. Once in the plasma membrane, anchorage requires enzyme activity, which suggests co-synthesis of chitin and 1,3-β-glucan polysaccharides at sites of exocytosis. Thus, delivery of cell-wall-forming enzymes in Mcs1 vesicles ensures local foci of fungal cell wall formation. PMID:27563844

  13. Co-delivery of cell-wall-forming enzymes in the same vesicle for coordinated fungal cell wall formation.

    PubMed

    Schuster, Martin; Martin-Urdiroz, Magdalena; Higuchi, Yujiro; Hacker, Christian; Kilaru, Sreedhar; Gurr, Sarah J; Steinberg, Gero

    2016-01-01

    Fungal cells are surrounded by an extracellular cell wall. This complex matrix of proteins and polysaccharides protects against adverse stresses and determines the shape of fungal cells. The polysaccharides of the fungal wall include 1,3-β-glucan and chitin, which are synthesized by membrane-bound synthases at the growing cell tip. A hallmark of filamentous fungi is the class V chitin synthase, which carries a myosin-motor domain. In the corn smut fungus Ustilago maydis, the myosin-chitin synthase Mcs1 moves to the plasma membrane in secretory vesicles, being delivered by kinesin-1 and myosin-5. The myosin domain of Mcs1 enhances polar secretion by tethering vesicles at the site of exocytosis. It remains elusive, however, how other cell-wall-forming enzymes are delivered and how their activity is coordinated post secretion. Here, we show that the U. maydis class VII chitin synthase and 1,3-β-glucan synthase travel in Mcs1-containing vesicles, and that their apical secretion depends on Mcs1. Once in the plasma membrane, anchorage requires enzyme activity, which suggests co-synthesis of chitin and 1,3-β-glucan polysaccharides at sites of exocytosis. Thus, delivery of cell-wall-forming enzymes in Mcs1 vesicles ensures local foci of fungal cell wall formation.

  14. Molecular Rigidity in Dry and Hydrated Onion Cell Walls.

    PubMed

    Ha, M. A.; Apperley, D. C.; Jarvis, M. C.

    1997-10-01

    Solid-state nuclear magnetic resonance relaxation experiments can provide information on the rigidity of individual molecules within a complex structure such as a cell wall, and thus show how each polymer can potentially contribute to the rigidity of the whole structure. We measured the proton magnetic relaxation parameters T2 (spin-spin) and T1p (spin-lattice) through the 13C-nuclear magnetic resonance spectra of dry and hydrated cell walls from onion (Allium cepa L.) bulbs. Dry cell walls behaved as rigid solids. The form of their T2 decay curves varied on a continuum between Gaussian, as in crystalline solids, and exponential, as in more mobile materials. The degree of molecular mobility that could be inferred from the T2 and T1p decay patterns was consistent with a crystalline state for cellulose and a glassy state for dry pectins. The theory of composite materials may be applied to explain the rigidity of dry onion cell walls in terms of their components. Hydration made little difference to the rigidity of cellulose and most of the xyloglucan shared this rigidity, but the pectic fraction became much more mobile. Therefore, the cellulose/xyloglucan microfibrils behaved as solid rods, and the most significant physical distinction within the hydrated cell wall was between the microfibrils and the predominantly pectic matrix. A minor xyloglucan fraction was much more mobile than the microfibrils and probably corresponded to cross-links between them. Away from the microfibrils, pectins expanded upon hydration into a nonhomogeneous, but much softer, almost-liquid gel. These data are consistent with a model for the stress-bearing hydrated cell wall in which pectins provide limited stiffness across the thickness of the wall, whereas the cross-linked microfibril network provides much greater rigidity in other directions.

  15. Molecular Rigidity in Dry and Hydrated Onion Cell Walls.

    PubMed

    Ha, M. A.; Apperley, D. C.; Jarvis, M. C.

    1997-10-01

    Solid-state nuclear magnetic resonance relaxation experiments can provide information on the rigidity of individual molecules within a complex structure such as a cell wall, and thus show how each polymer can potentially contribute to the rigidity of the whole structure. We measured the proton magnetic relaxation parameters T2 (spin-spin) and T1p (spin-lattice) through the 13C-nuclear magnetic resonance spectra of dry and hydrated cell walls from onion (Allium cepa L.) bulbs. Dry cell walls behaved as rigid solids. The form of their T2 decay curves varied on a continuum between Gaussian, as in crystalline solids, and exponential, as in more mobile materials. The degree of molecular mobility that could be inferred from the T2 and T1p decay patterns was consistent with a crystalline state for cellulose and a glassy state for dry pectins. The theory of composite materials may be applied to explain the rigidity of dry onion cell walls in terms of their components. Hydration made little difference to the rigidity of cellulose and most of the xyloglucan shared this rigidity, but the pectic fraction became much more mobile. Therefore, the cellulose/xyloglucan microfibrils behaved as solid rods, and the most significant physical distinction within the hydrated cell wall was between the microfibrils and the predominantly pectic matrix. A minor xyloglucan fraction was much more mobile than the microfibrils and probably corresponded to cross-links between them. Away from the microfibrils, pectins expanded upon hydration into a nonhomogeneous, but much softer, almost-liquid gel. These data are consistent with a model for the stress-bearing hydrated cell wall in which pectins provide limited stiffness across the thickness of the wall, whereas the cross-linked microfibril network provides much greater rigidity in other directions. PMID:12223827

  16. Simulated microgravity inhibits cell wall regeneration of Penicillium decumbens protoplasts

    NASA Astrophysics Data System (ADS)

    Zhao, C.; Sun, Y.; Yi, Z. C.; Rong, L.; Zhuang, F. Y.; Fan, Y. B.

    2010-09-01

    This work compares cell wall regeneration from protoplasts of the fungus Penicillium decumbens under rotary culture (simulated microgravity) and stationary cultures. Using an optimized lytic enzyme mixture, protoplasts were successfully released with a yield of 5.3 × 10 5 cells/mL. Under simulated microgravity conditions, the protoplast regeneration efficiency was 33.8%, lower than 44.9% under stationary conditions. Laser scanning confocal microscopy gave direct evidence for reduced formation of polysaccharides under simulated conditions. Scanning electron microscopy showed the delayed process of cell wall regeneration by simulated microgravity. The delayed regeneration of P. decumbens cell wall under simulated microgravity was likely caused by the inhibition of polysaccharide synthesis. This research contributes to the understanding of how gravitational loads affect morphological and physiological processes of fungi.

  17. Force and compliance: rethinking morphogenesis in walled cells.

    PubMed

    Harold, Franklin M

    2002-12-01

    In the turgid cells of plants, protists, fungi, and bacteria, walls resist swelling; they also confer shape on the cell. These two functions are not unrelated: cell physiologists have generally agreed that morphogenesis turns on the deformation of existing wall and the deposition of new wall, while turgor pressure produces the work of expansion. In 1990, I summed up consensus in a phrase: "localized compliance with the global force of turgor pressure." My purpose here is to survey the impact of recent discoveries on the traditional conceptual framework. Topics include the recognition of a cytoskeleton in bacteria; the tide of information and insight about budding in yeast; the role of the Spitzenkörper in hyphal extension; calcium ions and actin dynamics in shaping a tip; and the interplay of protons, expansins and cellulose fibrils in cells of higher plants.

  18. Characterization of O-mannosyltransferase family in Schizosaccharomyces pombe.

    PubMed

    Tanaka, Naotaka; Fujita, Yasuko; Suzuki, Shotaro; Morishita, Masayo; Giga-Hama, Yuko; Shimoda, Chikashi; Takegawa, Kaoru

    2005-05-13

    Protein O-glycosylation is an essential protein modification in eukaryotic cells. In Saccharomyces cerevisiae, O-mannosylation is initiated in the lumen of the endoplasmic reticulum by O-mannosyltransferase gene products (Pmt1p-7p). A search of the Schizosaccharomyces pombe genome database revealed a total of three O-glycoside mannosyltransferase homologs (ogm1+, ogm2+, and ogm4+), closely related to Saccharomyces cerevisiae PMT1, PMT2, and PMT4. Although individual ogm genes were not found to be essential, ogm1Delta and ogm4Delta mutants exhibited aberrant morphology and failed to agglutinate during mating. The phenotypes of the ogm4Delta mutant were not complemented by overexpression of ogm1+ or ogm2+, suggesting that each of the Ogm proteins does not have overlapping functions. Heterologous expression of a chitinase from S. cerevisiae in the ogm mutants revealed that O-glycosylation of chitinase had decreased in ogm1Delta cells. A GFP-tagged Fus1p from S. cerevisiae was specifically not glycosylated and accumulated in the Golgi in ogm4Delta cells. These results indicate that O-glycosylation initiated by Ogm proteins plays crucial physiological roles and can serve as a sorting determinant for protein transport of membrane glycoproteins in S. pombe.

  19. Chromosome and cell wall segregation in Streptococcus faecium ATCC 9790

    SciTech Connect

    Higgins, M.L.; Glaser, D.; Dicker, D.T.; Zito, E.T.

    1989-01-01

    Segregation was studied by measuring the positions of autoradiographic grain clusters in chains formed from single cells containing on average less than one radiolabeled chromosome strand. The degree to which chromosomal and cell wall material cosegregated was quantified by using the methods of S. Cooper and M. Weinberger, dividing the number of chains labeled at the middle. This analysis indicated that in contrast to chromosomal segregation in Escherichia coli and, in some studies, to that in gram-positive rods, chromosomal segregation in Streptococcus faecium was slightly nonrandom and did not vary with growth rate. Results were not significantly affected by strand exchange. In contrast, labeled cell wall segregated predominantly nonrandomly.

  20. 15. View of interior, north wall of hot cell featuring ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    15. View of interior, north wall of hot cell featuring radioactive materials containment box, facing east - Nevada Test Site, Reactor Maintenance & Disassembly Complex, Junior Hot Cell, Jackass Flats, Area 25, South of intersection of Roads F & G, Mercury, Nye County, NV

  1. Brachypodium distachyon grain: characterization of endosperm cell walls.

    PubMed

    Guillon, Fabienne; Bouchet, Brigitte; Jamme, Frédéric; Robert, Paul; Quéméner, Bernard; Barron, Cécile; Larré, Colette; Dumas, Paul; Saulnier, Luc

    2011-01-01

    The wild grass Brachypodium distachyon has been proposed as an alternative model species for temperate cereals. The present paper reports on the characterization of B. distachyon grain, placing emphasis on endosperm cell walls. Brachypodium distachyon is notable for its high cell wall polysaccharide content that accounts for ∼52% (w/w) of the endosperm in comparison with 2-7% (w/w) in other cereals. Starch, the typical storage polysaccharide, is low [<10% (w/w)] in the endosperm where the main polysaccharide is (1-3) (1-4)-β-glucan [40% (w/w) of the endosperm], which in all likelihood plays a role as a storage compound. In addition to (1-3) (1-4)-β-glucan, endosperm cells contain cellulose and xylan in significant amounts. Interestingly, the ratio of ferulic acid to arabinoxylan is higher in B. distachyon grain than in other investigated cereals. Feruloylated arabinoxylan is mainly found in the middle lamella and cell junction zones of the storage endosperm, suggesting a potential role in cell-cell adhesion. The present results indicate that B. distachyon grains contain all the cell wall polysaccharides encountered in other cereal grains. Thus, due to its fully sequenced genome, its short life cycle, and the genetic tools available for mutagenesis/transformation, B. distachyon is a good model to investigate cell wall polysaccharide synthesis and function in cereal grains.

  2. 47. ARAI. Interior view of operating wall of hot cell ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    47. ARA-I. Interior view of operating wall of hot cell in ARA-626. Note stands for operators at viewing windows. Manipulators with hand grips extend cables and other controls into hot cell through ducts above windows. Ineel photo no. 81-27. - Idaho National Engineering Laboratory, Army Reactors Experimental Area, Scoville, Butte County, ID

  3. Membrane-wall attachments in plasmolysed plant cells.

    PubMed

    Lang, I; Barton, D A; Overall, R L

    2004-12-01

    Field emission scanning electron microscopy of plasmolysed Tradescantia virginiana leaf epidermal cells gave novel insights into the three-dimensional architecture of Hechtian strands, Hechtian reticulum, and the inner surface of the cell wall without the need for extraction. At high magnification, we observed fibres that pin the plasma membrane to the cell wall after plasmolysis. Treatment with cellulase caused these connecting fibres to be lost and the pinned out plasma membrane of the Hechtian reticulum to disintegrate into vesicles with diameters of 100-250 nm. This suggests that the fibres may be cellulose. After 4 h of plasmolysis, a fibrous meshwork that labelled with anti-callose antibodies was observed within the space between the plasmolysed protoplast and the cell wall by field emission scanning electron microscopy. Interestingly, macerase-pectinase treatment resulted in the loss of this meshwork, suggesting that it was stabilised by pectins. We suggest that cellulose microfibrils extending from strands of the Hechtian reticulum and entwining into the cell wall matrix act as anchors for the plasma membrane as it moves away from the wall during plasmolysis.

  4. Purification and characterization of a soybean cell wall protein

    SciTech Connect

    San Francisco, S.; Tierney, M.L. )

    1989-04-01

    Plant cell wall composition is thought to reflect cellular responses to developmental and environmental signals. We have purified a 33 kDa protein from cell wall extracts of soybean seedlings which is most abundant in extracts from the hook region of the hypocotyl and is rich in proline and hydroxypyroline. In vivo {sup 3}H-proline labelling of hypocotyl tissues indicates that the hook tissue is the predominant site for synthesis of this protein. In unwounded hook, label is incorporated into a 33 kDa protein, while in wounded hook this and additional proteins rich in proline are synthesized. Similarly treated cell wall extracts analyzed by Western blot analysis, using a polyclonal antibody raised against this 33kD protein, showed that the 33 kDa protein is most abundant in cell wall extracts from the hook region of unwounded seedlings and does not increase upon wounding. An immunologically related 35kD protein is also apparent in extracts from wounded hooks and appears to co-migrate with one of the labelled proteins extractable from this tissue. These data indicate that there are two related, proline-rich cell wall proteins in the hook region of soybean seedlings, one of which (33 kDa) is prominent during seedling development and another (35 kDa) which is wound inducible.

  5. Structure, function, and biosynthesis of plant cell walls: proceedings of the seventh annual symposium in botany

    SciTech Connect

    Dugger, W.M.; Bartnicki-Garcia, S.

    1984-01-01

    Papers in the following areas were included in these symposium proceedings: (1) cell wall chemistry and biosynthesis; (2) cell wall hydrolysis and associated physiology; (3) cellular events associated with cell wall biosynthesis; and (4) interactions of plant cell walls with pathogens and related responses. Papers have been individually abstracted for the data base. (ACR)

  6. The Structure of Plant Cell Walls: II. The Hemicellulose of the Walls of Suspension-cultured Sycamore Cells.

    PubMed

    Bauer, W D; Talmadge, K W; Keegstra, K; Albersheim, P

    1973-01-01

    The molecular structure, chemical properties, and biological function of the xyloglucan polysaccharide isolated from cell walls of suspension-cultured sycamore (Acer pseudoplatanus) cells are described. The sycamore wall xyloglucan is compared to the extracellular xyloglucan secreted by suspension-cultured sycamore cells into their culture medium and is also compared to the seed "amyloid" xyloglucans.Xyloglucan-or fragments of xyloglucan-and acidic fragments of the pectic polysaccharides are released from endopolygalacturonase-pretreated sycamore walls by treatment of these walls with 8 m urea, endoglucanase, or 0.5 n NaOH. Some of the xyloglucan thus released is found to cochromatograph with the acidic pectic fragments on diethylaminoethyl Sephadex. The chemical or enzymic treatments required for the release of xyloglucan from the walls and the cochromatography of xyloglucan with the acidic pectic fragments indicate that xyloglucan is covalently linked to the pectic polysaccharides and is noncovalently bound to the cellulose fibrils of the sycamore cell wall.The molecular structure of sycamore xyloglucan was characterized by methylation analysis of the oligosaccharides obtained by endoglucanase treatment of the polymer. The structure of the polymer is based on a repeating heptasaccharide unit which consists of 4 residues of beta-1-4-linked glucose and 3 residues of terminal xylose. A single xylose residue is glycosidically linked to carbon 6 of 3 of the glucosyl residues.

  7. S. pombe strain maintenance and media.

    PubMed

    Forsburg, Susan L

    2003-11-01

    Fission yeast can be grown and maintained using similar culture methods to those employed for budding yeast. The optimal media for S. pombe is somewhat different than that employed for S. cerevisiae, although budding yeast media can be used if necessary. Good sterile technique is essential to prevent contamination by airborne molds or bacteria.

  8. Replication of centromere II of Schizosaccharomyces pombe.

    PubMed Central

    Smith, J G; Caddle, M S; Bulboaca, G H; Wohlgemuth, J G; Baum, M; Clarke, L; Calos, M P

    1995-01-01

    The centromeric DNAs of Schizosaccharomyces pombe chromosomes resemble those of higher eukaryotes in being large and composed predominantly of repeated sequences. To begin a detailed analysis of the mode of replication of a complex centromere, we examined whether any sequences within S. pombe centromere II (cen2) have the ability to mediate autonomous replication. We found a high density of segments with such activity, including at least eight different regions comprising most of the repeated and unique centromeric DNA elements. A physical mapping analysis using two-dimensional gels showed that autonomous replication initiated within the S. pombe sequences in each plasmid. A two-dimensional gel analysis of replication on the chromosomes revealed that the K and L repeat elements, which occur in multiple copies at all three centromeres and comprise approximately 70% of total centromeric DNA mass in S. pombe, are both sites of replication initiation. In contrast, the unique cen2 central core, which contains multiple segments that can support autonomous replication, appears to be repressed for initiation on the chromosome. We discuss the implications of these findings for our understanding of DNA replication and centromere function. PMID:7651433

  9. Cell Wall Alterations in the Arabidopsis emb30 Mutant

    PubMed Central

    Shevell, Diane E.; Kunkel, Tim; Chua, Nam-Hai

    2000-01-01

    The Arabidopsis EMB30 gene is essential for controlling the polarity of cell growth and for normal cell adhesion during seedling development. In this article, we show that emb30 mutations also affect the growth of undifferentiated plant cells and adult tissues. EMB30 possesses a Sec7 domain and, based on similarities to other proteins, presumably functions in the secretory pathway. The plant cell wall depends on the secretory pathway to deliver its complex polysaccharides. We show that emb30 mutants have a cell wall defect that sometimes allows material to be deposited into the interstitial space between cells instead of being restricted to cell corners. In addition, pectin, a complex polysaccharide important for cell adhesion, appears to be abnormally localized in emb30 plants. In contrast, localization of epitopes associated with xyloglucan or arabinogalactan was similar in wild-type and emb30 tissues, and the localization of a marker molecule to vacuoles appeared normal. Therefore, emb30 mutations do not cause a general defect in the secretory pathway. Together, these results suggest that emb30 mutations result in an abnormal cell wall, which in turn may account for the defects in cell adhesion and polar cell growth control observed in the mutants. PMID:11090208

  10. Cell growth pattern and wall microfibrillar arrangement: experiments with nitella.

    PubMed

    Gertel, E T; Green, P B

    1977-08-01

    In cylindrical cells growing throughout their length, over-all transverse reinforcement of the wall by microfibrils is believed to be required for cell elongation. The multinet theory states that in such cells microfibrils are deposited at the inner surface of the wall with transverse orientation and are then passively reoriented toward the longitudinal direction by the predominant longitudinal strain (surface expension). In the present study young Nitella cells were physically forced to grow in highly abnormal patterns: in length only, in girth only, or with localized suppression of growth. Subsequent gradients of microfibrillar arrangement within the wall cross-section were measured with polarized light and interference microscopes. The novel wall structures produced were in all cases explainable by passive reorientation, i.e. by the multinet theory. The study also showed that orientation of synthesis remains insensitive to several of the physical manipulations that strongly influence the passive behavior of wall microfibrils. Only the localized complete suppression of surface growth led to the deposition of nontransverse cellulose. These results suggest that the presence of strain is needed for continued oriented synthesis, but that the directional aspect of strain is not an "instructional" agent continuously guiding the orientation of synthesis, once this orientation has been established.

  11. Cell wall integrity signalling in human pathogenic fungi.

    PubMed

    Dichtl, Karl; Samantaray, Sweta; Wagener, Johannes

    2016-09-01

    Fungi are surrounded by a rigid structure, the fungal cell wall. Its plasticity and composition depend on active regulation of the underlying biosynthesis and restructuring processes. This involves specialised signalling pathways that control gene expression and activities of biosynthetic enzymes. The cell wall integrity (CWI) pathway is the central signalling cascade required for the adaptation to a wide spectrum of cell wall perturbing conditions, including heat, oxidative stress and antifungals. In the recent years, great efforts were made to analyse the CWI pathway of diverse fungi. It turned out that the CWI signalling cascade is mostly conserved in the fungal kingdom. In this review, we summarise as well as compare the current knowledge on the canonical CWI pathway in the human pathogenic fungi Candida albicans, Candida glabrata, Aspergillus fumigatus and Cryptococcus neoformans. Understanding the differences and similarities in the stress responses of these organisms could become a key to improving existing or developing new antifungal therapies. PMID:27155139

  12. Freezing stresses and hydration of isolated cell walls.

    PubMed

    Yoon, Yonghyeon; Pope, Jim; Wolfe, Joe

    2003-06-01

    The hydration of the cell walls of the giant alga Chara australis was measured as a function of temperature using quantitative deuterium nuclear magnetic resonance (NMR) of samples hydrated with D2O. At temperatures 23-5K below freezing, the hydration ratio (the ratio of mass of unfrozen water in microscopic phases in the cell wall to the dry mass) increases slowly with increasing temperature from about 0.2 to 0.4. It then rises rapidly with temperature in the few Kelvin below the freezing temperature. The linewidth of the NMR signal varies approximately linearly with the reciprocal of the hydration ratio, and with the freezing point depression or water potential. These empirical relations may be useful in estimating cell wall water contents in heterogeneous samples.

  13. A new method for extraction of pectin from cell walls

    SciTech Connect

    Maness, N.O.; Mort, A.J. )

    1991-05-01

    Pectin is often extracted from plant tissues using the Ca{sup ++} chelators ethylenediamine tetraacetate (EDTA) or cyclohexane-trans-1,2 diamine tetraacetate (CDTA). While these chelators are effective in solubilizing pectin, even after extensive dialysis against distilled water, EDTA or CDTA remains associated with the pectin. The authors have found that if 500 mM imidazole buffer, pH 7.0 is substituted for 50 mM CDTA, pH 6.5, and for equivalent extraction periods, an equivalent amount of pectin with the same sugar composition is extracted. But, the imidazole buffer can be dialyzed away completely into distilled water. Their alternative procedure for extraction of pectin from cell walls will be presented. Utilization of the procedure for extraction of whole cell walls or cell walls pretreated with liquid hydrogen fluoride is discussed.

  14. Transcriptional wiring of cell wall-related genes in Arabidopsis.

    PubMed

    Mutwil, Marek; Ruprecht, Colin; Giorgi, Federico M; Bringmann, Martin; Usadel, Björn; Persson, Staffan

    2009-09-01

    Transcriptional coordination, or co-expression, of genes may signify functional relatedness of the corresponding proteins. For example, several genes involved in secondary cell wall cellulose biosynthesis are co-expressed with genes engaged in the synthesis of xylan, which is a major component of the secondary cell wall. To extend these types of analyses, we investigated the co-expression relationships of all Carbohydrate-Active enZYmes (CAZy)-related genes for Arabidopsis thaliana. Thus, the intention was to transcriptionally link different cell wall-related processes to each other, and also to other biological functions. To facilitate easy manual inspection, we have displayed these interactions as networks and matrices, and created a web-based interface (http://aranet.mpimp-golm.mpg.de/corecarb) containing downloadable files for all the transcriptional associations.

  15. Modulation of Alternaria infectoria Cell Wall Chitin and Glucan Synthesis by Cell Wall Synthase Inhibitors

    PubMed Central

    Fernandes, Chantal; Anjos, Jorge; Walker, Louise A.; Silva, Branca M. A.; Cortes, Luísa; Mota, Marta; Munro, Carol A.; Gow, Neil A. R.

    2014-01-01

    The present work reports the effects of caspofungin, a β-1,3-glucan synthase inhibitor, and nikkomycin Z, an inhibitor of chitin synthases, on two strains of Alternaria infectoria, a melanized fungus involved in opportunistic human infections and respiratory allergies. One of the strains tested, IMF006, bore phenotypic traits that conferred advantages in resisting antifungal treatment. First, the resting cell wall chitin content was higher and in response to caspofungin, the chitin level remained constant. In the other strain, IMF001, the chitin content increased upon caspofungin treatment to values similar to basal IMF006 levels. Moreover, upon caspofungin treatment, the FKS1 gene was upregulated in IMF006 and downregulated in IMF001. In addition, the resting β-glucan content was also different in both strains, with higher levels in IMF001 than in IMF006. However, this did not provide any advantage with respect to echinocandin resistance. We identified eight different chitin synthase genes and studied relative gene expression when the fungus was exposed to the antifungals under study. In both strains, exposure to caspofungin and nikkomycin Z led to modulation of the expression of class V and VII chitin synthase genes, suggesting its importance in the robustness of A. infectoria. The pattern of A. infectoria phagocytosis and activation of murine macrophages by spores was not affected by caspofungin. Monotherapy with nikkomycin Z and caspofungin provided only fungistatic inhibition, while a combination of both led to fungal cell lysis, revealing a strong synergistic action between the chitin synthase inhibitor and the β-glucan synthase inhibitor against this fungus. PMID:24614372

  16. Particle Trajectories in Rotating Wall Cell Culture Devices

    NASA Technical Reports Server (NTRS)

    Ramachandran N.; Downey, J. P.

    1999-01-01

    Cell cultures are extremely important to the medical community since such cultures provide an opportunity to perform research on human tissue without the concerns inherent in experiments on individual humans. Development of cells in cultures has been found to be greatly influenced by the conditions of the culture. Much work has focused on the effect of the motions of cells in the culture relative to the solution. Recently rotating wall vessels have been used with success in achieving improved cellular cultures. Speculation and limited research have focused on the low shear environment and the ability of rotating vessels to keep cells suspended in solution rather than floating or sedimenting as the primary reasons for the improved cellular cultures using these devices. It is widely believed that the cultures obtained using a rotating wall vessel simulates to some degree the effect of microgravity on cultures. It has also been speculated that the microgravity environment may provide the ideal acceleration environment for culturing of cellular tissues due to the nearly negligible levels of sedimentation and shear possible. This work predicts particle trajectories of cells in rotating wall vessels of cylindrical and annular design consistent with the estimated properties of typical cellular cultures. Estimates of the shear encountered by cells in solution and the interactions with walls are studied. Comparisons of potential experiments in ground and microgravity environments are performed.

  17. MECHANISM OF CELL WALL PENETRATION BY VIRUSES

    PubMed Central

    Puck, Theodore T.; Lee, Howard H.

    1954-01-01

    Treatment of radioactively labelled host cells with T1 or T2 bacteriophages induces a leakage of cellular P and S into the medium. Evidence is presented showing that this increased cell permeability is not the result of complete lysis of a small fraction of the cells, but rather is made up of contributions from all or most of the infected population. This leakage of cellular constituents exhibits the following characteristics: (a) Infection of a cell with a single virus suffices to evoke the reaction; (b) Increasing the multiplicity up to 7 to 8 virus particles per cell does not affect the extent of leakage produced; (c) Some leakage does occur at 0°C., but much less than at 37°C.; (d) Infection by T1 virus results in a smaller amount of leakage than in the case of T2, but the pattern of response to varying virus multiplicity is the same; (e) The P resulting from such leakage contains no DNA and chemically resembles that which elutes in smaller amounts from uninfected cells; (f) At 37°C. the virus-induced leakage reaction appears within a matter of seconds, and usually decreases after 2 to 3 minutes; (g) The reaction is inhibited by 0.025 M Mg++. Theoretical considerations are presented suggesting the place of this reaction in the sequence of events constituting the virus penetration reaction; its relationship to the phenomenon of lysis-from-without; and its resemblance to the leakage reaction produced by electrostatic binding of ionized compounds to cell surfaces. The existence of similar effects in avian-mammalian virus systems is noted. PMID:13163323

  18. Interactions of Condensed Tannins with Saccharomyces cerevisiae Yeast Cells and Cell Walls: Tannin Location by Microscopy.

    PubMed

    Mekoue Nguela, Julie; Vernhet, Aude; Sieczkowski, Nathalie; Brillouet, Jean-Marc

    2015-09-01

    Interactions between grape tannins/red wine polyphenols and yeast cells/cell walls was previously studied within the framework of red wine aging and the use of yeast-derived products as an alternative to aging on lees. Results evidenced a quite different behavior between whole cells (biomass grown to elaborate yeast-derived products, inactivated yeast, and yeast inactivated after autolysis) and yeast cell walls (obtained from mechanical disruption of the biomass). Briefly, whole cells exhibited a high capacity to irreversibly adsorb grape and wine tannins, whereas only weak interactions were observed for cell walls. This last point was quite unexpected considering the literature and called into question the real role of cell walls in yeasts' ability to fix tannins. In the present work, tannin location after interactions between grape and wine tannins and yeast cells and cell walls was studied by means of transmission electron microscopy, light epifluorescence, and confocal microscopy. Microscopy observations evidenced that if tannins interact with cell walls, and especially cell wall mannoproteins, they also diffuse freely through the walls of dead cells to interact with their plasma membrane and cytoplasmic components.

  19. Interactions of Condensed Tannins with Saccharomyces cerevisiae Yeast Cells and Cell Walls: Tannin Location by Microscopy.

    PubMed

    Mekoue Nguela, Julie; Vernhet, Aude; Sieczkowski, Nathalie; Brillouet, Jean-Marc

    2015-09-01

    Interactions between grape tannins/red wine polyphenols and yeast cells/cell walls was previously studied within the framework of red wine aging and the use of yeast-derived products as an alternative to aging on lees. Results evidenced a quite different behavior between whole cells (biomass grown to elaborate yeast-derived products, inactivated yeast, and yeast inactivated after autolysis) and yeast cell walls (obtained from mechanical disruption of the biomass). Briefly, whole cells exhibited a high capacity to irreversibly adsorb grape and wine tannins, whereas only weak interactions were observed for cell walls. This last point was quite unexpected considering the literature and called into question the real role of cell walls in yeasts' ability to fix tannins. In the present work, tannin location after interactions between grape and wine tannins and yeast cells and cell walls was studied by means of transmission electron microscopy, light epifluorescence, and confocal microscopy. Microscopy observations evidenced that if tannins interact with cell walls, and especially cell wall mannoproteins, they also diffuse freely through the walls of dead cells to interact with their plasma membrane and cytoplasmic components. PMID:26223789

  20. Anammox Planctomycetes have a peptidoglycan cell wall

    PubMed Central

    van Teeseling, Muriel C.F.; Mesman, Rob J.; Kuru, Erkin; Espaillat, Akbar; Cava, Felipe; Brun, Yves V.; VanNieuwenhze, Michael S.; Kartal, Boran; van Niftrik, Laura

    2015-01-01

    Planctomycetes are intriguing microorganisms that apparently lack peptidoglycan, a structure that controls the shape and integrity of almost all bacterial cells. Therefore, the planctomycetal cell envelope is considered exceptional and their cell plan uniquely compartmentalized. Anaerobic ammonium-oxidizing (anammox) Planctomycetes play a key role in the global nitrogen cycle by releasing fixed nitrogen back to the atmosphere as N2. Here using a complementary array of state-of-the-art techniques including continuous culturing, cryo-transmission electron microscopy, peptidoglycan-specific probes and muropeptide analysis, we show that the anammox bacterium Kuenenia stuttgartiensis contains peptidoglycan. On the basis of the thickness, composition and location of peptidoglycan in K. stuttgartiensis, we propose to redefine Planctomycetes as Gram-negative bacteria. Our results demonstrate that Planctomycetes are not an exception to the universal presence of peptidoglycan in bacteria. PMID:25962786

  1. A model of cell wall expansion based on thermodynamics of polymer networks

    NASA Technical Reports Server (NTRS)

    Veytsman, B. A.; Cosgrove, D. J.

    1998-01-01

    A theory of cell wall extension is proposed. It is shown that macroscopic properties of cell walls can be explained through the microscopic properties of interpenetrating networks of cellulose and hemicellulose. The qualitative conclusions of the theory agree with the existing experimental data. The dependence of the cell wall yield threshold on the secretion of the wall components is discussed.

  2. Wood Contains a Cell-Wall Structural Protein

    NASA Astrophysics Data System (ADS)

    Bao, Wuli; O'Malley, David M.; Sederoff, Ronald R.

    1992-07-01

    A pine extensin-like protein (PELP) has been localized in metabolically active cells of differentiating xylem and in mature wood of loblolly pine (Pinus taeda L.). This proline-rich glycosylated protein was purified from cell walls of differentiating xylem by differential solubility and gel electrophoresis. Polyclonal rabbit antibodies were raised against the deglycosylated purified protein (dPELP) and purified antibody was used for immunolocalization. Immunogold and alkaline phosphatase secondary antibody staining both show antigen in secondary cell walls of earlywood and less staining in latewood. Immunoassays of milled dry wood were developed and used to show increased availability of antigen after hydrogen fluoride or cellulase treatment and decreased antigen after chlorite treatment. The specificity of the antigen-antibody reaction was confirmed by competition assays and by preadsorption of antibody to the purified protein. We propose that extensin-like protein is present in xylem cell walls during lignification and that the protein remains as a structural component of cell walls in wood for many years after xylogenesis. We suggest that such structural proteins play important roles in the differentiation of xylem and thereby could affect the properties of wood.

  3. Extracellular proteases modify cell wall turnover in Bacillus subtilis.

    PubMed Central

    Jolliffe, L K; Doyle, R J; Streips, U N

    1980-01-01

    The rate of turnover of peptidoglycan in exponentially growing cultures of Bacillus subtilis was observed to be sensitive to extracellular protease. In protease-deficient mutants the rates of cell wall turnover were greater than that of wild-type strain 168, whereas hyperprotease-producing strains exhibited decreased rates of peptidoglycan turnover. The rate of peptidogylcan turnover in a protease-deficient strain was decreased when the mutant was grown in the presence of a hyperprotease-producing strain. The addition of phenylmethylsulfonyl fluoride, a serine protease inhibitor, to cultures of hyperprotease-producing strains increased their rates of cell wall turnover. Isolated cell walls of all protease mutants contained autolysin levels equal to or greater than that of wild-type strain 168. The presence of filaments, or cells with incomplete septa, was observed in hyperprotease-producing strains or when a protease-deficient strain was grown in the presence of subtilisin. The results suggest that the turnover of cell walls in B. subtilis may be regulated by extracellular proteases. Images PMID:6102558

  4. Site Specific Genetic Incorporation of Azidophenylalanine in Schizosaccharomyces pombe.

    PubMed

    Shao, Nan; Singh, N Sadananda; Slade, Susan E; Jones, Alexandra M E; Balasubramanian, Mohan K

    2015-01-01

    The diversity of protein functions is impacted in significant part by the chemical properties of the twenty amino acids, which are used as building blocks for nearly all proteins. The ability to incorporate unnatural amino acids (UAA) into proteins in a site specific manner can vastly expand the repertoire of protein functions and also allows detailed analysis of protein function. In recent years UAAs have been incorporated in a site-specific manner into proteins in a number of organisms. In nearly all cases, the amber codon is used as a sense codon, and an orthogonal tRNA/aminoacyl-tRNA synthetase (RS) pair is used to generate amber suppressing tRNAs charged with the UAA. In this work, we have developed tools to incorporate the cross-linking amino acid azido-phenylalanine (AzF) through the use of bacterial tRNA(Tyr) and a modified version of TyrRS, AzFRS, in Schizosaccharomyces pombe, which is an attractive model organism for the study of cell behavior and function. We have incorporated AzF into three different proteins. We show that the majority of AzF is modified to amino-phenyl alanine, but protein cross-linking was still observed. These studies set the stage for exploitation of this new technology for the analysis of S. pombe proteins.

  5. Site Specific Genetic Incorporation of Azidophenylalanine in Schizosaccharomyces pombe

    PubMed Central

    Shao, Nan; Singh, N. Sadananda; Slade, Susan E.; Jones, Alexandra M. E.; Balasubramanian, Mohan K.

    2015-01-01

    The diversity of protein functions is impacted in significant part by the chemical properties of the twenty amino acids, which are used as building blocks for nearly all proteins. The ability to incorporate unnatural amino acids (UAA) into proteins in a site specific manner can vastly expand the repertoire of protein functions and also allows detailed analysis of protein function. In recent years UAAs have been incorporated in a site-specific manner into proteins in a number of organisms. In nearly all cases, the amber codon is used as a sense codon, and an orthogonal tRNA/aminoacyl-tRNA synthetase (RS) pair is used to generate amber suppressing tRNAs charged with the UAA. In this work, we have developed tools to incorporate the cross-linking amino acid azido-phenylalanine (AzF) through the use of bacterial tRNATyr and a modified version of TyrRS, AzFRS, in Schizosaccharomyces pombe, which is an attractive model organism for the study of cell behavior and function. We have incorporated AzF into three different proteins. We show that the majority of AzF is modified to amino-phenyl alanine, but protein cross-linking was still observed. These studies set the stage for exploitation of this new technology for the analysis of S. pombe proteins. PMID:26597962

  6. The transcription factor Rap1p is required for tolerance to cell-wall perturbing agents and for cell-wall maintenance in Saccharomyces cerevisiae.

    PubMed

    Azad, Gajendra Kumar; Singh, Vikash; Baranwal, Shivani; Thakare, Mayur Jankiram; Tomar, Raghuvir S

    2015-01-01

    Yeast repressor activator protein (Rap1p) is involved in genomic stability and transcriptional regulation. We explored the function of Rap1p in yeast physiology using Rap1p truncation mutants. Our results revealed that the N-terminal truncation of Rap1p (Rap1ΔN) leads to hypersensitivity towards elevated temperature and cell-wall perturbing agents. Cell wall analysis showed an increase in the chitin and glucan content in Rap1ΔN cells as compared with wild type cells. Accordingly, mutant cells had a twofold thicker cell wall, as observed by electron microscopy. Furthermore, Rap1ΔN cells had increased levels of phosphorylated Slt2p, a MAP kinase of the cell wall integrity pathway. Mutant cells also had elevated levels of cell wall integrity response transcripts. Taken together, our findings suggest a connection between Rap1p and cell wall homeostasis.

