Protein expression changes during cotton fiber elongation in response to low temperature stress
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Keywords:
Expansin
Elongation
Anthesis
Q10
The cell wall not only provides basic skeleton to plant cell,but also controls the cell size,shape and growth.After mitosis,protoplast enlarges its size through absorbing water.In this phase,the cell wall is remodeled,with cellulose and other new wall materials synthesized and integrated.It finally causes cell growth.Phytohormones,such as IAA,GA,ET and BR,play vital roles in the process of cell expansion,which alters the expression and activity of cell wall-related enzymes such as cellulose synthase A(CESA),expansin(EXP) and xyloglucan endotran glucosylase /hydrolase(XET/XTH),and these factors regulate the cell wall expansion and finally promote cell growth.
Expansin
Xyloglucan
Protoplast
Tip growth
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Expansin
Turgor pressure
Plant cell
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We made use of EXLX1, an expansin from Bacillus subtilis, to investigate protein features essential for its plant cell wall binding and wall loosening activities. We found that the two expansin domains, D1 and D2, need to be linked for wall extension activity and that D2 mediates EXLX1 binding to whole cell walls and to cellulose via distinct residues on the D2 surface. Binding to cellulose is mediated by three aromatic residues arranged linearly on the putative binding surface that spans D1 and D2. Mutation of these three residues to alanine eliminated cellulose binding and concomitantly eliminated wall loosening activity measured either by cell wall extension or by weakening of filter paper but hardly affected binding to whole cell walls, which is mediated by basic residues located on other D2 surfaces. Mutation of these basic residues to glutamine reduced cell wall binding but not wall loosening activities. We propose domain D2 as the founding member of a new carbohydrate binding module family, CBM63, but its function in expansin activity apparently goes beyond simply anchoring D1 to the wall. Several polar residues on the putative binding surface of domain D1 are also important for activity, most notably Asp82, whose mutation to alanine or asparagine completely eliminated wall loosening activity. The functional insights based on this bacterial expansin may be extrapolated to the interactions of plant expansins with cell walls. We made use of EXLX1, an expansin from Bacillus subtilis, to investigate protein features essential for its plant cell wall binding and wall loosening activities. We found that the two expansin domains, D1 and D2, need to be linked for wall extension activity and that D2 mediates EXLX1 binding to whole cell walls and to cellulose via distinct residues on the D2 surface. Binding to cellulose is mediated by three aromatic residues arranged linearly on the putative binding surface that spans D1 and D2. Mutation of these three residues to alanine eliminated cellulose binding and concomitantly eliminated wall loosening activity measured either by cell wall extension or by weakening of filter paper but hardly affected binding to whole cell walls, which is mediated by basic residues located on other D2 surfaces. Mutation of these basic residues to glutamine reduced cell wall binding but not wall loosening activities. We propose domain D2 as the founding member of a new carbohydrate binding module family, CBM63, but its function in expansin activity apparently goes beyond simply anchoring D1 to the wall. Several polar residues on the putative binding surface of domain D1 are also important for activity, most notably Asp82, whose mutation to alanine or asparagine completely eliminated wall loosening activity. The functional insights based on this bacterial expansin may be extrapolated to the interactions of plant expansins with cell walls.
