Accelerating forward genetics for cell wall deconstruction

The elucidation of the genes involved in cell wall synthesis and assembly remains one of the biggest challenges of cell wall biology. Although traditional genetic approaches, using simple yet elegant screens, have identified components of the cell wall, many unknowns remain. Exhausting the genetic toolbox by performing sensitized screens, adopting chemical genetics or combining these with improved cell wall imaging, hold the promise of new gene discovery and function. With the recent introduction of next-generation sequencing technologies, it is now possible to quickly and efficiently map and clone genes of interest in record time. The combination of a classical genetics approach and cutting edge technology will propel cell wall biology in plants forward into the future

Melamine adsorption on carbon materials: impact of carbon texture and surface chemistry

In this work, a comparative study between three carbon materials has been carried out to investigate the impact of the micro/mesoporous structure of the carbon substrate on their adsorption capabilities. The study included two commercial carbons: Darco KB-G (AC), and Vulcan XC-72R (VC). The third carbon material was a mesoporous material (MC), with tailored micro/mesoporous structure and surface area obtained by carbonization of a resorcinol?formaldehyde (RF) polymer gel using both soft and hard template materials. Melamine was used as a model adsorbate in both acid and alkaline solutions. For all carbons, melamine adsorption was found to be pH dependent with higher adsorption from alkaline solutions than from acidic solutions. To the best of our knowledge, these are the first reported values for the adsorption of melamine to these carbon materials. Adsorption data obtained using the Langmuir model were compared with theoretical studies involving melamine as a building block in the self-assembly of molecular structures on carbon substrates, and analyzed using the results of several characterization studies carried out as part of this research work, some of which include nitrogen and CO2 adsorption isotherms, Raman spectroscopy, powder X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), and transmission electron microscopy (TEM).Fil: Hynes, Lucas. University of Ontario Institute of Technology; CanadáFil: Montiel, Gonzalo. Instituto Nacional de Tecnología Industrial; Argentina. Comisión Nacional de Energía Atómica. Centro Atómico Constituyentes; ArgentinaFil: Jones, Allison. University Of Ontario Institute Of Technology; CanadáFil: Riel, Donna. University Of Ontario Institute Of Technology; CanadáFil: Abdulaziz, Muna. University Of Ontario Institute Of Technology; CanadáFil: Viva, Federico Andrés. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Ciudad Universitaria. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia - Nodo Constituyentes | Comision Nacional de Energia Atomica. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia. Unidad Ejecutora Instituto de Nanociencia y Nanotecnologia - Nodo Constituyentes.; Argentina. Comisión Nacional de Energía Atómica. Gerencia de Área Investigaciones y Aplicaciones No Nucleares. Gerencia Física (CAC). Departamento de Física de la Materia Condensada; ArgentinaFil: Bonetta, Dario. University Of Ontario Institute Of Technology; CanadáFil: Vreugdenhil, Andrew. Trent University; CanadáFil: Trevani, Liliana. University Of Ontario Institute Of Technology; Canad

An analysis of ERA1 function in Arabidopsis

grantor: University of TorontoThe elaboration of multicellular structures is a complex balance between cell division, growth and differentiation. At the molecular level, this balance is maintained by a dynamic array of regulatory mechanisms which are tightly coordinated. This coordination is partly achieved through the post-translational modification of proteins by farnesyl lipid attachment. Since its initial description, protein farnesylation has figured an important mechanism for modifying protein function either by facilitating protein interaction with cell membranes or by mediating protein-protein interaction. Although a great deal of information has accumulated on the characteristics of the farnesyltransferases which catalyze the attachment of farnesyl to various proteins, the diversity of proteins that are post-translationally farnesylated has made it difficult to assess the utility of this type of modification in regulating cellular events. The common thread linking many farnesylated proteins, however, is their involvement in cell division and growth. In this regard, this study describes the isolation of an 'Arabidopsis' nucleosome assembly protein (NAP1) which is farnesylated and which interacts with ' Arabidopsis' cyclin B1. In addition, the analysis of 'era1 ' of 'Arabidopsis', which is currently the only farnesyltransferase loss-of-function mutant of any multicellular organism, has afforded the unique opportunity of assessing the role that farnesylation plays in regulating various processes during the development of a multicellular organism. This characterization indicates that farnesyltransferase activity is required for proper coordination of cell division and differentiation during 'Arabidopsis' vegetative and reproductive development. Loss of farnesylation affects both apical and axillary meristem development and these phenotypes are contingent on plant growth conditions. Farnesylation appears to negatively regulate cell-cycle gene expression in the lower central core of apical meristems. This region has been identified previously by both genetic and molecular studies to be essential for overall meristem organization. It appears farnesylation also plays a role in the organization of this center.Ph.D