  7. O-Acetylation of Plant Cell Wall Polysaccharides

    PubMed Central

    Gille, Sascha; Pauly, Markus

    2011-01-01

    Plant cell walls are composed of structurally diverse polymers, many of which are O-acetylated. How plants O-acetylate wall polymers and what its function is remained elusive until recently, when two protein families were identified in the model plant Arabidopsis that are involved in the O-acetylation of wall polysaccharides – the reduced wall acetylation (RWA) and the trichome birefringence-like (TBL) proteins. This review discusses the role of these two protein families in polysaccharide O-acetylation and outlines the differences and similarities of polymer acetylation mechanisms in plants, fungi, bacteria, and mammals. Members of the TBL protein family had been shown to impact pathogen resistance, freezing tolerance, and cellulose biosynthesis. The connection of TBLs to polysaccharide O-acetylation thus gives crucial leads into the biological function of wall polymer O-acetylation. From a biotechnological point understanding the O-acetylation mechanism is important as acetyl-substituents inhibit the enzymatic degradation of wall polymers and released acetate can be a potent inhibitor in microbial fermentations, thus impacting the economic viability of, e.g., lignocellulosic based biofuel production. PMID:22639638

  8. Cell wall structure and function in lactic acid bacteria

    PubMed Central

    2014-01-01

    The cell wall of Gram-positive bacteria is a complex assemblage of glycopolymers and proteins. It consists of a thick peptidoglycan sacculus that surrounds the cytoplasmic membrane and that is decorated with teichoic acids, polysaccharides, and proteins. It plays a major role in bacterial physiology since it maintains cell shape and integrity during growth and division; in addition, it acts as the interface between the bacterium and its environment. Lactic acid bacteria (LAB) are traditionally and widely used to ferment food, and they are also the subject of more and more research because of their potential health-related benefits. It is now recognized that understanding the composition, structure, and properties of LAB cell walls is a crucial part of developing technological and health applications using these bacteria. In this review, we examine the different components of the Gram-positive cell wall: peptidoglycan, teichoic acids, polysaccharides, and proteins. We present recent findings regarding the structure and function of these complex compounds, results that have emerged thanks to the tandem development of structural analysis and whole genome sequencing. Although general structures and biosynthesis pathways are conserved among Gram-positive bacteria, studies have revealed that LAB cell walls demonstrate unique properties; these studies have yielded some notable, fundamental, and novel findings. Given the potential of this research to contribute to future applied strategies, in our discussion of the role played by cell wall components in LAB physiology, we pay special attention to the mechanisms controlling bacterial autolysis, bacterial sensitivity to bacteriophages and the mechanisms underlying interactions between probiotic bacteria and their hosts. PMID:25186919

  9. Direct measurement of cell wall stress stiffening and turgor pressure in live bacterial cells.

    PubMed

    Deng, Yi; Sun, Mingzhai; Shaevitz, Joshua W

    2011-10-01

    We study intact and bulging Escherichia coli cells using atomic force microscopy to separate the contributions of the cell wall and turgor pressure to the overall cell stiffness. We find strong evidence of power-law stress stiffening in the E. coli cell wall, with an exponent of 1.22±0.12, such that the wall is significantly stiffer in intact cells (E=23±8  MPa and 49±20  MPa in the axial and circumferential directions) than in unpressurized sacculi. These measurements also indicate that the turgor pressure in living cells E. coli is 29±3  kPa.

  10. Direct Measurement of Cell Wall Stress Stiffening and Turgor Pressure in Live Bacterial Cells

    NASA Astrophysics Data System (ADS)

    Deng, Yi; Sun, Mingzhai; Shaevitz, Joshua W.

    2011-10-01

    We study intact and bulging Escherichia coli cells using atomic force microscopy to separate the contributions of the cell wall and turgor pressure to the overall cell stiffness. We find strong evidence of power-law stress stiffening in the E. coli cell wall, with an exponent of 1.22±0.12, such that the wall is significantly stiffer in intact cells (E=23±8MPa and 49±20MPa in the axial and circumferential directions) than in unpressurized sacculi. These measurements also indicate that the turgor pressure in living cells E. coli is 29±3kPa.

  11. Influence of the Cell Wall on Intracellular Delivery to Algal Cells by Electroporation and Sonication

    PubMed Central

    Azencott, Harold R.; Peter, Gary F.; Prausnitz, Mark R.

    2007-01-01

    To assess the cell wall’s role as a barrier to intracellular delivery, wild-type Chlamydomonas reinhardtii algal cells and mutant cells lacking a cell wall were exposed to electroporation or sonication. Flow cytometry determined intracellular uptake of calcein and bovine serum albumin (BSA) and loss of cell viability as functions of electroporation transmembrane potential and acoustic energy. Electroporation of wild-type cells increased calcein uptake with increasing transmembrane potential, but delivered much less BSA. Electroporation of wall-deficient cells had similar effects on calcein uptake, but increased BSA uptake as much as 7.5-fold relative to wild-type cells, which indicated that the cell wall was a significant barrier to BSA delivery during electroporation. Sonication of wild-type cells caused calcein and BSA uptake at similar levels. This suggests that the cell wall barrier to BSA delivery can be overcome by sonication. Increased electroporation transmembrane potential or acoustic energy also caused increased loss of cell viability, where wall-deficient cells were especially susceptible to lysis. Overall, we believe this is the first study to compare the effects of electroporation and sonication in a direct fashion in any cell type. Specifically, these findings suggest that electroporation primarily transports molecules across the plasma membrane, because its mechanism is specific to lipid bilayer disruption, whereas sonication transports molecules across both the plasma membrane and cell wall, because it non-specifically disrupts cell-surface barriers. PMID:17602827

  12. Medicago truncatula as a Model for Dicot Cell Wall Development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strong interest in renewable energy has promoted an upsurge of research on plant cell wall traits that influence the availability of lignocellulosic-derived sugars for fermentation in production of biofuels. We have initiated a genome-wide transcript profiling study using the model legume Medicago t...

  13. Roles of cell wall peroxidases in plant development.

    PubMed

    Francoz, Edith; Ranocha, Philippe; Nguyen-Kim, Huan; Jamet, Elisabeth; Burlat, Vincent; Dunand, Christophe

    2015-04-01

    Class III peroxidases (CIII Prxs) are plant specific proteins. Based on in silico prediction and experimental evidence, they are mainly considered as cell wall localized proteins. Thanks to their dual hydroxylic and peroxidative cycles, they can produce ROS as well as oxidize cell wall aromatic compounds within proteins and phenolics that are either free or linked to polysaccharides. Thus, they are tightly associated to cell wall loosening and stiffening. They are members of large multigenic families, mostly due to an elevated rate of gene duplication in higher plants, resulting in a high risk of functional redundancy between them. However, proteomic and (micro)transcriptomic analyses have shown that CIII Prx expression profiles are highly specific. Based on these omic analyses, several reverse genetic studies have demonstrated the importance of the spatio-temporal regulation of their expression and ability to interact with cell wall microdomains in order to achieve specific activity in vivo. Each CIII Prx isoform could have specific functions in muro and this could explain the conservation of a high number of genes in plant genomes.

  14. Polymer mobility in cell walls of cucumber hypocotyls

    NASA Technical Reports Server (NTRS)

    Fenwick, K. M.; Apperley, D. C.; Cosgrove, D. J.; Jarvis, M. C.

    1999-01-01

    Cell walls were prepared from the growing region of cucumber (Cucumis sativus) hypocotyls and examined by solid-state 13C NMR spectroscopy, in both enzymically active and inactivated states. The rigidity of individual polymer segments within the hydrated cell walls was assessed from the proton magnetic relaxation parameter, T2, and from the kinetics of cross-polarisation from 1H to 13C. The microfibrils, including most of the xyloglucan in the cell wall, as well as cellulose, behaved as very rigid solids. A minor xyloglucan fraction, which may correspond to cross-links between microfibrils, shared a lower level of rigidity with some of the pectic galacturonan. Other pectins, including most of the galactan side-chain residues of rhamnogalacturonan I, were much more mobile and behaved in a manner intermediate between the solid and liquid states. The only difference observed between the enzymically active and inactive cell walls, was the loss of a highly mobile, methyl-esterified galacturonan fraction, as the result of pectinesterase activity.

  15. Imaging of plant cell walls by confocal Raman microscopy.

    PubMed

    Gierlinger, Notburga; Keplinger, Tobias; Harrington, Michael

    2012-09-01

    Raman imaging of plant cell walls represents a nondestructive technique that can provide insights into chemical composition in context with structure at the micrometer level (<0.5 μm). The major steps of the experimental procedure are described: sample preparation (embedding and microcutting), setting the mapping parameters, and finally the calculation of chemical images on the basis of the acquired Raman spectra. Every Raman image is based on thousands of spectra, each being a spatially resolved molecular 'fingerprint' of the cell wall. Multiple components are analyzed within the native cell walls, and insights into polymer composition as well as the orientation of the cellulose microfibrils can be gained. The most labor-intensive step of this process is often the sample preparation, as the imaging approach requires a flat surface of the plant tissue with intact cell walls. After finishing the map (acquisition time is ∼10 min to 10 h, depending on the size of the region of interest and scanning parameters), many possibilities exist for the analysis of spectral data and image generation.

  16. Determination of carbohydrate profile in sugarbeet (Beta vulgaris) cell walls

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sugarbeet germplasms USH20, C869, EL55, EL54 were used, and different tissues at different developmental stages were sampled, including dry seeds, germinating seedlings, developing leaves, mature leaves, petioles, hypocotyls, mature roots, flowering stems and inflorescences. Cell Wall Composition An...

  17. A cytoplasmic peptidoglycan amidase homologue controls mycobacterial cell wall synthesis.

    PubMed

    Boutte, Cara C; Baer, Christina E; Papavinasasundaram, Kadamba; Liu, Weiru; Chase, Michael R; Meniche, Xavier; Fortune, Sarah M; Sassetti, Christopher M; Ioerger, Thomas R; Rubin, Eric J

    2016-01-01

    Regulation of cell wall assembly is essential for bacterial survival and contributes to pathogenesis and antibiotic tolerance in Mycobacterium tuberculosis (Mtb). However, little is known about how the cell wall is regulated in stress. We found that CwlM, a protein homologous to peptidoglycan amidases, coordinates peptidoglycan synthesis with nutrient availability. Surprisingly, CwlM is sequestered from peptidoglycan (PG) by localization in the cytoplasm, and its enzymatic function is not essential. Rather, CwlM is phosphorylated and associates with MurA, the first enzyme in PG precursor synthesis. Phosphorylated CwlM activates MurA ~30 fold. CwlM is dephosphorylated in starvation, resulting in lower MurA activity, decreased cell wall metabolism, and increased tolerance to multiple antibiotics. A phylogenetic analysis of cwlM implies that localization in the cytoplasm drove the evolution of this factor. We describe a system that controls cell wall metabolism in response to starvation, and show that this regulation contributes to antibiotic tolerance. PMID:27304077

  18. Hetero-oligomeric cell wall channels (porins) of Nocardia farcinica.

    PubMed

    Kläckta, Christian; Knörzer, Philipp; Riess, Franziska; Benz, Roland

    2011-06-01

    The cell wall of Nocardia farcinica contains a cation-selective cell wall channel, which may be responsible for the limited permeability of the cell wall of N. farcinica for negatively charged antibiotics. Based on partial sequencing of the protein responsible for channel formation derived from N. farcinica ATTC 3318 we were able to identify the corresponding genes (nfa15890 and nfa15900) within the known genome of N. farcinica IFM 10152. The corresponding genes of N. farcinica ATTC 3318 were separately expressed in the Escherichia coli BL21DE3Omp8 strain and the N-terminal His10-tagged proteins were purified to homogeneity using immobilized metal affinity chromatography. The pure proteins were designated NfpANHis and NfpBNHis, for N. farcinica porin A and N. farcinica porin B. The two proteins were checked separately for channel formation in lipid bilayers. Our results clearly indicate that the proteins NfpANHis and NfpBNHis expressed in E. coli could only together form a channel in lipid bilayer membranes. This means that the cell wall channel of N. farcinica is formed by a heterooligomer. NfpA and NfpB form together a channel that may structurally be related to MspA of Mycobacterium smegmatis based on amino acid comparison and renaturation procedure.

  19. Hetero-oligomeric cell wall channels (porins) of Nocardia farcinica.

    PubMed

    Kläckta, Christian; Knörzer, Philipp; Riess, Franziska; Benz, Roland

    2011-06-01

    The cell wall of Nocardia farcinica contains a cation-selective cell wall channel, which may be responsible for the limited permeability of the cell wall of N. farcinica for negatively charged antibiotics. Based on partial sequencing of the protein responsible for channel formation derived from N. farcinica ATTC 3318 we were able to identify the corresponding genes (nfa15890 and nfa15900) within the known genome of N. farcinica IFM 10152. The corresponding genes of N. farcinica ATTC 3318 were separately expressed in the Escherichia coli BL21DE3Omp8 strain and the N-terminal His10-tagged proteins were purified to homogeneity using immobilized metal affinity chromatography. The pure proteins were designated NfpANHis and NfpBNHis, for N. farcinica porin A and N. farcinica porin B. The two proteins were checked separately for channel formation in lipid bilayers. Our results clearly indicate that the proteins NfpANHis and NfpBNHis expressed in E. coli could only together form a channel in lipid bilayer membranes. This means that the cell wall channel of N. farcinica is formed by a heterooligomer. NfpA and NfpB form together a channel that may structurally be related to MspA of Mycobacterium smegmatis based on amino acid comparison and renaturation procedure. PMID:21092733

  20. A cytoplasmic peptidoglycan amidase homologue controls mycobacterial cell wall synthesis

    PubMed Central

    Boutte, Cara C; Baer, Christina E; Papavinasasundaram, Kadamba; Liu, Weiru; Chase, Michael R; Meniche, Xavier; Fortune, Sarah M; Sassetti, Christopher M; Ioerger, Thomas R; Rubin, Eric J

    2016-01-01

    Regulation of cell wall assembly is essential for bacterial survival and contributes to pathogenesis and antibiotic tolerance in Mycobacterium tuberculosis (Mtb). However, little is known about how the cell wall is regulated in stress. We found that CwlM, a protein homologous to peptidoglycan amidases, coordinates peptidoglycan synthesis with nutrient availability. Surprisingly, CwlM is sequestered from peptidoglycan (PG) by localization in the cytoplasm, and its enzymatic function is not essential. Rather, CwlM is phosphorylated and associates with MurA, the first enzyme in PG precursor synthesis. Phosphorylated CwlM activates MurA ~30 fold. CwlM is dephosphorylated in starvation, resulting in lower MurA activity, decreased cell wall metabolism, and increased tolerance to multiple antibiotics. A phylogenetic analysis of cwlM implies that localization in the cytoplasm drove the evolution of this factor. We describe a system that controls cell wall metabolism in response to starvation, and show that this regulation contributes to antibiotic tolerance. DOI: http://dx.doi.org/10.7554/eLife.14590.001 PMID:27304077

  1. Biosynthesis and assembly of cell wall polysaccharides in cereal grasses

    SciTech Connect

    Carpita, N.C.

    1991-04-01

    We have just completed the second year of a three-year project entitled Biosynthesis assembly of cell wall polysaccharides in cereal grasses.'' We made significant progress on two aspects of cell wall synthesis in grasses and greatly refined gas-liquid and high- performance liquid chromatographic techniques necessary to identify the products of synthesis in vitro and in vivo. First, Dr. David Gibeaut, a post-doctoral associate, devised a convenient procedure for the enrichment of Golgi membranes by flotation centrifugation following initial downward rate-zonal separation. Based on comparison of the IDPase marker enzyme, flotation centrifugation enriched the Golgi apparatus almost 7-fold after the initial downward separation. This system is now used in our studies of the synthesis in vitro of the mixed-linkage {beta}-D-glucan. Second, Gibeaut and I have devised a simple technique to feed radioactive sugars into intact growing seedlings and follow incorporation of radioactivity into and turnover from specific cell wall polysaccharides. The project has also provided a few spin-off projects that have been productive as well. First, in collaboration with the group of Prof. Peter Kaufman, University of Michigan, we examined changes in cell wall structure concomitant with reaction to gravistimulation in the gravisensing oat pulvinus. Second, Dr. Gibeaut developed a simple clean-up procedure for partially methylated alditol derivatives to remove a large amount of undesirable interfering compounds that confound separation of the derivatives by gas-liquid chromatography. 5 refs.

  2. Casein kinase 2 inhibits HomolD-directed transcription by Rrn7 in Schizosaccharomyces pombe.

    PubMed

    Moreira-Ramos, Sandra; Rojas, Diego A; Montes, Matías; Urbina, Fabiola; Miralles, Vicente J; Maldonado, Edio

    2015-02-01

    In Schizosaccharomyces pombe, ribosomal protein gene (RPG) promoters contain a TATA analogue element called the HomolD box. The HomolD-binding protein Rrn7 forms a complex with the RNA polymerase II machinery. Despite the importance of ribosome biogenesis to cell survival, the mechanisms involved in the regulation of transcription of eukaryotic RPGs are unknown. In this study, we identified Rrn7 as a new substrate of the pleiotropic casein kinase 2 (CK2), which is a regulator of basal transcription. Recombinant Rrn7 from S. pombe, which is often used as a model organism for studying eukaryotic transcription, interacted with CK2 in vitro and in vivo. Furthermore, CK2-mediated phosphorylation of Rrn7 inhibited its HomolD-directed transcriptional activity and ability to bind to an oligonucleotide containing a HomolD box in vitro. Mutation of Rrn7 at Thr67 abolished these effects, indicating that this residue is a critical CK2 phosphorylation site. Finally, Rrn7 interacted with the regulatory subunit of CK2 in vivo, inhibition of CK2 in vivo potentiated ribosomal protein gene transcription, and chromatin immunoprecipitation analyses identified that the catalytic subunit of CK2 was associated with the rpk5 gene promoter in S. pombe. Taken together, these data suggest that CK2 inhibits ribosomal protein gene transcription in S. pombe via phosphorylation of Rrn7 at Thr67.

  3. Molecular deformation mechanisms of the wood cell wall material.

    PubMed

    Jin, Kai; Qin, Zhao; Buehler, Markus J

    2015-02-01

    Wood is a biological material with outstanding mechanical properties resulting from its hierarchical structure across different scales. Although earlier work has shown that the cellular structure of wood is a key factor that renders it excellent mechanical properties at light weight, the mechanical properties of the wood cell wall material itself still needs to be understood comprehensively. The wood cell wall material features a fiber reinforced composite structure, where cellulose fibrils act as stiff fibers, and hemicellulose and lignin molecules act as soft matrix. The angle between the fiber direction and the loading direction has been found to be the key factor controlling the mechanical properties. However, how the interactions between theses constitutive molecules contribute to the overall properties is still unclear, although the shearing between fibers has been proposed as a primary deformation mechanism. Here we report a molecular model of the wood cell wall material with atomistic resolution, used to assess the mechanical behavior under shear loading in order to understand the deformation mechanisms at the molecular level. The model includes an explicit description of cellulose crystals, hemicellulose, as well as lignin molecules arranged in a layered nanocomposite. The results obtained using this model show that the wood cell wall material under shear loading deforms in an elastic and then plastic manner. The plastic regime can be divided into two parts according to the different deformation mechanisms: yielding of the matrix and sliding of matrix along the cellulose surface. Our molecular dynamics study provides insights of the mechanical behavior of wood cell wall material at the molecular level, and paves a way for the multi-scale understanding of the mechanical properties of wood.

  4. Assembling of the Mycobacterium tuberculosis Cell Wall Core.

    PubMed

    Grzegorzewicz, Anna E; de Sousa-d'Auria, Célia; McNeil, Michael R; Huc-Claustre, Emilie; Jones, Victoria; Petit, Cécile; Angala, Shiva Kumar; Zemanová, Júlia; Wang, Qinglan; Belardinelli, Juan Manuel; Gao, Qian; Ishizaki, Yoshimasa; Mikušová, Katarína; Brennan, Patrick J; Ronning, Donald R; Chami, Mohamed; Houssin, Christine; Jackson, Mary

    2016-09-01

    The unique cell wall of mycobacteria is essential to their viability and the target of many clinically used anti-tuberculosis drugs and inhibitors under development. Despite intensive efforts to identify the ligase(s) responsible for the covalent attachment of the two major heteropolysaccharides of the mycobacterial cell wall, arabinogalactan (AG) and peptidoglycan (PG), the enzyme or enzymes responsible have remained elusive. We here report on the identification of the two enzymes of Mycobacterium tuberculosis, CpsA1 (Rv3267) and CpsA2 (Rv3484), responsible for this function. CpsA1 and CpsA2 belong to the widespread LytR-Cps2A-Psr (LCP) family of enzymes that has been shown to catalyze a variety of glycopolymer transfer reactions in Gram-positive bacteria, including the attachment of wall teichoic acids to PG. Although individual cpsA1 and cpsA2 knock-outs of M. tuberculosis were readily obtained, the combined inactivation of both genes appears to be lethal. In the closely related microorganism Corynebacterium glutamicum, the ortholog of cpsA1 is the only gene involved in this function, and its conditional knockdown leads to dramatic changes in the cell wall composition and morphology of the bacteria due to extensive shedding of cell wall material in the culture medium as a result of defective attachment of AG to PG. This work marks an important step in our understanding of the biogenesis of the unique cell envelope of mycobacteria and opens new opportunities for drug development.

  5. Structure of Plant Cell Walls: XI. GLUCURONOARABINOXYLAN, A SECOND HEMICELLULOSE IN THE PRIMARY CELL WALLS OF SUSPENSION-CULTURED SYCAMORE CELLS.

    PubMed

    Darvill, J E; McNeil, M; Darvill, A G; Albersheim, P

    1980-12-01

    The isolation, purification, and partial characterization of a glucuronoarabinoxylan, a previously unobserved component of the primary cell walls of dicotyledonous plants, are described. The glucuronoarabinoxylan constitutes approximately 5% of the primary walls of suspension-cultured sycamore cells. This glucuronoarabinoxylan possesses many of the structural characteristics of analogous polysaccharides that have been isolated from the primary and secondary cell walls of monocots as well as from the secondary cell walls of dicots. The glucuronoarabinoxylan of primary dicot cell walls has a linear beta-1,4-linked d-xylopyranosyl backbone with both neutral and acidic sidechains attached at intervals along its length. The acidic sidechains are terminated with glucuronosyl or 4-O-methyl glucuronosyl residues, whereas the neutral sidechains are composed of arabinosyl and/or xylosyl residues.

  6. Cell-Wall Polysaccharides of Developing Flax Plants.

    PubMed Central

    Gorshkova, T. A.; Wyatt, S. E.; Salnikov, V. V.; Gibeaut, D. M.; Ibragimov, M. R.; Lozovaya, V. V.; Carpita, N. C.

    1996-01-01

    Flax (Linum usitatissimum L.) fibers originate from procambial cells of the protophloem and develop in cortical bundles that encircle the vascular cylinder. We determined the polysaccharide composition of the cell walls from various organs of the developing flax plant, from fiber-rich strips peeled from the stem, and from the xylem. Ammonium oxalate-soluble polysaccharides from all tissues contained 5-linked arabinans with low degrees of branching, rhamnogalacturonans, and polygalacturonic acid. The fiber-rich peels contained, in addition, substantial amounts of a buffer-soluble, 4-linked galactan branched at the 0-2 and 0-3 positions with nonreducing terminal-galactosyl units. The cross-linking glycans from all tissues were (fucogalacto)xyloglucan, typical of type-I cell walls, xylans containing (1->)-[beta]-D-xylosyl units branched exclusively at the xylosyl O-2 with t-(4-O-methyl)-glucosyluronic acid units, and (galacto)glucomannans. Tissues containing predominantly primary cell wall contained a larger proportion of xyloglucan. The xylem cells were composed of about 60% 4-xylans, 32% cellulose, and small amounts of pectin and the other cross-linking polysaccharides. The noncellulosic polysaccharides of flax exhibit an uncommonly low degree of branching compared to similar polysaccharides from other flowering plants. Although the relative abundance of the various noncellulosic polysaccharides varies widely among the different cell types, the linkage structure and degree of branching of several of the noncellulosic polysaccharides are invariant. PMID:12226214

  7. Nucleated assembly of Chlamydomonas and Volvox cell walls.

    PubMed

    Adair, W S; Steinmetz, S A; Mattson, D M; Goodenough, U W; Heuser, J E

    1987-11-01

    The Chlamydomonas reinhardtii cell wall is made up of hydroxyproline-rich glycoproteins, arranged in five distinct layers. The W6 (crystalline) layer contains three major glycoproteins (GP1, GP2, GP3), selectively extractable with chaotropic agents, that self-assemble into crystals in vitro. A system to study W6 assembly in a quantitative fashion was developed that employs perchlorate-extracted Chlamydomonas cells as nucleating agents. Wall reconstitution by biotinylated W6 monomers was monitored by FITC-streptavidin fluorescence and quick-freeze/deep-etch electron microscopy. Optimal reconstitution was obtained at monomer concentrations (0.2-0.3 mg/ml) well below those required for nonnucleated assembly. Assembly occurred from multiple nucleation sites, and faithfully reflected the structure of the intact W6 layer. Specificity of nucleated assembly was demonstrated using two cell-wall mutants (cw-2 and cw-15); neither served as a substrate for assembly of wild-type monomers. In addition, W6 sublayers were assembled from purified components: GP2 and GP3 coassembled to form the inner (W6A) sublayer; this then served as a substrate for self-assembly of GP1 into the outer (W6B) sublayer. Finally, evolutionary relationships between C. reinhardtii and two additional members of the Volvocales (Chlamydomonas eugametos and Volvox carteri) were explored by performing interspecific reconstitutions. Hybrid walls were obtained between C. reinhardtii and Volvox but not with C. eugametos, confirming taxonomic assignments based on structural criteria. PMID:3680387

  8. Crushing Strength of Aluminum Honeycomb with Thinning Cell Wall

    NASA Astrophysics Data System (ADS)

    Ogasawara, Nagahisa; Chiba, Norimasa; Kobayashi, Eiji; Kikuchi, Yuji

    To evaluate the crash safety of automobiles, various collision tests are performed by the auto industry. In the offset frontal collision test and the side collision test, the target is an aluminum honeycomb material which has thinning cell walls. In this study, based on the analyses of the shock absorption mechanism, a new crushing strength formula is proposed. First, load-displacement curves obtained from compression tests in quasi-static condition showed an almost linear relation between a thinning rate of cell walls and a crushing strength. Second, based on Wierzbicki's theory, a new formula was proposed, which can estimate a crushing strength of a honeycomb material with thinning wall. In addition, a correcting equation which considered an elastic deformation was also proposed. Third, parametric analyses were carried out with a FE model which can simulate a delamination between cell walls. The results obtained from the theory and FEM almost corresponded to each other for a wide range of the thinning rate. Fourth, impact tests were carried out, in which the weight was dropped freely at the speed used for the automobile tests. Those results almost agreed well with the sum of the theoretical crush strength and the inside air pressure.

  9. Engineering of plant cell walls for enhanced biofuel production.

    PubMed

    Loqué, Dominique; Scheller, Henrik V; Pauly, Markus

    2015-06-01

    The biomass of plants consists predominately of cell walls, a sophisticated composite material composed of various polymer networks including numerous polysaccharides and the polyphenol lignin. In order to utilize this renewable, highly abundant resource for the production of commodity chemicals such as biofuels, major hurdles have to be surpassed to reach economical viability. Recently, major advances in the basic understanding of the synthesis of the various wall polymers and its regulation has enabled strategies to alter the qualitative composition of wall materials. Such emerging strategies include a reduction/alteration of the lignin network to enhance polysaccharide accessibility, reduction of polymer derived processing inhibitors, and increases in polysaccharides with a high hexose/pentose ratio.

  10. Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells

    PubMed Central

    Liesche, Johannes; Marek, Magdalena; Günther-Pomorski, Thomas

    2015-01-01

    Yeast cells are protected by a cell wall that plays an important role in the exchange of substances with the environment. The cell wall structure is dynamic and can adapt to different physiological states or environmental conditions. For the investigation of morphological changes, selective staining with fluorescent dyes is a valuable tool. Furthermore, cell wall staining is used to facilitate sub-cellular localization experiments with fluorescently-labeled proteins and the detection of yeast cells in non-fungal host tissues. Here, we report staining of Saccharomyces cerevisiae cell wall with Trypan Blue, which emits strong red fluorescence upon binding to chitin and yeast glucan; thereby, it facilitates cell wall analysis by confocal and super-resolution microscopy. The staining pattern of Trypan Blue was similar to that of the widely used UV-excitable, blue fluorescent cell wall stain Calcofluor White. Trypan Blue staining facilitated quantification of cell size and cell wall volume when utilizing the optical sectioning capacity of a confocal microscope. This enabled the quantification of morphological changes during growth under anaerobic conditions and in the presence of chemicals, demonstrating the potential of this approach for morphological investigations or screening assays. PMID:25717323

  11. Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells.

    PubMed

    Liesche, Johannes; Marek, Magdalena; Günther-Pomorski, Thomas

    2015-01-01

    Yeast cells are protected by a cell wall that plays an important role in the exchange of substances with the environment. The cell wall structure is dynamic and can adapt to different physiological states or environmental conditions. For the investigation of morphological changes, selective staining with fluorescent dyes is a valuable tool. Furthermore, cell wall staining is used to facilitate sub-cellular localization experiments with fluorescently-labeled proteins and the detection of yeast cells in non-fungal host tissues. Here, we report staining of Saccharomyces cerevisiae cell wall with Trypan Blue, which emits strong red fluorescence upon binding to chitin and yeast glucan; thereby, it facilitates cell wall analysis by confocal and super-resolution microscopy. The staining pattern of Trypan Blue was similar to that of the widely used UV-excitable, blue fluorescent cell wall stain Calcofluor White. Trypan Blue staining facilitated quantification of cell size and cell wall volume when utilizing the optical sectioning capacity of a confocal microscope. This enabled the quantification of morphological changes during growth under anaerobic conditions and in the presence of chemicals, demonstrating the potential of this approach for morphological investigations or screening assays.

  12. Caffeine stabilizes Cdc25 independently of Rad3 in Schizosaccharomyces pombe contributing to checkpoint override.

    PubMed

    Alao, John P; Sjölander, Johanna J; Baar, Juliane; Özbaki-Yagan, Nejla; Kakoschky, Bianca; Sunnerhagen, Per

    2014-05-01

    Cdc25 is required for Cdc2 dephosphorylation and is thus essential for cell cycle progression. Checkpoint activation requires dual inhibition of Cdc25 and Cdc2 in a Rad3-dependent manner. Caffeine is believed to override activation of the replication and DNA damage checkpoints by inhibiting Rad3-related proteins in both Schizosaccharomyces pombe and mammalian cells. In this study, we have investigated the impact of caffeine on Cdc25 stability, cell cycle progression and checkpoint override. Caffeine induced Cdc25 accumulation in S. pombe independently of Rad3. Caffeine delayed cell cycle progression under normal conditions but advanced mitosis in cells treated with replication inhibitors and DNA-damaging agents. In the absence of Cdc25, caffeine inhibited cell cycle progression even in the presence of hydroxyurea or phleomycin. Caffeine induces Cdc25 accumulation in S. pombe by suppressing its degradation independently of Rad3. The induction of Cdc25 accumulation was not associated with accelerated progression through mitosis, but rather with delayed progression through cytokinesis. Caffeine-induced Cdc25 accumulation appears to underlie its ability to override cell cycle checkpoints. The impact of Cdc25 accumulation on cell cycle progression is attenuated by Srk1 and Mad2. Together our findings suggest that caffeine overrides checkpoint enforcement by inducing the inappropriate nuclear localization of Cdc25.

  13. Messenger Functions of the Bacterial Cell Wall-derived Muropeptides

    PubMed Central

    Boudreau, Marc A.; Fisher, Jed. F.; Mobashery, Shahriar

    2012-01-01

    Bacterial muropeptides are soluble peptidoglycan structures central to recycling of the bacterial cell wall, and messengers in diverse cell-signaling events. Bacteria sense muropeptides as signals that antibiotics targeting cell-wall biosynthesis are present, and eukaryotes detect muropeptides during the innate immune response to bacterial infection. This review summarizes the roles of bacterial muropeptides as messengers, with a special emphasis on bacterial muropeptide structures and the relationship of structure to the biochemical events that the muropeptides elicit. Muropeptide sensing and recycling in both Gram-positive and Gram-negative bacteria is discussed, followed by muropeptide sensing by eukaryotes as a crucial event to the innate immune response of insects (via peptidoglycan-recognition proteins) and mammals (through Nod-like receptors) to bacterial invasion. PMID:22409164

  14. Resistance to antibiotics targeted to the bacterial cell wall

    PubMed Central

    Nikolaidis, I; Favini-Stabile, S; Dessen, A

    2014-01-01

    Peptidoglycan is the main component of the bacterial cell wall. It is a complex, three-dimensional mesh that surrounds the entire cell and is composed of strands of alternating glycan units crosslinked by short peptides. Its biosynthetic machinery has been, for the past five decades, a preferred target for the discovery of antibacterials. Synthesis of the peptidoglycan occurs sequentially within three cellular compartments (cytoplasm, membrane, and periplasm), and inhibitors of proteins that catalyze each stage have been identified, although not all are applicable for clinical use. A number of these antimicrobials, however, have been rendered inactive by resistance mechanisms. The employment of structural biology techniques has been instrumental in the understanding of such processes, as well as the development of strategies to overcome them. This review provides an overview of resistance mechanisms developed toward antibiotics that target bacterial cell wall precursors and its biosynthetic machinery. Strategies toward the development of novel inhibitors that could overcome resistance are also discussed. PMID:24375653

  15. TERRA promotes telomerase-mediated telomere elongation in Schizosaccharomyces pombe.

    PubMed

    Moravec, Martin; Wischnewski, Harry; Bah, Amadou; Hu, Yan; Liu, Na; Lafranchi, Lorenzo; King, Megan C; Azzalin, Claus M

    2016-07-01

    Telomerase-mediated telomere elongation provides cell populations with the ability to proliferate indefinitely. Telomerase is capable of recognizing and extending the shortest telomeres in cells; nevertheless, how this mechanism is executed remains unclear. Here, we show that, in the fission yeast Schizosaccharomyces pombe, shortened telomeres are highly transcribed into the evolutionarily conserved long noncoding RNA TERRA A fraction of TERRA produced upon telomere shortening is polyadenylated and largely devoid of telomeric repeats, and furthermore, telomerase physically interacts with this polyadenylated TERRA in vivo We also show that experimentally enhanced transcription of a manipulated telomere promotes its association with telomerase and concomitant elongation. Our data represent the first direct evidence that TERRA stimulates telomerase recruitment and activity at chromosome ends in an organism with human-like telomeres.

  16. Identification of Cell Wall Synthesis Regulatory Genes Controlling Biomass Characteristics and Yield in Rice (Oryza Sativa)

    SciTech Connect

    Peng, Zhaohua PEng; Ronald, Palmela; Wang, Guo-Liang

    2013-04-26

    This project aims to identify the regulatory genes of rice cell wall synthesis pathways using a cell wall removal and regeneration system. We completed the gene expression profiling studies following the time course from cell wall removal to cell wall regeneration in rice suspension cells. We also completed, total proteome, nuclear subproteome and histone modification studies following the course from cell wall removal and cell wall regeneration process. A large number of differentially expressed regulatory genes and proteins were identified. Meanwhile, we generated RNAi and over-expression transgenic rice for 45 genes with at least 10 independent transgenic lines for each gene. In addition, we ordered T-DNA and transposon insertion mutants for 60 genes from Korea, Japan, and France and characterized the mutants. Overall, we have mutants and transgenic lines for over 90 genes, exceeded our proposed goal of generating mutants for 50 genes. Interesting Discoveries a) Cell wall re-synthesis in protoplasts may involve a novel cell wall synthesis mechanism. The synthesis of the primary cell wall is initiated in late cytokinesis with further modification during cell expansion. Phragmoplast plays an essential role in cell wall synthesis. It services as a scaffold for building the cell plate and formation of a new cell wall. Only one phragmoplast and one new cell wall is produced for each dividing cell. When the cell wall was removed enzymatically, we found that cell wall re-synthesis started from multiple locations simultaneously, suggesting that a novel mechanism is involved in cell wall re-synthesis. This observation raised many interesting questions, such as how the starting sites of cell wall synthesis are determined, whether phragmoplast and cell plate like structures are involved in cell wall re-synthesis, and more importantly whether the same set of enzymes and apparatus are used in cell wall re-synthesis as during cytokinesis. Given that many known cell wall

  17. Ectopic lignification in primary cellulose-deficient cell walls of maize cell suspension cultures.

    PubMed

    Mélida, Hugo; Largo-Gosens, Asier; Novo-Uzal, Esther; Santiago, Rogelio; Pomar, Federico; García, Pedro; García-Angulo, Penélope; Acebes, José Luis; Álvarez, Jesús; Encina, Antonio

    2015-04-01

    Maize (Zea mays L.) suspension-cultured cells with up to 70% less cellulose were obtained by stepwise habituation to dichlobenil (DCB), a cellulose biosynthesis inhibitor. Cellulose deficiency was accompanied by marked changes in cell wall matrix polysaccharides and phenolics as revealed by Fourier transform infrared (FTIR) spectroscopy. Cell wall compositional analysis indicated that the cellulose-deficient cell walls showed an enhancement of highly branched and cross-linked arabinoxylans, as well as an increased content in ferulic acid, diferulates and p-coumaric acid, and the presence of a polymer that stained positive for phloroglucinol. In accordance with this, cellulose-deficient cell walls showed a fivefold increase in Klason-type lignin. Thioacidolysis/GC-MS analysis of cellulose-deficient cell walls indicated the presence of a lignin-like polymer with a Syringyl/Guaiacyl ratio of 1.45, which differed from the sensu stricto stress-related lignin that arose in response to short-term DCB-treatments. Gene expression analysis of these cells indicated an overexpression of genes specific for the biosynthesis of monolignol units of lignin. A study of stress signaling pathways revealed an overexpression of some of the jasmonate signaling pathway genes, which might trigger ectopic lignification in response to cell wall integrity disruptions. In summary, the structural plasticity of primary cell walls is proven, since a lignification process is possible in response to cellulose impoverishment. PMID:25735403

  18. Ectopic lignification in primary cellulose-deficient cell walls of maize cell suspension cultures.

    PubMed

    Mélida, Hugo; Largo-Gosens, Asier; Novo-Uzal, Esther; Santiago, Rogelio; Pomar, Federico; García, Pedro; García-Angulo, Penélope; Acebes, José Luis; Álvarez, Jesús; Encina, Antonio

    2015-04-01

    Maize (Zea mays L.) suspension-cultured cells with up to 70% less cellulose were obtained by stepwise habituation to dichlobenil (DCB), a cellulose biosynthesis inhibitor. Cellulose deficiency was accompanied by marked changes in cell wall matrix polysaccharides and phenolics as revealed by Fourier transform infrared (FTIR) spectroscopy. Cell wall compositional analysis indicated that the cellulose-deficient cell walls showed an enhancement of highly branched and cross-linked arabinoxylans, as well as an increased content in ferulic acid, diferulates and p-coumaric acid, and the presence of a polymer that stained positive for phloroglucinol. In accordance with this, cellulose-deficient cell walls showed a fivefold increase in Klason-type lignin. Thioacidolysis/GC-MS analysis of cellulose-deficient cell walls indicated the presence of a lignin-like polymer with a Syringyl/Guaiacyl ratio of 1.45, which differed from the sensu stricto stress-related lignin that arose in response to short-term DCB-treatments. Gene expression analysis of these cells indicated an overexpression of genes specific for the biosynthesis of monolignol units of lignin. A study of stress signaling pathways revealed an overexpression of some of the jasmonate signaling pathway genes, which might trigger ectopic lignification in response to cell wall integrity disruptions. In summary, the structural plasticity of primary cell walls is proven, since a lignification process is possible in response to cellulose impoverishment.

  19. Penium margaritaceum as a model organism for cell wall analysis of expanding plant cells.

    PubMed

    Rydahl, Maja G; Fangel, Jonatan U; Mikkelsen, Maria Dalgaard; Johansen, I Elisabeth; Andreas, Amanda; Harholt, Jesper; Ulvskov, Peter; Jørgensen, Bodil; Domozych, David S; Willats, William G T

    2015-01-01

    The growth of a plant cell encompasses a complex set of subcellular components interacting in a highly coordinated fashion. Ultimately, these activities create specific cell wall structural domains that regulate the prime force of expansion, internally generated turgor pressure. The precise organization of the polymeric networks of the cell wall around the protoplast also contributes to the direction of growth, the shape of the cell, and the proper positioning of the cell in a tissue. In essence, plant cell expansion represents the foundation of development. Most studies of plant cell expansion have focused primarily upon late divergent multicellular land plants and specialized cell types (e.g., pollen tubes, root hairs). Here, we describe a unicellular green alga, Penium margaritaceum (Penium), which can serve as a valuable model organism for understanding cell expansion and the underlying mechanics of the cell wall in a single plant cell.