Expansin
Secondary cell wall
Alanine
Binding selectivity
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Growing plant cell walls characteristically exhibit a property known as 'acid growth', by which we mean they are more extensible at low pH (< 5) (1). The plant hormone auxin rapidly stimulates cell elongation in young stems and similar tissues at least in part by an acid-growth mechanism (2, 3). Auxin activates a H(+) pump in the plasma membrane, causing acidification of the cell wall solution. Wall acidification activates expansins, which are endogenous cell wall-loosening proteins (4), causing the cell wall to yield to the wall tensions created by cell turgor pressure. As a result, the cell begins to enlarge rapidly. This 'acid growth' phenomenon is readily measured in isolated (nonliving) cell wall specimens. The ability of cell walls to undergo acid-induced extension is not simply the result of the structural arrangement of the cell wall polysaccharides (e.g. pectins), but depends on the activity of expansins (5). Expansins do not have any known enzymatic activity and the only way to assay for expansin activity is to measure their induction of cell wall extension. This video report details the sources and preparation techniques for obtaining suitable wall materials for expansin assays and goes on to show acid-induced extension and expansin-induced extension of wall samples prepared from growing cucumber hypocotyls. To obtain suitable cell wall samples, cucumber seedlings are grown in the dark, the hypocotyls are cut and frozen at -80 degrees C. Frozen hypocotyls are abraded, flattened, and then clamped at constant tension in a special cuvette for extensometer measurements. To measure acid-induced extension, the walls are initially buffered at neutral pH, resulting in low activity of expansins that are components of the native cell walls. Upon buffer exchange to acidic pH, expansins are activated and the cell walls extend rapidly. We also demonstrate expansin activity in a reconstitution assay. For this part, we use a brief heat treatment to denature the native expansins in the cell wall samples. These inactivated cell walls do not extend even in acidic buffer, but addition of expansins to the cell walls rapidly restores their ability to extend.
Expansin
Turgor pressure
Elongation
Epicotyl
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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.
Turgor pressure
Expansin
Microfibril
Secondary cell wall
Pectin
Matrix (chemical analysis)
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The plant primary cell wall is a three-dimensional interwoven network of cellulose microfibrils, cross-linked by xyloglucan and dispersed in a pectin matrix. It has been suggested that in the wall of growing plant cells, xyloglucan is bound to the rigid cellulose microfibrils by hydrogen bonds and holds the microfibrils together by forming molecular tethers, which is referred to as the ‘sticky network’ model. Plant growth occurs when these tethers are peeled from the microfibrils by expansins or broken by glycosidases or transglycosylases. A number of researchers have presented theoretical difficulties and observations inconsistent with this model and a new hypothesis has been proposed, claiming that the cellulose – xyloglucan cross-links may act as ‘scaffolds’ holding the microfibrils apart. Analogies with synthetic polymers suggests that the spacing between the cellulose microfibrils may be an important determinant of the mechanical properties of the cell wall and the results presented in this thesis support this hypothesis. Water contents of Acetobacter xylinus synthesized cellulose based cell wall analogues (as a mimic of primary cell wall) and sunflower hypocotyl cell walls were altered using high molecular weight polyethylene glycol (PEG) solution, and their extension under a constant load was measured using a creep extensiometer and showed that there were clear reduction (30-35%) in extensibility suggesting that water content of the wall and therefore the cell wall free volume directly influence wall extensibility. When hydration of A. xylinus cellulose composite pellicles was reduced using PEG 6000 solution and re-hydrated in buffer solution, followed by treatment with ?-expansin or snail acetone powder extract, it was found that expansin and snail powder extracts caused a rapid rehydration of the composites and that the pellicles only returned to their original weights after these treatments, suggesting that expansin and snail powder can increase the free volume of the wall perhaps contributing to the increases in extensibility that they cause. Assays on cell wall fragments also indicated that expansin increased the cell wall free volume, demonstrated by changes of the turbidity of fragment suspensions. The role of pectic polysaccharide, RG-II, in cell wall biomechanics was also investigated using mechanical and biochemical testing of available Arabidopsis thaliana cell wall mutants and by incorporating RG-II (purified from red wine) with Acetobacter cellulose. It was demonstrated that RG-II significantly increased the hydration of cellulose composite; hydration rate was 15 -16% more than the composite without RG-II and thus increased the pellicle extensibility.
From the results, it is evidenced that cell wall extension is not only the consequences of breaking hydrogen bonds between cellulose microfibrils and xyloglucan by expansins or glycosidases and transglycosylases, but also a wider range of factors are involved including cell wall water content, cell wall free volume and the pectic polymers, especially RG-II.
