Increased Pectin Methylesterase Activity Research Articles

Assessing the impact of isochoric freezing as a preservation method on the quality attributes of orange juice.

Isochoric (constant volume) freezing is a novel food preservation technology that has demonstrated the ability to preserve food products at subfreezing temperatures in an unfrozen state, thereby avoiding the detrimental effects of ice formation. It minimizes the quality loss of fresh fruits and juices, increases their nutrient content, and reduces microbial counts. Orange juice (OJ) samples were subjected to conventional freezing (CF) and isochoric freezing (IF) for 7 days and then stored at 4°C for an additional 7 days. We evaluated the microbiological and physicochemical quality of CF and IF OJ before and after storage. The IF was performed at three different conditions: -5°C/73MPa, -10°C/93MPa, and -15°C/143MPa. The results indicate that the total aerobic count of OJ remained below the detection limit after heat treatment, 7 days of CF and 7 days of IF. Yeast and mold counts increased in fresh and CF OJ after 7 days of storage at 4°C, whereas IF OJ remained below the detection limit. Less color difference was observed in IF (-15°C/143MPa) OJ compared to heat-treated and CF OJ. Heat treatment inactivated 42% of pectin methylesterase (PME), whereas 7-day long IF increased PME activity up to 150%. Additionally, IF (-15°C/143MPa) OJ showed reduced pulp sedimentation, which can be advantageous, as sedimentation in juices has been a recognized technological issue in the juice industry. Ascorbic acid level was significantly higher in IF (-15°C/143MPa) OJ compared to fresh and CF OJ after storage.

Epigenetic modification of a pectin methylesterase gene activates apoplastic iron reutilization in tomato roots.

Iron (Fe) distribution and reutilization are crucial for maintaining Fe homeostasis in plants. Here, we demonstrate that the tomato (Solanum lycopersicum) Colorless non-ripening (Cnr) epimutant exhibits increased Fe retention in cell wall pectin due to an increase in pectin methylesterase (PME) activity. This ultimately leads to Fe deficiency responses even under Fe-sufficient conditions when compared to the wild type (WT). Whole-genome bisulfite sequencing revealed that modifications to cell wall-related genes, especially CG hypermethylation in the intron region of PECTIN METHYLESTERASE53 (SlPME53), are involved in the Cnr response to Fe deficiency. When this intron hypermethylation of SlPME53 was artificially induced in WT, we found that elevated SlPME53 expression was accompanied by increased PME activity and increased pectin-Fe retention. The manipulation of SlPME53, either through overexpression in WT or knockdown in Cnr, influenced levels of pectin methylesterification and accumulation of apoplast Fe in roots. Moreover, CG hypermethylation mediated by METHYLTRANSFERASE1 (SlMET1) increased SlPME53 transcript abundance, resulting in greater PME activity and higher Fe retention in cell wall pectin. Therefore, we conclude that the Cnr mutation epigenetically modulates SlPME53 expression by SlMET1-mediated CG hypermethylation, and thus the capacity of the apoplastic Fe pool, creating opportunities for genetic improvement of crop mineral nutrition.

Aqueous ozone treatment inhibited degradation of cellwall polysaccharides in fresh-cut apple during cold storage

The effects of aqueous ozone (AO) treatment on the firmness, cellwall polysaccharide content, enzymatic activity, and gene expression of cell wall-degrading enzymes were investigated in fresh-cut apple during cold storage. AO treatment (1.4 mg L−1 for 5 min) delayed degradation of cellwall polysaccharides to reduce softening of fresh-cut apple. Water-soluble pectin content increased more slowly, while protopectin content, 4% KOH-soluble fraction, and cellulose content decreased at a lower rate in AO-treated fresh-cut apple compared with the control. However, no significant influence was shown on the 24% KOH-soluble fraction. AO treatment promoted increased pectin methylesterase activity, and distinctly inhibited β-galactosidase and α-arabinofuranosidase activities during storage. Polygalacturonase activity was not affected by AO treatment. Meanwhile, AO maintained the intactness of cell walls through repression of Mdβ-Gal and Mdα-Af transcript levels, thereby preserving the textural quality of fresh-cut apple during cold storage.