  20. Viscoelastic properties of cell walls of single living plant cells determined by dynamic nanoindentation.

    PubMed

    Hayot, Céline M; Forouzesh, Elham; Goel, Ashwani; Avramova, Zoya; Turner, Joseph A

    2012-04-01

    Plant development results from controlled cell divisions, structural modifications, and reorganizations of the cell wall. Thereby, regulation of cell wall behaviour takes place at multiple length scales involving compositional and architectural aspects in addition to various developmental and/or environmental factors. The physical properties of the primary wall are largely determined by the nature of the complex polymer network, which exhibits time-dependent behaviour representative of viscoelastic materials. Here, a dynamic nanoindentation technique is used to measure the time-dependent response and the viscoelastic behaviour of the cell wall in single living cells at a micron or sub-micron scale. With this approach, significant changes in storage (stiffness) and loss (loss of energy) moduli are captured among the tested cells. The results reveal hitherto unknown differences in the viscoelastic parameters of the walls of same-age similarly positioned cells of the Arabidopsis ecotypes (Col 0 and Ws 2). The technique is also shown to be sensitive enough to detect changes in cell wall properties in cells deficient in the activity of the chromatin modifier ATX1. Extensive computational modelling of the experimental measurements (i.e. modelling the cell as a viscoelastic pressure vessel) is used to analyse the influence of the wall thickness, as well as the turgor pressure, at the positions of our measurements. By combining the nanoDMA technique with finite element simulations quantifiable measurements of the viscoelastic properties of plant cell walls are achieved. Such techniques are expected to find broader applications in quantifying the influence of genetic, biological, and environmental factors on the nanoscale mechanical properties of the cell wall.

  1. Chemical modification of the surfaces of bacterial cell walls.

    PubMed

    Neihof, R A; Echols, W H

    1978-01-01

    The surfaces of the isolated cell walls of four bacterial species were studied by microelectrophoresis following chemical treatments intended to remove specific charged groups. Acid-base titrations of the walls were used to assess specificity and extent of the modifications. Carboxyl groups were specifically and completely modified by activation with a water-soluble carbodiimide and subsequent reaction with a nucleophile, such as glycinamide, to give an uncharged pH-stable product. Aqueous media and mild reaction conditions make the method suitable for modifying carboxyl groups on cell surfaces too labile to withstand the harsh conditions required for conventional esterification reactions. Use of the carbodiimide-mediated reaction for discharging carboxyl groups, along with fluorodinitrobenzene for discharging amino groups and extraction procedures for removing constituents carrying phosphoester groups (teichoic acids), made it possible to obtain information about the spatial arrangement of charged groups on the wall surfaces. Removal of the exterior negative charge dominating wall surfaces allowed underlying amino groups to become electrokinetically effective and, in the case of E. coli, also revealed a lipophilic region with an affinity for a cationic surfactant.

  2. Cell Wall Architecture of the Elongating Maize Coleoptile1

    PubMed Central

    Carpita, Nicholas C.; Defernez, Marianne; Findlay, Kim; Wells, Brian; Shoue, Douglas A.; Catchpole, Gareth; Wilson, Reginald H.; McCann, Maureen C.

    2001-01-01

    The primary walls of grasses are composed of cellulose microfibrils, glucuronoarabinoxylans (GAXs), and mixed-linkage β-glucans, together with smaller amounts of xyloglucans, glucomannans, pectins, and a network of polyphenolic substances. Chemical imaging by Fourier transform infrared microspectroscopy revealed large differences in the distributions of many chemical species between different tissues of the maize (Zea mays) coleoptile. This was confirmed by chemical analyses of isolated outer epidermal tissues compared with mesophyll-enriched preparations. Glucomannans and esterified uronic acids were more abundant in the epidermis, whereas β-glucans were more abundant in the mesophyll cells. The localization of β-glucan was confirmed by immunocytochemistry in the electron microscope and quantitative biochemical assays. We used field emission scanning electron microscopy, infrared microspectroscopy, and biochemical characterization of sequentially extracted polymers to further characterize the cell wall architecture of the epidermis. Oxidation of the phenolic network followed by dilute NaOH extraction widened the pores of the wall substantially and permitted observation by scanning electron microscopy of up to six distinct microfibrillar lamellae. Sequential chemical extraction of specific polysaccharides together with enzymic digestion of β-glucans allowed us to distinguish two distinct domains in the grass primary wall. First, a β-glucan-enriched domain, coextensive with GAXs of low degrees of arabinosyl substitution and glucomannans, is tightly associated around microfibrils. Second, a GAX that is more highly substituted with arabinosyl residues and additional glucomannan provides an interstitial domain that interconnects the β-glucan-coated microfibrils. Implications for current models that attempt to explain the biochemical and biophysical mechanism of wall loosening during cell growth are discussed. PMID:11598229

  3. (Rapid regulatory control of plant cell expansion and wall relaxation)

    SciTech Connect

    Cosgrove, D.J.

    1990-01-01

    This section presents a brief overview of accomplishments related to this project in the past 3-year period. Our work has focused on the basic mechanisms of plant cell expansion, particularly on the interrelations of water and solute transport with cell wall relaxation and expansion. To study these processes, we have developed new methods and used these methods to analyze the dynamic behavior of growth processes and to examine how various agents (GA, drought, light, genetic lesions) alter the growth machinery of the cell.

  4. Quantitative Fitness Analysis Identifies exo1∆ and Other Suppressors or Enhancers of Telomere Defects in Schizosaccharomyces pombe

    PubMed Central

    Narayanan, Siddharth; Dubarry, Marion; Lawless, Conor; Banks, A. Peter; Wilkinson, Darren J.; Whitehall, Simon K.; Lydall, David

    2015-01-01

    Synthetic genetic array (SGA) has been successfully used to identify genetic interactions in S. cerevisiae and S. pombe. In S. pombe, SGA methods use either cycloheximide (C) or heat shock (HS) to select double mutants before measuring colony size as a surrogate for fitness. Quantitative Fitness Analysis (QFA) is a different method for determining fitness of microbial strains. In QFA, liquid cultures are spotted onto solid agar and growth curves determined for each spot by photography and model fitting. Here, we compared the two S. pombe SGA methods and found that the HS method was more reproducible for us. We also developed a QFA procedure for S. pombe. We used QFA to identify genetic interactions affecting two temperature sensitive, telomere associated query mutations (taz1Δ and pot1-1). We identify exo1∆ and other gene deletions as suppressors or enhancers of S. pombe telomere defects. Our study identifies known and novel gene deletions affecting the fitness of strains with telomere defects. The interactions we identify may be relevant in human cells. PMID:26168240

  5. Quantitative Fitness Analysis Identifies exo1∆ and Other Suppressors or Enhancers of Telomere Defects in Schizosaccharomyces pombe.

    PubMed

    Narayanan, Siddharth; Dubarry, Marion; Lawless, Conor; Banks, A Peter; Wilkinson, Darren J; Whitehall, Simon K; Lydall, David

    2015-01-01

    Synthetic genetic array (SGA) has been successfully used to identify genetic interactions in S. cerevisiae and S. pombe. In S. pombe, SGA methods use either cycloheximide (C) or heat shock (HS) to select double mutants before measuring colony size as a surrogate for fitness. Quantitative Fitness Analysis (QFA) is a different method for determining fitness of microbial strains. In QFA, liquid cultures are spotted onto solid agar and growth curves determined for each spot by photography and model fitting. Here, we compared the two S. pombe SGA methods and found that the HS method was more reproducible for us. We also developed a QFA procedure for S. pombe. We used QFA to identify genetic interactions affecting two temperature sensitive, telomere associated query mutations (taz1Δ and pot1-1). We identify exo1∆ and other gene deletions as suppressors or enhancers of S. pombe telomere defects. Our study identifies known and novel gene deletions affecting the fitness of strains with telomere defects. The interactions we identify may be relevant in human cells.

  6. Orbital wall infarction in child with sickle cell disease.

    PubMed

    Janssens, C; Claeys, L; Maes, P; Boiy, T; Wojciechowski, M

    2015-12-01

    We present the case of a 17-year-old boy, known with homozygous sickle cell disease, who was admitted because of generalised pain. He developed bilateral periorbital oedema and proptosis, without pain or visual disturbances. In addition to hyperhydration, oxygen and analgesia IV antibiotics were started, to cover a possible osteomyelitis. Patients with sickle cell disease are at risk for vaso-occlusive crises, when the abnormally shaped red blood cells aggregate and block the capillaries. Such a crisis typically presents at a location with high bone marrow activity, as the vertebrae and long bones. At an early age, the bone marrow is still active at other sites, for example the orbital wall, and thus infarction can also occur there. Thus, in young persons with sickle cell disease, it is important to consider orbital wall infarction in the differential diagnosis, since the approach is different from osteomyelitis. If the disease is complicated by an orbital compression syndrome, corticosteroids or surgical intervention may be necessary to preserve the vision. In our patient, an MRI of the orbitae demonstrated periorbital oedema with bone anomalies in the orbital and frontal bones, confirming orbital wall infarction. Ophthalmological examination revealed no signs of pressure on the nervus opticus. The patient recovered gradually with conservative treatment. PMID:26790559

  7. Orbital wall infarction in child with sickle cell disease.

    PubMed

    Janssens, C; Claeys, L; Maes, P; Boiy, T; Wojciechowski, M

    2015-12-01

    We present the case of a 17-year-old boy, known with homozygous sickle cell disease, who was admitted because of generalised pain. He developed bilateral periorbital oedema and proptosis, without pain or visual disturbances. In addition to hyperhydration, oxygen and analgesia IV antibiotics were started, to cover a possible osteomyelitis. Patients with sickle cell disease are at risk for vaso-occlusive crises, when the abnormally shaped red blood cells aggregate and block the capillaries. Such a crisis typically presents at a location with high bone marrow activity, as the vertebrae and long bones. At an early age, the bone marrow is still active at other sites, for example the orbital wall, and thus infarction can also occur there. Thus, in young persons with sickle cell disease, it is important to consider orbital wall infarction in the differential diagnosis, since the approach is different from osteomyelitis. If the disease is complicated by an orbital compression syndrome, corticosteroids or surgical intervention may be necessary to preserve the vision. In our patient, an MRI of the orbitae demonstrated periorbital oedema with bone anomalies in the orbital and frontal bones, confirming orbital wall infarction. Ophthalmological examination revealed no signs of pressure on the nervus opticus. The patient recovered gradually with conservative treatment.

  8. Sexual reproduction as a response to H sub 2 O sub 2 damage in Schizosaccharomyces pombe

    SciTech Connect

    Bernstein, C.; Johns, V. )

    1989-04-01

    Although sexual reproduction is widespread, its adaptive advantage over asexual reproduction is unclear. One major advantage of sex may be its promotion of recombinational repair of DNA damage during meiosis. This idea predicts that treatment of the asexual form of a facultatively sexual-asexual eucaryote with a DNA-damaging agent may cause it to enter the sexual cycle more frequently. Endogenous hydrogen peroxide is a major natural source of DNA damage. Thus, the authors treated vegetative cells of Schizosaccharomyces pombe with hydrogen peroxide to test if sexual reproduction increases. Among untreated stationary-phase S. pombe populations the sexual spores produced by meiosis represented about 1% of the total cells. However, treatment of late-exponential-phase vegetative cells with hydrogen peroxide increased the percentage of meiotic spores in the stationary phase by 4- to 18-fold. Oxidative damage therefore induces sexual reproduction in a facultatively sexual organism, a result expected by the hypothesis that sex promotes DNA repair.

  9. Compounds active against cell walls of medically important fungi.

    PubMed Central

    Hector, R F

    1993-01-01

    A number of substances that directly or indirectly affect the cell walls of fungi have been identified. Those that actively interfere with the synthesis or degradation of polysaccharide components share the property of being produced by soil microbes as secondary metabolites. Compounds specifically interfering with chitin or beta-glucan synthesis have proven effective in studies of preclinical models of mycoses, though they appear to have a restricted spectrum of coverage. Semisynthetic derivatives of some of the natural products have offered improvements in activity, toxicology, or pharmacokinetic behavior. Compounds which act on the cell wall indirectly or by a secondary mechanism of action, such as the azoles, act against diverse fungi but are usually fungistatic in nature. Overall, these compounds are attractive candidates for further development. PMID:8457977

  10. Cell wall bound anionic peroxidases from asparagus byproducts.

    PubMed

    Jaramillo-Carmona, Sara; López, Sergio; Vazquez-Castilla, Sara; Jimenez-Araujo, Ana; Rodriguez-Arcos, Rocio; Guillen-Bejarano, Rafael

    2014-10-01

    Asparagus byproducts are a good source of cationic soluble peroxidases (CAP) useful for the bioremediation of phenol-contaminated wastewaters. In this study, cell wall bound peroxidases (POD) from the same byproducts have been purified and characterized. The covalent forms of POD represent >90% of the total cell wall bound POD. Isoelectric focusing showed that whereas the covalent fraction is constituted primarily by anionic isoenzymes, the ionic fraction is a mixture of anionic, neutral, and cationic isoenzymes. Covalently bound peroxidases were purified by means of ion exchange chromatography and affinity chromatography. In vitro detoxification studies showed that although CAP are more effective for the removal of 4-CP and 2,4-DCP, anionic asparagus peroxidase (AAP) is a better option for the removal of hydroxytyrosol (HT), the main phenol present in olive mill wastewaters.

  11. Cytoplasmic streaming in plant cells: the role of wall slip.

    PubMed

    Wolff, K; Marenduzzo, D; Cates, M E

    2012-06-01

    We present a computer simulation study, via lattice Boltzmann simulations, of a microscopic model for cytoplasmic streaming in algal cells such as those of Chara corallina. We modelled myosin motors tracking along actin lanes as spheres undergoing directed motion along fixed lines. The sphere dimension takes into account the fact that motors drag vesicles or other organelles, and, unlike previous work, we model the boundary close to which the motors move as walls with a finite slip layer. By using realistic parameter values for actin lane and myosin density, as well as for endoplasmic and vacuole viscosity and the slip layer close to the wall, we find that this simplified view, which does not rely on any coupling between motors, cytoplasm and vacuole other than that provided by viscous Stokes flow, is enough to account for the observed magnitude of streaming velocities in intracellular fluid in living plant cells.

  12. Plant cell walls: Protecting the barrier from degradation by microbial enzymes.

    PubMed

    Lagaert, Stijn; Beliën, Tim; Volckaert, Guido

    2009-12-01

    Plant cell walls are predominantly composed of polysaccharides, which are connected in a strong, yet resilient network. They determine the size and shape of plant cells and form the interface between the cell and its often hostile environment. To penetrate the cell wall and thus infect plants, most phytopathogens secrete numerous cell wall degrading enzymes. Conversely, as a first line of defense, plant cell walls contain an array of inhibitors of these enzymes. Scientific knowledge on these inhibitors significantly progressed in the past years and this review is meant to give a comprehensive overview of plant inhibitors against microbial cell wall degrading enzymes and their role in plant protection.

  13. Dynamics of cell wall elasticity pattern shapes the cell during yeast mating morphogenesis.

    PubMed

    Goldenbogen, Björn; Giese, Wolfgang; Hemmen, Marie; Uhlendorf, Jannis; Herrmann, Andreas; Klipp, Edda

    2016-09-01

    The cell wall defines cell shape and maintains integrity of fungi and plants. When exposed to mating pheromone, Saccharomyces cerevisiae grows a mating projection and alters in morphology from spherical to shmoo form. Although structural and compositional alterations of the cell wall accompany shape transitions, their impact on cell wall elasticity is unknown. In a combined theoretical and experimental approach using finite-element modelling and atomic force microscopy (AFM), we investigated the influence of spatially and temporally varying material properties on mating morphogenesis. Time-resolved elasticity maps of shmooing yeast acquired with AFM in vivo revealed distinct patterns, with soft material at the emerging mating projection and stiff material at the tip. The observed cell wall softening in the protrusion region is necessary for the formation of the characteristic shmoo shape, and results in wider and longer mating projections. The approach is generally applicable to tip-growing fungi and plants cells. PMID:27605377

  14. Dynamics of cell wall elasticity pattern shapes the cell during yeast mating morphogenesis

    PubMed Central

    Goldenbogen, Björn; Giese, Wolfgang; Hemmen, Marie; Uhlendorf, Jannis; Herrmann, Andreas

    2016-01-01

    The cell wall defines cell shape and maintains integrity of fungi and plants. When exposed to mating pheromone, Saccharomyces cerevisiae grows a mating projection and alters in morphology from spherical to shmoo form. Although structural and compositional alterations of the cell wall accompany shape transitions, their impact on cell wall elasticity is unknown. In a combined theoretical and experimental approach using finite-element modelling and atomic force microscopy (AFM), we investigated the influence of spatially and temporally varying material properties on mating morphogenesis. Time-resolved elasticity maps of shmooing yeast acquired with AFM in vivo revealed distinct patterns, with soft material at the emerging mating projection and stiff material at the tip. The observed cell wall softening in the protrusion region is necessary for the formation of the characteristic shmoo shape, and results in wider and longer mating projections. The approach is generally applicable to tip-growing fungi and plants cells. PMID:27605377

  15. Single-molecule imaging reveals modulation of cell wall synthesis dynamics in live bacterial cells

    PubMed Central

    Lee, Timothy K.; Meng, Kevin; Shi, Handuo; Huang, Kerwyn Casey

    2016-01-01

    The peptidoglycan cell wall is an integral organelle critical for bacterial cell shape and stability. Proper cell wall construction requires the interaction of synthesis enzymes and the cytoskeleton, but it is unclear how the activities of individual proteins are coordinated to preserve the morphology and integrity of the cell wall during growth. To elucidate this coordination, we used single-molecule imaging to follow the behaviours of the two major peptidoglycan synthases in live, elongating Escherichia coli cells and after perturbation. We observed heterogeneous localization dynamics of penicillin-binding protein (PBP) 1A, the synthase predominantly associated with cell wall elongation, with individual PBP1A molecules distributed between mobile and immobile populations. Perturbations to PBP1A activity, either directly through antibiotics or indirectly through PBP1A's interaction with its lipoprotein activator or other synthases, shifted the fraction of mobile molecules. Our results suggest that multiple levels of regulation control the activity of enzymes to coordinate peptidoglycan synthesis. PMID:27774981

  16. Dynamics of cell wall elasticity pattern shapes the cell during yeast mating morphogenesis.

    PubMed

    Goldenbogen, Björn; Giese, Wolfgang; Hemmen, Marie; Uhlendorf, Jannis; Herrmann, Andreas; Klipp, Edda

    2016-09-01

    The cell wall defines cell shape and maintains integrity of fungi and plants. When exposed to mating pheromone, Saccharomyces cerevisiae grows a mating projection and alters in morphology from spherical to shmoo form. Although structural and compositional alterations of the cell wall accompany shape transitions, their impact on cell wall elasticity is unknown. In a combined theoretical and experimental approach using finite-element modelling and atomic force microscopy (AFM), we investigated the influence of spatially and temporally varying material properties on mating morphogenesis. Time-resolved elasticity maps of shmooing yeast acquired with AFM in vivo revealed distinct patterns, with soft material at the emerging mating projection and stiff material at the tip. The observed cell wall softening in the protrusion region is necessary for the formation of the characteristic shmoo shape, and results in wider and longer mating projections. The approach is generally applicable to tip-growing fungi and plants cells.

  17. Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption.

    PubMed

    Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

    2015-01-01

    The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry. PMID:26295574

  18. Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption

    PubMed Central

    Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

    2015-01-01

    The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry. PMID:26295574

  19. Enzyme Amplified Detection of Microbial Cell Wall Components

    NASA Technical Reports Server (NTRS)

    Wainwright, Norman R.

    2004-01-01

    This proposal is MBL's portion of NASA's Johnson Space Center's Astrobiology Center led by Principal Investigator, Dr. David McKay, entitled: 'Institute for the Study of Biomarkers in Astromaterials.' Dr. Norman Wainwright is the principal investigator at MBL and is responsible for developing methods to detect trace quantities of microbial cell wall chemicals using the enzyme amplification system of Limulus polyphemus and other related methods.

  20. Life behind cell walls: paradigm lost, paradigm regained.

    PubMed

    Lamport, D T

    2001-09-01

    This review of the living cell wall and its protein components is in two parts. The first is anecdotal. A personal account spanning over 40 years research may perhaps be an antidote to one stereotypical view of scientists as detached and humorless. The second part deals with the meaning of function, particularly as it applies to hydroxyproline-rich glycoproteins. Function is a difficult word to define objectively. However, with help from such luminaries as Humpty Dumpty: "A word means what I want it to mean, neither more nor less," and Wittgenstein: "Giving examples of usage ... is the only way to talk about meaning," it is possible to construct a ziggurat representing increasingly complex levels of organization from molecular structure to ecology. Forty years ago I suggested that hydroxyproline-rich structural proteins played a key role in cell wall functioning. But because the bulk of the wall is carbohydrate, there has been an understandable resistance to paradigm change. Expansins, paradoxically, contribute greatly to this resistance because their modus operandi as cell-wall-loosening proteins is based on the idea that they break hydrogen bonds between polysaccharide chains allowing slippage. However, this view is not consistent with the recent discovery [Grobe et al. (1999) Eur. J. Biochem 263: 33-40] that beta-expansins may be proteases, as it implies that the extensin network is not a straightjacket but a substrate for expansin in muro. Such a direct role for extensins in both negative and positive regulation of cell expansion and elongation may constitute a major morphogenetic mechanism operating at all levels of plant growth and development.

  1. Proteomic Analysis to Identify Tightly-Bound Cell Wall Protein in Rice Calli.

    PubMed

    Cho, Won Kyong; Hyun, Tae Kyung; Kumar, Dhinesh; Rim, Yeonggil; Chen, Xiong Yan; Jo, Yeonhwa; Kim, Suwha; Lee, Keun Woo; Park, Zee-Yong; Lucas, William J; Kim, Jae-Yean

    2015-08-01

    Rice is a model plant widely used for basic and applied research programs. Plant cell wall proteins play key roles in a broad range of biological processes. However, presently, knowledge on the rice cell wall proteome is rudimentary in nature. In the present study, the tightly-bound cell wall proteome of rice callus cultured cells using sequential extraction protocols was developed using mass spectrometry and bioinformatics methods, leading to the identification of 1568 candidate proteins. Based on bioinformatics analyses, 389 classical rice cell wall proteins, possessing a signal peptide, and 334 putative non-classical cell wall proteins, lacking a signal peptide, were identified. By combining previously established rice cell wall protein databases with current data for the classical rice cell wall proteins, a comprehensive rice cell wall proteome, comprised of 496 proteins, was constructed. A comparative analysis of the rice and Arabidopsis cell wall proteomes revealed a high level of homology, suggesting a predominant conservation between monocot and eudicot cell wall proteins. This study importantly increased information on cell wall proteins, which serves for future functional analyses of these identified rice cell wall proteins. PMID:26194822

  2. Proteomic Analysis to Identify Tightly-Bound Cell Wall Protein in Rice Calli

    PubMed Central

    Cho, Won Kyong; Hyun, Tae Kyung; Kumar, Dhinesh; Rim, Yeonggil; Chen, Xiong Yan; Jo, Yeonhwa; Kim, Suwha; Lee, Keun Woo; Park, Zee-Yong; Lucas, William J.; Kim, Jae-Yean

    2015-01-01

    Rice is a model plant widely used for basic and applied research programs. Plant cell wall proteins play key roles in a broad range of biological processes. However, presently, knowledge on the rice cell wall proteome is rudimentary in nature. In the present study, the tightly-bound cell wall proteome of rice callus cultured cells using sequential extraction protocols was developed using mass spectrometry and bioinformatics methods, leading to the identification of 1568 candidate proteins. Based on bioinformatics analyses, 389 classical rice cell wall proteins, possessing a signal peptide, and 334 putative non-classical cell wall proteins, lacking a signal peptide, were identified. By combining previously established rice cell wall protein databases with current data for the classical rice cell wall proteins, a comprehensive rice cell wall proteome, comprised of 496 proteins, was constructed. A comparative analysis of the rice and Arabidopsis cell wall proteomes revealed a high level of homology, suggesting a predominant conservation between monocot and eudicot cell wall proteins. This study importantly increased information on cell wall proteins, which serves for future functional analyses of these identified rice cell wall proteins. PMID:26194822

  3. Proteomic Analysis to Identify Tightly-Bound Cell Wall Protein in Rice Calli.

    PubMed

    Cho, Won Kyong; Hyun, Tae Kyung; Kumar, Dhinesh; Rim, Yeonggil; Chen, Xiong Yan; Jo, Yeonhwa; Kim, Suwha; Lee, Keun Woo; Park, Zee-Yong; Lucas, William J; Kim, Jae-Yean

    2015-08-01

    Rice is a model plant widely used for basic and applied research programs. Plant cell wall proteins play key roles in a broad range of biological processes. However, presently, knowledge on the rice cell wall proteome is rudimentary in nature. In the present study, the tightly-bound cell wall proteome of rice callus cultured cells using sequential extraction protocols was developed using mass spectrometry and bioinformatics methods, leading to the identification of 1568 candidate proteins. Based on bioinformatics analyses, 389 classical rice cell wall proteins, possessing a signal peptide, and 334 putative non-classical cell wall proteins, lacking a signal peptide, were identified. By combining previously established rice cell wall protein databases with current data for the classical rice cell wall proteins, a comprehensive rice cell wall proteome, comprised of 496 proteins, was constructed. A comparative analysis of the rice and Arabidopsis cell wall proteomes revealed a high level of homology, suggesting a predominant conservation between monocot and eudicot cell wall proteins. This study importantly increased information on cell wall proteins, which serves for future functional analyses of these identified rice cell wall proteins.

  4. Aspergillus Enzymes Involved in Degradation of Plant Cell Wall Polysaccharides

    PubMed Central

    de Vries, Ronald P.; Visser, Jaap

    2001-01-01

    Degradation of plant cell wall polysaccharides is of major importance in the food and feed, beverage, textile, and paper and pulp industries, as well as in several other industrial production processes. Enzymatic degradation of these polymers has received attention for many years and is becoming a more and more attractive alternative to chemical and mechanical processes. Over the past 15 years, much progress has been made in elucidating the structural characteristics of these polysaccharides and in characterizing the enzymes involved in their degradation and the genes of biotechnologically relevant microorganisms encoding these enzymes. The members of the fungal genus Aspergillus are commonly used for the production of polysaccharide-degrading enzymes. This genus produces a wide spectrum of cell wall-degrading enzymes, allowing not only complete degradation of the polysaccharides but also tailored modifications by using specific enzymes purified from these fungi. This review summarizes our current knowledge of the cell wall polysaccharide-degrading enzymes from aspergilli and the genes by which they are encoded. PMID:11729262

  5. Lignin variability in plant cell walls: contribution of new models.

    PubMed

    Neutelings, Godfrey

    2011-10-01

    Lignin is a major component of certain plant cell walls. The enzymes and corresponding genes associated with the metabolic pathway leading to the production of this complex phenolic polymer have been studied for many years now and are relatively well characterized. The use of genetically modified model plants (Arabidopsis, tobacco, poplar.) and mutants has contributed greatly to our current understanding of this process. The recent utilisation and/or development of a number of dedicated genomic and transcriptomic tools for other species opens new perspectives for advancing our knowledge of the biological role of this important polymer in less typical situations and/or species. In this context, studies on the formation of hypolignified G-type fibres in angiosperm tension wood, and the natural hypolignification of secondary cell walls in plant bast fibre species such as hemp (Cannabis sativa), flax (Linum usitatissimum) or ramie (Boehmeria nivea) are starting to provide novel information about how plants control secondary cell wall formation. Finally, other biologically interesting species for which few molecular resources currently exist could also represent interesting future models.

  6. Progress toward the tomato fruit cell wall proteome

    PubMed Central

    Ruiz-May, Eliel; Rose, Jocelyn K. C.

    2013-01-01

    The plant cell wall (CW) compartment, or apoplast, is host to a highly dynamic proteome, comprising large numbers of both enzymatic and structural proteins. This reflects its importance as the interface between adjacent cells and the external environment, the presence of numerous extracellular metabolic and signaling pathways, and the complex nature of wall structural assembly and remodeling during cell growth and differentiation. Tomato fruit ontogeny, with its distinct phases of rapid growth and ripening, provides a valuable experimental model system for CW proteomic studies, in that it involves substantial wall assembly, remodeling, and coordinated disassembly. Moreover, diverse populations of secreted proteins must be deployed to resist microbial infection and protect against abiotic stresses. Tomato fruits also provide substantial amounts of biological material, which is a significant advantage for many types of biochemical analyses, and facilitates the detection of lower abundance proteins. In this review, we describe a variety of orthogonal techniques that have been applied to identify CW localized proteins from tomato fruit, including approaches that: target the proteome of the CW and the overlying cuticle; functional “secretome” screens; lectin affinity chromatography; and computational analyses to predict proteins that enter the secretory pathway. Each has its merits and limitations, but collectively they are providing important insights into CW proteome composition and dynamics, as well as some potentially controversial issues, such as the prevalence of non-canonical protein secretion. PMID:23755055

  7. Cell wall proteins in seedling cotyledons of Prosopis chilensis.

    PubMed

    Rodríguez, J G; Cardemil, L

    1994-01-01

    Four cell wall proteins of cotyledons of Prosopis chilensis seedlings were characterized by PAGE and Western analyses using a polyclonal antibody, generated against soybean seed coat extensin. These proteins had M(r)s of 180,000, 126,000, 107,000 and 63,000, as determined by SDS-PAGE. The proteins exhibited a fluorescent positive reaction with dansylhydrazine suggesting that they are glycoproteins; they did not show peroxidase activity. The cell wall proteins were also characterized by their amino acid composition and by their amino-terminal sequence. These analyses revealed that there are two groups of related cell wall proteins in the cotyledons. The first group comprises the proteins of M(r)s 180,000, 126,000, 107,000 which are rich in glutamic acid/glutamine and aspartic acid/asparagine and they have almost identical NH2-terminal sequences. The second group comprises the M(r) 63,000 protein which is rich in proline, glycine, valine and tyrosine, with an NH2-terminal sequence which was very similar to that of soybean proline-rich proteins.

  8. Synchronizing Progression of Schizosaccharomyces pombe Cells from G2 through Repeated Rounds of Mitosis and S Phase with cdc25-22 Arrest Release.

    PubMed

    Hagan, Iain M; Grallert, Agnes; Simanis, Viesturs

    2016-01-01

    Transient inactivation of the cdc25(+) gene product by manipulation of the culture temperature for cdc25-22 cells is the most commonly exploited approach to mitotic synchronization in fission yeast. Because Cdc25 removes the inhibitory phosphate placed on Cdk1 by Wee1, inactivation of Cdc25 arrests cells at the G2/M boundary. Incubation at the restrictive temperature of 36°C for just over one generation time forces all cells in the culture to accumulate at the G2/M boundary. Restoration of Cdc25 function via a return to the permissive temperature or chemical inhibition of Wee1 activity at 36°C can then promote a highly synchronous wave of cell division throughout the culture. These approaches can be performed on any scale and thus support simultaneous assessment of numerous events within a single culture. After describing this simple and widely applicable procedure, we discuss frequently overlooked issues that can have a considerable impact on the interpretation of data from cdc25-22 induction-synchronized cultures. PMID:27480720

  9. Modeling of thin, back-wall silicon solar cells

    NASA Technical Reports Server (NTRS)

    Baraona, C. R.

    1979-01-01

    The performance of silicon solar cells with p-n junctions on the nonilluminated surface (i.e., upside-down or back-wall cells) was calculated. These structures consisted of a uniformly shaped p-type substrate layer, a p(+)-type field layer on the front (illuminated) surface, and a shallow, n-type junction on the back (nonilluminated) surface. A four-layer solar cell model was used to calculate efficiency, open-circuit voltage, and short-circuit current. The effect on performance of p-layer thickness and resistivity was determined. The diffusion length was varied to simulate the effect of radiation damage. The results show that peak initial efficiencies greater than 15 percent are possible for cell thicknesses or 100 micrometers or less. After 10 years of radiation damage in geosynchronous orbit, thin (25 to 50 micrometers thick) cells made from 10 to 100 ohm cm material show the smallest decrease (approximately 10 percent) in performance.

  10. Localization of Boron in Cell Walls of Squash and Tobacco and Its Association with Pectin (Evidence for a Structural Role of Boron in the Cell Wall).

    PubMed Central

    Hu, H.; Brown, P. H.

    1994-01-01

    B deficiency results in a rapid inhibition of plant growth, and yet the form and function of B in plants remains unclear. In this paper we provide evidence that B is chemically localized and structurally important in the cell wall of plants. The localization and chemical fractionation of B was followed in squash plants (Curcurbita pepo L.) and cultured tobacco cells (Nicotiana tabacum) grown in B-replete or B-deficient medium. As squash plants and cultured tobacco cells became deficient, an increasingly large proportion of cellular B was found to be localized in the cell wall. Cytoplasmic B concentrations were reduced to essentially zero as plants became deficient, whereas cell wall B concentration remained at or above 10 [mu]g B/g cell wall dry weight in all experiments. Chemical and enzymic fractionation studies suggest that the majority of cell B is associated with pectins within the cell wall. Physical analysis of B-deficient tissue indicates that cell wall plastic extensibility is greatly reduced under B deficiency, and anatomical observations indicate that B deficiency impairs normal cell elongation in growing plant tissue. In plants in which B deficiency had inhibited all plant growth, tissues remained green and did not show any additional visible symptoms for at least 1 week with no additional B. This occurred even though cytoplasmic B had been reduced to extremely low levels (<0.2 [mu]g/g). This suggests that B in these species is largely associated with the cell wall and that any cytoplasmic role for B is satisfied by very low concentrations of B. The localization of B in the cell wall, its association with cell wall pectins, and the contingent effects of B on cell wall extensibility suggest that B plays a critical, although poorly defined, role in the cell wall structure of higher plants. PMID:12232235

  11. Evaluating fundamental position-dependent differences in wood cell wall adhesion using nanoindentation

    PubMed Central

    Obersriebnig, Michael; Konnerth, Johannes; Gindl-Altmutter, Wolfgang

    2013-01-01

    Spruce wood specimens were bonded with one-component polyurethane (PUR) and urea-formaldehyde (UF) adhesive, respectively. The adhesion of the adhesives to the wood cell wall was evaluated at two different locations by means of a new micromechanical assay based on nanoindentation. One location tested corresponded to the interface between the adhesive and the natural inner cell wall surface of the secondary cell wall layer 3 (S3), whereas the second location corresponded to the interface between the adhesive and the freshly cut secondary cell wall layer 2 (S2). Overall, a trend towards reduced cell wall adhesion was found for PUR compared to UF. Position-resolved examination revealed excellent adhesion of UF to freshly cut cell walls (S2) but significantly diminished adhesion to the inner cell wall surface (S3). In contrast, PUR showed better adhesion to the inner cell wall surface and less adhesion to freshly cut cell walls. Atomic force microscopy revealed a less polar character for the inner cell wall surface (S3) compared to freshly cut cell walls (S2). It is proposed that differences in the polarity of the used adhesives and the surface chemistry of the two cell wall surfaces examined account for the observed trends. PMID:27570321

  12. Evidence for 'silicon' within the cell walls of suspension-cultured rice cells.

    PubMed

    He, Congwu; Wang, Lijun; Liu, Jian; Liu, Xin; Li, Xiuli; Ma, Jie; Lin, Yongjun; Xu, Fangsen

    2013-11-01

    Despite the ubiquity and beneficial role of silicon (Si) in plant biology, structural and chemical mechanisms operating at the single-cell level have not been extensively studied. To obtain insights regarding the effect of Si on individual cells, we cultivated suspended rice (Oryza sativa) cells in the absence and presence of Si and analyzed single cells using a combination of physical techniques including atomic force microscopy (AFM). Si is naturally present as a constituent of the cell walls, where it is firmly bound to the cell wall matrix rather than occurring within intra- or extracellular silica deposition, as determined by using inductively coupled plasma mass spectrometry (ICP-MS) and X-ray photoelectron spectroscopy (XPS). This species of Si, linked with the cell wall matrix, improves the structural stability of cell walls during their expansion and subsequent cell division. Maintaining cell shape is thereby enhanced, which may be crucial for the function and survival of cells. This study provides further evidence that organosilicon is present in plant cell walls, which broadens our understanding of the chemical nature of 'anomalous Si' in plant biology.

  13. Stress analysis for wall structure in mobile hot cell design

    NASA Astrophysics Data System (ADS)

    Bahrin, Muhammad Hannan; Rahman, Anwar Abdul; Hamzah, Mohd Arif; Mamat, Mohd Rizal; Azman, Azraf; Hasan, Hasni

    2016-01-01

    Malaysian Nuclear Agency is developing a Mobile Hot Cell (MHC) in order to handle and manage Spent High Activity Radioactive Sources (SHARS) such as teletherapy heads and irradiators. At present, there are only two units of MHC in the world, in South Africa and China. Malaysian Mobile Hot cell is developed by Malaysian Nuclear Agency with the assistance of IAEA expert, based on the design of South Africa and China, but with improved features. Stress analysis has been performed on the design in order to fulfil the safety requirement in operation of MHC. This paper discusses the loading analysis effect from the sand to the MHC wall structure.

  14. Isolation of Cytokinetic Actomyosin Rings from Saccharomyces cerevisiae and Schizosaccharomyces pombe

    PubMed Central

    Palani, Saravanan; Chew, Ting Gang; Balasubramanian, Mohan K.

    2016-01-01

    Cytokinesis is the final stage of cell division, through which cellular constituents of mother cells are partitioned into two daughter cells resulting in the increase in cell number. In animal and fungal cells cytokinesis is mediated by an actomyosin contractile ring, which is attached to the overlying cell membrane. Contraction of this ring after chromosome segregation physically severs the mother cell into two daughters. Here we describe methods for the isolation and partial purification of the actomyosin ring from the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae, which can serve as in vitro systems to facilitate biochemical and ultrastructural analysis of cytokinesis in these genetically tractable model systems. PMID:26519310

  15. Isolation of Cytokinetic Actomyosin Rings from Saccharomyces cerevisiae and Schizosaccharomyces pombe.

    PubMed

    Huang, Junqi; Mishra, Mithilesh; Palani, Saravanan; Chew, Ting Gang; Balasubramanian, Mohan K

    2016-01-01

    Cytokinesis is the final stage of cell division, through which cellular constituents of mother cells are partitioned into two daughter cells resulting in the increase in cell number. In animal and fungal cells cytokinesis is mediated by an actomyosin contractile ring, which is attached to the overlying cell membrane. Contraction of this ring after chromosome segregation physically severs the mother cell into two daughters. Here we describe methods for the isolation and partial purification of the actomyosin ring from the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae, which can serve as in vitro systems to facilitate biochemical and ultrastructural analysis of cytokinesis in these genetically tractable model systems.

  16. Response to Arsenate Treatment in Schizosaccharomyces pombe and the Role of Its Arsenate Reductase Activity

    PubMed Central

    Matia-González, Ana M.; Sotelo, Jael; Zarco-Fernández, Sonia; Muñoz-Olivas, Riansares; Cámara, Carmen; Rodríguez-Gabriel, Miguel A.

    2012-01-01

    Arsenic toxicity has been studied for a long time due to its effects in humans. Although epidemiological studies have demonstrated multiple effects in human physiology, there are many open questions about the cellular targets and the mechanisms of response to arsenic. Using the fission yeast Schizosaccharomyces pombe as model system, we have been able to demonstrate a strong activation of the MAPK Spc1/Sty1 in response to arsenate. This activation is dependent on Wis1 activation and Pyp2 phosphatase inactivation. Using arsenic speciation analysis we have also demonstrated the previously unknown capacity of S. pombe cells to reduce As (V) to As (III). Genetic analysis of several fission yeast mutants point towards the cell cycle phosphatase Cdc25 as a possible candidate to carry out this arsenate reductase activity. We propose that arsenate reduction and intracellular accumulation of arsenite are the key mechanisms of arsenate tolerance in fission yeast. PMID:22912829

  17. In situ analysis of cell wall polymers associated with phloem fibre cells in stems of hemp, Cannabis sativa L.

    PubMed

    Blake, Anthony W; Marcus, Susan E; Copeland, James E; Blackburn, Richard S; Knox, J Paul

    2008-06-01

    A study of stem anatomy and the sclerenchyma fibre cells associated with the phloem tissues of hemp (Cannabis sativa L.) plants is of interest for both understanding the formation of secondary cell walls and for the enhancement of fibre utility as industrial fibres and textiles. Using a range of molecular probes for cell wall polysaccharides we have surveyed the presence of cell wall components in stems of hemp in conjunction with an anatomical survey of stem and phloem fibre development. The only polysaccharide detected to occur abundantly throughout the secondary cell walls of phloem fibres was cellulose. Pectic homogalacturonan epitopes were detected in the primary cell walls/intercellular matrices between the phloem fibres although these epitopes were present at a lower level than in the surrounding parenchyma cell walls. Arabinogalactan-protein glycan epitopes displayed a diversity of occurrence in relation to fibre development and the JIM14 epitope was specific to fibre cells, binding to the inner surface of secondary cell walls, throughout development. Xylan epitopes were found to be present in the fibre cells (and xylem secondary cell walls) and absent from adjacent parenchyma cell walls. Analysis of xylan occurrence in the phloem fibre cells of hemp and flax indicated that xylan epitopes were restricted to the primary cell walls of fibre cells and were not present in the secondary cell walls of these cells.