With an Ear Up against the Wall: An Update on Mechanoperception in Arabidopsis

Cells interpret mechanical signals and adjust their physiology or development appropriately. In plants, the interface with the outside world is the cell wall, a structure that forms a continuum with the plasma membrane and the cytoskeleton. Mechanical stress from cell wall damage or deformation is interpreted to elicit compensatory responses, hormone signalling, or immune responses. Our understanding of how this is achieved is still evolving; however, we can refer to examples from animals and yeast where more of the details have been worked out. Here, we provide an update on this changing story with a focus on candidate mechanosensitive channels and plasma membrane-localized receptors

Characterization of pellicle inhibition in Gluconacetobacter xylinus 53582 by a small molecule, pellicin, identified by a chemical genetics screen.

Pellicin ([2E]-3-phenyl-1-[2,3,4,5-tetrahydro-1,6-benzodioxocin-8-yl]prop-2-en-1-one) was identified in a chemical genetics screen of 10,000 small molecules for its ability to completely abolish pellicle production in Gluconacetobacter xylinus. Cells grown in the presence of pellicin grew 1.5 times faster than untreated cells. Interestingly, growth in pellicin also caused G. xylinus cells to elongate. Measurement of cellulose synthesis in vitro showed that cellulose synthase activity was not directly inhibited by pellicin. Rather, when cellulose synthase activity was measured in cells that were pre-treated with the compound, the rate of cellulose synthesis increased eight-fold over that observed for untreated cells. This phenomenon was also apparent in the rapid production of cellulose when cells grown in the presence of pellicin were washed and transferred to media lacking the inhibitor. The rate at which cellulose was produced could not be accounted for by growth of the organism. Pellicin was not detected when intracellular contents were analyzed. Furthermore, it was found that pellicin exerts its effect extracellularly by interfering with the crystallization of pre-cellulosic tactoidal aggregates. This interference of the crystallization process resulted in enhanced production of cellulose II as evidenced by the ratio of acid insoluble to acid soluble product in in vitro assays and confirmed in vivo by scanning electron microscopy and powder X-ray diffraction. The relative crystallinity index, RCI, of pellicle produced by untreated G. xylinus cultures was 70% while pellicin-grown cultures had RCI of 38%. Mercerized pellicle of untreated cells had RCI of 42%, which further confirms the mechanism of action of pellicin as an inhibitor of the cellulose I crystallization process. Pellicin is a useful tool for the study of cellulose biosynthesis in G. xylinus

The FRIABLE1 Gene Product Affects Cell Adhesion in Arabidopsis

Cell adhesion in plants is mediated predominantly by pectins, a group of complex cell wall associated polysaccharides. An Arabidopsis mutant, friable1 (frb1), was identified through a screen of T-DNA insertion lines that exhibited defective cell adhesion. Interestingly, the frb1 plants displayed both cell and organ dissociations and also ectopic defects in organ separation. The FRB1 gene encodes a Golgi-localized, plant specific protein with only weak sequence similarities to known proteins (DUF246). Unlike other cell adhesion deficient mutants, frb1 mutants do not have reduced levels of adhesion related cell wall polymers, such as pectins. Instead, FRB1 affects the abundance of galactose- and arabinose-containing oligosaccharides in the Golgi. Furthermore, frb1 mutants displayed alteration in pectin methylesterification, cell wall associated extensins and xyloglucan microstructure. We propose that abnormal FRB1 action has pleiotropic consequences on wall architecture, affecting both the extensin and pectin matrices, with consequent changes to the biomechanical properties of the wall and middle lamella, thereby influencing cell-cell adhesion

Pellicin affects the cell length of <i>G. xylinus</i>.

<p>Box plot of cell lengths of <i>G. xylinus</i> grown in the presence and absence of pellicin. The box signifies the upper and lower quartiles for each growth condition and the median is represented by the black line within each box. Outliers are denoted by (○). Extreme outliers are denoted by (*). Cell lengths of 500 cells from each growth condition were measured (p<0.001, independent samples <i>t</i>-test).</p

Pellicin remains in the extracellular environment.

<p>High performance liquid chromatography of organic extracts of A) <i>G. xylinus</i> in the absence and B) presence of pellicin. Data shown is representative of triplicate experiments: (red dashed line) pellicin standard, (black line) extracellular supernatant, (blue line) pellet wash prior to cellulase treatment, (green line) supernatant from cellulase treated cells, (orange line) cell free lysate, (purple line) cellular debris analyzed at 220 nm, the optimum wavelength for pellicin detection.</p