Xyloglucan
Expansin
Turgor pressure
Bacterial Cellulose
Microfibril
Pectin
Secondary cell wall
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Abstract Plant cell growth is regulated through manipulation of the cell wall network, which consists of oriented cellulose microfibrils embedded within a ground matrix incorporating pectin and hemicellulose components. There remain many unknowns as to how this manipulation occurs. Experiments have shown that cellulose reorients in cell walls as the cell expands, while recent data suggest that growth is controlled by distinct collections of hemicellulose called biomechanical hotspots, which join the cellulose molecule together. The enzymes expansin and Cel12A have both been shown to induce growth of the cell wall; however, while Cel12A’s wall-loosening action leads to a reduction in the cell wall strength, expansin’s has been shown to increase the strength of the cell wall. In contrast, members of the XTH enzyme family hydrolyse hemicellulose but do not appear to cause wall creep. This experimentally observed behaviour still awaits a full explanation. We derive and analyse a mathematical model for the effective mechanical properties of the evolving cell wall network, incorporating cellulose microfibrils, which reorient with cell growth and are linked via biomechanical hotspots made up of regions of crosslinking hemicellulose. Assuming a visco-elastic response for the cell wall and using a continuum approach, we calculate the total stress resultant of the cell wall for a given overall growth rate. By changing appropriate parameters affecting breakage rate and viscous properties, we provide evidence for the biomechanical hotspot hypothesis and develop mechanistic understanding of the growth-inducing enzymes. Graphic Abstract
Hemicellulose
Expansin
Secondary cell wall
Xyloglucan
Turgor pressure
Microfibril
Plant cell
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Cell wall integrity is tightly regulated and maintained given that non-physiological modification of cell walls could render plants vulnerable to biotic and/or abiotic stresses. Expansins are plant cell wall-modifying proteins active during many developmental and physiological processes, but they can also be produced by bacteria and fungi during interaction with plant hosts. Cell wall alteration brought about by ectopic expression, overexpression, or exogenous addition of expansins from either eukaryote or prokaryote origin can in some instances provide resistance to pathogens, while in other cases plants become more susceptible to infection. In these circumstances altered cell wall mechanical properties might be directly responsible for pathogen resistance or susceptibility outcomes. Simultaneously, through membrane receptors for enzymatically released cell wall fragments or by sensing modified cell wall barrier properties, plants trigger intracellular signaling cascades inducing defense responses and reinforcement of the cell wall, contributing to various infection phenotypes, in which expansins might also be involved. Here, we review the plant immune response activated by cell wall surveillance mechanisms, cell wall fragments identified as responsible for immune responses, and expansin's roles in resistance and susceptibility of plants to pathogen attack.
Expansin
Cell membrane
Eukaryote
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Pectate lyase
Expansin
Xyloglucan
Pectin
Secondary cell wall
Arabinogalactan
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Plant cell enlargement is controlled by the ability of the constraining cell wall to expand. This ability has been postulated to be under the control of polysaccharide hydrolases or transferases that weaken or rearrange the loadbearing polymeric networks in the wall. We recently identified a family of wall proteins, called expansins, that catalyze the extension of isolated plant cell walls. Here we report that these proteins mechanically weaken pure cellulose paper in extension assays and stress relaxation assays, without detectable cellulase activity (exo- or endo- type). Because paper derives its mechanical strength from hydrogen bonding between cellulose microfibrils, we conclude that expansins can disrupt hydrogen bonding between cellulose fibers. This conclusion is further supported by experiments in which expansin-mediated wall extension (i) was increased by 2 M urea (which should weaken hydrogen bonding between wall polymers) and (ii) was decreased by replacement of water with deuterated water, which has a stronger hydrogen bond. The temperature sensitivity of expansin-mediated wall extension suggests that units of 3 or 4 hydrogen bonds are broken by the action of expansins. In the growing cell wall, expansin action is likely to catalyze slippage between cellulose microfibrils and the polysaccharide matrix, and thereby catalyze wall stress relaxation, followed by wall surface expansion and plant cell enlargement.
Expansin
Biopolymer
Secondary cell wall
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Citations (564)