  18. Biosynthesis of Branched-Chain Amino Acids in Schizosaccharomyces pombe: Properties of Acetohydroxy Acid Synthetase1

    PubMed Central

    McDonald, Roderick A.; Satyanarayana, T.; Kaplan, J. G.

    1973-01-01

    The regulatory properties of acetohydroxy acid synthetase (AHAS), the first enzyme in the biosynthetic pathway to valine and the second in the isoleucine pathway, were investigated in the fission yeast Schizosaccharomyces pombe. The enzyme was partially purified from crude extracts by protamine sulfate treatment, ammonium sulfate fractionation, and gel filtration through Sephadex G-25. AHAS from S. pombe is unique in that its activity shows a single peak around pH 6.5; high sensitivity to feedback inhibition by valine at this pH (Ki = 0.1 mM) indicates that the enzyme is involved in valine biosynthesis. Pyruvate saturation kinetics of AHAS extracted from cells grown on glycerol as sole carbon and energy source were normal and hyperbolic. In contrast, the enzyme from glucose-grown cells exhibited sigmoidal saturation kinetics, an effect which disappeared when the synthetase from such cells was partially purified. This phenomenon was shown to be due to competition for pyruvate between AHAS and pyruvate decarboxylase; the latter enzyme is present in large amounts in cells fermenting glucose. Valine inhibition is noncompetitive in nature, and this effector exhibits homotropic cooperative effects; isoleucine is a less-potent inhibitor of AHAS activity. Mercurial treatment reversibly desensitized the enzyme to valine inhibition. On the basis of these data, the S. pombe AHAS appears to be an allosteric regulatory enzyme with the properties of a negative V system. PMID:4698210

  19. Local Nanomechanical Motion of the Cell Wall of Saccharomyces cerevisiae

    NASA Astrophysics Data System (ADS)

    Pelling, Andrew E.; Sehati, Sadaf; Gralla, Edith B.; Valentine, Joan S.; Gimzewski, James K.

    2004-08-01

    We demonstrate that the cell wall of living Saccharomyces cerevisiae (baker's yeast) exhibits local temperature-dependent nanomechanical motion at characteristic frequencies. The periodic motions in the range of 0.8 to 1.6 kHz with amplitudes of ~3 nm were measured using the cantilever of an atomic force microscope (AFM). Exposure of the cells to a metabolic inhibitor causes the periodic motion to cease. From the strong frequency dependence on temperature, we derive an activation energy of 58 kJ/mol, which is consistent with the cell's metabolism involving molecular motors such as kinesin, dynein, and myosin. The magnitude of the forces observed (~10 nN) suggests concerted nanomechanical activity is operative in the cell.

  20. Enzymology and molecular biology of cell wall biosynthesis. Progress report

    SciTech Connect

    Ray, P.M.

    1993-03-20

    In order to be able to explore the control of cell wall polysaccharide synthesis at the molecular level, which inter alia might eventually lead to means for useful modification of plant biomass polysaccharide production, the immediate goals of this project are to identify polypeptides responsible for wall polysaccharide synthase activities and to obtain clones of the genes that encode them. We are concentrating on plasma membraneassociated (1,3)-{beta}-glucan synthase (glucan synthase-II or GS-II) and Golgi-associated (1,4)-{beta}-glucan synthase (glucan synthase-I or GS-I), of growing pea stem tissue. Our progress has been much more rapid with respect to GS-II than regarding GS-I.

  1. Cell wall pH and auxin transport velocity

    NASA Technical Reports Server (NTRS)

    Hasenstein, K. H.; Rayle, D.

    1984-01-01

    According to the chemiosmotic polar diffusion hypothesis, auxin pulse velocity and basal secretion should increase with decreasing cell wall pH. Experiments were designed to test this prediction. Avena coleoptile sections were preincubated in either fusicoccin (FC), cycloheximide, pH 4.0, or pH 8.0 buffer and subsequently their polar transport capacities were determined. Relative to controls, FC enhanced auxin (IAA) uptake while CHI and pH 8.0 buffer reduced IAA uptake. Nevertheless, FC reduced IAA pulse velocity while cycloheximide increased velocity. Additional experiments showed that delivery of auxin to receivers is enhanced by increased receiver pH. This phenomenon was overcome by a pretreatment of the tissue with IAA. Our data suggest that while acidic wall pH values facilitate cellular IAA uptake, they do not enhance pulse velocity or basal secretion. These findings are inconsistent with the chemiosmotic hypothesis for auxin transport.

  2. Stipe cell wall architecture varies with the stipe elongation of the mushroom Coprinopsis cinerea.

    PubMed

    Niu, Xin; Liu, Zhonghua; Zhou, Yajun; Wang, Jun; Zhang, Wenming; Yuan, Sheng

    2015-10-01

    A large amount of granular protrusions overlie the outer cell wall surfaces in both elongating and non-elongating stipe regions but overlie the inner cell wall surfaces only in non-elongating stipe regions. Removal of granular protrusions using alkali, amorphous materials overlying on both the inner and outer cell wall surfaces were explored in the non-elongating stipe regions. β-1,3-Glucanase treatment not only removed above those granular protrusions and underlying amorphous materials on the wall surfaces but also removed wall matrices embedding chitin microfibrils on the cell walls of most stipe regions, except for the outer cell wall surfaces of the non-elongating stipe regions where most of the wall matrices remained. The chitin microfibrils were closely and transversely arranged on both the inner and outer cell wall surfaces in the elongating apical stipe region, whereas they were loosely and transversely arranged on the inner cell wall surfaces and further became sparser and even randomly arranged on the outer cell wall surface in the non-elongating stipe regions. We propose that the surface deposition of granular protrusions and amorphous materials and the change of microfibril architecture and wall matrices may cause loss of wall plasticity and cessation of stipe elongation.

  3. Cloning and characterisation of the S. pombe rad15 gene, a homologue to the S. cerevisiae RAD3 and human ERCC2 genes.

    PubMed Central

    Murray, J M; Doe, C L; Schenk, P; Carr, A M; Lehmann, A R; Watts, F Z

    1992-01-01

    The RAD3 gene of Saccharomyces cerevisiae encodes an ATP-dependent 5'-3' DNA helicase, which is involved in excision repair of ultraviolet radiation damage. By hybridisation of a Schizosaccharomyces pombe genomic library with a RAD3 gene probe we have isolated the S. pombe homologue of RAD3. We have also cloned the rad15 gene of S. pombe by complementation of radiation-sensitive phenotype of the rad15 mutant. Comparison of the restriction map and DNA sequence, shows that the S. pombe rad15 gene is identical to the gene homologous to S. cerevisiae RAD3, identified by hybridisation. The S. pombe rad15.P mutant is highly sensitive to UV radiation, but only slightly sensitive to ionising radiation, as expected for a mutant defective in excision repair. DNA sequence analysis of the rad15 gene indicates an open reading frame of 772 amino acids, and this is consistent with a transcript size of 2.6 kb as detected by Northern analysis. The predicted rad15 protein has 65% identity to RAD3 and 55% identity to the human homologue ERCC2. This homology is particularly striking in the regions identified as being conserved in a group of DNA helicases. Gene deletion experiments indicate that, like the S. cerevisiae RAD3 gene, the S. pombe rad15 gene is essential for viability, suggesting that the protein product has a role in cell proliferation and not solely in DNA repair. Images PMID:1319571

  4. Change in wall composition of transfer and aleurone cells during wheat grain development.

    PubMed

    Robert, P; Jamme, F; Barron, C; Bouchet, B; Saulnier, L; Dumas, P; Guillon, F

    2011-02-01

    In addition to the starchy endosperm, a specialized tissue accumulating storage material, the endosperm of wheat grain, comprises the aleurone layer and the transfer cells next to the crease. The transfer cells, located at the ventral region of the grain, are involved in nutrient transfer from the maternal tissues to the developing endosperm. Immunolabeling techniques, Raman spectroscopy, and synchrotron infrared micro-spectroscopy were used to study the chemistry of the transfer cell walls during wheat grain development. The kinetic depositions of the main cell wall polysaccharides of wheat grain endosperm, arabinoxylan, and (1-3)(1-4)-β-glucan in transfer cell walls were different from kinetics previously observed in the aleurone cell walls. While (1-3)(1-4)-β-glucan appeared first in the aleurone cell walls at 90°D, arabinoxylan predominated in the transfer cell walls from 90 to 445°D. Both aleurone and transfer cell walls were enriched in (1-3)(1-4)-β-glucan at the mature stage of wheat grain development. Arabinoxylan was more substituted in the transfer cell walls than in the aleurone walls. However, arabinoxylan was more feruloylated in the aleurone than in the transfer cell walls, whatever the stage of grain development. In the transfer cells, the ferulic acid was less abundant in the outer periclinal walls while para-coumarate was absent. Possible implications of such differences are discussed.

  5. Measuring the Mechanical Properties of Plant Cell Walls.

    PubMed

    Vogler, Hannes; Felekis, Dimitrios; Nelson, Bradley J; Grossniklaus, Ueli

    2015-03-25

    The size, shape and stability of a plant depend on the flexibility and integrity of its cell walls, which, at the same time, need to allow cell expansion for growth, while maintaining mechanical stability. Biomechanical studies largely vanished from the focus of plant science with the rapid progress of genetics and molecular biology since the mid-twentieth century. However, the development of more sensitive measurement tools renewed the interest in plant biomechanics in recent years, not only to understand the fundamental concepts of growth and morphogenesis, but also with regard to economically important areas in agriculture, forestry and the paper industry. Recent advances have clearly demonstrated that mechanical forces play a crucial role in cell and organ morphogenesis, which ultimately define plant morphology. In this article, we will briefly review the available methods to determine the mechanical properties of cell walls, such as atomic force microscopy (AFM) and microindentation assays, and discuss their advantages and disadvantages. But we will focus on a novel methodological approach, called cellular force microscopy (CFM), and its automated successor, real-time CFM (RT-CFM).

  6. Measuring the Mechanical Properties of Plant Cell Walls

    PubMed Central

    Vogler, Hannes; Felekis, Dimitrios; Nelson, Bradley J.; Grossniklaus, Ueli

    2015-01-01

    The size, shape and stability of a plant depend on the flexibility and integrity of its cell walls, which, at the same time, need to allow cell expansion for growth, while maintaining mechanical stability. Biomechanical studies largely vanished from the focus of plant science with the rapid progress of genetics and molecular biology since the mid-twentieth century. However, the development of more sensitive measurement tools renewed the interest in plant biomechanics in recent years, not only to understand the fundamental concepts of growth and morphogenesis, but also with regard to economically important areas in agriculture, forestry and the paper industry. Recent advances have clearly demonstrated that mechanical forces play a crucial role in cell and organ morphogenesis, which ultimately define plant morphology. In this article, we will briefly review the available methods to determine the mechanical properties of cell walls, such as atomic force microscopy (AFM) and microindentation assays, and discuss their advantages and disadvantages. But we will focus on a novel methodological approach, called cellular force microscopy (CFM), and its automated successor, real-time CFM (RT-CFM). PMID:27135321

  7. Measuring the Mechanical Properties of Plant Cell Walls.

    PubMed

    Vogler, Hannes; Felekis, Dimitrios; Nelson, Bradley J; Grossniklaus, Ueli

    2015-01-01

    The size, shape and stability of a plant depend on the flexibility and integrity of its cell walls, which, at the same time, need to allow cell expansion for growth, while maintaining mechanical stability. Biomechanical studies largely vanished from the focus of plant science with the rapid progress of genetics and molecular biology since the mid-twentieth century. However, the development of more sensitive measurement tools renewed the interest in plant biomechanics in recent years, not only to understand the fundamental concepts of growth and morphogenesis, but also with regard to economically important areas in agriculture, forestry and the paper industry. Recent advances have clearly demonstrated that mechanical forces play a crucial role in cell and organ morphogenesis, which ultimately define plant morphology. In this article, we will briefly review the available methods to determine the mechanical properties of cell walls, such as atomic force microscopy (AFM) and microindentation assays, and discuss their advantages and disadvantages. But we will focus on a novel methodological approach, called cellular force microscopy (CFM), and its automated successor, real-time CFM (RT-CFM). PMID:27135321

  8. [Hydroxyproline: Rich glycoproteins of the plant and cell wall

    SciTech Connect

    Varner, J.E.

    1993-01-01

    Since xylem tissue includes the main cell types which are lignified, we are interested in gene expression of glycine-rich proteins and proline-rich proteins, and other proteins which are involved in secondary cell wall thickening during xylogenesis. Since the main feature of xylogenesis is the deposition of additional wall components, study of the mechanism of xylogenesis will greatly advance our knowledge of the synthesis and assembly of wall macromolecules. We are using the in vitro xylogenesis system from isolated Zinnia mesophyll cells to isolate genes which are specifically expressed during xylogenesis. We have used subtractive hybridization methods to isolate a number of cDNA clones for differentially regulated genes from the cells after hormonal induction. So far, we have partially characterized 18 different cDNA clones from 239 positive clones. These differentially regulated genes can be divided into three sets according to the characteristics of gene expression in the induction medium and the control medium. The first set is induced in both the induction medium and the control medium without hormones. The second set is induced mainly in the induction medium and in the control medium with the addition of NAA alone. Two of thesegenes are exclusively induced by auxin. The third set of genes is induced mainly in the induction medium. Since these genes are not induced by either auxin or cytokinin alone, they may be directly involved in the process of xylogenesis. Our experiments on the localization of H[sub 2]O[sub 2] production reinforce the earlier ideas of others that H[sub 2]O[sub 2] is involved in normal lignification.

  9. Monoclonal antibodies, carbohydrate-binding modules, and the detection of polysaccharides in plant cell walls.

    PubMed

    Hervé, Cécile; Marcus, Susan E; Knox, J Paul

    2011-01-01

    Plant cell walls are diverse composites of complex polysaccharides. Molecular probes such as monoclonal antibodies (MABs) and carbohydrate-binding modules (CBMs) are important tools to detect and dissect cell wall structures in plant materials. We provide an account of methods that can be used to detect cell wall polysaccharide structures (epitopes) in plant materials and also describe treatments that can provide information on the masking of sets of polysaccharides that may prevent detection. These masking -phenomena may indicate potential interactions between sets of cell wall polysaccharides, and methods to uncover them are an important aspect of cell wall immunocytochemistry.

  10. Comparative structure and biomechanics of plant primary and secondary cell walls.

    PubMed

    Cosgrove, Daniel J; Jarvis, Michael C

    2012-01-01

    Recent insights into the physical biology of plant cell walls are reviewed, summarizing the essential differences between primary and secondary cell walls and identifying crucial gaps in our knowledge of their structure and biomechanics. Unexpected parallels are identified between the mechanism of expansion of primary cell walls during growth and the mechanisms by which hydrated wood deforms under external tension. There is a particular need to revise current "cartoons" of plant cell walls to be more consistent with data from diverse approaches and to go beyond summarizing limited aspects of cell walls, serving instead as guides for future experiments and for the application of new techniques.

  11. Comparative structure and biomechanics of plant primary and secondary cell walls

    PubMed Central

    Cosgrove, Daniel J.; Jarvis, Michael C.

    2012-01-01

    Recent insights into the physical biology of plant cell walls are reviewed, summarizing the essential differences between primary and secondary cell walls and identifying crucial gaps in our knowledge of their structure and biomechanics. Unexpected parallels are identified between the mechanism of expansion of primary cell walls during growth and the mechanisms by which hydrated wood deforms under external tension. There is a particular need to revise current “cartoons” of plant cell walls to be more consistent with data from diverse approaches and to go beyond summarizing limited aspects of cell walls, serving instead as guides for future experiments and for the application of new techniques. PMID:22936943

  12. The Schizosaccharomyces pombe Hsp104 Disaggregase Is Unable to Propagate the [PSI+] Prion

    PubMed Central

    Sénéchal, Patrick; Lindquist, Susan; Rokeach, Luis A.

    2009-01-01

    The molecular chaperone Hsp104 is a crucial factor in the acquisition of thermotolerance in yeast. Under stress conditions, the disaggregase activity of Hsp104 facilitates the reactivation of misfolded proteins. Hsp104 is also involved in the propagation of fungal prions. For instance, the well-characterized [PSI+] prion of Saccharomyces cerevisiae does not propagate in Δhsp104 cells or in cells overexpressing Hsp104. In this study, we characterized the functional homolog of Hsp104 from Schizosaccharomyces pombe (Sp_Hsp104). As its S. cerevisiae counterpart, Sp_hsp104+ is heat-inducible and required for thermotolerance in S. pombe. Sp_Hsp104 displays low disaggregase activity and cannot propagate the [PSI+] prion in S. cerevisiae. When overexpressed in S. cerevisiae, Sp_Hsp104 confers thermotolerance to Δhsp104 cells and reactivates heat-aggregated proteins. However, overexpression of Sp_Hsp104 does not propagate nor eliminate [PSI+]. Strikingly, [PSI+] was cured by overexpression of a chimeric chaperone bearing the C-terminal domain (CTD) of the S. cerevisiae Hsp104 protein. Our study demonstrates that the ability to untangle aggregated proteins is conserved between the S. pombe and S. cerevisiae Hsp104 homologs, and points to a role of the CTD in the propagation of the S. cerevisiae [PSI+] prion. PMID:19759825

  13. Quantitative PCR for detection of DNA damage in mitochondrial DNA of the fission yeast Schizosaccharomyces pombe.

    PubMed

    Senoo, Takanori; Yamanaka, Mayumi; Nakamura, Atori; Terashita, Tomoki; Kawano, Shinji; Ikeda, Shogo

    2016-08-01

    Quantitative polymerase chain reaction (QPCR) has been employed to detect DNA damage and repair in mitochondrial DNA (mtDNA) of human and several model organisms. The assay also permits the quantitation of relative mtDNA copy number in cells. Here, we developed the QPCR assay primers and reaction conditions for the fission yeast Schizosaccharomyces pombe, an important model of eukaryote biology, not previously described. Under these conditions, long targets (approximately 10kb) in mtDNA were quantitatively amplified using 0.1ng of crude DNA templates without isolation of mitochondria and mtDNA. Quantitative detection of oxidative DNA damage in mtDNA was illustrated by using a DNA template irradiated with UVA in the presence of riboflavin. The damage to mtDNA in S. pombe cells treated with hydrogen peroxide and paraquat was also quantitatively measured. Finally, we found that mtDNA copy number in S. pombe cells increased after transition into a stationary phase and that the damage to mtDNA due to endogenous cellular processes accumulated during chronological aging.

  14. Architecture-based multiscale computational modeling of plant cell wall mechanics to examine the hydrogen-bonding hypothesis of cell wall network structure model

    SciTech Connect

    Yi, Hojae; Puri, Virendra M.

    2012-11-01

    A primary plant cell wall network was computationally modeled using the finite element approach to study the hypothesis of hemicellulose (HC) tethering with the cellulose microfibrils (CMFs) as one of the major load-bearing mechanisms of the growing cell wall. A computational primary cell wall network fragment (10 × 10 μm) comprising typical compositions and properties of CMFs and HC was modeled with well-aligned CMFs. The tethering of HC to CMFs is modeled in accordance with the strength of the hydrogen bonding by implementing a specific load-bearing connection (i.e. the joint element). The introduction of the CMF-HC interaction to the computational cell wall network model is a key to the quantitative examination of the mechanical consequences of cell wall structure models, including the tethering HC model. When the cell wall network models with and without joint elements were compared, the hydrogen bond exhibited a significant contribution to the overall stiffness of the cell wall network fragment. When the cell wall network model was stretched 1% in the transverse direction, the tethering of CMF-HC via hydrogen bonds was not strong enough to maintain its integrity. When the cell wall network model was stretched 1% in the longitudinal direction, the tethering provided comparable strength to maintain its integrity. This substantial anisotropy suggests that the HC tethering with hydrogen bonds alone does not manifest sufficient energy to maintain the integrity of the cell wall during its growth (i.e. other mechanisms are present to ensure the cell wall shape).

  15. Proteomic Analysis of Cell Walls of Two Developmental Stages of Alfalfa Stems

    PubMed Central

    Verdonk, Julian C.; Hatfield, Ronald D.; Sullivan, Michael L.

    2012-01-01

    Cell walls are important for the growth and development of all plants. They are also valuable resources for feed and fiber, and more recently as a potential feedstock for bioenergy production. Cell wall proteins comprise only a fraction of the cell wall, but play important roles in establishing the walls and in the chemical interactions (e.g., crosslinking) of cell wall components. This crosslinking provides structure, but restricts digestibility of cell wall complex carbohydrates, limiting available energy in animal and bioenergy production systems. Manipulation of cell wall proteins could be a strategy to improve digestibility. An analysis of the cell wall proteome of apical alfalfa stems (less mature, more digestible) and basal alfalfa stems (more mature, less digestible) was conducted using a recently developed low-salt/density gradient method for the isolation of cell walls. Walls were subsequently subjected to a modified extraction utilizing EGTA to remove pectins, followed by a LiCl extraction to isolate more tightly bound proteins. Recovered proteins were identified using shotgun proteomics. We identified 272 proteins in the alfalfa stem cell wall proteome, 153 of which had not previously been identified in cell wall proteomic analyses. Nearly 70% of the identified proteins were predicted to be secreted, as would be expected for most cell wall proteins, an improvement over previously published studies using traditional cell wall isolation methods. A comparison of our and several other cell wall proteomic studies indicates little overlap in identified proteins among them, which may be largely due to differences in the tissues used as well as differences in experimental approach. PMID:23248635

  16. Bacterial cell wall-induced arthritis: chemical composition and tissue distribution of four Lactobacillus strains.

    PubMed

    Simelyte, E; Rimpiläinen, M; Lehtonen, L; Zhang, X; Toivanen, P

    2000-06-01

    To study what determines the arthritogenicity of bacterial cell walls, cell wall-induced arthritis in the rat was applied, using four strains of Lactobacillus. Three of the strains used proved to induce chronic arthritis in the rat; all were Lactobacillus casei. The cell wall of Lactobacillus fermentum did not induce chronic arthritis. All arthritogenic bacterial cell walls had the same peptidoglycan structure, whereas that of L. fermentum was different. Likewise, all arthritogenic cell walls were resistant to lysozyme degradation, whereas the L. fermentum cell wall was lysozyme sensitive. Muramic acid was observed in the liver, spleen, and lymph nodes in considerably larger amounts after injection of an arthritogenic L. casei cell wall than following injection of a nonarthritogenic L. fermentum cell wall. The L. casei cell wall also persisted in the tissues longer than the L. fermentum cell wall. The present results, taken together with those published previously, underline the possibility that the chemical structure of peptidoglycan is important in determining the arthritogenicity of the bacterial cell wall. PMID:10816508

  17. SUMO Chain Formation Is Required for Response to Replication Arrest in S. pombe

    PubMed Central

    Skilton, Andrew; Ho, Jenny C. Y.; Mercer, Brenda; Outwin, Emily; Watts, Felicity Z.

    2009-01-01

    SUMO is a ubiquitin-like protein that is post-translationally attached to one or more lysine residues on target proteins. Despite having only 18% sequence identity with ubiquitin, SUMO contains the conserved ββαββαβ fold present in ubiquitin. However, SUMO differs from ubiquitin in having an extended N-terminus. In S. pombe the N-terminus of SUMO/Pmt3 is significantly longer than those of SUMO in S. cerevisiae, human and Drosophila. Here we investigate the role of this N-terminal region. We have used two dimensional gel electrophoresis to demonstrate that S. pombe SUMO/Pmt3 is phosphorylated, and that this occurs on serine residues at the extreme N-terminus of the protein. Mutation of these residues (in pmt3-1) results in a dramatic reduction in both the levels of high Mr SUMO-containing species and of total SUMO/Pmt3, indicating that phosphorylation of SUMO/Pmt3 is required for its stability. Despite the significant reduction in high Mr SUMO-containing species, pmt3-1 cells do not display an aberrant cell morphology or sensitivity to genotoxins or stress. Additionally, we demonstrate that two lysine residues in the N-terminus of S. pombe SUMO/Pmt3 (K14 and K30) can act as acceptor sites for SUMO chain formation in vitro. Inability to form SUMO chains results in aberrant cell and nuclear morphologies, including stretched and fragmented chromatin. SUMO chain mutants are sensitive to the DNA synthesis inhibitor, hydroxyurea (HU), but not to other genotoxins, such as UV, MMS or CPT. This implies a role for SUMO chains in the response to replication arrest in S. pombe. PMID:19707600

  18. Cell migration and division in amoeboid-like fission yeast

    PubMed Central

    Flor-Parra, Ignacio; Bernal, Manuel; Zhurinsky, Jacob; Daga, Rafael R.

    2014-01-01

    Summary Yeast cells are non-motile and are encased in a cell wall that supports high internal turgor pressure. The cell wall is also essential for cellular morphogenesis and cell division. Here, we report unexpected morphogenetic changes in a Schizosaccharomyces pombe mutant defective in cell wall biogenesis. These cells form dynamic cytoplasmic protrusions caused by internal turgor pressure and also exhibit amoeboid-like cell migration resulting from repeated protrusive cycles. The cytokinetic ring responsible for cell division in wild-type yeast often fails in these cells; however, they were still able to divide using a ring-independent alternative mechanism relying on extrusion of the cell body through a hole in the cell wall. This mechanism of cell division may resemble an ancestral mode of division in the absence of cytokinetic machinery. Our findings highlight how a single gene change can lead to the emergence of different modes of cell growth, migration and division. PMID:24357230

  19. KRE5 Suppression Induces Cell Wall Stress and Alternative ER Stress Response Required for Maintaining Cell Wall Integrity in Candida glabrata.

    PubMed

    Tanaka, Yutaka; Sasaki, Masato; Ito, Fumie; Aoyama, Toshio; Sato-Okamoto, Michiyo; Takahashi-Nakaguchi, Azusa; Chibana, Hiroji; Shibata, Nobuyuki

    2016-01-01

    The maintenance of cell wall integrity in fungi is required for normal cell growth, division, hyphae formation, and antifungal tolerance. We observed that endoplasmic reticulum stress regulated cell wall integrity in Candida glabrata, which possesses uniquely evolved mechanisms for unfolded protein response mechanisms. Tetracycline-mediated suppression of KRE5, which encodes a predicted UDP-glucose:glycoprotein glucosyltransferase localized in the endoplasmic reticulum, significantly increased cell wall chitin content and decreased cell wall β-1,6-glucan content. KRE5 repression induced endoplasmic reticulum stress-related gene expression and MAP kinase pathway activation, including Slt2p and Hog1p phosphorylation, through the cell wall integrity signaling pathway. Moreover, the calcineurin pathway negatively regulated cell wall integrity, but not the reduction of β-1,6-glucan content. These results indicate that KRE5 is required for maintaining both endoplasmic reticulum homeostasis and cell wall integrity, and that the calcineurin pathway acts as a regulator of chitin-glucan balance in the cell wall and as an alternative mediator of endoplasmic reticulum stress in C. glabrata. PMID:27548283

  20. KRE5 Suppression Induces Cell Wall Stress and Alternative ER Stress Response Required for Maintaining Cell Wall Integrity in Candida glabrata

    PubMed Central

    Sasaki, Masato; Ito, Fumie; Aoyama, Toshio; Sato-Okamoto, Michiyo; Takahashi-Nakaguchi, Azusa; Chibana, Hiroji; Shibata, Nobuyuki

    2016-01-01

    The maintenance of cell wall integrity in fungi is required for normal cell growth, division, hyphae formation, and antifungal tolerance. We observed that endoplasmic reticulum stress regulated cell wall integrity in Candida glabrata, which possesses uniquely evolved mechanisms for unfolded protein response mechanisms. Tetracycline-mediated suppression of KRE5, which encodes a predicted UDP-glucose:glycoprotein glucosyltransferase localized in the endoplasmic reticulum, significantly increased cell wall chitin content and decreased cell wall β-1,6-glucan content. KRE5 repression induced endoplasmic reticulum stress-related gene expression and MAP kinase pathway activation, including Slt2p and Hog1p phosphorylation, through the cell wall integrity signaling pathway. Moreover, the calcineurin pathway negatively regulated cell wall integrity, but not the reduction of β-1,6-glucan content. These results indicate that KRE5 is required for maintaining both endoplasmic reticulum homeostasis and cell wall integrity, and that the calcineurin pathway acts as a regulator of chitin-glucan balance in the cell wall and as an alternative mediator of endoplasmic reticulum stress in C. glabrata. PMID:27548283

  1. Profiling the Hydrolysis of Isolated Grape Berry Skin Cell Walls by Purified Enzymes.

    PubMed

    Zietsman, Anscha J J; Moore, John P; Fangel, Jonatan U; Willats, William G T; Vivier, Melané A

    2015-09-23

    The unraveling of crushed grapes by maceration enzymes during winemaking is difficult to study because of the complex and rather undefined nature of both the substrate and the enzyme preparations. In this study we simplified both the substrate, by using isolated grape skin cell walls, and the enzyme preparations, by using purified enzymes in buffered conditions, to carefully follow the impact of the individual and combined enzymes on the grape skin cell walls. By using cell wall profiling techniques we could monitor the compositional changes in the grape cell wall polymers due to enzyme activity. Extensive enzymatic hydrolysis, achieved with a preparation of pectinases or pectinases combined with cellulase or hemicellulase enzymes, completely removed or drastically reduced levels of pectin polymers, whereas less extensive hydrolysis only opened up the cell wall structure and allowed extraction of polymers from within the cell wall layers. Synergistic enzyme activity was detectable as well as indications of specific cell wall polymer associations.

  2. Schizosaccharomyces pombe encodes a mutated AP endonuclease 1.

    PubMed

    Laerdahl, Jon K; Korvald, Hanne; Nilsen, Line; Dahl-Michelsen, Kristin; Rognes, Torbjørn; Bjørås, Magnar; Alseth, Ingrun

    2011-03-01

    Mutagenic and cytotoxic apurinic/apyrimidinic (AP) sites are among the most frequent lesions in DNA. Repair of AP sites is initiated by AP endonucleases and most organisms possess two or more of these enzymes. Saccharomyces cerevisiae has AP endonuclease 1 (Apn1) as the major enzymatic activity with AP endonuclease 2 (Apn2) being an important backup. Schizosaccharomyces pombe also encodes two potential AP endonucleases, and Apn2 has been found to be the main repair activity, while Apn1 has no, or only a limited role in AP site repair. Here we have identified a new 5' exon (exon 1) in the apn1 gene and show that the inactivity of S. pombe Apn1 is due to a nonsense mutation in the fifth codon of this new exon. Reversion of this mutation restored the AP endonuclease activity of S. pombe Apn1. Interestingly, the apn1 nonsense mutation was only found in laboratory strains derived from L972 h(-) and not in unrelated isolates of S. pombe. Since all S. pombe laboratory strains originate from L972 h(-), it appears that all experiments involving S. pombe have been conducted in an apn1(-) mutant strain with a corresponding DNA repair deficiency. These observations have implications both for future research in S. pombe and for the interpretation of previously conducted epistatis analysis.

  3. Pneumococcal cell wall phosphorylcholine elicits polyclonal antibody secretion in mice.

    PubMed

    Bach, M A; Beckmann, E; Levitt, D

    1984-07-01

    Immunization of mice with phosphorylcholine (PC)-bearing Staphylococcus pneumoniae Type 2, strain 36a (R36a) results in both a PC-specific and a polyclonal increase in splenic plaque-forming cells. The polyclonal increase was observed in all strains tested, including those bearing an X-linked immune defect resulting in an undetectable anti-PC immune response. The magnitude of the polyclonal response is directly related to the amount of bacterial surface PC as detected by enzyme-linked immunosorbent assay. Congenitally athymic (nude) mice mount an anti-PC plaque-forming cell response after R36a immunization but fail to produce a significant polyclonal response. From our results it appears that PC on the cell wall of a bacterium acts both as a polyclonal activator and a specific antigen, stimulating each by different mechanisms.

  4. Cellulose-hemicellulose interaction in wood secondary cell-wall

    NASA Astrophysics Data System (ADS)

    Zhang, Ning; Li, Shi; Xiong, Liming; Hong, Yu; Chen, Youping

    2015-12-01

    The wood cell wall features a tough and relatively rigid fiber reinforced composite structure. It acts as a pressure vessel, offering protection against mechanical stress. Cellulose microfibrils, hemicellulose and amorphous lignin are the three major components of wood. The structure of secondary cell wall could be imagined as the same as reinforced concrete, in which cellulose microfibrils acts as reinforcing steel bar and hemicellulose-lignin matrices act as the concrete. Therefore, the interface between cellulose and hemicellulose/lignin plays a significant role in determine the mechanical behavior of wood secondary cell wall. To this end, we present a molecular dynamics (MD) simulation study attempting to quantify the strength of the interface between cellulose microfibrils and hemicellulose. Since hemicellulose binds with adjacent cellulose microfibrils in various patterns, the atomistic models of hemicellulose-cellulose composites with three typical binding modes, i.e. bridge, loop and random binding modes are constructed. The effect of the shape of hemicellulose chain on the strength of hemicellulose-cellulose composites under shear loadings is investigated. The contact area as well as hydrogen bonds between cellulose and hemicellulose, together with the covalent bonds in backbone of hemicellulose chain are found to be the controlling parameters which determine the strength of the interfaces in the composite system. For the bridge binding model, the effect of shear loading direction on the strength of the cellulose material is also studied. The obtained results suggest that the shear strength of wood-inspired engineering composites can be optimized through maximizing the formations of the contributing hydrogen bonds between cellulose and hemicellulose.

  5. A screen for genes involved in respiration control and longevity in Schizosaccharomyces pombe.

    PubMed

    Roux, Antoine E; Arseneault, Geneviève; Chartrand, Pascal; Ferbeyre, Gerardo; Rokeach, Luis A

    2010-06-01

    We present results showing that glucose signaling has proaging effects in the yeast Schizosaccharomyces pombe. Deletion of the receptor that senses extracellular glucose (Git3) increases the life span of S. pombe, while constitutive activation of the Galpha subunit acting downstream of this receptor (Gpa2) shortens its life span. The latter mutant is also impaired for growth under respiration conditions. We have used this phenotype in a selection strategy to identify genes that when overexpressed can rescue the respiratory defect of constitutively active Galpha subunit mutants. Here, we report an extended version of the work we presented at the IABG meeting and the results of this screen. This strategy allowed us to isolate four genes: psp1(+)/moc1(+), cka1(+), adh1(+), and rpb10(+). Interestingly, the overexpression of these genes was also capable of increasing the chronological life span of wild-type yeast cells.

  6. Ultrastructure of the cell wall of Bacillus polymyxa.

    PubMed

    Nermut, M V; Murray, R G

    1967-06-01

    The macromolecular arrangement on the surface of Bacillus polymyxa was revealed by metal shadowing of whole cells and wall fragments; it consisted of a rectangular array of 70-A globules with a repeating interval of 100 A. The substructure was studied in plan with phosphotungstic acid (pH 6) or uranyl acetate as negative stains of fragments and was studied also in profile with sections of embedded material. Staining of sections of cells fixed with glutaraldehyde showed that layering (approx. 80-A dense, 40-A light, and 120-A dense layers, outermost layer first) could be demonstrated in the cell wall with lead or uranyl acetate, used together or separately. The outer "dense" layer corresponded to the regularly arrayed structure (RS); it was removed by guanidine hydrochloride, sodium lauryl sulfate, cold formamide, and by trypsin. The RS layer (isolated by a hydrogen bond breaking reagent, guanidine hydrochloride) was disrupted by agents such as sodium lauryl sulfate or damaged by 3 m sodium chloride. Qualitative chemical tests, ultraviolet absorption, and removal by trypsin indicated that the structured layer consisted mainly of protein, but exact characterization was not attempted. The globular units making up the layer consisted of a small number of subunits, imperfectly resolved by negative staining. The underlying polysaccharide appeared to be covalently bound to the deepest (probably mucopeptide) layer since it required "hot" formamide for its removal. A survey of species was not made.

  7. A radioimmunoassay for lignin in plant cell walls

    SciTech Connect

    Dawley, R.M.

    1989-01-01

    Lignin detection and determination in herbaceous tissue requires selective, specific assays which are not currently available. A radioimmunoassay (RIA) was developed to study lignin metabolism in these tissues. A {beta}-aryl ether lignin model compound was synthesized, linked to keyhole limpet hemocyanin using a water-soluble carbodiimide, and injected into rabbits. The highest titer of the antiserum obtained was 34 {eta}g/mL of model derivatized BSA. An in vitro system was developed to characterize the RIA. The model compound was linked to amino activated polyacrylamide beads to mimic lignin in the cell walls. {sup 125}I Radiolabelled protein A was used to detect IgG antibody binding. The RIA was shown in the in vitro system to exhibit saturable binding. The amount of antibody bound decreased when the serum was diluted. Immunoelectrophoresis and competitive binding experiments confirmed that both aromatic rings of the lignin model compound had been antigenic. Chlorogenic acid, a phenolic known to be present in plant cells, did not compete for antibody binding. The RIA was used to measure lignin in milled plant samples and barley seedlings. Antiserum binding to wheat cell walls and stressed barley segments was higher than preimmune serum binding. Antibody binding to stressed barley tissue decreased following NaClO{sub 2} delignification. The RIA was found to be less sensitive than expected, so several avenues for improving the method are discussed.

  8. Ultrastructure of the Cell Wall of Bacillus polymyxa

    PubMed Central

    Nermut, M. V.; Murray, R. G. E.

    1967-01-01

    The macromolecular arrangement on the surface of Bacillus polymyxa was revealed by metal shadowing of whole cells and wall fragments; it consisted of a rectangular array of 70-A globules with a repeating interval of 100 A. The substructure was studied in plan with phosphotungstic acid (pH 6) or uranyl acetate as negative stains of fragments and was studied also in profile with sections of embedded material. Staining of sections of cells fixed with glutaraldehyde showed that layering (approx. 80-A dense, 40-A light, and 120-A dense layers, outermost layer first) could be demonstrated in the cell wall with lead or uranyl acetate, used together or separately. The outer “dense” layer corresponded to the regularly arrayed structure (RS); it was removed by guanidine hydrochloride, sodium lauryl sulfate, cold formamide, and by trypsin. The RS layer (isolated by a hydrogen bond breaking reagent, guanidine hydrochloride) was disrupted by agents such as sodium lauryl sulfate or damaged by 3 m sodium chloride. Qualitative chemical tests, ultraviolet absorption, and removal by trypsin indicated that the structured layer consisted mainly of protein, but exact characterization was not attempted. The globular units making up the layer consisted of a small number of subunits, imperfectly resolved by negative staining. The underlying polysaccharide appeared to be covalently bound to the deepest (probably mucopeptide) layer since it required “hot” formamide for its removal. A survey of species was not made. Images PMID:6025307

  9. Structure, cell wall elasticity and polysaccharide properties of living yeast cells, as probed by AFM

    NASA Astrophysics Data System (ADS)

    Alsteens, David; Dupres, Vincent; McEvoy, Kevin; Wildling, Linda; Gruber, Hermann J.; Dufrêne, Yves F.