Pivotal role for root cell wall polysaccharides in cultivar-dependent cadmium accumulation in Brassica chinensis L.

Polysaccharides are the main components of plant cell walls in which they make an important contribution to cadmium (Cd) fixation. However, knowledge regarding the role of root cell wall polysaccharides in Cd accumulation in low-Cd cultivars is limited. Here, we compared the differences in root cell wall polysaccharides between two cultivars of Brassica chinensis L. (pakchoi) with different Cd accumulation abilities. A hydroponic experiment was conducted using low- (Huajun 2) and high-Cd (Hanlv) pakchoi cultivars. We investigated Cd subcellular distribution and Cd accumulation in cell wall polysaccharides and examined polysaccharide modifications in root cell walls by Fourier transform infrared spectroscopy. A Cd adsorption kinetics experiment was conducted to examine the connection between Cd-induced polysaccharide modifications and Cd fixation by cell walls. Amounts of Cd were significantly higher and more Cd was bound to cell walls in the roots of Huajun 2 than in those of Hanlv. These results indicated that the greater Cd retention capacity of the root cell wall in Huajun 2 accounted for the low Cd accumulation in the shoot. Up to 79.4% and 32.1% of cell-wall-bound Cd was found in the pectin and hemicellulose 1, respectively, and higher amounts of Cd were found in these cell wall components of Huajun 2 than in those of Hanlv. Exposure to Cd significantly increased amounts of pectin and hemicellulose 1 in both pakchoi cultivars, but the pectin levels were significantly higher in Huajun 2 than in Hanlv. Huajun 2 had higher pectin methylesterase (PME) activity and a lower degree of pectin methyl-esterification (DM) than Hanlv, although Cd treatments resulted in increased PME activity and decreased DM in both cultivars. The higher Cd treatment (44.5μM) resulted in enhanced Cd-binding capacity in root cell walls of the two cultivars with higher Cd adsorption levels in the root cell wall of Huajun 2. These results indicate that differences in the amount of cell wall polysaccharide and DM play key roles in establishing the genotypic differences underlying Cd accumulation in pakchoi. These findings conduce to a better understanding of the physiological mechanisms underlying low Cd accumulation in pakchoi and the breeding of new, low-Cd pakchoi cultivars.

The Auxin-Regulated CrRLK1L Kinase ERULUS Controls Cell Wall Composition during Root Hair Tip Growth

Root hairs facilitate a plant's ability to acquire soil anchorage and nutrients. Root hair growth is regulated by the plant hormone auxin and dependent on localized synthesis, secretion, and modification of the root hair tip cell wall. However, the exact cell wall regulators in root hairs controlled by auxin have yet to be determined. In this study, we describe the characterization of ERULUS (ERU), an auxin-induced Arabidopsis receptor-like kinase, whose expression is directly regulated by ARF7 and ARF19 transcription factors. ERU belongs to the Catharanthus roseus RECEPTOR-LIKE KINASE 1-LIKE (CrRLK1L) subfamily of putative cell wall sensor proteins. Imaging of a fluorescent fusion protein revealed that ERU is localized to the apical root hair plasma membrane. ERU regulates cell wall composition in root hairs and modulates pectin dynamics through negative control of pectin methylesterase (PME) activity. Mutant eru (-/-) root hairs accumulate de-esterified homogalacturonan and exhibit aberrant pectin Ca2+-binding site oscillations and increased PME activity. Up to 80% of the eru root hair phenotype is rescued by pharmacological supplementation with a PME-inhibiting catechin extract. ERU transcription is altered in specific cell wall-related root hair mutants, suggesting that it is a targetfor feedback regulation. Loss of ERU alters the phosphorylation status of FERONIA and H+-ATPases 1/2, regulators of apoplastic pH. Furthermore, H+-ATPases 1/2 and ERU are differentially phosphorylated in response to auxin. We conclude that ERULUS is a key auxin-controlled regulator of cell wall composition and pectin dynamics during root hair tip growth.

The Necrotrophic Fungus Macrophomina phaseolina Promotes Charcoal Rot Susceptibility in Grain Sorghum Through Induced Host Cell-Wall-Degrading Enzymes.