    2008-09-01

    Although the chemical composition of yeast cell walls is known, the organization, assembly, and interactions of the various macromolecules remain poorly understood. Here, we used in situ atomic force microscopy (AFM) in three different modes to probe the ultrastructure, cell wall elasticity and polymer properties of two brewing yeast strains, i.e. Saccharomyces carlsbergensis and S. cerevisiae. Topographic images of the two strains revealed smooth and homogeneous cell surfaces, and the presence of circular bud scars on dividing cells. Nanomechanical measurements demonstrated that the cell wall elasticity of S. carlsbergensis is homogeneous. By contrast, the bud scar of S. cerevisiae was found to be stiffer than the cell wall, presumably due to the accumulation of chitin. Notably, single molecule force spectroscopy with lectin-modified tips revealed major differences in polysaccharide properties of the two strains. Polysaccharides were clearly more extended on S. cerevisiae, suggesting that not only oligosaccharides, but also polypeptide chains of the mannoproteins were stretched. Consistent with earlier cell surface analyses, these findings may explain the very different aggregation properties of the two organisms. This study demonstrates the power of using multiple complementary AFM modalities for probing the organization and interactions of the various macromolecules of microbial cell walls.

  10. Mass Spectrometry for Characterizing Plant Cell Wall Polysaccharides

    PubMed Central

    Bauer, Stefan

    2012-01-01

    Mass spectrometry is a selective and powerful technique to obtain identification and structural information on compounds present in complex mixtures. Since it requires only small sample amount it is an excellent tool for researchers interested in detecting changes in composition of complex carbohydrates of plants. This mini-review gives an overview of common mass spectrometry techniques applied to the analysis of plant cell wall carbohydrates. It presents examples in which mass spectrometry has been used to elucidate the structure of oligosaccharides derived from hemicelluloses and pectins and illustrates how information on sequence, linkages, branching, and modifications are obtained from characteristic fragmentation patterns. PMID:22645587

  11. Theoretical investigation on breaking plant cell wall by laser

    NASA Astrophysics Data System (ADS)

    Chen, Liang-cai; Wang, Jin-ji; Ma, Peng; Zuo, Du-luo; Wang, Xin-bing; Cheng, Zu-hai

    2011-11-01

    The experiment collected some spinach leaves which were irradiated by pulsed CO2 laser with energy 5.6J, 8.0J and 9.5J respectively. Each of them was soaked in three kinds of solvents (water, ethanol, the mixture of ethanol and petroleum ether) respectively. The experiment shows that the ethanol solution which contains the irradiated leaves turn dark green than the ethanol solution which contains the intact leaves and the color of solution with the leaves irradiated by CO2 laser with 9.5J changes the most significantly. Further, selective excitation on the molecular level of the cell wall were used to explain the phenomenon.

  12. Theoretical investigation on breaking plant cell wall by laser

    NASA Astrophysics Data System (ADS)

    Chen, Liang-cai; Wang, Jin-ji; Ma, Peng; Zuo, Du-luo; Wang, Xin-bing; Cheng, Zu-hai

    2012-03-01

    The experiment collected some spinach leaves which were irradiated by pulsed CO2 laser with energy 5.6J, 8.0J and 9.5J respectively. Each of them was soaked in three kinds of solvents (water, ethanol, the mixture of ethanol and petroleum ether) respectively. The experiment shows that the ethanol solution which contains the irradiated leaves turn dark green than the ethanol solution which contains the intact leaves and the color of solution with the leaves irradiated by CO2 laser with 9.5J changes the most significantly. Further, selective excitation on the molecular level of the cell wall were used to explain the phenomenon.

  13. Immunofluorescence Microscopy of Schizosaccharomyces pombe Using Chemical Fixation.

    PubMed

    Hagan, Iain M

    2016-01-01

    Establishing the subcellular distribution of molecules of interest and the dynamics of their spatial control underpins all areas of cell and developmental biology. Although the ability to monitor the distribution of fluorescent fusion proteins has revolutionized cell and developmental biology, indirect immunofluorescence microscopy of fixed samples remains an essential complement to this approach. Immunofluorescence is often a more appropriate approach for the study of subcellular architecture. It avoids potential artifacts caused by studying fusion proteins, which might show altered function under stressful imaging conditions. Furthermore, the quantitative analysis of multiple cells in an unperturbed population by immunofluorescence invariably provides a more accurate assessment of the spatial and temporal control of a particular process than does the analysis of individual cells that is the hallmark of live-cell imaging. Parallel studies of living and fixed cells often provide complementary data sets, both of which can be considered necessary for a comprehensive understanding of molecular function. This protocol provides a method for the visualization of the Schizosaccharomyces pombe microtubule cytoskeleton by indirect immunofluorescence microscopy following chemical fixation with formaldehyde and glutaraldehyde. It includes discussion of common modifications used to monitor the distribution of other fission yeast antigens and forms a basis from which to develop protocols to localize new molecules of interest. PMID:27371599

  14. In situ microscopic observation of chitin and fungal cells with chitinous cell walls in hydrothermal conditions.

    PubMed

    Deguchi, Shigeru; Tsujii, Kaoru; Horikoshi, Koki

    2015-07-07

    Recent findings of intact chitin in fossil records suggest surprisingly high recalcitrance of this biopolymer during hydrothermal treatments. We also know in the experience of everyday life that mushroom, cells of which have chitinous cell walls, do not fall apart however long they are simmered. We used in situ optical microscopy to examine chitin and fungal cells with chitinous cell walls during hydrothermal treatments, and obtained direct evidence that they remained undegraded at temperatures well over 200 °C. The results show very hot and compressed water is needed to make mushrooms mushy.

  15. In situ microscopic observation of chitin and fungal cells with chitinous cell walls in hydrothermal conditions

    PubMed Central

    Deguchi, Shigeru; Tsujii, Kaoru; Horikoshi, Koki

    2015-01-01

    Recent findings of intact chitin in fossil records suggest surprisingly high recalcitrance of this biopolymer during hydrothermal treatments. We also know in the experience of everyday life that mushroom, cells of which have chitinous cell walls, do not fall apart however long they are simmered. We used in situ optical microscopy to examine chitin and fungal cells with chitinous cell walls during hydrothermal treatments, and obtained direct evidence that they remained undegraded at temperatures well over 200 °C. The results show very hot and compressed water is needed to make mushrooms mushy. PMID:26148792

  16. A Geographically Diverse Collection of Schizosaccharomyces pombe Isolates Shows Limited Phenotypic Variation but Extensive Karyotypic Diversity

    PubMed Central

    Brown, William R. A.; Liti, Gianni; Rosa, Carlos; James, Steve; Roberts, Ian; Robert, Vincent; Jolly, Neil; Tang, Wen; Baumann, Peter; Green, Carter; Schlegel, Kristina; Young, Jonathan; Hirchaud, Fabienne; Leek, Spencer; Thomas, Geraint; Blomberg, Anders; Warringer, Jonas

    2011-01-01

    The fission yeast Schizosaccharomyces pombe has been widely used to study eukaryotic cell biology, but almost all of this work has used derivatives of a single strain. We have studied 81 independent natural isolates and 3 designated laboratory strains of Schizosaccharomyces pombe. Schizosaccharomyces pombe varies significantly in size but shows only limited variation in proliferation in different environments compared with Saccharomyces cerevisiae. Nucleotide diversity, π, at a near neutral site, the central core of the centromere of chromosome II is approximately 0.7%. Approximately 20% of the isolates showed karyotypic rearrangements as detected by pulsed field gel electrophoresis and filter hybridization analysis. One translocation, found in 6 different isolates, including the type strain, has a geographically widespread distribution and a unique haplotype and may be a marker of an incipient speciation event. All of the other translocations are unique. Exploitation of this karyotypic diversity may cast new light on both the biology of telomeres and centromeres and on isolating mechanisms in single-celled eukaryotes. PMID:22384373

  17. A simplified vector system for visualization of localized RNAs in Schizosaccharomyces pombe.

    PubMed

    Takeuchi-Andoh, Tomoko; Ohba, Sayaka; Shinoda, Yu; Fuchita, Ayako; Hayashi, Sachiko; Nishiyoshi, Emi; Terouchi, Nobuyuki; Tani, Tokio

    2016-07-01

    RNA localization is an important event that is essential for the polarization and differentiation of a cell. Although several methods are currently used to detect localized RNAs, a simplified detection system has not yet been developed for Schizosaccharomyces pombe. In the present study, we describe a new vector system for the visualization of localized RNAs in S. pombe using a U1A-tag-GFP system. A pREP1-U1A-tag vector plasmid to express U1A-tagged RNA and a pREP2-U1AGFP plasmid to produce a U1A-GFP fusion protein were constructed for this system. Since the U1A-GFP protein binds U1A-tagged RNA, fluorescence is observed at the location of U1A-tagged RNA in cells expressing both of these. The nucleolar localization of U3 snoRNA was successfully detected using this system, and a novel RNA localized at the DNA region of the nucleus was found by screening localized RNAs. This system will accelerate the study of localized RNAs in S. pombe.

  18. A simplified vector system for visualization of localized RNAs in Schizosaccharomyces pombe.

    PubMed

    Takeuchi-Andoh, Tomoko; Ohba, Sayaka; Shinoda, Yu; Fuchita, Ayako; Hayashi, Sachiko; Nishiyoshi, Emi; Terouchi, Nobuyuki; Tani, Tokio

    2016-07-01

    RNA localization is an important event that is essential for the polarization and differentiation of a cell. Although several methods are currently used to detect localized RNAs, a simplified detection system has not yet been developed for Schizosaccharomyces pombe. In the present study, we describe a new vector system for the visualization of localized RNAs in S. pombe using a U1A-tag-GFP system. A pREP1-U1A-tag vector plasmid to express U1A-tagged RNA and a pREP2-U1AGFP plasmid to produce a U1A-GFP fusion protein were constructed for this system. Since the U1A-GFP protein binds U1A-tagged RNA, fluorescence is observed at the location of U1A-tagged RNA in cells expressing both of these. The nucleolar localization of U3 snoRNA was successfully detected using this system, and a novel RNA localized at the DNA region of the nucleus was found by screening localized RNAs. This system will accelerate the study of localized RNAs in S. pombe. PMID:26979837

  19. De novo biosynthesis of vanillin in fission yeast (Schizosaccharomyces pombe) and baker's yeast (Saccharomyces cerevisiae).

    PubMed

    Hansen, Esben H; Møller, Birger Lindberg; Kock, Gertrud R; Bünner, Camilla M; Kristensen, Charlotte; Jensen, Ole R; Okkels, Finn T; Olsen, Carl E; Motawia, Mohammed S; Hansen, Jørgen

    2009-05-01

    Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast, Saccharomyces cerevisiae. Productivities were 65 and 45 mg/liter, after introduction of three and four heterologous genes, respectively. The engineered pathways involve incorporation of 3-dehydroshikimate dehydratase from the dung mold Podospora pauciseta, an aromatic carboxylic acid reductase (ACAR) from a bacterium of the Nocardia genus, and an O-methyltransferase from Homo sapiens. In S. cerevisiae, the ACAR enzyme required activation by phosphopantetheinylation, and this was achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol was achieved by knockout of the host alcohol dehydrogenase ADH6. In S. pombe, the biosynthesis was further improved by introduction of an Arabidopsis thaliana family 1 UDP-glycosyltransferase, converting vanillin into vanillin beta-D-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts. These de novo pathways represent the first examples of one-cell microbial generation of these valuable compounds from glucose. S. pombe yeast has not previously been metabolically engineered to produce any valuable, industrially scalable, white biotech commodity.

  20. Heterogeneity in the chemistry, structure and function of plant cell walls.

    PubMed

    Burton, Rachel A; Gidley, Michael J; Fincher, Geoffrey B

    2010-10-01

    Higher plants resist the forces of gravity and powerful lateral forces through the cumulative strength of the walls that surround individual cells. These walls consist mainly of cellulose, noncellulosic polysaccharides and lignin, in proportions that depend upon the specific functions of the cell and its stage of development. Spatially and temporally controlled heterogeneity in the physicochemical properties of wall polysaccharides is observed at the tissue and individual cell levels, and emerging in situ technologies are providing evidence that this heterogeneity also occurs across a single cell wall. We consider the origins of cell wall heterogeneity and identify contributing factors that are inherent in the molecular mechanisms of polysaccharide biosynthesis and are crucial for the changing biological functions of the wall during growth and development. We propose several key questions to be addressed in cell wall biology, together with an alternative two-phase model for the assembly of noncellulosic polysaccharides in plants.

  1. Binding of /sup 18/F by cell membranes and cell walls of Streptococcus mutans

    SciTech Connect

    Yotis, W.W.; Zeb, M.; McNulty, J.; Kirchner, F.; Reilly, C.; Glendenin, L.

    1983-07-01

    The binding of /sup 18/F to isolated cell membranes and cell walls of Streptococcus mutans GS-5 or other bacteria was assayed. The attachment of /sup 18/F to these cell envelopes proceeded slowly and reached equilibrium within 60 min. /sup 18/F binding was stimulated by Ca/sup 2 +/ (1 mM). The binding of /sup 18/F to cellular components was dependent upon the pH, as well as the amount of /sup 18/F and dose of the binder employed. The binding of /sup 18/F by cell walls prepared from fluoride-sensitive and fluoride-resistant cells of S. salivarius and S. mutans did not differ significantly. The pretreatment of cell walls or cell membranes for 60 min at 30 degrees C with 1 mg of RNase, DNase, or trypsin per ml did not influence the binding of /sup 18/F by the walls and membranes of S. mutans GS-5. However, prior exposure of cell membranes to sodium dodecyl sulfate caused a significant reduction in the number of /sup 18/F atoms bound by the membranes. In saturated assay systems, cell membranes of S. mutans GS-5 bound 10(15) to 10(16) atoms of /sup 18/F per mg (dry weight), whereas cell walls from S. mutans GS-5, FA-1, and HS-6 or Actinomyces viscosus T14V and T14AV bound 10(12) to 10(13) atoms of /sup 18/F per mg (dry weight). /sup 18/F in this quantity (10(12) to 10(13) atoms) cannot be detected with the fluoride electrode. The data provide, for the first time, a demonstration of /sup 18/F binding by cell membranes and walls of oral flora.

  2. Daughter cell separation is controlled by cytokinetic ring-activated cell wall hydrolysis.

    PubMed

    Uehara, Tsuyoshi; Parzych, Katherine R; Dinh, Thuy; Bernhardt, Thomas G

    2010-04-21

    During bacterial cytokinesis, hydrolytic enzymes are used to split wall material shared by adjacent daughter cells to promote their separation. Precise control over these enzymes is critical to prevent breaches in wall integrity that can cause cell lysis. How these potentially lethal hydrolases are regulated has remained unknown. Here, we investigate the regulation of cell wall turnover at the Escherichia coli division site. We show that two components of the division machinery with LytM domains (EnvC and NlpD) are direct regulators of the cell wall hydrolases (amidases) responsible for cell separation (AmiA, AmiB and AmiC). Using in vitro cell wall cleavage assays, we show that EnvC activates AmiA and AmiB, whereas NlpD activates AmiC. Consistent with these findings, we show that an unregulated EnvC mutant requires functional AmiA or AmiB but not AmiC to induce cell lysis, and that the loss of NlpD phenocopies an AmiC(-) defect. Overall, our results suggest that cellular amidase activity is regulated spatially and temporally by coupling their activation to the assembly of the cytokinetic ring.

  3. The MAP kinase Pmk1 and protein kinase A are required for rotenone resistance in the fission yeast, Schizosaccharomyces pombe

    SciTech Connect

    Wang, Yiwei; Gulis, Galina; Buckner, Scott; Johnson, P. Connor; Sullivan, Daniel; Busenlehner, Laura; Marcus, Stevan

    2010-08-20

    Research highlights: {yields} Rotenone induces generation of ROS and mitochondrial fragmentation in fission yeast. {yields} The MAPK Pmk1 and PKA are required for rotenone resistance in fission yeast. {yields} Pmk1 and PKA are required for ROS clearance in rotenone treated fission yeast cells. {yields} PKA plays a role in ROS clearance under normal growth conditions in fission yeast. -- Abstract: Rotenone is a widely used pesticide that induces Parkinson's disease-like symptoms in rats and death of dopaminergic neurons in culture. Although rotenone is a potent inhibitor of complex I of the mitochondrial electron transport chain, it can induce death of dopaminergic neurons independently of complex I inhibition. Here we describe effects of rotenone in the fission yeast, Schizosaccharomyces pombe, which lacks complex I and carries out rotenone-insensitive cellular respiration. We show that rotenone induces generation of reactive oxygen species (ROS) as well as fragmentation of mitochondrial networks in treated S. pombe cells. While rotenone is only modestly inhibitory to growth of wild type S. pombe cells, it is strongly inhibitory to growth of mutants lacking the ERK-type MAP kinase, Pmk1, or protein kinase A (PKA). In contrast, cells lacking the p38 MAP kinase, Spc1, exhibit modest resistance to rotenone. Consistent with these findings, we provide evidence that Pmk1 and PKA, but not Spc1, are required for clearance of ROS in rotenone treated S. pombe cells. Our results demonstrate the usefulness of S. pombe for elucidating complex I-independent molecular targets of rotenone as well as mechanisms conferring resistance to the toxin.

  4. The cell-wall glycoproteins of the green alga Scenedesmus obliquus. The predominant cell-wall polypeptide of Scenedesmus obliquus is related to the cell-wall glycoprotein gp3 of Chlamydomonas reinhardtii.

    PubMed

    Voigt, Jürgen; Stolarczyk, Adam; Zych, Maria; Malec, Przemysław; Burczyk, Jan

    2014-02-01

    The green alga Scenedesmus obliquus contains a multilayered cell wall, ultrastructurally similar to that of Chlamydomonas reinhardtii, although its proportion of hydroxyproline is considerably lower. Therefore, we have investigated the polypeptide composition of the insoluble and the chaotrope-soluble wall fractions of S. obliquus. The polypeptide pattern of the chaotrope-soluble wall fraction was strongly modified by chemical deglycosylation with anhydrous hydrogen fluoride (HF) in pyridine indicating that most of these polypeptides are glycosylated. Polypeptide constituents of the chaotrope-soluble cell-wall fraction with apparent molecular masses of 240, 270, 265, and 135 kDa cross-reacted with a polyclonal antibody raised against the 100 kDa deglycosylation product of the C. reinhardtii cell-wall glycoprotein GP3B. Chemical deglycosylation of the chaotrope-soluble wall fraction resulted in a 135 kDa major polypeptide and a 106 kDa minor component reacting with the same antibody. This antibody recognized specific peptide epitopes of GP3B. When the insoluble wall fraction of S. obliquus was treated with anhydrous HF/pyridine, three polypeptides with apparent molecular masses of 144, 135, and 65 kDa were solubilized, which also occured in the deglycosylated chaotrope-soluble wall fraction. These findings indicate that theses glycoproteins are cross-linked to the insoluble wall fraction via HF-sensitive bonds.

  5. The cell-wall glycoproteins of the green alga Scenedesmus obliquus. The predominant cell-wall polypeptide of Scenedesmus obliquus is related to the cell-wall glycoprotein gp3 of Chlamydomonas reinhardtii.

    PubMed

    Voigt, Jürgen; Stolarczyk, Adam; Zych, Maria; Malec, Przemysław; Burczyk, Jan

    2014-02-01

    The green alga Scenedesmus obliquus contains a multilayered cell wall, ultrastructurally similar to that of Chlamydomonas reinhardtii, although its proportion of hydroxyproline is considerably lower. Therefore, we have investigated the polypeptide composition of the insoluble and the chaotrope-soluble wall fractions of S. obliquus. The polypeptide pattern of the chaotrope-soluble wall fraction was strongly modified by chemical deglycosylation with anhydrous hydrogen fluoride (HF) in pyridine indicating that most of these polypeptides are glycosylated. Polypeptide constituents of the chaotrope-soluble cell-wall fraction with apparent molecular masses of 240, 270, 265, and 135 kDa cross-reacted with a polyclonal antibody raised against the 100 kDa deglycosylation product of the C. reinhardtii cell-wall glycoprotein GP3B. Chemical deglycosylation of the chaotrope-soluble wall fraction resulted in a 135 kDa major polypeptide and a 106 kDa minor component reacting with the same antibody. This antibody recognized specific peptide epitopes of GP3B. When the insoluble wall fraction of S. obliquus was treated with anhydrous HF/pyridine, three polypeptides with apparent molecular masses of 144, 135, and 65 kDa were solubilized, which also occured in the deglycosylated chaotrope-soluble wall fraction. These findings indicate that theses glycoproteins are cross-linked to the insoluble wall fraction via HF-sensitive bonds. PMID:24388513

  6. Penium margaritaceum: A Unicellular Model Organism for Studying Plant Cell Wall Architecture and Dynamics

    PubMed Central

    Domozych, David S.

    2014-01-01

    Penium margaritaceum is a new and valuable unicellular model organism for studying plant cell wall structure and developmental dynamics. This charophyte has a cell wall composition remarkably similar to the primary cell wall of many higher plants and clearly-defined inclusive zones containing specific polymers. Penium has a simple cylindrical phenotype with a distinct region of focused wall synthesis. Specific polymers, particularly pectins, can be identified using monoclonal antibodies raised against polymers of higher plant cell walls. Immunofluorescence-based labeling is easily performed using live cells that subsequently can be returned to culture and monitored. This feature allows for rapid assessment of wall expansion rates and identification of multiple polymer types in the wall microarchitecture during the cell cycle. Cryofixation by means of spray freezing provides excellent transmission electron microscopy imaging of the cell, including its elaborate endomembrane and cytoskeletal systems, both integral to cell wall development. Penium’s fast growth rate allows for convenient microarray screening of various agents that alter wall biosynthesis and metabolism. Finally, recent successful development of transformed cell lines has allowed for non-invasive imaging of proteins in cells and for RNAi reverse genetics that can be used for cell wall biosynthesis studies. PMID:27135519

  7. Properties of lead deposits in cell walls of radish (Raphanus sativus) roots.

    PubMed

    Inoue, Hiroshi; Fukuoka, Daisuke; Tatai, Yuri; Kamachi, Hiroyuki; Hayatsu, Manabu; Ono, Manami; Suzuki, Suechika

    2013-01-01

    Various mechanisms are involved in detoxification of heavy metals such as lead (Pb) in plant cells. Most of the Pb taken up by plants accumulates in their roots. However, the detailed properties of Pb complexes in roots remain unclear. We have investigated the properties of Pb deposits in root cell walls of radish (Raphanus sativus L.) seedlings grown on glass beads bed containing Pb pellets, which are the source of Pb-contamination in shooting range soils. Pb deposits were tightly bound to cell walls. Cell wall fragments containing about 50,000 ppm Pb were prepared from the roots. After extracting Pb from the cell wall fragments using HCl, Pb ions were recombined with the Pb-extracted cell wall fragments in a solution containing Pb acetate. When the cell wall fragments were treated with pectinase (E.C. 3.2.1.15) and were chemically modified with 1-ethyl-3-dimethylamino-propylcarboimide, the Pb-rebinding ability of the treated cell wall fragments decreased. When acid-treated cell wall fragments were incubated in a solution containing Pb(2+) and excess amounts of a chelating agent, Pb recombined with the cell wall fragments were measured to estimate the affinity between Pb(2+) and the cell wall fragments. Our data show that Pb(2+) binds to carboxyl groups of cell walls. The source of the carboxyl groups is suggested to be pectic compounds. A stability constant of the Pb-cell wall complex was estimated to be about 10(8). The role of root cell walls in the mechanism underlying heavy metal tolerance was discussed.

  8. Biosynthesis of non-cellulosic polysaccharides of plant cell walls.

    PubMed

    Dhugga, Kanwarpal S

    2012-02-01

    Enzymes that make the polymer backbones of plant cell wall polysaccharides have proven to be recalcitrant to biochemical purification. Availability of mutational genetics and genomic tools paved the way for rapid progress in identifying genes encoding various cell wall glycan synthases. Mutational genetics, the primary tool used in unraveling cellulose biosynthesis, was ineffective in assigning function to any of the hemicellulosic, polymerizing glycan synthases. A combination of comparative genomics and functional expression in a heterologous system allowed identification of various cellulose synthase-like (Csl) sequences as being involved in the formation of β-1,4-mannan, β-1,4-glucan, and mixed-linked glucan. A number of xylose-deficient mutants have led to a variety of genes, none of which thus far possesses the motifs known to be conserved among polymerizing β-glycan synthases. Except for xylan synthase, which appears to be an agglomerate of proteins just like cellulose synthase, Golgi glycan synthases already identified suggest that the catalytic polypeptide by itself is sufficient for enzyme activity, most likely as a homodimer. Several of the Csl genes remain to be assigned a function. The possibility of the involvement of various Csl genes in making more than one product remains.

  9. Chemical Profiling of the Plant Cell Wall through Raman Microspectroscopy

    SciTech Connect

    Han, Ju; Singh, Seema; Sun, Lan; Simmons, Blake; Auer, Manfred; Parvin, Bahram

    2010-03-02

    This paper presents a computational framework for chemical pro.ling of the plant cell wall through the Raman spectroscopy. The system enables query of known spectral signatures and clustering of spectral data based on intrinsic properties. As a result, presence and relative concentration of speci.c chemical bonds can be quanti.ed. The primary contribution of this paper is in representation of raman pro.le in terms of .uorescence background and multiscale peak detection at each grid point (voxel). Such a representation allows ef.cient spatial segmentation based on the coupling between high-level salient properties and low-level symbolic representation at each voxel. The high-level salient properties refer to preferred peaks and their attributes for the entire image. The low-level symbolic representations are based on .uorescence background, spectral peak locations, and their attributes. We present results on a corn stover tissue section that is imaged through Raman microscopy, and the results are consistent with the literature. In addition, automatic clustering indicates several distinct layers of the cell walls with different spectral signatures.

  10. The toughness of secondary cell wall and woody tissue

    PubMed Central

    Lucas, P. W.; Tan, H. T. W.; Cheng, P. Y.

    1997-01-01

    The 'across grain' toughness of 51 woods has been determined on thin wet sections using scissors. The moisture content of sections and the varying sharpness of the scissor blades had little effect on the results. In thin sections (less than 0.6mm), toughness rose linearly with section thickness. The intercept toughness at zero thickness, estimated from regression analysis, was proportional to relative density, consistent with values reported for non-woody plant tissues. Extrapolation of the intercept toughness of these woods and other plant tissues/materials to a relative density of 1.0 predicted a toughness of 3.45kJ m-2 , which we identify with the intrinsic toughness of the cell wall. This quantity appears to predict published results from KIC tests on woods and is related to the propensity for crack deflection. The slope of the relationship between section thickness and toughness, describing the work of plastic buckling of cells, was not proportional to relative density, the lightest (balsa) and heaviest (lignum vitae) woods fracturing with less plastic work than predicted. The size of the plastic zone around the crack tip was estimated to be 0.5mm in size. From this, the hypothetical overall toughness of a thick (greater than 1 mm) block of solid cell wall material was calculated as 39.35 kJ m-2, due to both cell wall resistance (10 per cent) and the plastic buckling of cells (90 per cent). This value successfully predicts the toughness of most commercial woods (of relative densities between 0.2 and 0.8) from 'work area' tests in tension and bending. Though density was the most important factor, both fibre width/fibre length (in hardwoods) and lignin/cellulose ratios were negatively correlated with the work of plastic buckling, after correcting for density. At low densities the work of plastic buckling in the longitudinal radial (LR) direction exceeded that in longitudinal tangential (LT), but the reverse was true for relative densities above 0.25. This could

  11. Molecular Mechanisms for Vascular Development and Secondary Cell Wall Formation.

    PubMed

    Yang, Jung Hyun; Wang, Huanzhong

    2016-01-01

    Vascular tissues are important for transporting water and nutrients throughout the plant and as physical support of upright growth. The primary constituents of vascular tissues, xylem, and phloem, are derived from the meristematic vascular procambium and cambium. Xylem cells develop secondary cell walls (SCWs) that form the largest part of plant lignocellulosic biomass that serve as a renewable feedstock for biofuel production. For the last decade, research on vascular development and SCW biosynthesis has seen rapid progress due to the importance of these processes to plant biology and to the biofuel industry. Plant hormones, transcriptional regulators and peptide signaling regulate procambium/cambium proliferation, vascular patterning, and xylem differentiation. Transcriptional regulatory pathways play a pivot role in SCW biosynthesis. Although most of these discoveries are derived from research in Arabidopsis, many genes have shown conserved functions in biofuel feedstock species. Here, we review the recent advances in our understanding of vascular development and SCW formation and discuss potential biotechnological uses. PMID:27047525

  12. Cell wall proteins of Sporothrix schenckii as immunoprotective agents.

    PubMed

    Alba-Fierro, Carlos A; Pérez-Torres, Armando; López-Romero, Everardo; Cuéllar-Cruz, Mayra; Ruiz-Baca, Estela

    2014-01-01

    Sporothrix schenckii is the etiological agent of sporotrichosis, an endemic subcutaneous mycosis in Latin America. Cell wall (CW) proteins located on the cell surface are inducers of cellular and humoral immune responses, potential candidates for diagnosis purposes and to generate vaccines to prevent fungal infections. This mini-review emphasizes the potential use of S. schenckii CW proteins as protective and therapeutic immune response inducers against sporotrichosis. A number of pathogenic fungi display CW components that have been characterized as inducers of protective cellular and humoral immune responses against the whole pathogen from which they were originally purified. The isolation and characterization of immunodominant protein components of the CW of S. schenckii have become relevant because of their potential in the development of protective and therapeutic immune responses against sporotrichosis. This manuscript is part of the series of works presented at the "V International Workshop: Molecular genetic approaches to the study of human pathogenic fungi" (Oaxaca, Mexico, 2012).

  13. Rapid regulatory control of plant cell expansion and wall relaxation

    SciTech Connect

    Cosgrove, D.J.

    1991-08-14

    The aim of this project is to elucidate the biophysical and cellular mechanisms that control plant cell expansion. At present we are attempting to characterize the kinetics of the system(s) responsible for regulatory and compensatory behavior of growing cells and tissues. This work is significantly because it indicates that biochemical loosening and biophysical stress relaxation of the wall are part of a feedback loop controlling growth. This report briefly summarizes the efforts and results of the past 12 months. In large part, we have been trying to analyze the nature of growth rate noise,'' i.e. spontaneous and often erratic variations in growth rate. We are obtaining evidence that such noise'' is not random, but rather reveals an underlying growth mechanism with complex dynamics.

  14. Molecular Mechanisms for Vascular Development and Secondary Cell Wall Formation

    PubMed Central

    Yang, Jung Hyun; Wang, Huanzhong

    2016-01-01

    Vascular tissues are important for transporting water and nutrients throughout the plant and as physical support of upright growth. The primary constituents of vascular tissues, xylem, and phloem, are derived from the meristematic vascular procambium and cambium. Xylem cells develop secondary cell walls (SCWs) that form the largest part of plant lignocellulosic biomass that serve as a renewable feedstock for biofuel production. For the last decade, research on vascular development and SCW biosynthesis has seen rapid progress due to the importance of these processes to plant biology and to the biofuel industry. Plant hormones, transcriptional regulators and peptide signaling regulate procambium/cambium proliferation, vascular patterning, and xylem differentiation. Transcriptional regulatory pathways play a pivot role in SCW biosynthesis. Although most of these discoveries are derived from research in Arabidopsis, many genes have shown conserved functions in biofuel feedstock species. Here, we review the recent advances in our understanding of vascular development and SCW formation and discuss potential biotechnological uses. PMID:27047525

  15. Sugarcane cell wall structure and lignin distribution investigated by confocal and electron microscopy.

    PubMed

    Sant'Anna, Celso; Costa, Lilian T; Abud, Yuri; Biancatto, Lucas; Miguens, Flávio Costa; de Souza, Wanderley

    2013-08-01

    Lignocellulosic plant cell wall is considered a potential source for second generation biofuels. The plant cell wall is a highly complex structure mainly composed of cellulose, hemicelluloses, and lignin that form a network of crosslinked fibers. The structural organization of the sugarcane cell wall has not been previously analyzed in detail, and this analysis is a prerequisite for further studies on the recalcitrance and deconstruction of its biomass. In this work, cellulose and lignin localization were investigated by confocal laser scanning microscopy. In addition, the internode sugarcane cell wall structural organization was analyzed by electron microscopy. Internode stem anatomy showed a typical monocot structure consisting of epidermis, hypoderm, and vascular bundles scattered throughout ground parenchyma tissue and surrounded by sclerenchyma fibers. Confocal images of safranin labeled sugarcane showed that lignin distribution was predominant in the vessel elements, cell wall corners (CC), and middle lamella (ML), while cellulose-rich cell walls were randomly distributed in the ML and organized in the other cell wall layers. KMnO4 cytochemistry revealed that lignin was predominantly distributed in secondary cell walls, ML and CC. Cell wall sublayers (S1, S2, and S3) were identified and measured by transmission electron microscopy. Our results provide insights that may help further understanding of sugarcane cell wall organization, which is crucial for the research and technology of plant-based biofuel production. PMID:23733560

  16. Reduction in Young`s modulus of aluminum foams due to cell wall curvature and corrugation

    SciTech Connect

    Sanders, W.; Gibson, L.J.

    1998-12-31

    Measurements of the Young`s modulus and compressive strength of several closed-cell aluminum foams indicate that they are lower than expected from models for foam behavior. Microstructural characterization has revealed that there are a number of defects in the cell structure which may contribute to the reduction in mechanical properties. These include: cell wall curvature, cell wall corrugations, density variations and non-equiaxed cell shape. Finite element analysis of a closed-cell tetrakaidecahedral unit cell with idealized curved or corrugated cell walls indicates that these two types of defects can reduce the Young`s modulus and compressive strength by up to 70%. In this paper the authors report the results of measurements of the curvature of the cell walls and of the amplitude and frequency of corrugations in the cell walls and use simple bounds to estimate the reduction in modulus that they are responsible for.

  17. The connection of cytoskeletal network with plasma membrane and the cell wall

    PubMed Central

    Liu, Zengyu; Persson, Staffan; Zhang, Yi

    2015-01-01

    The cell wall provides external support of the plant cells, while the cytoskeletons including the microtubules and the actin filaments constitute an internal framework. The cytoskeletons contribute to the cell wall biosynthesis by spatially and temporarily regulating the transportation and deposition of cell wall components. This tight control is achieved by the dynamic behavior of the cytoskeletons, but also through the tethering of these structures to the plasma membrane. This tethering may also extend beyond the plasma membrane and impact on the cell wall, possibly in the form of a feedback loop. In this review, we discuss the linking components between the cytoskeletons and the plasma membrane, and/or the cell wall. We also discuss the prospective roles of these components in cell wall biosynthesis and modifications, and aim to provide a platform for further studies in this field. PMID:25693826

  18. Adaptations of higher plant cell walls to water loss: drought vs desiccation.

    PubMed

    Moore, John P; Vicré-Gibouin, Mäite; Farrant, Jill M; Driouich, Azeddine

    2008-10-01

    Water-deficit stress poses unique challenges to plant cells dependent on a hydrostatic skeleton and a polysaccharide-rich cell wall for growth and development. How the plant cell wall is adapted to loss of water is of interest in developing a general understanding of water stress tolerance in plants and of relevance in strategies related to crop improvement. Drought tolerance involves adaptations to growth under reduced water potential and the concomitant restructuring of the cell wall that allow growth processes to occur at lower water contents. Desiccation tolerance, by contrast, is the evolution of cell walls that are capable of losing the majority of cellular water without suffering permanent and irreversible damage to cell wall structure and polymer organization. This minireview highlights common features and differences between these two water-deficit responses observed in plants, emphasizing the role of the cell wall, while suggesting future research avenues that could benefit fundamental understanding in this area.

  19. The connection of cytoskeletal network with plasma membrane and the cell wall.

    PubMed

    Liu, Zengyu; Persson, Staffan; Zhang, Yi

    2015-04-01

    The cell wall provides external support of the plant cells, while the cytoskeletons including the microtubules and the actin filaments constitute an internal framework. The cytoskeletons contribute to the cell wall biosynthesis by spatially and temporarily regulating the transportation and deposition of cell wall components. This tight control is achieved by the dynamic behavior of the cytoskeletons, but also through the tethering of these structures to the plasma membrane. This tethering may also extend beyond the plasma membrane and impact on the cell wall, possibly in the form of a feedback loop. In this review, we discuss the linking components between the cytoskeletons and the plasma membrane, and/or the cell wall. We also discuss the prospective roles of these components in cell wall biosynthesis and modifications, and aim to provide a platform for further studies in this field.

  20. A cell wall damage response mediated by a sensor kinase/response regulator pair enables beta-lactam tolerance.

    PubMed

    Dörr, Tobias; Alvarez, Laura; Delgado, Fernanda; Davis, Brigid M; Cava, Felipe; Waldor, Matthew K

    2016-01-12

    The bacterial cell wall is critical for maintenance of cell shape and survival. Following exposure to antibiotics that target enzymes required for cell wall synthesis, bacteria typically lyse. Although several cell envelope stress response systems have been well described, there is little knowledge of systems that modulate cell wall synthesis in response to cell wall damage, particularly in Gram-negative bacteria. Here we describe WigK/WigR, a histidine kinase/response regulator pair that enables Vibrio cholerae, the cholera pathogen, to survive exposure to antibiotics targeting cell wall synthesis in vitro and during infection. Unlike wild-type V. cholerae, mutants lacking wigR fail to recover following exposure to cell-wall-acting antibiotics, and they exhibit a drastically increased cell diameter in the absence of such antibiotics. Conversely, overexpression of wigR leads to cell slimming. Overexpression of activated WigR also results in increased expression of the full set of cell wall synthesis genes and to elevated cell wall content. WigKR-dependent expression of cell wall synthesis genes is induced by various cell-wall-acting antibiotics as well as by overexpression of an endogenous cell wall hydrolase. Thus, WigKR appears to monitor cell wall integrity and to enhance the capacity for increased cell wall production in response to damage. Taken together, these findings implicate WigKR as a regulator of cell wall synthesis that controls cell wall homeostasis in response to antibiotics and likely during normal growth as well.

  1. Discovery of Novel Cell Wall-Active Compounds Using PywaC, a Sensitive Reporter of Cell Wall Stress, in the Model Gram-Positive Bacterium Bacillus subtilis

    PubMed Central

    Czarny, T. L.; Perri, A. L.; French, S.

    2014-01-01

    The emergence of antibiotic resistance in recent years has radically reduced the clinical efficacy of many antibacterial treatments and now poses a significant threat to public health. One of the earliest studied well-validated targets for antimicrobial discovery is the bacterial cell wall. The essential nature of this pathway, its conservation among bacterial pathogens, and its absence in human biology have made cell wall synthesis an attractive pathway for new antibiotic drug discovery. Herein, we describe a highly sensitive screening methodology for identifying chemical agents that perturb cell wall synthesis, using the model of the Gram-positive bacterium Bacillus subtilis. We report on a cell-based pilot screen of 26,000 small molecules to look for cell wall-active chemicals in real time using an autonomous luminescence gene cluster driven by the promoter of ywaC, which encodes a guanosine tetra(penta)phosphate synthetase that is expressed under cell wall stress. The promoter-reporter system was generally much more sensitive than growth inhibition testing and responded almost exclusively to cell wall-active antibiotics. Follow-up testing of the compounds from the pilot screen with secondary assays to verify the mechanism of action led to the discovery of 9 novel cell wall-active compounds. PMID:24687489

  2. Characterization of nuclear pore complex components in fission yeast Schizosaccharomyces pombe.

    PubMed

    Asakawa, Haruhiko; Yang, Hui-Ju; Yamamoto, Takaharu G; Ohtsuki, Chizuru; Chikashige, Yuji; Sakata-Sogawa, Kumiko; Tokunaga, Makio; Iwamoto, Masaaki; Hiraoka, Yasushi; Haraguchi, Tokuko

    2014-01-01

    The nuclear pore complex (NPC) is an enormous proteinaceous complex composed of multiple copies of about 30 different proteins called nucleoporins. In this study, we analyzed the composition of the NPC in the model organism Schizosaccharomyces pombe using strains in which individual nucleoporins were tagged with GFP. We identified 31 proteins as nucleoporins by their localization to the nuclear periphery. Gene disruption analysis in previous studies coupled with gene disruption analysis in the present study indicates that 15 of these nucleoporins are essential for vegetative cell growth and the other 16 nucleoporins are non-essential. Among the 16 non-essential nucleoporins, 11 are required for normal progression through meiosis and their disruption caused abnormal spore formation or poor spore viability. Based on fluorescence measurements of GFP-fused nucleoporins, we estimated the composition of the NPC in S. pombe and found that the organization of the S. pombe NPC is largely similar to that of other organisms; a single NPC was estimated as being 45.8-47.8 MDa in size. We also used fluorescence measurements of single NPCs and quantitative western blotting to analyze the composition of the Nup107-Nup160 subcomplex, which plays an indispensable role in NPC organization and function. Our analysis revealed low amounts of Nup107 and Nup131 and high amounts of Nup132 in the Nup107-Nup160 subcomplex, suggesting that the composition of this complex in S. pombe may differ from that in S. cerevisiae and humans. Comparative analysis of NPCs in various organisms will lead to a comprehensive understanding of the functional architecture of the NPC.

  3. Characterization of nuclear pore complex components in fission yeast Schizosaccharomyces pombe

    PubMed Central

    Asakawa, Haruhiko; Yang, Hui-Ju; Yamamoto, Takaharu G; Ohtsuki, Chizuru; Chikashige, Yuji; Sakata-Sogawa, Kumiko; Tokunaga, Makio; Iwamoto, Masaaki; Hiraoka, Yasushi; Haraguchi, Tokuko

    2014-01-01

    The nuclear pore complex (NPC) is an enormous proteinaceous complex composed of multiple copies of about 30 different proteins called nucleoporins. In this study, we analyzed the composition of the NPC in the model organism Schizosaccharomyces pombe using strains in which individual nucleoporins were tagged with GFP. We identified 31 proteins as nucleoporins by their localization to the nuclear periphery. Gene disruption analysis in previous studies coupled with gene disruption analysis in the present study indicates that 15 of these nucleoporins are essential for vegetative cell growth and the other 16 nucleoporins are non-essential. Among the 16 non-essential nucleoporins, 11 are required for normal progression through meiosis and their disruption caused abnormal spore formation or poor spore viability. Based on fluorescence measurements of GFP-fused nucleoporins, we estimated the composition of the NPC in S. pombe and found that the organization of the S. pombe NPC is largely similar to that of other organisms; a single NPC was estimated as being 45.8–47.8 MDa in size. We also used fluorescence measurements of single NPCs and quantitative western blotting to analyze the composition of the Nup107-Nup160 subcomplex, which plays an indispensable role in NPC organization and function. Our analysis revealed low amounts of Nup107 and Nup131 and high amounts of Nup132 in the Nup107-Nup160 subcomplex, suggesting that the composition of this complex in S. pombe may differ from that in S. cerevisiae and humans. Comparative analysis of NPCs in various organisms will lead to a comprehensive understanding of the functional architecture of the NPC. PMID:24637836

  4. New vectors for epitope tagging and gene disruption in Schizosaccharomyces pombe

    PubMed Central

    Gadaleta, Mariana C.; Iwasaki, Osamu; Noguchi, Chiaki; Noma, Ken-ichi; Noguchi, Eishi

    2013-01-01

    We describe a series of new vectors for PCR-based epitope tagging and gene disruption in the fission yeast Schizosaccharomyces pombe, an exceptional model organism for the study of cellular processes. The vectors are designed for amplification of gene-targeting DNA cassettes and integration into specific genetic loci, allowing expression of proteins fused to 12 tandem copies of the Pk (V5) epitope or 5 tandem copies of the FLAG epitope with a glycine linker. These vectors are available with various antibiotic or nutritional markers and are useful for protein studies using biochemical and cell biological methods. We also describe new vectors for fluorescent protein-tagging and gene disruption using ura4MX6, LEU2MX6, and his3MX6 selection markers, allowing researchers in the S. pombe community to disrupt genes and manipulate genomic loci using primer sets already available for the widely used pFA6a-MX6 system. Our new vectors may also be useful for gene manipulation in Saccharomyces cerevisiae. Method Summary Our 12Pk-tagging vectors for genomic integration significantly improve the sensitivity of protein detection by Western blotting. We also describe genomic 5FLAG-tagging vectors with a glycine linker, which allows flexibility between the epitope and the protein, enhancing immunoprecipitation efficiency. This report also describes vectors for fluorescent-tagging and gene deletion useful in S. pombe. PMID:24215641

  5. Nitric oxide signaling and its role in oxidative stress response in Schizosaccharomyces pombe.

    PubMed

    Astuti, Rika Indri; Watanabe, Daisuke; Takagi, Hiroshi

    2016-01-30

    In the fission yeast Schizosaccharomyces pombe, we found that the putative NO dioxygenase SPAC869.02c (named Yhb1) and the S-nitrosoglutathione reductase Fmd2 cooperatively reduced intracellular NO levels as NO-detoxification enzymes. Although both mRNA and protein levels were increased with exogenous NO, their expression patterns were different during growth phases. While treatment with an NO synthase inhibitor in the log phase abrogated both NO production and Yhb1 expression, induction of Fmd2 in the stationary phase was correlated with elevated mitochondrial respiratory chain (MRC) activity, confirmed by the fact that inhibition of MRC complex III led to a decrease in Fmd2 and NO levels. Moreover, NO was localized in the mitochondria in the stationary phase, suggesting that there are two distinctive types of NO signaling in S. pombe. For mitochondria, pretreatment with an NO donor rescued cell growth by repressing generation of reactive oxygen species (ROS) under oxidative stress. DNA microarray analysis revealed that exogenous NO contributes to tolerance to hydrogen peroxide (H2O2) by (i) inhibition of Fe(3+) to Fe(2+) conversion, (ii) upregulation of the H2O2-detoxifying enzymes, and (iii) downregulation of the MRC genes, suggesting that NO plays a pivotal role in the negative feedback system to regulate ROS levels in S. pombe.

  6. Nitric oxide signaling and its role in oxidative stress response in Schizosaccharomyces pombe.

    PubMed

    Astuti, Rika Indri; Watanabe, Daisuke; Takagi, Hiroshi

    2016-01-30

    In the fission yeast Schizosaccharomyces pombe, we found that the putative NO dioxygenase SPAC869.02c (named Yhb1) and the S-nitrosoglutathione reductase Fmd2 cooperatively reduced intracellular NO levels as NO-detoxification enzymes. Although both mRNA and protein levels were increased with exogenous NO, their expression patterns were different during growth phases. While treatment with an NO synthase inhibitor in the log phase abrogated both NO production and Yhb1 expression, induction of Fmd2 in the stationary phase was correlated with elevated mitochondrial respiratory chain (MRC) activity, confirmed by the fact that inhibition of MRC complex III led to a decrease in Fmd2 and NO levels. Moreover, NO was localized in the mitochondria in the stationary phase, suggesting that there are two distinctive types of NO signaling in S. pombe. For mitochondria, pretreatment with an NO donor rescued cell growth by repressing generation of reactive oxygen species (ROS) under oxidative stress. DNA microarray analysis revealed that exogenous NO contributes to tolerance to hydrogen peroxide (H2O2) by (i) inhibition of Fe(3+) to Fe(2+) conversion, (ii) upregulation of the H2O2-detoxifying enzymes, and (iii) downregulation of the MRC genes, suggesting that NO plays a pivotal role in the negative feedback system to regulate ROS levels in S. pombe. PMID:26645666

  7. Fractionation and Structural Characterization of Arabinogalactan-Proteins from the Cell Wall of Rose Cells.

    PubMed Central

    Serpe, M. D.; Nothnagel, E. A.

    1995-01-01

    Arabinogalactan-proteins (AGPs) have been purified from Paul's Scarlet rose (Rosa sp.) cell walls. As estimated by gel permeation chromatography, the apparent molecular masses of the two major cell-wall AGP fractions were 130 and 242 kD. Since the 130-kD AGP had a ratio of arabinose/glucuronic acid that was 12 times higher than that of the 242-kD AGP, the fractions were named cell-wall AGP1 (CW-AGP1) and glucuronogalactan-protein (GGP), respectively. CW-AGP1 and GGP contained predominantly t-arabinofuranosyl residues; 3-linked, 6-linked, and 3,6-branched galactopyranosyl residues; and 4-linked and t-glucuronopyranosyl residues. The 1H-nuclear magnetic resonance spectra of CW-AGP1 and GGP showed that the arabinofuranosyl and galactopyranosyl residues were predominantly in [alpha]- and [beta]-anomeric configuration, respectively, and that GGP contained a few O-acetyl residues. The protein moieties of CW-AGP1 and GGP were both rich in hydroxyproline and alanine but differed in the percentage of various amino acids, including hydroxyproline, alanine, serine, and glycine. Cell-wall AGPs bound to ([beta]-D-glucosyl)3 Yariv phenylglycoside, but the stoichiometry of binding was about 6 times greater in GGP than in other Rosa AGPs. GGP seems to be peculiar to the cell wall, since no similar molecule was found in the culture medium. PMID:12228648

  8. Dynein Promotes Achiasmate Segregation in Schizosaccharomyces pombe

    PubMed Central

    Davis, Luther; Smith, Gerald R.

    2005-01-01

    Most organisms use crossovers (chiasmata) to maintain physical connections between homologous chromosomes that ensure their proper segregation at the first meiotic division. The fission yeast Schizosaccharomyces pombe has a residual ability to segregate homologous chromosomes in the absence of meiotic recombination (achiasmate segregation). Using cytologically tagged chromosomes, we established a role for the microtubule motor dynein in meiotic chromosome segregation. Dhc1, the motor subunit of dynein, is required for chromosome segregation in both the presence and the absence of recombination. Dlc1, a member of the Tctex-1 dynein light-chain family, preferentially affects the segregation of achiasmate chromosomes. Dlc1 is the first identified protein, outside of Drosophila, that preferentially affects achiasmate chromosome segregation. We discuss possible roles of the dynein motor in this process. PMID:15802518

  9. The plant cell wall in the feeding sites of cyst nematodes

    PubMed Central

    Bohlmann, Holger; Sobczak, Miroslaw

    2014-01-01

    Plant parasitic cyst nematodes (genera Heterodera and Globodera) are serious pests for many crops. They enter the host roots as migratory second stage juveniles (J2) and migrate intracellularly toward the vascular cylinder using their stylet and a set of cell wall degrading enzymes produced in the pharyngeal glands. They select an initial syncytial cell (ISC) within the vascular cylinder or inner cortex layers to induce the formation of a multicellular feeding site called a syncytium, which is the only source of nutrients for the parasite during its entire life. A syncytium can consist of more than hundred cells whose protoplasts are fused together through local cell wall dissolutions. While the nematode produces a cocktail of cell wall degrading and modifying enzymes during migration through the root, the cell wall degradations occurring during syncytium development are due to the plants own cell wall modifying and degrading proteins. The outer syncytial cell wall thickens to withstand the increasing osmotic pressure inside the syncytium. Furthermore, pronounced cell wall ingrowths can be formed on the outer syncytial wall at the interface with xylem vessels. They increase the surface of the symplast-apoplast interface, thus enhancing nutrient uptake into the syncytium. Processes of cell wall degradation, synthesis and modification in the syncytium are facilitated by a variety of plant proteins and enzymes including expansins, glucanases, pectate lyases and cellulose synthases, which are produced inside the syncytium or in cells surrounding the syncytium. PMID:24678316

  10. Nanoindentation techniques for the cell walls of wood

    NASA Astrophysics Data System (ADS)

    Jakes, Joseph Eugene

    There is a recognized need in forest products research to better understand how the mechanical properties of wood derive from the basic polymer components that make up the wood. For development of new engineered wood products there is the need to understand how chemical additives and adhesives interact with wood polymers and influence properties at the cellular level. To meet these needs I have developed nanoindentation techniques for probing the mechanical properties of the cell walls in wood. There are two, key results of this research. The first is a newly invented structural compliance method for isolating the properties of local regions within materials and excluding artifacts brought about by neighboring edges including free edges and interfaces between dissimilar cell wall layers. The second consists of methods to obtain viscoplastic and viscoelastic data over as wide a range of deformation rate as possible. The broadband nanoindentation creep (BNC) technique assesses the viscoplastic properties over 5 orders of magnitude in deformation rate (-10-4 to 10 s-1). Viscoelastic measurements can be made with unloading times ranging from 0.01 to 100 s, resulting in viscoelastic data that span four orders of magnitude in frequency or inverse time (˜10-3 to 10 s-1). To demonstrate the efficacy of these techniques, experiments are performed on a range of materials including fused silica, silicon, molybdenum, siliconon-insulator layered specimen, poly (methylmetacrylate), polycarbonate, polystyrene, wood cells in loblolly pine (Pinus taeda ), and a polypropylene-wood composite. Finally, the structural compliance method and BNC are combined to explore polymeric methylene diphenyl diisocyanate (pMDI)-wood interactions. The data suggest that pMDI polymerizes in situ to create an interpenetrating polymer network.

  11. Role of calcium in the mechanical strength of soybean hypocotyl cell walls

    SciTech Connect

    Virk, S.S.; Cleland, R.E.

    1986-04-01

    Calcium ions inhibit auxin-induced growth in both dicot stems and coleoptiles. In coleoptiles calcium does not directly stiffen cell walls. The authors have tested here whether calcium might alter the mechanical strength of a dicot cell wall, the soybean hypocotyl. Sections were longitudinally bisected, boiled or frozen-thawed, incubated in solutions and then the mechanical strength was determined with an Instron. The calcium content was also measured. Removal of calcium by EGTA or by acidic buffers such as K-Pi-citrate resulted in a proportional increase in wall extensibility. Addition of calcium, on the other hand, stiffened the walls. These changes were reversible. It was concluded that calcium crosslinks make a significant contribution to the strength of dicot stem cell walls, and that in vivo, removal of calcium from the wall by uptake into the cell could result in wall loosening and thus enhanced growth.

  12. Identifying cytoplasmic input to the cell wall of growing Chara corallina.

    PubMed

    Proseus, Timothy E; Boyer, John S

    2006-01-01

    Plants enlarge mostly because the walls of certain cells enlarge, with accompanying input of wall constituents and other factors from the cytoplasm. However, the enlargement can occur without input, suggesting an uncertain relationship between cytoplasmic input and plant growth. Therefore, the role of the input was investigated by quantitatively comparing growth in isolated walls (no input) with that in living cells (input occurring). Cell walls were isolated from growing internodes of Chara corallina and filled with pressurized oil to control turgor pressure while elongation was monitored. Turgor pressure in living cells was similarly controlled and monitored by adding/removing cell solution. Temperature was varied in some experiments. At all pressures and temperatures, isolated walls displayed turgor-driven growth indistinguishable in every respect from that in living cells, except the rate decelerated in the isolated walls while the living cells grew rapidly. The growth in the isolated walls was highly responsive to temperature, in contrast to the elastic extension that has been shown to be insensitive to similar temperatures. Consequently, strong intermolecular bonds were responsible for growth and weak bonds for elastic extension. Boiling the walls gave the same results, indicating that enzyme activities were not controlling these bonds. However, pectin added to isolated walls reversed their growth deceleration and returned the rate to that in the living cells. The pectin was similar to that normally produced by the cytoplasm and deposited in the wall, suggesting that continued cytoplasmic input of pectin may play a role in sustaining turgor-driven growth in Chara.

  13. Bacterial cell wall biogenesis is mediated by SEDS and PBP polymerase families functioning semi-autonomously.

    PubMed

    Cho, Hongbaek; Wivagg, Carl N; Kapoor, Mrinal; Barry, Zachary; Rohs, Patricia D A; Suh, Hyunsuk; Marto, Jarrod A; Garner, Ethan C; Bernhardt, Thomas G

    2016-01-01

    Multi-protein complexes organized by cytoskeletal proteins are essential for cell wall biogenesis in most bacteria. Current models of the wall assembly mechanism assume that class A penicillin-binding proteins (aPBPs), the targets of penicillin-like drugs, function as the primary cell wall polymerases within these machineries. Here, we use an in vivo cell wall polymerase assay in Escherichia coli combined with measurements of the localization dynamics of synthesis proteins to investigate this hypothesis. We find that aPBP activity is not necessary for glycan polymerization by the cell elongation machinery, as is commonly believed. Instead, our results indicate that cell wall synthesis is mediated by two distinct polymerase systems, shape, elongation, division, sporulation (SEDS)-family proteins working within the cytoskeletal machines and aPBP enzymes functioning outside these complexes. These findings thus necessitate a fundamental change in our conception of the cell wall assembly process in bacteria. PMID:27643381

  14. Reversible swelling of the cell wall of poplar biomass by ionic liquid at room temperature

    PubMed Central

    Lucas, Marcel; Wagner, Greg L.; Nishiyama, Yoshiharu; Hanson, Leif; Samayam, Indira P.; Schall, Constance A.; Langan, Paul; Rector, Kirk D.

    2012-01-01

    Time-resolved autofluorescence, Raman microspectroscopy, and scanning microprobe X-ray diffraction were combined in order to characterize lignocellulosic biomass from poplar trees and how it changes during treatment with the ionic liquid 1-n-ethyl-3-methylimidazolium acetate (EMIMAC) at room temperature. The EMIMAC penetrates the cell wall from the lumen, swelling the cell wall by about a factor of two towards the empty lumen. However, the middle lamella remains unchanged, preventing the cell wall from swelling outwards. During this swelling, most of the cellulose microfibrils are solubi-lized but chain migration is restricted and a small percentage of microfibrils persist. When the EMIMAC is expelled, the cellulose recrystallizes as microfibrils of cellulose I. There is little change in the relative chemical composition of the cell wall after treatment. The action of EMIMAC on the poplar cell wall at room temperature would therefore appear to be a reversible swelling and a reversible decrystallization of the cell wall. PMID:21247757

  15. Plant biomass recalcitrance: effect of hemicellulose composition on nanoscale forces that control cell wall strength.

    PubMed

    Silveira, Rodrigo L; Stoyanov, Stanislav R; Gusarov, Sergey; Skaf, Munir S; Kovalenko, Andriy

    2013-12-26

    Efficient conversion of lignocellulosic biomass to second-generation biofuels and valuable chemicals requires decomposition of resilient plant cell wall structure. Cell wall recalcitrance varies among plant species and even phenotypes, depending on the chemical composition of the noncellulosic matrix. Changing the amount and composition of branches attached to the hemicellulose backbone can significantly alter the cell wall strength and microstructure. We address the effect of hemicellulose composition on primary cell wall assembly forces by using the 3D-RISM-KH molecular theory of solvation, which provides statistical-mechanical sampling and molecular picture of hemicellulose arrangement around cellulose. We show that hemicellulose branches of arabinose, glucuronic acid, and especially glucuronate strengthen the primary cell wall by strongly coordinating to hydrogen bond donor sites on the cellulose surface. We reveal molecular forces maintaining the cell wall structure and provide directions for genetic modulation of plants and pretreatment design to render biomass more amenable to processing. PMID:24274712

  16. Relating the mechanics of the primary plant cell wall to morphogenesis.

    PubMed

    Bidhendi, Amir J; Geitmann, Anja

    2016-01-01

    Regulation of the mechanical properties of the cell wall is a key parameter used by plants to control the growth behavior of individual cells and tissues. Modulation of the mechanical properties occurs through the control of the biochemical composition and the degree and nature of interlinking between cell wall polysaccharides. Preferentially oriented cellulose microfibrils restrict cellular expansive growth, but recent evidence suggests that this may not be the trigger for anisotropic growth. Instead, non-uniform softening through the modulation of pectin chemistry may be an initial step that precedes stress-induced stiffening of the wall through cellulose. Here we briefly review the major cell wall polysaccharides and their implication for plant cell wall mechanics that need to be considered in order to study the growth behavior of the primary plant cell wall.

  17. Laccases Direct Lignification in the Discrete Secondary Cell Wall Domains of Protoxylem1[W][OPEN

    PubMed Central

    Schuetz, Mathias; Benske, Anika; Smith, Rebecca A.; Watanabe, Yoichiro; Tobimatsu, Yuki; Ralph, John; Demura, Taku; Ellis, Brian; Samuels, A. Lacey

    2014-01-01

    Plants precisely control lignin deposition in spiral or annular secondary cell wall domains during protoxylem tracheary element (TE) development. Because protoxylem TEs function to transport water within rapidly elongating tissues, it is important that lignin deposition is restricted to the secondary cell walls in order to preserve the plasticity of adjacent primary wall domains. The Arabidopsis (Arabidopsis thaliana) inducible VASCULAR NAC DOMAIN7 (VND7) protoxylem TE differentiation system permits the use of mutant backgrounds, fluorescent protein tagging, and high-resolution live-cell imaging of xylem cells during secondary cell wall development. Enzymes synthesizing monolignols, as well as putative monolignol transporters, showed a uniform distribution during protoxylem TE differentiation. By contrast, the oxidative enzymes LACCASE4 (LAC4) and LAC17 were spatially localized to secondary cell walls throughout protoxylem TE differentiation. These data support the hypothesis that precise delivery of oxidative enzymes determines the pattern of cell wall lignification. This view was supported by lac4lac17 mutant analysis demonstrating that laccases are necessary for protoxylem TE lignification. Overexpression studies showed that laccases are sufficient to catalyze ectopic lignin polymerization in primary cell walls when exogenous monolignols are supplied. Our data support a model of protoxylem TE lignification in which monolignols are highly mobile once exported to the cell wall, and in which precise targeting of laccases to secondary cell wall domains directs lignin deposition. PMID:25157028

  18. Area Expansivity Moduli of Regenerating Plant Protoplast Cell Walls Exposed to Shear Flows

    NASA Astrophysics Data System (ADS)

    Fujimura, Yuu; Iino, Masaaki; Watanabe, Ugai

    2005-05-01

    To control the elasticity of the plant cell wall, protoplasts isolated from cultured Catharanthus roseus cells were regenerated in shear flows of 115 s-1 (high shear) and 19.2 s-1 (low shear, as a control). The surface area expansivity modulus and the surface breaking strength of these regenerating protoplasts were measured by a micropipette aspiration technique. Cell wall synthesis was also measured using a cell wall-specific fluorescent dye. High shear exposure for 3 h doubled both the surface area modulus and breaking strength observed under low shear, significantly decreased cell wall synthesis, and roughly quadrupled the moduli of the cell wall. Based on the cell wall synthesis data, we estimated the three-dimensional modulus of the cell wall to be 4.1± 1.2 GPa for the high shear, and 0.35± 0.2 GPa for the low shear condition, using the surface area expansivity modulus divided by the cell wall thickness, which is identical with the Young’s modulus divided by 2(1-σ), where σ is Poisson's ratio. We concluded that high shear exposure considerably strengthens the newly synthesized cell wall.

  19. Modulation of Spc1 stress-activated protein kinase activity by methylglyoxal through inhibition of protein phosphatase in the fission yeast Schizosaccharomyces pombe

    SciTech Connect

    Takatsume, Yoshifumi; Izawa, Shingo; Inoue, Yoshiharu

    2007-11-30

    Methylglyoxal, a ubiquitous metabolite derived from glycolysis has diverse physiological functions in yeast cells. Previously, we have reported that extracellularly added methylglyoxal activates Spc1, a stress-activated protein kinase (SAPK), in the fission yeast Schizosaccharomyces pombe [Y. Takatsume, S. Izawa, Y. Inoue, J. Biol. Chem. 281 (2006) 9086-9092]. Phosphorylation of Spc1 by treatment with methylglyoxal in S. pombe cells defective in glyoxalase I, an enzyme crucial for the metabolism of methylglyoxal, continues for a longer period than in wild-type cells. Here we show that methylglyoxal inhibits the activity of the protein phosphatase responsible for the dephosphorylation of Spc1 in vitro. In addition, we found that methylglyoxal inhibits human protein tyrosine phosphatase 1B (PTP1B) also. We propose a model for the regulation of the activity of the Spc1-SAPK signaling pathway by methylglyoxal in S. pombe.

  20. New multi-purpose high copy number vector with greater mitotic stability for diverse applications in fission yeast Schizosaccharomyces pombe.

    PubMed

    Verma, Hemant Kumar; Singh, Jagmohan

    2012-11-01

    We have constructed a pUC19-based multipurpose ATG vector in Schizosaccharomyces pombe with higher copy number and mitotic stability possible with commonly used vectors. The vector, having an NdeI site in its polylinker to provide ATG site for expression, carries a greatly truncated version of URA3 gene, URA3m, of Saccharomyces cerevisiae as a selection marker. In addition, it contains the mat2P-right flank region (mat2P-RF) of S. pombe as an autonomous replicating sequence (ARS) and a polylinker with wider choice of restriction sites. While URA3m confers an increase in plasmid copy number up to 200 copies/cell, mat2P-RF imparts greater mitotic stability than the standard ars1 element of S. pombe. Finally, the vector also includes the transcription termination signal of the nmt1 gene (Tnmt1). This basic vector should serve as a versatile tool for studies of gene function in S. pombe.

  1. Starting to Gel: How Arabidopsis Seed Coat Epidermal Cells Produce Specialized Secondary Cell Walls

    PubMed Central

    Voiniciuc, Cătălin; Yang, Bo; Schmidt, Maximilian Heinrich-Wilhelm; Günl, Markus; Usadel, Björn

    2015-01-01

    For more than a decade, the Arabidopsis seed coat epidermis (SCE) has been used as a model system to study the synthesis, secretion and modification of cell wall polysaccharides, particularly pectin. Our detailed re-evaluation of available biochemical data highlights that Arabidopsis seed mucilage is more than just pectin. Typical secondary wall polymers such as xylans and heteromannans are also present in mucilage. Despite their low abundance, these components appear to play essential roles in controlling mucilage properties, and should be further investigated. We also provide a comprehensive community resource by re-assessing the mucilage phenotypes of almost 20 mutants using the same conditions. We conduct an in-depth functional evaluation of all the SCE genes described in the literature and propose a revised model for mucilage production. Further investigation of SCE cells will improve our understanding of plant cell walls. PMID:25658798

  2. Starting to gel: how Arabidopsis seed coat epidermal cells produce specialized secondary cell walls.

    PubMed

    Voiniciuc, Cătălin; Yang, Bo; Schmidt, Maximilian Heinrich-Wilhelm; Günl, Markus; Usadel, Björn

    2015-02-04

    For more than a decade, the Arabidopsis seed coat epidermis (SCE) has been used as a model system to study the synthesis, secretion and modification of cell wall polysaccharides, particularly pectin. Our detailed re-evaluation of available biochemical data highlights that Arabidopsis seed mucilage is more than just pectin. Typical secondary wall polymers such as xylans and heteromannans are also present in mucilage. Despite their low abundance, these components appear to play essential roles in controlling mucilage properties, and should be further investigated. We also provide a comprehensive community resource by re-assessing the mucilage phenotypes of almost 20 mutants using the same conditions. We conduct an in-depth functional evaluation of all the SCE genes described in the literature and propose a revised model for mucilage production. Further investigation of SCE cells will improve our understanding of plant cell walls.

  3. Two cationic peroxidases from cell walls of Araucaria araucana seeds.

    PubMed

    Riquelme, A; Cardemil, L

    1995-05-01

    We have previously reported the purification and partial characterization of two cationic peroxidases from the cell walls of seeds and seedlings of the South American conifer, Araucaria araucana. In this work, we have studied the amino acid composition and NH2-terminal sequences of both enzymes. We also compare the data obtained from these analyses with those reported for other plant peroxidases. The two peroxidases are similar in their amino acid compositions. Both are particularly rich in glycine, which comprises more than 30% of the amino acid residues. The content of serine is also high, ca 17%. The two enzymes are different in their content of arginine, alanine, valine, phenylalanine and threonine. Both peroxidases have identical NH2-terminal sequences, indicating that the two proteins are genetically related and probably are isoforms of the same kind of peroxidase. The amino acid composition and NH2-terminal sequence analyses showed marked differences from the cationic peroxidases from turnip and horseradish. PMID:7786490

  4. Single Wall Carbon Nanotube-polymer Solar Cells

    NASA Technical Reports Server (NTRS)

    Bailey, Sheila G.; Castro, Stephanie L.; Landi, Brian J.; Gennett, Thomas; Raffaelle, Ryne P.

    2005-01-01

    Investigation of single wall carbon nanotube (SWNT)-polymer solar cells has been conducted towards developing alternative lightweight, flexible devices for space power applications. Photovoltaic devices were constructed with regioregular poly(3-octylthiophene)-(P3OT) and purified, >95% w/w, laser-generated SWNTs. The P3OT composites were deposited on ITO-coated polyethylene terapthalate (PET) and I-V characterization was performed under simulated AM0 illumination. Fabricated devices for the 1.0% w/w SWNT-P3OT composites showed a photoresponse with an open-circuit voltage (V(sub oc)) of 0.98 V and a short-circuit current density (I(sub sc)) of 0.12 mA/sq cm. Optimization of carrier transport within these novel photovoltaic systems is proposed, specifically development of nanostructure-SWNT complexes to enhance exciton dissociation.

  5. Composition and architecture of the cell walls of grasses and the mechanisms of synthesis of cell wall polysaccharides. Final report for period September 1, 1988 - April 30, 2001

    SciTech Connect

    Carpita, Nicholas C.

    2001-10-18

    This program was devoted toward complete understanding of the polysaccharide structure and architecture of the primary cell walls grasses and cereals, and the biosynthesis of the mixed-linkage beta-glucane, a cellulose interacting polymer that is synthesized uniquely by grass species and close relatives. With these studies as focal point, the support from DOE was instrumental in the development of new analytical means that enabled us to characterize carbohydrate structure, to reveal new features of cell wall dynamics during cell growth, and to apply these techniques in other model organisms. The support by DOE in these basic studies was acknowledged on numerous occasions in review articles covering current knowledge of cell wall structure, architecture, dynamics, biosynthesis, and in all genes related to cell wall biogenesis.

  6. Pectinous cell wall thickenings formation - A common defense strategy of plants to cope with Pb.

    PubMed

    Krzesłowska, Magdalena; Rabęda, Irena; Basińska, Aneta; Lewandowski, Michał; Mellerowicz, Ewa J; Napieralska, Anna; Samardakiewicz, Sławomir; Woźny, Adam

    2016-07-01

    Lead, one of the most abundant and hazardous trace metals affecting living organisms, has been commonly detected in plant cell walls including some tolerant plants, mining ecotypes and hyperaccumulators. We have previously shown that in tip growing Funaria sp. protonemata cell wall is remodeled in response to lead by formation of thickenings rich in low-methylesterified pectins (pectin epitope JIM5 - JIM5-P) able to bind metal ions, which accumulate large amounts of Pb. Hence, it leads to the increase of cell wall capacity for Pb compartmentalization. Here we show that diverse plant species belonging to different phyla (Arabidopsis, hybrid aspen, star duckweed), form similar cell wall thickenings in response to Pb. These thickenings are formed in tip growing cells such as the root hairs, and in diffuse growing cells such as meristematic and root cap columella cells of root apices in hybrid aspen and Arabidopsis and in mesophyll cells in star duckweed fronds. Notably, all analyzed cell wall thickenings were abundant in JIM5-P and accumulated high amounts of Pb. In addition, the co-localization of JIM5-P and Pb commonly occurred in these cells. Hence, cell wall thickenings formed the extra compartment for Pb accumulation. In this way plant cells increased cell wall capacity for compartmentalization of this toxic metal, protecting protoplast from its toxicity. As cell wall thickenings occurred in diverse plant species and cell types differing in the type of growth we may conclude that pectinous cell wall thickenings formation is a widespread defense strategy of plants to cope with Pb. Moreover, detection of natural defense strategy, increasing plant cell walls capacity for metal accumulation, reveals a promising direction for enhancing plant efficiency in phytoremediation. PMID:27107260

  7. Pectinous cell wall thickenings formation - A common defense strategy of plants to cope with Pb.

    PubMed

    Krzesłowska, Magdalena; Rabęda, Irena; Basińska, Aneta; Lewandowski, Michał; Mellerowicz, Ewa J; Napieralska, Anna; Samardakiewicz, Sławomir; Woźny, Adam

    2016-07-01

    Lead, one of the most abundant and hazardous trace metals affecting living organisms, has been commonly detected in plant cell walls including some tolerant plants, mining ecotypes and hyperaccumulators. We have previously shown that in tip growing Funaria sp. protonemata cell wall is remodeled in response to lead by formation of thickenings rich in low-methylesterified pectins (pectin epitope JIM5 - JIM5-P) able to bind metal ions, which accumulate large amounts of Pb. Hence, it leads to the increase of cell wall capacity for Pb compartmentalization. Here we show that diverse plant species belonging to different phyla (Arabidopsis, hybrid aspen, star duckweed), form similar cell wall thickenings in response to Pb. These thickenings are formed in tip growing cells such as the root hairs, and in diffuse growing cells such as meristematic and root cap columella cells of root apices in hybrid aspen and Arabidopsis and in mesophyll cells in star duckweed fronds. Notably, all analyzed cell wall thickenings were abundant in JIM5-P and accumulated high amounts of Pb. In addition, the co-localization of JIM5-P and Pb commonly occurred in these cells. Hence, cell wall thickenings formed the extra compartment for Pb accumulation. In this way plant cells increased cell wall capacity for compartmentalization of this toxic metal, protecting protoplast from its toxicity. As cell wall thickenings occurred in diverse plant species and cell types differing in the type of growth we may conclude that pectinous cell wall thickenings formation is a widespread defense strategy of plants to cope with Pb. Moreover, detection of natural defense strategy, increasing plant cell walls capacity for metal accumulation, reveals a promising direction for enhancing plant efficiency in phytoremediation.

  8. Mechanisms for shaping, orienting, positioning and patterning plant secondary cell walls.

    PubMed

    Pesquet, Edouard; Korolev, Andrey V; Calder, Grant; Lloyd, Clive W

    2011-06-01

    Xylem vessels are cells that develop a specifically ornamented secondary cell wall to ensure their vascular function, conferring both structural strength and impermeability. Further plasticity is given to these vascular cells by a range of different patterns described by their secondary cell walls that-as for the growth of all plant organs-are developmentally regulated. Microtubules and their associated proteins, named MAPs, are essential to define the shape, the orientation, the position and the overall pattern of these secondary cell walls. Key actors in this process are the land-plant specific MAP70 proteins which not only allow the secondary cell wall to be positioned at the cell cortex but also determine the overall pattern described by xylem vessel secondary cell walls

  9. Lead sulfide nanoparticles increase cell wall chitin content and induce apoptosis in Saccharomyces cerevisiae.

    PubMed

    Sun, Meiqing; Yu, Qilin; Hu, Mengyuan; Hao, Zhenwei; Zhang, Chengdong; Li, Mingchun

    2014-05-30

    Although there have been numerous studies on bacterial toxicity, the cytotoxicity of nanoparticles toward fungi remains poorly understood. We investigated the toxicity of various sizes of lead sulfide particles against the important model fungus, Saccharomyces cerevisiae. The smallest particle exerted the highest toxicity, inhibiting cell growth and decreasing cell viability, likely reflecting reduced sedimentation and persistent cell wall attack. In response to cell wall stress, S. cerevisiae showed an increase in the cell wall chitin content and the overexpression of FKS2 and PRM5, two genes of the cell wall integrity signaling pathway. Cell wall stress increased the concentration of intracellular reactive oxygen species, leading to mitochondrial dysfunction and cell apoptosis. The contribution of dissolved lead ions to the overall toxicity was negligible. These findings provide the first demonstration of the physiological protective response of a fungus toward nanoparticles, thereby contributing useful information to the assessment of the environmental impact of metal nanoparticles.

  10. Plant cell walls throughout evolution: towards a molecular understanding of their design principles.

    PubMed

    Sarkar, Purbasha; Bosneaga, Elena; Auer, Manfred

    2009-01-01

    Throughout their life, plants typically remain in one location utilizing sunlight for the synthesis of carbohydrates, which serve as their sole source of energy as well as building blocks of a protective extracellular matrix, called the cell wall. During the course of evolution, plants have repeatedly adapted to their respective niche, which is reflected in the changes of their body plan and the specific design of cell walls. Cell walls not only changed throughout evolution but also are constantly remodelled and reconstructed during the development of an individual plant, and in response to environmental stress or pathogen attacks. Carbohydrate-rich cell walls display complex designs, which together with the presence of phenolic polymers constitutes a barrier for microbes, fungi, and animals. Throughout evolution microbes have co-evolved strategies for efficient breakdown of cell walls. Our current understanding of cell walls and their evolutionary changes are limited as our knowledge is mainly derived from biochemical and genetic studies, complemented by a few targeted yet very informative imaging studies. Comprehensive plant cell wall models will aid in the re-design of plant cell walls for the purpose of commercially viable lignocellulosic biofuel production as well as for the timber, textile, and paper industries. Such knowledge will also be of great interest in the context of agriculture and to plant biologists in general. It is expected that detailed plant cell wall models will require integrated correlative multimodal, multiscale imaging and modelling approaches, which are currently underway.

  11. Discovery of Recurrent Sequence Motifs in Saccharomyces cerevisiae Cell Wall Proteins

    PubMed Central

    Coronado, Juan E.; Epstein, Susan L.; Qiu, Wei-Gang; Lipke, Peter N.

    2008-01-01

    This paper describes a procedure for the discovery of recurrent substrings in amino acid sequences of proteins, and its application to fungal cell walls. The evolutionary origins of fungal cell walls are an open biological question. This question can be approached by studies of similarity among the sequences and sub-sequences of fungal wall proteins and by comparison to proteins in animals. We describe here how we have discovered building blocks, represented as recurrent sequence motifs (sub-sequences), within fungal cell wall proteins. These motifs have not been systematically identified before, because the low Shannon entropy of the cell wall sequences has hindered searches for local sequence similarities by sequence alignments. Nonetheless, our new, composition-based scoring matrices for local alignment searches now support statistically valid alignments for such low entropy sequences (Coronado et al. 2006. Euk. Cell 5: 628–637). We have now searched for similarities in a set of 171 known and putative cell wall proteins from baker’s yeast, Saccharomyces cerevisiae. The aligned segments were repeatedly subdivided and catalogued to identify 217 recurrent sequence motifs of length 8 amino acids or greater. 95% of these motifs occur in more than one cell wall protein. The median length of the motifs is 22 amino acid residues, considerably shorter than protein domains. For many cell wall proteins, these motifs collectively account for more than half of their amino acids. The prevalence of these motifs supports the idea of fungal cell wall proteins as assemblies of recurrent building blocks. PMID:19430580

  12. Plant cell walls throughout evolution: towards a molecular understanding of their design principles

    SciTech Connect

    Sarkar, Purbasha; Bosneaga, Elena; Auer, Manfred

    2009-02-16

    Throughout their life, plants typically remain in one location utilizing sunlight for the synthesis of carbohydrates, which serve as their sole source of energy as well as building blocks of a protective extracellular matrix, called the cell wall. During the course of evolution, plants have repeatedly adapted to their respective niche,which is reflected in the changes of their body plan and the specific design of cell walls. Cell walls not only changed throughout evolution but also are constantly remodelled and reconstructed during the development of an individual plant, and in response to environmental stress or pathogen attacks. Carbohydrate-rich cell walls display complex designs, which together with the presence of phenolic polymers constitutes a barrier for microbes, fungi, and animals. Throughout evolution microbes have co-evolved strategies for efficient breakdown of cell walls. Our current understanding of cell walls and their evolutionary changes are limited as our knowledge is mainly derived from biochemical and genetic studies, complemented by a few targeted yet very informative imaging studies. Comprehensive plant cell wall models will aid in the re-design of plant cell walls for the purpose of commercially viable lignocellulosic biofuel production as well as for the timber, textile, and paper industries. Such knowledge will also be of great interest in the context of agriculture and to plant biologists in general. It is expected that detailed plant cell wall models will require integrated correlative multimodal, multiscale imaging and modelling approaches, which are currently underway.

  13. Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy

    SciTech Connect

    Donohoe, B. S.; Selig, M. J.; Viamajala, S.; Vinzant, T. B.; Adney, W. S.; Himmel, M. E.

    2009-06-15

    In general, pretreatments are designed to enhance the accessibility of cellulose to enzymes, allowing for more efficient conversion. In this study, we have detected the penetration of major cellulases present in a commercial enzyme preparation (Spezyme CP) into corn stem cell walls following mild-, moderate- and high-severity dilute sulfuric acid pretreatments. The Trichoderma reesei enzymes, Cel7A (CBH I) and Cel7B (EG I), as well as the cell wall matrix components xylan and lignin were visualized within digested corn stover cell walls by immuno transmission electron microscopy (TEM) using enzyme- and polymer-specific antibodies. Low severity dilute-acid pretreatment (20 min at 100 C) enabled <1% of the thickness of secondary cell walls to be penetrated by enzyme, moderate severity pretreatment at (20 min at 120 C) allowed the enzymes to penetrate {approx}20% of the cell wall, and the high severity (20 min pretreatment at 150 C) allowed 100% penetration of even the thickest cell walls. These data allow direct visualization of the dramatic effect dilute-acid pretreatment has on altering the condensed ultrastructure of biomass cell walls. Loosening of plant cell wall structure due to pretreatment and the subsequently improved access by cellulases has been hypothesized by the biomass conversion community for over two decades, and for the first time, this study provides direct visual evidence to verify this hypothesis. Further, the high-resolution enzyme penetration studies presented here provide insight into the mechanisms of cell wall deconstruction by cellulolytic enzymes.

  14. Functional analysis of putative phosphoenolpyruvate transporters localized to the Golgi apparatus in Schizosaccharomyces pombe.

    PubMed

    Yoritsune, Ken-ichi; Higuchi, Yujiro; Matsuzawa, Tomohiko; Takegawa, Kaoru

    2014-11-01

    The cell surface of Schizosaccharomyces pombe is negatively charged due to the presence of pyruvylated oligosaccharides, which is important for cell-cell recognition. However, the mechanism of pyruvate supply to oligosaccharides is not clearly understood. Here, we analyzed three putative phosphoenolpyruvate (PEP) transporter genes (pet1(+) , pet2(+) , and pet3(+) ) in S. pombe, identified by sequence homology search against the Arabidopsis thaliana PEP transporter AtPPT1. Schizosaccharomyces pombe strain carrying a disruption in pet1(+) (pet1Δ) or in pet2(+) (pet2Δ), but not the strain carrying a disruption in pet3(+) (pet3Δ), showed reduced pyruvate level on the cell surface. This reduction in pyruvate level was restored to the control level by expressing green fluorescent protein (GFP)-tagged Pet1p and Pet2p in respective disruptants. Fluorescence microscope studies revealed that GFP-tagged Pet1p and Pet2p were localized to the Golgi apparatus. Although expression of neither AtPPT1 nor AtPPT2 suppressed the pet1Δ phenotype, that of chimeric constructs, where the N-terminal regions of AtPPT1 and AtPPT2 were replaced by the N-terminal region of Pet1p, partially suppressed the pet1Δ phenotype. Furthermore, the reduction in cell surface negative charge in pet1Δ cells was restored by incubating these cells with recombinant Pvg1p and PEP. Thus, Pet1p and Pet2p are likely involved in transporting PEP from the cytoplasm into the Golgi.

  15. Early cell-wall modifications of maize cell cultures during habituation to dichlobenil.

    PubMed

    de Castro, María; Largo-Gosens, Asier; Alvarez, Jesús Miguel; García-Angulo, Penélope; Acebes, José Luis

    2014-01-15

    Studies involving the habituation of plant cell cultures to cellulose biosynthesis inhibitors have achieved significant progress as regards understanding the structural plasticity of cell walls. However, since habituation studies have typically used high concentrations of inhibitors and long-term habituation periods, information on initial changes associated with habituation has usually been lost. This study focuses on monitoring and characterizing the short-term habituation process of maize (Zea mays) cell suspensions to dichlobenil (DCB). Cellulose quantification and FTIR spectroscopy of cell walls from 20 cell lines obtained during an incipient DCB-habituation process showed a reduction in cellulose levels which tended to revert depending on the inhibitor concentration and the length of time that cells were in contact with it. Variations in the cellulose content were concomitant with changes in the expression of several ZmCesA genes, mainly involving overexpression of ZmCesA7 and ZmCesA8. In order to explore these changes in more depth, a cell line habituated to 1.5μM DCB was identified as representative of incipient DCB habituation and selected for further analysis. The cells of this habituated cell line grew more slowly and formed larger clusters. Their cell walls were modified, showing a 33% reduction in cellulose content, that was mainly counteracted by an increase in arabinoxylans, which presented increased extractability. This result was confirmed by immunodot assays graphically plotted by heatmaps, since habituated cell walls had a more extensive presence of epitopes for arabinoxylans and xylans, but also for homogalacturonan with a low degree of esterification and for galactan side chains of rhamnogalacturonan I. Furthermore, a partial shift of xyloglucan epitopes toward more easily extractable fractions was found. However, other epitopes, such as these specific for arabinan side chains of rhamnogalacturonan I or homogalacturonan with a high degree of

  16. Analysis of Cell Wall Teichoic Acids in Staphylococcus aureus.

    PubMed

    Covas, Gonçalo; Vaz, Filipa; Henriques, Gabriela; Pinho, Mariana G; Filipe, Sérgio R

    2016-01-01

    Most bacterial cells are surrounded by a surface composed mainly of peptidoglycan (PGN), a glycopolymer responsible for ensuring the bacterial shape and a telltale molecule that betrays the presence of bacteria to the host immune system. In Staphylococcus aureus, as in most gram-positive bacteria, peptidoglycan is concealed by covalently linked molecules of wall teichoic acids (WTA)-phosphate rich molecules made of glycerol and ribitol phosphates which may be tailored by different amino acids and sugars.In order to analyze and compare the composition of WTA produced by different S. aureus strains, we describe methods to: (1) quantify the total amount of WTA present at the bacterial cell surface, through the determination of the inorganic phosphate present in phosphodiester linkages of WTA; (2) identify which sugar constituents are present in the assembled WTA molecules, by detecting the monosaccharides, released by acid hydrolysis, through an high-performance anion exchange chromatography analysis coupled with pulsed amperometric detection (HPAEC-PAD) and (3) compare the polymerization degree of WTA found at the cell surface of different S. aureus strains, through their different migration in a polyacrylamide gel electrophoresis (PAGE). PMID:27311674

  17. Wall extensibility: its nature, measurement and relationship to plant cell growth

    NASA Technical Reports Server (NTRS)

    Cosgrove, D. J.

    1993-01-01

    Expansive growth of plant cells is controlled principally by processes that loosen the wall and enable it to expand irreversibly. The central role of wall relaxation for cell expansion is reviewed. The most common methods for assessing the extension properties of plant cell walls ( wall extensibility') are described, categorized and assessed critically. What emerges are three fundamentally different approaches which test growing cells for their ability (a) to enlarge at different values of turgor, (b) to induce wall relaxation, and (c) to deform elastically or plastically in response to an applied tensile force. Analogous methods with isolated walls are similarly reviewed. The results of these different assays are related to the nature of plant cell growth and pertinent biophysical theory. I argue that the extensibilities' measured by these assays are fundamentally different from one another and that some are more pertinent to growth than others.

  18. Habituation of Bean (Phaseolus vulgaris) Cell Cultures to Quinclorac and Analysis of the Subsequent Cell Wall Modifications

    PubMed Central

    Alonso-Simón, Ana; García-Angulo, Penélope; Encina, Antonio; Acebes, José Luis; Álvarez, Jesús

    2008-01-01

    Background and Aims The herbicide quinclorac has been reported to inhibit incorporation of glucose both into cellulose and other cell wall polysaccharides. However, further work has failed to detect any apparent effect of this herbicide on the synthesis of the wall. In order to elucidate whether quinclorac elicits the inhibition of cellulose biosynthesis directly, in this study bean cell calli were habituated to grow on lethal concentrations of the herbicide and the modifications in cell wall composition due to the habituation process were analysed. Methods Fourier transform infrared spectroscopy associated with multivariate analysis, cell wall fractionation techniques, biochemical analyses and the immunolocation of different cell wall components with specific monoclonal antibodies were used to characterize the cell walls of quinclorac-habituated cells. Key Results Quinclorac-habituated cells were more irregularly shaped than non-habituated cells and they accumulated an extracellular material, which was more abundant as the level of habituation rose. Habituated cells did not show any decrease in cellulose content, but cell wall fractionation revealed that changes occurred in the distribution and post-depositional modifications of homogalacturonan and rhamnogalacturonan I during the habituation process. Therefore, since the action of quinclorac on the cell wall does not seem to be due to a direct inhibition of any cell wall component, it is suggested that the effect of quinclorac on the cell wall could be due to a side-effect of the herbicide. Conclusions Long-term modifications of the cell wall caused by the habituation of bean cell cultures to quinclorac did not resemble those of bean cells habituated to the well-known cellulose biosynthesis inhibitors dichlobenil or isoxaben. Quinclorac does not seem to act primarily as an inhibitor of cellulose biosynthesis. PMID:18408242

  19. DBIO Best Thesis Award: Mechanics, Dynamics, and Organization of the Bacterial Cytoskeleton and Cell Wall

    NASA Astrophysics Data System (ADS)

    Wang, Siyuan

    2012-02-01

    Bacteria come in a variety of shapes. While the peptidoglycan (PG) cell wall serves as an exoskeleton that defines the static cell shape, the internal bacterial cytoskeleton mediates cell shape by recruiting PG synthesis machinery and thus defining the pattern of cell-wall synthesis. While much is known about the chemistry and biology of the cytoskeleton and cell wall, much of their biophysics, including essential aspects of the functionality, dynamics, and organization, remain unknown. This dissertation aims to elucidate the detailed biophysical mechanisms of cytoskeleton guided wall synthesis. First, I find that the bacterial cytoskeleton MreB contributes nearly as much to the rigidity of an Escherichia coli cell as the cell wall. This conclusion implies that the cytoskeletal polymer MreB applies meaningful force to the cell wall, an idea favored by theoretical modeling of wall growth, and suggests an evolutionary origin of cytoskeleton-governed cell rigidity. Second, I observe that MreB rotates around the long axis of E. coli, and the motion depends on wall synthesis. This is the first discovery of a cell-wall assembly driven molecular motor in bacteria. Third, I prove that both cell-wall synthesis and the PG network have chiral ordering, which is established by the spatial pattern of MreB. This work links the molecular structure of the cytoskeleton and of the cell wall with organismal-scale behavior. Finally, I develop a mathematical model of cytoskeleton-cell membrane interactions, which explains the preferential orientation of different cytoskeleton components in bacteria.

  20. Vesicles between plasma membrane and cell wall prior to visible senescence of Iris and Dendrobium flowers.

    PubMed

    Kamdee, Channatika; Kirasak, Kanjana; Ketsa, Saichol; van Doorn, Wouter G

    2015-09-01

    Cut Iris flowers (Iris x hollandica, cv. Blue Magic) show visible senescence about two days after full opening. Epidermal cells of the outer tepals collapse due to programmed cell death (PCD). Transmission electron microscopy (TEM) showed irregular swelling of the cell walls, starting prior to cell collapse. Compared to cells in flowers that had just opened, wall thickness increased up to tenfold prior to cell death. Fibrils were visible in the swollen walls. After cell death very little of the cell wall remained. Prior to and during visible wall swelling, vesicles (paramural bodies) were observed between the plasma membrane and the cell walls. The vesicles were also found in groups and were accompanied by amorphous substance. They usually showed a single membrane, and had a variety of diameters and electron densities. Cut Dendrobium hybrid cv. Lucky Duan flowers exhibited visible senescence about 14 days after full flower opening. Paramural bodies were also found in Dendrobium tepal epidermis and mesophyll cells, related to wall swelling and degradation. Although alternative explanations are well possible, it is hypothesized that paramural bodies carry enzymes involved in cell wall breakdown. The literature has not yet reported such bodies in association with senescence/PCD.

  1. Traffic monitors at the cell periphery: the role of cell walls during early female reproductive cell differentiation in plants.

    PubMed

    Tucker, Matthew R; Koltunow, Anna M G

    2014-02-01

    The formation of female gametes in plants occurs within the ovule, a floral organ that is also the precursor of the seed. Unlike animals, plants lack a typical germline separated from the soma early in development and rely on positional signals, including phytohormones, mobile mRNAs and sRNAs, to direct diploid somatic precursor cells onto a reproductive program. In addition, signals moving between plant cells must overcome the architectural limitations of a cell wall which surrounds the plasma membrane. Recent studies have addressed the molecular and histological signatures of young ovule cells and indicate that dynamic cell wall changes occur over a short developmental window. These changes in cell wall properties impact signal flow and ovule cell identity, thereby aiding the establishment of boundaries between reproductive and somatic ovule domains.

  2. Changes in levels of cell wall constituents in wheat seedlings grown under continuous hypergravity conditions

    NASA Astrophysics Data System (ADS)

    Wakabayashi, K.; Soga, K.; Kamisaka, S.; Hoson, T.

    Effects of continuous hypergravity stimuli on the amounts and composition of cell wall constituents were investigated in wheat shoots. Hypergravity (300 g) treatment for three days after germination increased the net amount of cell wall polysaccharides such as hemicellulose and cellulose, but reduced the shoot elongation. As a result, the amount of cell wall polysaccharides per unit length of shoot increased under hypergravity. The hemicellulose fraction contained polysaccharides in the middle and low molecular mass range (5 kDa-1 MDa) and increased in response to hypergravity. Also, the amounts of arabinose (Ara) and xylose (Xyl), the major sugar components of the hemicellulose fraction, increased under hypergravity conditions. In addition to wall polysaccharides, hypergravity increased the amounts of cell wall-bound phenolic acids, such as ferulic acid (FA) and diferulic acid (DFA). Furthermore, the activity of phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) was enhanced under hypergravity conditions. These results suggest that continuous hypergravity stimulates the synthesis of cell wall constituents, especially hemicellulosic arabinoxylans and cell wall-bound FA and DFA in wheat shoots. The increased PAL activity may promote the formation of FA and DFA. These changes in cell wall architecture may be involved in making rigid and tough cell walls under hypergravity conditions and thereby contribute to the ability of plant to sustain their structures against gravitational stimuli.

  3. Toward one step analysis of cellular lipidomes using liquid chromatography coupled with mass spectrometry: application to Saccharomyces cerevisiae and Schizosaccharomyces pombe lipidomics.

    PubMed

    Shui, Guanghou; Guan, Xue Li; Low, Choon Pei; Chua, Gek Huey; Goh, Joyce Sze Yuin; Yang, Hongyuan; Wenk, Markus R

    2010-06-01

    Recent rapid growth of lipidomics is mainly attributed to technological advances in mass spectrometry. Development of soft ionization techniques, in combination with computational tools, has spurred subsequent development of various methods for lipid analysis. However, none of these existing approaches can cover major cellular lipids in a single run. Here we demonstrate that a single method of liquid chromatography coupled with mass spectrometry (LCMS) can be used for simultaneous profiling of major cellular lipids including glycerophospholipids (PLs), sphingolipids (SPLs), waxes, sterols (ST) and mono-, di- as well as triacylglycerides (MAG, DAG, TAG). We applied this approach to analyze these lipids in various organisms including Saccharomyces cerevisiae and Schizosaccharomyces pombe. While phospholipids and triacylglycerides of S. pombe mainly contain 18 : 1 fatty acyls, those of S. cerevisiae contain 16 : 1, 16 : 0 and 18 : 1 fatty acyls. S. cerevisiae and S. pombe contain distinct sphingolipid profiles. S. cerevisiae has abundant inositol phytoceramides (IPC), while S. pombe contains high levels of free phytoceramides as well as short chain phytoceramides (t18:1/20 : 0-B) and IPC (t18:1/20 : 0-B). In S. cerevisiae, our results demonstrated accumulation of ergosterol esters in tgl1Delta cells and accumulation of various TAG species in tgl3Delta cells, which are consistent with the function of the respective enzymes. Furthermore, we, for the first time, systematically characterized lipids in S. pombe and measured their dynamic changes in Deltaplh1Deltadga1 cells at different growth phases. We further discussed dynamic changes of phospholipids, sphingolipids and neutral lipids in the progress of programmed cell death in Deltaplh1Deltadga1 cells of S. pombe.

  4. The Cell Wall Protein Ecm33 of Candida albicans is Involved in Chronological Life Span, Morphogenesis, Cell Wall Regeneration, Stress Tolerance, and Host-Cell Interaction.

    PubMed

    Gil-Bona, Ana; Reales-Calderon, Jose A; Parra-Giraldo, Claudia M; Martinez-Lopez, Raquel; Monteoliva, Lucia; Gil, Concha

    2016-01-01

    Ecm33 is a glycosylphosphatidylinositol-anchored protein in the human pathogen Candida albicans. This protein is known to be involved in fungal cell wall integrity (CWI) and is also critical for normal virulence in the mouse model of hematogenously disseminated candidiasis, but its function remains unknown. In this work, several phenotypic analyses of the C. albicans ecm33/ecm33 mutant (RML2U) were performed. We observed that RML2U displays the inability of protoplast to regenerate the cell wall, activation of the CWI pathway, hypersensitivity to temperature, osmotic and oxidative stresses and a shortened chronological lifespan. During the exponential and stationary culture phases, nuclear and actin staining revealed the possible arrest of the cell cycle in RML2U cells. Interestingly, a "veil growth," never previously described in C. albicans, was serendipitously observed under static stationary cells. The cells that formed this structure were also observed in cornmeal liquid cultures. These cells are giant, round cells, without DNA, and contain large vacuoles, similar to autophagic cells observed in other fungi. Furthermore, RML2U was phagocytozed more than the wild-type strain by macrophages at earlier time points, but the damage caused to the mouse cells was less than with the wild-type strain. Additionally, the percentage of RML2U apoptotic cells after interaction with macrophages was fewer than in the wild-type strain.

  5. The Cell Wall Protein Ecm33 of Candida albicans is Involved in Chronological Life Span, Morphogenesis, Cell Wall Regeneration, Stress Tolerance, and Host–Cell Interaction

    PubMed Central

    Gil-Bona, Ana; Reales-Calderon, Jose A.; Parra-Giraldo, Claudia M.; Martinez-Lopez, Raquel; Monteoliva, Lucia; Gil, Concha

    2016-01-01

    Ecm33 is a glycosylphosphatidylinositol-anchored protein in the human pathogen Candida albicans. This protein is known to be involved in fungal cell wall integrity (CWI) and is also critical for normal virulence in the mouse model of hematogenously disseminated candidiasis, but its function remains unknown. In this work, several phenotypic analyses of the C. albicans ecm33/ecm33 mutant (RML2U) were performed. We observed that RML2U displays the inability of protoplast to regenerate the cell wall, activation of the CWI pathway, hypersensitivity to temperature, osmotic and oxidative stresses and a shortened chronological lifespan. During the exponential and stationary culture phases, nuclear and actin staining revealed the possible arrest of the cell cycle in RML2U cells. Interestingly, a “veil growth,” never previously described in C. albicans, was serendipitously observed under static stationary cells. The cells that formed this structure were also observed in cornmeal liquid cultures. These cells are giant, round cells, without DNA, and contain large vacuoles, similar to autophagic cells observed in other fungi. Furthermore, RML2U was phagocytozed more than the wild-type strain by macrophages at earlier time points, but the damage caused to the mouse cells was less than with the wild-type strain. Additionally, the percentage of RML2U apoptotic cells after interaction with macrophages was fewer than in the wild-type strain. PMID:26870022

  6. Glycoproteins from the cell wall of Phaseolus coccineus.

    PubMed

    O'Neill, M A; Selvendran, R R

    1980-04-01

    1. The use of a modified sodium chlorite/acetic acid delignification procedure for the solubilization of a hydroxyproline-rich glycoprotein fraction from the depectinated cell walls of Phaseolus coccineus is described. 2. The crude glycoprotein was associated with some pectic material; hydroxyproline and serine were the most abundant amino acids, and arabinose, galactose and galacturonic acid the predominant monosaccharides. 3. The bulk of the hydroxyproline is O-glycosidically substituted with tetra- and tri-arabinofuranosides. From methylation analysis the linkages in these arabinosides could be inferred. 4. Ion-exchange chromatography of the crude glycoprotein gave one major and two minor hydroxyproline-rich fractions, with similar amino acid but different monosaccharide composition. 5. In the major fraction, serine appears to be O-glycosidically substituted with a single galactopyranoside residue that can be removed by the action of alpha-galactosidase but not beta-galactosidase. Removal of arabinofuranoside residues by partial acid hydrolysis greatly enhanced the action of alpha-galactosidase. 6. Methylation followed by carboxy reduction with LiAl2H4 has shown the presence of (1 leads to 4)-linked galacturonic acid in the crude glycoprotein fraction but not in the major fraction from the ion-exchange column. Hence the bulk of the pectic material is not associated with the major glycoprotein component. It is suggested that the glycoprotein is held in the wall by phenolic cross-links. 7. Similarities with the glycopeptide moiety of potato lectin provides further evidence for a class of hydroxyproline-rich glycoproteins with common features.

  7. Construction of the first compendium of chemical-genetic profiles in the fission yeast Schizosaccharomyces pombe and comparative compendium approach

    SciTech Connect

    Han, Sangjo; Lee, Minho; Chang, Hyeshik; Nam, Miyoung; Park, Han-Oh; Kwak, Youn-Sig; Ha, Hye-jeong; Kim, Dongsup; Hwang, Sung-Ook; Hoe, Kwang-Lae; Kim, Dong-Uk

    2013-07-12

    Highlights: •The first compendium of chemical-genetic profiles form fission yeast was generated. •The first HTS of drug mode-of-action in fission yeast was performed. •The first comparative chemical genetic analysis between two yeasts was conducted. -- Abstract: Genome-wide chemical genetic profiles in Saccharomyces cerevisiae since the budding yeast deletion library construction have been successfully used to reveal unknown mode-of-actions of drugs. Here, we introduce comparative approach to infer drug target proteins more accurately using two compendiums of chemical-genetic profiles from the budding yeast S. cerevisiae and the fission yeast Schizosaccharomyces pombe. For the first time, we established DNA-chip based growth defect measurement of genome-wide deletion strains of S. pombe, and then applied 47 drugs to the pooled heterozygous deletion strains to generate chemical-genetic profiles in S. pombe. In our approach, putative drug targets were inferred from strains hypersensitive to given drugs by analyzing S. pombe and S. cerevisiae compendiums. Notably, many evidences in the literature revealed that the inferred target genes of fungicide and bactericide identified by such comparative approach are in fact the direct targets. Furthermore, by filtering out the genes with no essentiality, the multi-drug sensitivity genes, and the genes with less eukaryotic conservation, we created a set of drug target gene candidates that are expected to be directly affected by a given drug in human cells. Our study demonstrated that it is highly beneficial to construct the multiple compendiums of chemical genetic profiles using many different species. The fission yeast chemical-genetic compendium is available at (http://pombe.kaist.ac.kr/compendium)

  8. Endomembrane proteomics reveals putative enzymes involved in cell wall metabolism in wheat grain outer layers

    PubMed Central

    Chateigner-Boutin, Anne-Laure; Suliman, Muhtadi; Bouchet, Brigitte; Alvarado, Camille; Lollier, Virginie; Rogniaux, Hélène; Guillon, Fabienne; Larré, Colette

    2015-01-01

    Cereal grain outer layers fulfil essential functions for the developing seed such as supplying energy and providing protection. In the food industry, the grain outer layers called ‘the bran’ is valuable since it is rich in dietary fibre and other beneficial nutriments. The outer layers comprise several tissues with a high content in cell wall material. The cell wall composition of the grain peripheral tissues was investigated with specific probes at a stage of active cell wall synthesis. Considerable wall diversity between cell types was revealed. To identify the cellular machinery involved in cell wall synthesis, a subcellular proteomic approach was used targeting the Golgi apparatus where most cell wall polysaccharides are synthesized. The tissues were dissected into outer pericarp and intermediate layers where 822 and 1304 proteins were identified respectively. Many carbohydrate-active enzymes were revealed: some in the two peripheral grain fractions, others only in one tissue. Several protein families specific to one fraction and with characterized homologs in other species might be related to the specific detection of a polysaccharide in a particular cell layer. This report provides new information on grain cell walls and its biosynthesis in the valuable outer tissues, which are poorly studied so far. A better understanding of the mechanisms controlling cell wall composition could help to improve several quality traits of cereal products (e.g. dietary fibre content, biomass conversion to biofuel). PMID:25769308

  9. Polysaccharide composition of unlignified cell walls of pineapple [Ananas comosus (L.) Merr.] fruit.

    PubMed Central

    Smith, B G; Harris, P J

    1995-01-01

    The polysaccharides of cell walls isolated from the fleshy, edible part of the fruit of the monocotyledon pineapple [Ananas comosus (L.) Merr.] (family Bromeliaceae) were analyzed chemically. These cell walls were derived mostly from parenchyma cells and were shown histochemically to be unlignified, but they contained ester-linked ferulic acid. The analyses indicated that the noncellulosic polysaccharide composition of the cell walls was intermediate between that of unlignified cell walls of species of the monocotyledon family Poaceae (grasses and cereals) and that of unlignified cell walls of dicotyledons. Glucuronoarabinoxylans were the major non-cellulosic polysaccharides in the pineapple cell walls. Xyloglucans were also present, together with small amounts of pectic polysaccharides and glucomannans (or galactoglucomannans). The large amounts of glucuronoarabinoxylans and small amounts of pectic polysaccharides resemble the noncellulosic polysaccharide composition of the unlignified cell walls of the Poaceae. However, the absence of (1-->3,1-->4)-beta-glucans, the presence of relatively large amounts of xyloglucans, and the possible structure of the xyloglucans resemble the noncellulosic polysaccharide composition of the unlignified cell walls of dicotyledons. PMID:7770529

  10. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens.

    PubMed

    Berry, Elizabeth A; Tran, Mai L; Dimos, Christos S; Budziszek, Michael J; Scavuzzo-Duggan, Tess R; Roberts, Alison W

    2016-01-01

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants.

  11. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens.

    PubMed

    Berry, Elizabeth A; Tran, Mai L; Dimos, Christos S; Budziszek, Michael J; Scavuzzo-Duggan, Tess R; Roberts, Alison W

    2016-01-01

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants. PMID:27014284

  12. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens

    DOE PAGES

    Berry, Elizabeth A.; Tran, Mai L.; Dimos, Christos S.; Budziszek, Michael J.; Scavuzzo-Duggan, Tess R.; Roberts, Alison W.

    2016-03-08

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into severalmore » different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants.« less

  13. Immuno and Affinity Cytochemical Analysis of Cell Wall Composition in the Moss Physcomitrella patens

    PubMed Central

    Berry, Elizabeth A.; Tran, Mai L.; Dimos, Christos S.; Budziszek, Michael J.; Scavuzzo-Duggan, Tess R.; Roberts, Alison W.

    2016-01-01

    In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalactuonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants. PMID:27014284

  14. Autolysis of cell walls from polygalacturonase-antisense tomato fruit in simulated apoplastic solutions.

    PubMed

    Almeida, Domingos P F; Huber, Donald J

    2011-06-01

    Autolysis of cell walls from polygalacturonase (PG)-antisense tomato fruit was studied in a conventional buffer designed to maximize the catalytic activity of PG (30 mM sodium acetate, 150 mM NaCl, pH 4.5), and in solutions mimicking the pH and mineral composition of the fruit apoplast at the mature-green and ripe stages. Autolytic release of uronic acids was very limited under simulated apoplastic conditions compared with the conventional buffer, but minimal differences in the release of reducing groups were observed among the incubation conditions. Autolytic release of uronic acids from active walls was lower than solubilization from enzymically inactive walls. Uronic acids that remained ionically bound to the cell walls during autolysis were subsequently extracted and analyzed by size exclusion chromatography. The elution profiles of ionically bound uronic acids from cell walls incubated under optimal conditions were similar for all ripening stages. In solutions mimicking the pH and mineral composition of the apoplast of mature-green and ripe fruit, uronic acids extracted from pink and ripe fruit cell walls showed a decrease in average molecular mass compared with polymers from mature-green cell walls. The results suggest that the composition of the incubation solution exert strong influence on PG-independent cell wall autolysis and that enzymically active walls restrain PG-independent pectin solubilization.

  15. The architecture of the Schizosaccharomyces pombe CCR4-NOT complex.

    PubMed

    Ukleja, Marta; Cuellar, Jorge; Siwaszek, Aleksandra; Kasprzak, Joanna M; Czarnocki-Cieciura, Mariusz; Bujnicki, Janusz M; Dziembowski, Andrzej; Valpuesta, Jose M

    2016-01-01

    CCR4-NOT is a large protein complex present both in cytoplasm and the nucleus of eukaryotic cells. Although it is involved in a variety of distinct processes related to expression of genetic information such as poly(A) tail shortening, transcription regulation, nuclear export and protein degradation, there is only fragmentary information available on some of its nine subunits. Here we show a comprehensive structural characterization of the native CCR4-NOT complex from Schizosaccharomyces pombe. Our cryo-EM 3D reconstruction of the complex, combined with techniques such as immunomicroscopy, RNA-nanogold labelling, docking of the available high-resolution structures and models of different subunits and domains, allow us to propose its full molecular architecture. We locate all functionally defined domains endowed with deadenylating and ubiquitinating activities, the nucleus-specific RNA-interacting subunit Mmi1, as well as surfaces responsible for protein-protein interactions. This information provides insight into cooperation of the different CCR4-NOT complex functions.

  16. The architecture of the Schizosaccharomyces pombe CCR4-NOT complex

    PubMed Central

    Ukleja, Marta; Cuellar, Jorge; Siwaszek, Aleksandra; Kasprzak, Joanna M.; Czarnocki-Cieciura, Mariusz; Bujnicki, Janusz M.; Dziembowski, Andrzej; M. Valpuesta, Jose

    2016-01-01

    CCR4-NOT is a large protein complex present both in cytoplasm and the nucleus of eukaryotic cells. Although it is involved in a variety of distinct processes related to expression of genetic information such as poly(A) tail shortening, transcription regulation, nuclear export and protein degradation, there is only fragmentary information available on some of its nine subunits. Here we show a comprehensive structural characterization of the native CCR4-NOT complex from Schizosaccharomyces pombe. Our cryo-EM 3D reconstruction of the complex, combined with techniques such as immunomicroscopy, RNA-nanogold labelling, docking of the available high-resolution structures and models of different subunits and domains, allow us to propose its full molecular architecture. We locate all functionally defined domains endowed with deadenylating and ubiquitinating activities, the nucleus-specific RNA-interacting subunit Mmi1, as well as surfaces responsible for protein–protein interactions. This information provides insight into cooperation of the different CCR4-NOT complex functions. PMID:26804377

  17. Detection of 2 immunoreactive antigens in the cell wall of Sporothrix brasiliensis and Sporothrix globosa.

    PubMed

    Ruiz-Baca, Estela; Hernández-Mendoza, Gustavo; Cuéllar-Cruz, Mayra; Toriello, Conchita; López-Romero, Everardo; Gutiérrez-Sánchez, Gerardo

    2014-07-01

    The cell wall of members of the Sporothrix schenckii complex contains highly antigenic molecules which are potentially useful for the diagnosis and treatment of sporotrichosis. In this study, 2 immunoreactive antigens of 60 (Gp60) and 70 kDa (Gp70) were detected in the cell wall of the yeast morphotypes of Sporothrix brasiliensis and Sporothrix globosa.

  18. Changes in Cell Wall Polysaccharides of Green Bean Pods during Development

    PubMed Central

    Stolle-Smits, Trinette; Beekhuizen, Jan Gerard; Kok, Matthieu T.C.; Pijnenburg, Mary; Recourt, Kees; Derksen, Jan; Voragen, Alphons G.J.

    1999-01-01

    The changes in cell wall polysaccharides and selected cell wall-modifying enzymes were studied during the development of green bean (Phaseolus vulgaris L.) pods. An overall increase of cell wall material on a dry-weight basis was observed during pod development. Major changes were detected in the pectic polymers. Young, exponentially growing cell walls contained large amounts of neutral, sugar-rich pectic polymers (rhamnogalacturonan), which were water insoluble and relatively tightly connected to the cell wall. During elongation, more galactose-rich pectic polymers were deposited into the cell wall. In addition, the level of branched rhamnogalacturonan remained constant, while the level of linear homogalacturonan steadily increased. During maturation of the pods, galactose-rich pectic polymers were degraded, while the accumulation of soluble homogalacturonan continued. During senescence there was an increase in the amount of ionically complexed pectins, mainly at the expense of freely soluble pectins. The most abundant of the enzymes tested for was pectin methylesterase. Peroxidase, β-galactosidase, and α-arabinosidase were also detected in appreciable amounts. Polygalacturonase was detected only in very small amounts throughout development. The relationship between endogenous enzyme levels and the properties of cell wall polymers is discussed with respect to cell wall synthesis and degradation. PMID:10517827

  19. [Mechanism of manganese binding to leaf cell wall of Phytolacca americana L].

    PubMed

    Xu, Xiang-Hua; Liu, Cui-Ying; Li, Ping; Lang, Man; Zhao, Xiao-Yan; Yang, Jian-Jun; Gong, Min

    2015-02-01

    Phytolacca americana L. (P. americana) is a manganese (Mn) hyperaccumulator and cell wall plays an important role in the accumulation and detoxicity of Mn. We studied the impact of pH and Mn initial concentration on the binding of Mn by the leaf cell wall of P. americana, and explored the binding mechanisms by Fourier Transform Infrared Spectroscopy (FTIR) and synchrotron-based X-ray Absorption Fine Structure (XAFS) Spectroscopy. The results show that the optimum pH of Mn bingding for the leaf cell wall is between 5 and 6. The adsorption behavior of leaf cell wall can be described by Langmuir equation (R2 = 0.978 5) and the maximum adsorption of Mn on the leaf cell wall is 62.50 μmol x g(-1). Hydronyl and carbonyl groups are involved in the binding of Mn on the leaf cell wall. The Mn absorbed on the leaf cell wall is bonded by 6.3 oxygen around, and the bond length of Mn-O is 0.216 nm, which indicates the binding mechasnism of Mn to cell wall was inner-sphere complexation.

  20. Ethanol yields and cell wall properties in divergently bred switchgrass genotypes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetic modification of herbaceous plant cell walls to increase biofuels yields from harvested biomass is a primary bioenergy research goal. The focus of much of this research has been on cell wall lignin concentration. Using switchgrass genotypes developed by divergent breeding for ruminant diges...

  1. Distinct cell wall architectures in seed endosperms in representatives of the Brassicaceae and Solanaceae.

    PubMed

    Lee, Kieran J D; Dekkers, Bas J W; Steinbrecher, Tina; Walsh, Cherie T; Bacic, Antony; Bentsink, Leónie; Leubner-Metzger, Gerhard; Knox, J Paul

    2012-11-01

    In some species, a crucial role has been demonstrated for the seed endosperm during germination. The endosperm has been shown to integrate environmental cues with hormonal networks that underpin dormancy and seed germination, a process that involves the action of cell wall remodeling enzymes (CWREs). Here, we examine the cell wall architectures of the endosperms of two related Brassicaceae, Arabidopsis (Arabidopsis thaliana) and the close relative Lepidium (Lepidium sativum), and that of the Solanaceous species, tobacco (Nicotiana tabacum). The Brassicaceae species have a similar cell wall architecture that is rich in pectic homogalacturonan, arabinan, and xyloglucan. Distinctive features of the tobacco endosperm that are absent in the Brassicaceae representatives are major tissue asymmetries in cell wall structural components that reflect the future site of radicle emergence and abundant heteromannan. Cell wall architecture of the micropylar endosperm of tobacco seeds has structural components similar to those seen in Arabidopsis and Lepidium endosperms. In situ and biomechanical analyses were used to study changes in endosperms during seed germination and suggest a role for mannan degradation in tobacco. In the case of the Brassicaceae representatives, the structurally homogeneous cell walls of the endosperm can be acted on by spatially regulated CWRE expression. Genetic manipulations of cell wall components present in the Arabidopsis seed endosperm demonstrate the impact of cell wall architectural changes on germination kinetics.

  2. Silicon-induced changes in viscoelastic properties of sorghum root cell walls.

    PubMed

    Hattori, Taiichiro; Inanaga, Shinobu; Tanimoto, Eiichi; Lux, Alexander; Luxová, Miroslava; Sugimoto, Yukihiro

    2003-07-01

    Silicon is deposited in the endodermal tissue in sorghum (Sorghum bicolor L. Moench) roots. Its deposition is thought to protect vascular tissues in the stele against invasion by parasites and drying soil via hardening of endodermal cells. We studied the silicon-induced changes in mechanical properties of cell walls to clarify the role of silicon in sorghum root. Sorghum seedlings were grown in nutrient solution with or without silicon. The mechanical properties of cell walls were measured in three separated root zones: basal, apical and subapical. Silicon treatment decreased cell-wall extensibility in the basal zone of isolated stele tissues covered by endodermal inner tangential walls. The silicon-induced hardening of cell walls was also measured with increases in elastic moduli (E) and viscosity coefficients (eta). These results provided new evidence that silicon deposition might protect the stele as a mechanical barrier by hardening the cell walls of stele and endodermal tissues. In contrast to the basal zone, silicon treatment increased cell-wall extensibility in the apical and subapical zones with concomitant decrease in E and eta. Simultaneously, silicon promoted root elongation. When root elongation is promoted by silicon, one of the causal factors maybe the silicon-enhanced extensibility of cell walls in the growing zone.

  3. Experimental approaches to study plant cell walls during plant-microbe interactions

    PubMed Central

    Xia, Ye; Petti, Carloalberto; Williams, Mark A.; DeBolt, Seth

    2014-01-01

    Plant cell walls provide physical strength, regulate the passage of bio-molecules, and act as the first barrier of defense against biotic and abiotic stress. In addition to providing structural integrity, plant cell walls serve an important function in connecting cells to their extracellular environment by sensing and transducing signals to activate cellular responses, such as those that occur during pathogen infection. This mini review will summarize current experimental approaches used to study cell wall functions during plant-pathogen interactions. Focus will be paid to cell imaging, spectroscopic analyses, and metabolic profiling techniques. PMID:25352855

  4. Experimental approaches to study plant cell walls during plant-microbe interactions.

    PubMed

    Xia, Ye; Petti, Carloalberto; Williams, Mark A; DeBolt, Seth

    2014-01-01

    Plant cell walls provide physical strength, regulate the passage of bio-molecules, and act as the first barrier of defense against biotic and abiotic stress. In addition to providing structural integrity, plant cell walls serve an important function in connecting cells to their extracellular environment by sensing and transducing signals to activate cellular responses, such as those that occur during pathogen infection. This mini review will summarize current experimental approaches used to study cell wall functions during plant-pathogen interactions. Focus will be paid to cell imaging, spectroscopic analyses, and metabolic profiling techniques.

  5. Pectate chemistry links cell expansion to wall deposition in Chara corallina.

    PubMed

    Proseus, Timothy E; Boyer, John S

    2012-11-01

    Pectate (polygalacturonic acid) acts as a chelator to bind calcium and form cross-links that hold adjacent pectate polymers and thus plant cell walls together. When under tension from turgor pressure in the cell, the cross-links appear to distort and weaken. New pectate supplied by the cytoplasm is undistorted and removes wall calcium preferentially from the weakened bonds, loosening the wall and accelerating cell expansion. The new pectate now containing the removed calcium can bind to the wall, strengthening it and linking expansion to wall deposition. But new calcium needs to be added as well to replenish the calcium lost from the vacated wall pectate.  A recent report demonstrated that growth was disrupted if new calcium was unavailable.  The present addendum highlights this conclusion by reviewing an experiment from before the chelation chemistry was understood. Using cell wall labeling, a direct link appeared between wall expansion and wall deposition. Together, these experiments support the concept that newly supplied pectate has growth activity on its way to deposition in the wall. Growth rate is thus controlled by signals affecting the rate of pectate release. After release, the coordination of expansion and deposition arises naturally from chelation chemistry when polymers are under tension from turgor pressure. 

  6. Structure of Plant Cell Walls : XXVI. The Walls of Suspension-Cultured Sycamore Cells Contain a Family of Rhamnogalacturonan-I-Like Pectic Polysaccharides.

    PubMed

    Ishii, T; Thomas, J; Darvill, A; Albersheim, P

    1989-02-01

    Considerable information has been obtained about the primary structures of suspension-cultured sycamore (Acer pseudoplatanus) cell-wall pectic polysaccharides, i.e. rhamnogalacturonan I, rhamnogalacturonan II, and homogalacturonan. However, these polysaccharides, which are solubilized from the walls by endo-alpha-1,4-polygalacturonase, account for only about half of the pectic polysaccharides known to be present in sycamore cell walls. We now report that, after exhaustive treatment with endo-alpha-1,4-polygalacturonase, additional pectic polysaccharides were extracted from sycamore cell walls by treatment with Na(2)CO(3) at 1 and 22 degrees C. These previously uncharacterized polysaccharides accounted for approximately 4% of the cell wall. Based on the glycosyl and glycosyl-linkage compositions and the nature of the products obtained by treating the quantitatively predominant NaCO(3)-extracted polysaccharides with lithium metal dissolved in ethylenediamine, the polysaccharides were found to strongly resemble rhamnogalacturonan I. However, unlike rhamnogalacturonan I that characteristically had equal amounts of 2- and 2,4-linked rhamnosyl residues in its backbone, the polysaccharides extracted in Na(2)CO(3) at 1 degrees C had markedly disparate ratios of 2- to 2,4-linked rhamnosyl residues. We concluded that polysaccharides similar to rhamnogalacturonan I but with different degrees of branching are present in the walls of suspension-cultured sycamore cells.

  7. Measurements of cell wall mechanical properties using optically trapped fluorescent microspheres

    NASA Astrophysics Data System (ADS)

    Ermilov, Sergey; Qian, Feng; Murdock, David; Brownell, William E.; Anvari, Bahman

    2004-10-01

    Information on plasma membrane (PM) and cell wall mechanical properties is important for many biophysical applications, especially for those, which involve cells, undergoing significant mechanical stress (red blood cells, outer hair cells, fibrocytes, etc.). Optical tweezers is frequently used to study PM mechanics, particularly by pulling long PM tethers. One of the limitations on using optical tweezers to study cell wall mechanics is associated with transillumination technique of the trapped object position sensing, which prevents accurate mechanical testing in the proximity to the cell. In this work we use an optical tweezers in conjunction with a position-sensing system, which spectrally separates signals from the trapped fluorescent microsphere and imaging background. We have used this setup to study mechanics of the cell wall and PM separated from the underlying cytoskeleton on human embryonic kidney cells. We measured the force exerted by the cell on the trapped microsphere as a function of the cell wall displacement during the process of tether formation, and as a function of time during the process of tether growth and relaxation. Tethering force - cell wall displacement profiles have shown a behavior, implying that tether formation process starts with elastic deformation of the intact cell wall, followed by the plastic deformations and sliding of the PM over the underlying cytoskeleton, and ends with the local separation of a PM. Tethering force - cell wall displacement profiles have been used to estimate tether formation force, stiffness parameter of the cell wall and the works of tether formation, elastic and plastic deformations of the cell wall, related to the mechanical properties of a composite cell wall and cell wall - plasma membrane association strength. Temporal steady-state and relaxation tethering force profiles have been similar to the ones measured using transillumination position sensing, however average force values have been smaller in

  8. Histology and cell wall biochemistry of stone cells in the physical defence of conifers against insects.

    PubMed

    Whitehill, Justin G A; Henderson, Hannah; Schuetz, Mathias; Skyba, Oleksandr; Yuen, Macaire Man Saint; King, John; Samuels, A Lacey; Mansfield, Shawn D; Bohlmann, Jörg

    2016-08-01

    Conifers possess an array of physical and chemical defences against stem-boring insects. Stone cells provide a physical defence associated with resistance against bark beetles and weevils. In Sitka spruce (Picea sitchensis), abundance of stone cells in the cortex of apical shoots is positively correlated with resistance to white pine weevil (Pissodes strobi). We identified histological, biochemical and molecular differences in the stone cell phenotype of weevil resistant (R) or susceptible (S) Sitka spruce genotypes. R trees displayed significantly higher quantities of cortical stone cells near the apical shoot node, the primary site for weevil feeding. Lignin, cellulose, xylan and mannan were the most abundant components of stone cell secondary walls, respectively. Lignin composition of stone cells isolated from R trees contained a higher percentage of G-lignin compared with S trees. Transcript profiling revealed higher transcript abundance in the R genotype of coumarate 3-hydroxylase, a key monolignol biosynthetic gene. Developing stone cells in current year apical shoots incorporated fluorescent-tagged monolignol into the secondary cell wall, while mature stone cells of previous year apical shoots did not. Stone cell development is an ephemeral process, and fortification of shoot tips in R trees is an effective strategy against insect feeding.

  9. Degradation of 14C-labeled streptococcal cell walls by egg white lysozyme and lysosomal enzymes.

    PubMed Central

    Gallis, H A; Miller, S E; Wheat, R W

    1976-01-01

    The resistance of native and trypsin-treated [14C] glucose-labeled cell walls to degradation by lysozyme and human lysosomal enzymes was confirmed. In contrast, chemically N-acetylated cell walls undergo significant degradation by these enzymes in the pH range of 4.5 to 5.5 without prior removal of the group-specific carbohydrate. N-acetylation after removal of the group A carbohydrate by formamide extraction renders the cell walls considerably more susceptible to these enzymes than by formamaide extraction alone. It appears, therefore, that unless N-acetylation can occur in vivo, streptococcal cell walls are minimally degraded, if at all, by human peripheral blood leukocytes or lysozyme. Examination of leukocyte extracts from normal subjects and patients with post-streptococcal syndromes revealed no qualitative differences in ability to dissolve streptococcal cell walls. Images PMID:773836

  10. Imaging of polysaccharides in the tomato cell wall with Raman microspectroscopy

    PubMed Central

    2014-01-01

    Background The primary cell wall of fruits and vegetables is a structure mainly composed of polysaccharides (pectins, hemicelluloses, cellulose). Polysaccharides are assembled into a network and linked together. It is thought that the percentage of components and of plant cell wall has an important influence on mechanical properties of fruits and vegetables. Results In this study the Raman microspectroscopy technique was introduced to the visualization of the distribution of polysaccharides in cell wall of fruit. The methodology of the sample preparation, the measurement using Raman microscope and multivariate image analysis are discussed. Single band imaging (for preliminary analysis) and multivariate image analysis methods (principal component analysis and multivariate curve resolution) were used for the identification and localization of the components in the primary cell wall. Conclusions Raman microspectroscopy supported by multivariate image analysis methods is useful in distinguishing cellulose and pectins in the cell wall in tomatoes. It presents how the localization of biopolymers was possible with minimally prepared samples. PMID:24917885

  11. Non-invasive imaging of cellulose microfibril orientation within plant cell walls by polarized Raman microspectroscopy.

    PubMed

    Sun, Lan; Singh, Seema; Joo, Michael; Vega-Sanchez, Miguel; Ronald, Pamela; Simmons, Blake A; Adams, Paul; Auer, Manfred

    2016-01-01

    Cellulose microfibrils represent the major scaffold of plant cell walls. Different packing and orientation of the microfibrils at the microscopic scale determines the macroscopic properties of cell walls and thus affect their functions with a profound effect on plant survival. We developed a polarized Raman microspectroscopic method to determine cellulose microfibril orientation within rice plant cell walls. Employing an array of point measurements as well as area imaging and subsequent Matlab-assisted data processing, we were able to characterize the distribution of cellulose microfibril orientation in terms of director angle and anisotropy magnitude. Using this approach we detected differences between wild type rice plants and the rice brittle culm mutant, which shows a more disordered cellulose microfibril arrangement, and differences between different tissues of a wild type rice plant. This novel non-invasive Raman imaging approach allows for quantitative assessment of cellulose fiber orientation in cell walls of herbaceous plants, an important advancement in cell wall characterization.

  12. Amyloid-like properties of Saccharomyces cerevisiae cell wall glucantransferase Bgl2p

    PubMed Central

    Plotnikova, Tatyana A; Gorkovskii, Anton A; Selyakh, Irina O; Galzitskaya, Oxana V; Bezsonov, Evgeniy E; Gellissen, Gerd; Kulaev, Igor S

    2008-01-01

    Glucantransferase Bgl2p is a major conserved cell wall constituent described for a wide range of yeast species. In the baker's yeast Saccharomyces cerevisiae it is the only non-covalently bound cell wall protein that cannot be released from cell walls by sequential SDS and trypsin treatment. It contains seven amyloidogenic determinants. Circular dichroism analysis and fluorescence spectroscopy with thioflavin T indicate the presence of β-sheet structures in Bgl2p isolates. Bgl2p forms fibrils, a process that is enforced in the presence of other cell wall components. Thus the data obtained is the first evidence for amyloid-like properties of yeast cell wall protein—glucantransferase Bgl2p. PMID:19098439

  13. Clinostation influence on regeneration of cell wall in Solanum Tuberosum L. protoplasts

    NASA Astrophysics Data System (ADS)

    Nedukha, Elena M.; Sidorov, V. A.; Samoylov, V. M.

    1994-08-01

    Regeneration of cell walls in protoplasts was investigated using light- and electronmicroscopic methods. The protoplasts were isolated from mesophyll of Solanum tuberosum leaves and were cultivated on the horizontal low rotating clinostat (2 rpm) and in control for 10 days. Using a fluorescent method (with Calcofluor white) it was demonstrated that changes in vector gravity results in an regeneration inhibition of cell wall. With electron-microscopical and electro-cytochemical methods (staining with alcianum blue) dynamics of the regeneration of cell walls in protoplasts was studied; carbohydrate matrix of cell walls is deposited at the earliest stages of this process. The influence of microgravity on the cell wall regeneration is discussed in higher plants.

  14. Interactions between grape skin cell wall material and commercial enological tannins. Practical implications.

    PubMed

    Bautista-Ortín, Ana Belén; Cano-Lechuga, Mario; Ruiz-García, Yolanda; Gómez-Plaza, Encarna

    2014-01-01

    Commercial enological tannins were used to investigate the role that cell wall material plays in proanthocyanidin adsorption. Insoluble cell wall material, prepared from the skin of Vitis vinifera L. cv. Monastrell berries, was combined with solutions containing six different commercial enological tannins (proanthocyanidin-type tannins). Analysis of the proanthocyanidins in the solution, after fining with cell wall material, using phloroglucinolysis and size exclusion chromatography, provided quantitative and qualitative information on the non-adsorbed compounds. Cell wall material showed strong affinity for the proanthocyanidins, one of the commercial tannins being bound up to 61% in the experiment. Comparison of the molecular mass distribution of the commercial enological tannins in solution, before and after fining, suggested that cell walls affinity for proanthocyanidins was more related with the proanthocyanidin molecular mass than with their percentage of galloylation. These interactions may have some enological implications, especially as regards the time of commercial tannins addition to the must/wine.

  15. Generation of hydroxyl radical in isolated pea root cell wall, and the role of cell wall-bound peroxidase, Mn-SOD and phenolics in their production.

    PubMed

    Kukavica, Biljana; Mojovic, Milos; Vuccinic, Zeljko; Maksimovic, Vuk; Takahama, Umeo; Jovanovic, Sonja Veljovic

    2009-02-01

    The hydroxyl radical produced in the apoplast has been demonstrated to facilitate cell wall loosening during cell elongation. Cell wall-bound peroxidases (PODs) have been implicated in hydroxyl radical formation. For this mechanism, the apoplast or cell walls should contain the electron donors for (i) H(2)O(2) formation from dioxygen; and (ii) the POD-catalyzed reduction of H(2)O(2) to the hydroxyl radical. The aim of the work was to identify the electron donors in these reactions. In this report, hydroxyl radical (.OH) generation in the cell wall isolated from pea roots was detected in the absence of any exogenous reductants, suggesting that the plant cell wall possesses the capacity to generate .OH in situ. Distinct POD and Mn-superoxide dismutase (Mn-SOD) isoforms different from other cellular isoforms were shown by native gel electropho-resis to be preferably bound to the cell walls. Electron paramagnetic resonance (EPR) spectroscopy of cell wall isolates containing the spin-trapping reagent, 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO), was used for detection of and differentiation between .OH and the superoxide radical (O(2)(-).). The data obtained using POD inhibitors confirmed that tightly bound cell wall PODs are involved in DEPMPO/OH adduct formation. A decrease in DEPMPO/OH adduct formation in the presence of H(2)O(2) scavengers demonstrated that this hydroxyl radical was derived from H(2)O(2). During the generation of .OH, the concentration of quinhydrone structures (as detected by EPR spectroscopy) increased, suggesting that the H(2)O(2) required for the formation of .OH in isolated cell walls is produced during the reduction of O(2) by hydroxycinnamic acids. Cell wall isolates in which the proteins have been denaturated (including the endogenous POD and SOD) did not produce .OH. Addition of exogenous H(2)O(2) again induced the production of .OH, and these were shown to originate from the Fenton reaction with tightly bound metal ions

  16. [Transfer of T-DNA from agrobacteria into plant cells through cell walls and membranes].

    PubMed

    Chumakov, M I

    2001-01-01

    Discusses probable routes of agrobacterial penetration through the plant integumental tissues, cell wall, and plant cell plasmodesma. Analyzes the contribution of extracellular structures of agrobacteria in penetration through barriers of a plant cell, primary contact (adhesion), and during DNA transfer from bacterial (E. coli, A. tumefaciens) to recipient (bacterial or plant) cells. Discusses the relationship between donor cell adhesion to recipient cell surface and the infectious and conjugation processes. Considers the probable role of piles in conjugative transfer of agrobacterial DNA through membranes of donor and recipient (bacterial and plant) cells. Analyzes the contribution of the plant cell cytoskeleton to T-DNA transfer. Suggests a model of transport of T-DNA-VirD2 complex and VirE2 proteins through independent channels consisting of vir-coded proteins. PMID:11236737

  17. Immunoprofiling reveals unique cell-specific patterns of wall epitopes in the expanding Arabidopsis stem.

    PubMed

    Hall, Hardy C; Cheung, Jingling; Ellis, Brian E

    2013-04-01

    The Arabidopsis inflorescence stem undergoes rapid directional growth, requiring massive axial cell-wall extension in all its tissues, but, at maturity, these tissues are composed of cell types that exhibit markedly different cell-wall structures. It is not clear whether the cell-wall compositions of these cell types diverge rapidly following axial growth cessation, or whether compositional divergence occurs at earlier stages in differentiation, despite the common requirement for cell-wall extensibility. To examine this question, seven cell types were assayed for the abundance and distribution of 18 major cell-wall glycan classes at three developmental stages along the developing inflorescence stem, using a high-throughput immunolabelling strategy. These stages represent a phase of juvenile growth, a phase displaying the maximum rate of stem extension, and a phase in which extension growth is ceasing. The immunolabelling patterns detected demonstrate that the cell-wall composition of most stem tissues undergoes pronounced changes both during and after rapid extension growth. Hierarchical clustering of the immunolabelling signals identified cell-specific binding patterns for some antibodies, including a sub-group of arabinogalactan side chain-directed antibodies whose epitope targets are specifically associated with the inter-fascicular fibre region during the rapid cell expansion phase. The data reveal dynamic, cell type-specific changes in cell-wall chemistry across diverse cell types during cell-wall expansion and maturation in the Arabidopsis inflorescence stem, and highlight the paradox between this structural diversity and the uniform anisotropic cell expansion taking place across all tissues during stem growth.

  18. Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae

    PubMed Central

    Chardwiriyapreecha, Soracom; Manabe, Kunio; Iwaki, Tomoko; Kawano-Kawada, Miyuki; Sekito, Takayuki; Lunprom, Siriporn; Akiyama, Koichi; Takegawa, Kaoru; Kakinuma, Yoshimi

    2015-01-01

    In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles. PMID:26083598

  19. Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae.

    PubMed

    Chardwiriyapreecha, Soracom; Manabe, Kunio; Iwaki, Tomoko; Kawano-Kawada, Miyuki; Sekito, Takayuki; Lunprom, Siriporn; Akiyama, Koichi; Takegawa, Kaoru; Kakinuma, Yoshimi

    2015-01-01

    In Saccharomyces cerevisiae, Avt3p and Avt4p mediate the extrusion of several amino acids from the vacuolar lumen into the cytosol. SpAvt3p of Schizosaccharomyces pombe, a homologue of these vacuolar amino acid transporters, has been indicated to be involved in spore formation. In this study, we confirmed that GFP-SpAvt3p localized to the vacuolar membrane in S. pombe. The amounts of various amino acids increased significantly in the vacuolar pool of avt3Δ cells, but decreased in that of avt3+-overexpressing avt3Δ cells. These results suggest that SpAvt3p participates in the vacuolar compartmentalization of amino acids in S. pombe. To examine the export activity of SpAvt3p, we expressed the avt3+ gene in S. cerevisiae cells. We found that the heterologously overproduced GFP-SpAvt3p localized to the vacuolar membrane in S. cerevisiae. Using the vacuolar membrane vesicles isolated from avt3+-overexpressing S. cerevisiae cells, we detected the export activities of alanine and tyrosine in an ATP-dependent manner. These activities were inhibited by the addition of a V-ATPase inhibitor, concanamycin A, thereby suggesting that the activity of SpAvt3p is dependent on a proton electrochemical gradient generated by the action of V-ATPase. In addition, the amounts of various amino acids in the vacuolar pools of S. cerevisiae cells were decreased by the overproduction of SpAvt3p, which indicated that SpAvt3p was functional in S. cerevisiae cells. Thus, SpAvt3p is a vacuolar transporter that is involved in the export of amino acids from S. pombe vacuoles.

  20. Differential recognition of plant cell walls by microbial xylan-specific carbohydrate-binding modules.

    PubMed

    McCartney, Lesley; Blake, Anthony W; Flint, James; Bolam, David N; Boraston, Alisdair B; Gilbert, Harry J; Knox, J Paul

    2006-03-21

    Glycoside hydrolases that degrade plant cell walls have complex molecular architectures in which one or more catalytic modules are appended to noncatalytic carbohydrate-binding modules (CBMs). CBMs promote binding to polysaccharides and potentiate enzymic hydrolysis. Although there are diverse sequence-based families of xylan-binding CBMs, these modules, in general, recognize both decorated and unsubstituted forms of the target polysaccharide, and thus the evolutionary rationale for this diversity is unclear. Using immunohistochemistry to interrogate the specificity of six xylan-binding CBMs for their target polysaccharides in cell walls has revealed considerable differences in the recognition of plant materials between these protein modules. Family 2b and 15 CBMs bind to xylan in secondary cell walls in a range of dicotyledon species, whereas family 4, 6, and 22 CBMs display a more limited capability to bind to secondary cell walls. A family 35 CBM, which displays more restricted ligand specificity against purified xylans than the other five protein modules, reveals a highly distinctive binding pattern to plant material including the recognition of primary cell walls of certain dicotyledons, a feature shared with CBM15. Differences in the specificity of the CBMs toward walls of wheat grain and maize coleoptiles were also evident. The variation in CBM specificity for ligands located in plant cell walls provides a biological rationale for the repertoire of structurally distinct xylan-binding CBMs present in nature, and points to the utility of these modules in probing the molecular architecture of cell walls.

  1. Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria

    PubMed Central

    Mistou, Michel-Yves; Sutcliffe, Iain C.; van Sorge, Nina M.

    2016-01-01

    The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall teichoic acids (WTA). The presence and chemical structure of many rhamnose-containing cell wall polysaccharides (RhaCWP) has sometimes been known for decades. In contrast to WTA, insight into the biosynthesis and functional role of RhaCWP has been lacking. Recent studies in human streptococcal and enterococcal pathogens have highlighted critical roles for these complex polysaccharides in bacterial cell wall architecture and pathogenesis. In this review, we provide an overview of the RhaCWP with regards to their biosynthesis, genetics and biological function in species most relevant to human health. We also briefly discuss how increased knowledge in this field can provide interesting leads for new therapeutic compounds and improve biotechnological applications. PMID:26975195

  2. Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria.

    PubMed

    Mistou, Michel-Yves; Sutcliffe, Iain C; van Sorge, Nina M

    2016-07-01

    The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall teichoic acids (WTA). The presence and chemical structure of many rhamnose-containing cell wall polysaccharides (RhaCWP) has sometimes been known for decades. In contrast to WTA, insight into the biosynthesis and functional role of RhaCWP has been lacking. Recent studies in human streptococcal and enterococcal pathogens have highlighted critical roles for these complex polysaccharides in bacterial cell wall architecture and pathogenesis. In this review, we provide an overview of the RhaCWP with regards to their biosynthesis, genetics and biological function in species most relevant to human health. We also briefly discuss how increased knowledge in this field can provide interesting leads for new therapeutic compounds and improve biotechnological applications.

  3. Cotton fiber tips have diverse morphologies and show evidence of apical cell wall synthesis

    PubMed Central

    Stiff , Michael R.; Haigler, Candace H.

    2016-01-01

    Cotton fibers arise through highly anisotropic expansion of a single seed epidermal cell. We obtained evidence that apical cell wall synthesis occurs through examining the tips of young elongating Gossypium hirsutum (Gh) and G. barbadense (Gb) fibers. We characterized two tip types in Gh fiber (hemisphere and tapered), each with distinct apical diameter, central vacuole location, and distribution of cell wall components. The apex of Gh hemisphere tips was enriched in homogalacturonan epitopes, including a relatively high methyl-esterified form associated with cell wall pliability. Other wall components increased behind the apex including cellulose and the α-Fuc-(1,2)-β-Gal epitope predominantly found in xyloglucan. Gb fibers had only one narrow tip type featuring characters found in each Gh tip type. Pulse-labeling of cell wall glucans indicated wall synthesis at the apex of both Gh tip types and in distal zones. Living Gh hemisphere and Gb tips ruptured preferentially at the apex upon treatment with wall degrading enzymes, consistent with newly synthesized wall at the apex. Gh tapered tips ruptured either at the apex or distantly. Overall, the results reveal diverse cotton fiber tip morphologies and support primary wall synthesis occurring at the apex and discrete distal regions of the tip. PMID:27301434

  4. Single-Wall Carbon Nanotube Anodes for Lithium Cells

    NASA Technical Reports Server (NTRS)

    Hepp, Aloysius F.; Raffaelle, Ryne; Gennett, Tom; Kumta, Prashant; Maranchi, Jeff; Heben, Mike

    2006-01-01

    In recent experiments, highly purified batches of single-wall carbon nanotubes (SWCNTs) have shown promise as superior alternatives to the graphitic carbon-black anode materials heretofore used in rechargeable thin-film lithium power cells. The basic idea underlying the experiments is that relative to a given mass of graphitic carbon-black anode material, an equal mass of SWCNTs can be expected to have greater lithium-storage and charge/discharge capacities. The reason for this expectation is that whereas the microstructure and nanostructure of a graphitic carbon black is such as to make most of the interior of the material inaccessible for intercalation of lithium, a batch of SWCNTs can be made to have a much more open microstructure and nanostructure, such that most of the interior of the material is accessible for intercalation of lithium. Moreover, the greater accessibility of SWCNT structures can be expected to translate to greater mobilities for ion-exchange processes and, hence, an ability to sustain greater charge and discharge current densities.

  5. Breakdown of Cell Wall Nanostructure in Dilute Acid Pretreated Biomass

    SciTech Connect

    Pingali, Sai Venkatesh; Urban, Volker S; Heller, William T; McGaughey, Joseph; O'Neill, Hugh Michael; Foston, Marcus B; Myles, Dean A A; Ragauskas, Arthur J; Evans, Barbara R

    2010-01-01

    The generation of bioethanol from lignocellulosic biomass holds great promise for renewable and clean energy production. A better understanding of the complex mechanisms of lignocellulose breakdown during various pretreatment methods is needed to realize this potential in a cost and energy efficient way. Here, we use small-angle neutron scattering (SANS) to characterize morphological changes in switchgrass lignocellulose across molecular to sub-micron length scales resulting from the industrially-relevant dilute acid pretreatment method. Our results demonstrate that dilute acid pretreatment increases the cross-sectional radius of the crystalline cellulose fibril. This change is accompanied by removal of hemicellulose and the formation of Rg ~ 135 lignin aggregates. The structural signature of smooth cell wall surfaces is observed at length scales larger than 1000 , and it remains remarkably invariable during pretreatment. This study elucidates the interplay of the different biomolecular components in the break down process of switchgrass by dilute acid pretreatment. The results are important for the development of efficient strategies of biomass to biofuel conversion.

  6. Modifications of Saccharomyces pastorianus cell wall polysaccharides with brewing process.

    PubMed

    Bastos, Rita; Coelho, Elisabete; Coimbra, Manuel A

    2015-06-25

    The cell wall polysaccharides of brewers spent yeast Saccharomyces pastorianus (BSY) and the inoculum yeast (IY) were studied in order to understand the changes induced by the brewing process. The hot water and alkali extractions performed solubilized mainly mannoproteins, more branched for BSY than those of IY. Also, (31)P solid state NMR showed that the BSY mannoproteins were 3 times more phosphorylated. By electron microscopy it was observed that the final residues of alkali sequential extraction until 4M KOH preserved the yeast three-dimensional structure. The final residues, composed mainly by glucans (92%), showed that the BSY, when compared with IY, contained higher amount of (1→4)-linked Glc (43% for BSY and 16% for IY) and lower (1→3)-linked Glc (17% for BSY and 42% for IY). The enzymatic treatment of final residue showed that both BSY and IY had (α1→4)-linked Glc and (β1→4)-linked Glc, in a 2:1 ratio, showing that S. pastorianus increases their cellulose-like linkages with the brewing process.

  7. Deciphering the Transcriptional-Regulatory Network of Flocculation in Schizosaccharomyces pombe

    PubMed Central

    Kwon, Eun-Joo Gina; Laderoute, Amy; Chatfield-Reed, Kate; Vachon, Lianne; Karagiannis, Jim; Chua, Gordon

    2012-01-01

    In the fission yeast Schizosaccharomyces pombe, the transcriptional-regulatory network that governs flocculation remains poorly understood. Here, we systematically screened an array of transcription factor deletion and overexpression strains for flocculation and performed microarray expression profiling and ChIP–chip analysis to identify the flocculin target genes. We identified five transcription factors that displayed novel roles in the activation or inhibition of flocculation (Rfl1, Adn2, Adn3, Sre2, and Yox1), in addition to the previously-known Mbx2, Cbf11, and Cbf12 regulators. Overexpression of mbx2+ and deletion of rfl1+ resulted in strong flocculation and transcriptional upregulation of gsf2+/pfl1+ and several other putative flocculin genes (pfl2+–pfl9+). Overexpression of the pfl+ genes singly was sufficient to trigger flocculation, and enhanced flocculation was observed in several combinations of double pfl+ overexpression. Among the pfl1+ genes, only loss of gsf2+ abrogated the flocculent phenotype of all the transcription factor mutants and prevented flocculation when cells were grown in inducing medium containing glycerol and ethanol as the carbon source, thereby indicating that Gsf2 is the dominant flocculin. In contrast, the mild flocculation of adn2+ or adn3+ overexpression was likely mediated by the transcriptional activation of cell wall–remodeling genes including gas2+, psu1+, and SPAC4H3.03c. We also discovered that Mbx2 and Cbf12 displayed transcriptional autoregulation, and Rfl1 repressed gsf2+ expression in an inhibitory feed-forward loop involving mbx2+. These results reveal that flocculation in S. pombe is regulated by a complex network of multiple transcription factors and target genes encoding flocculins and cell wall–remodeling enzymes. Moreover, comparisons between the flocculation transcriptional-regulatory networks of Saccharomyces cerevisiae and S. pombe indicate substantial rewiring of transcription factors and cis

  8. Turgor Pressure Moves Polysaccharides into Growing Cell Walls of Chara corallina

    PubMed Central

    PROSEUS, TIMOTHY E.; BOYER, JOHN S.

    2005-01-01

    • Background and Aims Plant growth involves pressure-driven cell enlargement generally accompanied by deposition of new cell wall. New polysaccharides are secreted by the plasma membrane but their subsequent entry into the wall is obscure. Therefore, polysaccharides and gold colloids of various sizes were presented to the inner wall face as though they were secreted by the plasma membrane. • Methods Primary cell walls were isolated from growing internodes of Chara corallina and one end was attached to a glass capillary. Solutions of dextran or suspensions of gold colloids were pushed into the lumen by oil in the capillary. The oil did not enter the wall, and the solution or suspension was pressed against the inner wall face, pressurized at various ‘artificial’ P (turgor pressure), and polymer or colloid movement through the wall was monitored. • Key Results Interstices in the wall matrix had a diameter of about 4·6 nm measured at high P with gold colloids. Small solute (0·8 nm) readily moved through these interstices unaffected by P. Dextrans of 3·5 nm diameter moved faster at higher P while dextran of 9 nm scarcely entered unless high P was present. Dextran of 11 nm did not enter unless P was above a threshold, and dextran of 27 nm did not enter at P as high as 0·5 MPa. The walls filtered the dextrans, which became concentrated against the inner wall face, and most polymer movement occurred after P stabilized and bulk flow ended. • Conclusions P created a steep gradient in concentration and mechanical force at the inner wall face that moved large polymers into small wall openings apparently by starting a polymer end or deforming the polymer mechanically at the inner wall face. This movement occurred at P generally accepted to extend the walls for growth. PMID:15760911

  9. Phosphatase activity on the cell wall of Fonsecaea pedrosoi.

    PubMed

    Kneipp, L F; Palmeira, V F; Pinheiro, A A S; Alviano, C S; Rozental, S; Travassos, L R; Meyer-Fernandes, J R

    2003-12-01

    The activity of a phosphatase was characterized in intact mycelial forms of Fonsecaea pedrosoi, a pathogenic fungus that causes chromoblastomycosis. At pH 5.5, this fungus hydrolyzed p-nitrophenylphosphate (p-NPP) to p-nitrophenol (p-NP) at a rate of 12.78 +/- 0.53 nmol p-NP per h per mg hyphal dry weight. The values of Vmax and apparent Km for p-NPP hydrolyses were measured as 17.89 +/- 0.92 nmol p-NP per h per mg hyphal dry weight and 1.57 +/- 0.26 mmol/l, respectively. This activity was inhibited at increased pH, a finding compatible with an acid phosphatase. The enzymatic activity was strongly inhibited by classical inhibitors of acid phosphatases such as sodium orthovanadate (Ki = 4.23 micromol/l), sodium molybdate (Ki = 7.53 micromol/l) and sodium fluoride (Ki = 126.78 micromol/l) in a dose-dependent manner. Levamizole (1 mmol/l) and sodium tartrate (10 mmol/l), had no effect on the enzyme activity. Cytochemical localization of the acid phosphatase showed electrondense cerium phosphate deposits on the cell wall, as visualized by transmission electron microscopy. Phosphatase activity in F. pedrosoi seems to be associated with parasitism, as sclerotic cells, which are the fungal forms mainly detected in chromoblastomycosis lesions, showed much higher activities than conidia and mycelia did. A strain of F. pedrosoi recently isolated from a human case of chromoblastomycosis also showed increased enzyme activity, suggesting that the expression of surface phosphatases may be stimulated by interaction with the host.

  10. Shared catalysis in virus entry and bacterial cell wall depolymerization

    PubMed Central

    Cohen, Daniel N.; Sham, Yuk Y.; Haugstad, Greg D.; Xiang, Ye; Rossmann, Michael G.; Anderson, Dwight L.; Popham, David L.

    2009-01-01

    Summary Bacterial virus entry and cell wall depolymerization require the breakdown of peptidoglycan (PG), the peptide cross-linked polysaccharide matrix that surrounds bacterial cells. Structural studies of lysostaphin, a PG lytic enzyme (autolysin), have suggested that residues in the active site facilitate hydrolysis, but a clear mechanism for this reaction has remained unsolved. The active site residues and a structural pattern of β-sheets are conserved among lysostaphin homologs (such as LytM of Staphylococcus aureus) and the C-terminal domain of gene product 13 (gp13), a protein at the tail tip of the Bacillus subtilis bacteriophage φ29. gp13 activity on PG and muropeptides was assayed using high performance liquid chromatography, and gp13 was found to be a D,D-endopeptidase that cleaved the peptide cross-link. Computational modeling of the B. subtilis cross-linked peptide into the gp13 active site suggested that Asp195 may facilitate scissile bond activation and His247 is oriented to mediate nucleophile generation. This is the first model of a Zn2+-metallopeptidase and its substrate to our knowledge. Residue Asp195 of gp13 was found to be critical for Zn2+-binding and catalysis by substitution mutagenesis with Ala or Cys. Circular dichroism and particle induced X-ray emission spectroscopy showed that the general protein folding and Zn2+-binding was maintained in the Cys mutant but reduced in the Ala mutant. These findings together support a model where the Asp195 and His247 in gp13 and homologous residues in the LytM and lysostaphin active sites facilitate hydrolysis of the peptide substrate that cross-links PG. Thus, these autolysins and phage entry enzymes have a shared chemical mechanism of action. PMID:19361422

  11. Loss of cell wall alpha(1–3) glucan affects Cryptococcus neoformans from ultrastructure to virulence

    PubMed Central

    Reese, Amy J.; Yoneda, Aki; Breger, Julia A.; Beauvais, Anne; Liu, Hong; Griffith, Cara L.; Bose, Indrani; Kim, Myoung-Ju; Skau, Colleen; Yang, Sarah; Sefko, Julianne A.; Osumi, Masako; Latge, Jean-Paul; Mylonakis, Eleftherios; Doering, Tamara L.

    2007-01-01

    SUMMARY Yeast cell walls are critical for maintaining cell integrity, particularly in the face of challenges such as growth in mammalian hosts. The pathogenic fungus Cryptococcus neoformans additionally anchors its polysaccharide capsule to the cell surface via α(1–3) glucan in the wall. Cryptococcal cells disrupted in their alpha glucan synthase gene were sensitive to stresses, including temperature, and showed difficulty dividing. These cells lacked surface capsule, although they continued to shed capsule material into the environment. Electron microscopy showed that the alpha glucan that is usually localized to the outer portion of the cell wall was absent, the outer region of the wall was highly disorganized, and the inner region was hypertrophic. Analysis of cell wall composition demonstrated complete loss of alpha glucan accompanied by a compensatory increase in chitin/chitosan and a redistribution of beta glucan between cell wall fractions. The mutants were unable to grow in a mouse model of infection, but caused death in nematodes. These studies integrate morphological and biochemical investigations of the role of alpha glucan in the cryptococcal cell wall. PMID:17244196

  12. Sequence conservation of the rad21 Schizosaccharomyces pombe DNA double-strand break repair gene in human and mouse

    SciTech Connect

    McKay, M.J.; Troelstra, C.; Kanaar, R.

    1996-09-01

    The rad21 gene of Schizosaccharomyces pombe is involved in the repair of ionizing radiation-induced DNA double-strand breaks. The isolation of mouse and human putative homologs of rad21 is reported here. Alignment of the predicted amino acid sequence of Rad21 with the mammalian proteins showed that the similarity was distributed across the length of the proteins, with more highly conserved regions at both termini. The mHR21{sup sp} (mouse homolog of Rad21, S. pombe) and hHR21{sup sp} (human homolog of Rad21, S. pombe) predicted proteins were 96% identical, whereas the human and S. pombe proteins were 25% identical and 47% similar. RNA blot analysis showed that mHR21{sup sp} mRNA was abundant in all adult mouse tissues examined, with highest expression in testis and thymus. In addition to a 3.1-kb constitutive mRNA transcript, a 2.2-kb transcript was present at a high level in postmeiotic spermatids, while expression of the 3.1-kb mRNA in testis was confined to the meiotic compartment. hHR21{sup sp} mRNA was cell-cycle regulated in human cells, increasing in late S phase to a peak in G2 phase. The level of hHR21{sup sp} transcripts was not altered by exposure of normal diploid fibroblasts to 10 Gy ionizing radiation. In situ hybridization showed that mHR21{sup sp} resided on chromosome 15D3, whereas hHR21{sup sp} localized to the syntenic 8q24 region. Elevated expression of mHR21{sup sp} in testis and thymus supports a possible role for the rad21 mammalian homologs in V(D)J and meiotic recombination, respectively. Cell cycle regulation of rad21, retained from S. pombe to human, is consistent with a conservation of function between S. pombe and human rad21 genes. 62 refs., 8 figs., 1 tab.

  13. Wall Extensibility and Cell Hydraulic Conductivity Decrease in Enlarging Stem Tissues at Low Water Potentials 1

    PubMed Central

    Nonami, Hiroshi; Boyer, John S.

    1990-01-01

    Measurements with a guillotine psychrometer (H Nonami, JS Boyer [1990] Plant Physiol 94: 1601-1609) indicate that the inhibition of stem growth at low water potentials (low ψw) is accompanied by decreases in cell wall extensibility and tissue hydraulic conductance to water that eventually limit growth rate in soybean (Glycine max L. Merr.). To check this conclusion, we measured cell wall properties and cell hydraulic conductivities with independent techniques in soybean seedlings grown and treated the same way, i.e. grown in the dark and exposed to low ψw by transplanting dark grown seedlings to vermiculite of low water content. Wall properties were measured with an extensiometer modified for intact plants, and conductances were measured with a cell pressure probe in intact plants. Theory was developed to relate the wall measurements to those with the psychrometer. In the elongation zone, the plastic deformability of the walls decreased when measured with the extensiometer while growth was inhibited at low ψw. It increased during a modest growth recovery. This behavior was the same as that for the wall extensibility observed previously with the psychrometer. Tissue that was killed before measurement with the extensiometer also showed a similar response, indicating that changes in wall extensibility represented changes in wall physical properties and not rates of wall biosynthesis. The elastic compliance (reciprocal of bulk elastic modulus) did not change in the elongating or mature tissue. The hydraulic conductivity of cortical cells decreased in the elongating tissue and increased slightly during growth recovery in a response similar to that observed with the psychrometer. We conclude that the plastic properties of the cell walls and the conductance of the cells to water were decreased at low ψw but that the elastic properties of the walls were of little consequence in this response. PMID:16667664

  14. A chimeric NST repressor has the potential to improve glucose productivity from plant cell walls.

    PubMed

    Iwase, Akira; Hideno, Akihiro; Watanabe, Keiji; Mitsuda, Nobutaka; Ohme-Takagi, Masaru

    2009-07-15

    Bioethanol might be produced more economically and with less ecological impact (with reduced exploitation of food crops) if we could increase the production of glucose from the cellulosic materials in plant cell walls. However, plant cell walls are relatively resistant to enzymatic and physicochemical hydrolysis and, therefore, it is necessary to develop methods for reducing such resistance. Changes in plant cell wall materials, by genetic engineering, that render them more easily hydrolyzable to glucose might be a valuable approach to this problem. We showed previously that, in Arabidopsis, NAC secondary wall thickening-promoting factor1 (NST1) and NST3 are key regulators of secondary wall formation. We report here that transgenic Arabidopsis plants that expressed a chimeric repressor derived from NST1 produced cell wall materials that were twice as susceptible to both enzymatic and physicochemical hydrolysis as those from wild-type plants. The yields of glucose from both fresh and dry biomass were increased in the chimeric repressor lines. Use of the NST1 chimeric repressor might enhance production of glucose from plant cell walls, by changing the nature of the cell walls themselves.

  15. Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

    PubMed

    Wang, Yanting; Fan, Chunfen; Hu, Huizhen; Li, Ying; Sun, Dan; Wang, Youmei; Peng, Liangcai

    2016-01-01

    Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology.

  16. Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

    PubMed

    Wang, Yanting; Fan, Chunfen; Hu, Huizhen; Li, Ying; Sun, Dan; Wang, Youmei; Peng, Liangcai

    2016-01-01

    Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology. PMID:27269671

  17. Immunogold localization of xyloglucan and rhamnogalacturonan I in the cell walls of suspension-cultured sycamore cells.

    PubMed

    Moore, P