The cell-wall-degrading enzymes (CWDE) secreted by necrotrophs are important virulence factors. Although not unequivocally demonstrated, it has been suggested that necrotrophs induce hosts to cooperate in disease development through manipulation of host CWDE. The necrotrophic fungus Macrophomina phaseolina causes charcoal rot disease in Sorghum bicolor. An RNA-seq experiment was conducted to investigate the behavior of sorghum CWDE-encoding genes after M. phaseolina inoculation. Results revealed M. phaseolina's ability to significantly upregulate pectin methylesterase-, polygalacturonase-, cellulase-, endoglucanase-, and glycosyl hydrolase-encoding genes in a charcoal rot-susceptible sorghum genotype (Tx7000) but not in a resistant genotype (SC599). For functional validation, crude enzyme mixtures were extracted from M. phaseolina- and mock-inoculated charcoal-rot-resistant (SC599 and SC35) and -susceptible (Tx7000 and BTx3042) sorghum genotype stalks. A gel diffusion assay (pectin substrate) revealed significantly increased pectin methylesterase activity in M. phaseolina-inoculated Tx7000 and BTx3042. Polygalacturonase activity was determined using a ruthenium red absorbance assay (535 nm). Significantly increased polygalacturonase activity was observed in two susceptible genotypes after M. phaseolina inoculation. The activity of cellulose-degrading enzymes was determined using a 2-cyanoacetamide fluorimetric assay (excitation and emission maxima at 331 and 383 nm, respectively). The assay revealed significantly increased cellulose-degrading enzyme activity in M. phaseolina-inoculated Tx7000 and BTx3042. These findings revealed M. phaseolina's ability to promote charcoal rot susceptibility in grain sorghum through induced host CWDE.

Elevation of NO production increases Fe immobilization in the Fe-deficiency roots apoplast by decreasing pectin methylation of cell wall.

Cell wall is the major component of root apoplast which is the main reservoir for iron in roots, while nitric oxide (NO) is involved in regulating the synthesis of cell wall. However, whether such regulation could influence the reutilization of iron stored in root apoplast remains unclear. In this study, we observed that iron deficiency elevated NO level in tomato (Solanum lycopersicum) roots. However, application of S-nitrosoglutathione, a NO donor, significantly enhanced iron retention in root apoplast of iron-deficient plants, accompanied with a decrease of iron level in xylem sap. Consequently, S-nitrosoglutathione treatment increased iron concentration in roots, but decreased it in shoots. The opposite was true for the NO scavenging treatment with 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). Interestingly, S-nitrosoglutathione treatment increased pectin methylesterase activity and decreased degree of pectin methylation in root cell wall of both iron-deficient and iron-sufficient plants, which led to an increased iron retention in pectin fraction, thus increasing the binding capacity of iron to the extracted cell wall. Altogether, these results suggested that iron-deficiency-induced elevation of NO increases iron immobilization in root apoplast by decreasing pectin methylation in cell wall.

Novel roles of hydrogen peroxide (H₂O₂) in regulating pectin synthesis and demethylesterification in the cell wall of rice (Oryza sativa) root tips.

Hydrogen peroxide (H₂O₂) has been reported to increase lignin formation, enhance cell wall rigidification, restrict cell expansion and inhibit root elongation. However, our results showed that it not only inhibited rice (Oryza sativa) root elongation, but also increased root diameter. No study has reported how and why H₂O₂ increases cell expansion and root diameter. Exogenous H₂O₂ and its scavenger 4-hydroxy-Tempo were applied to confirm the roles of H₂O₂. Immunofluorescence, fluorescence probe, ruthenium red staining, histological section and spectrophotometry were used to monitor changes in the degree of pectin methylesterification, pectin content, pectin methylesterase (PME) activity and H₂O₂ content. Exogenous H₂O₂ inhibited root elongation, but increased cell expansion and root diameter significantly. H₂O₂ not only increased the region of pectin synthesis and pectin content in root tips, but also increased PME activity and pectin demethylesterification. The scavenger 4-hydroxy-Tempo reduced root H₂O₂ content and recovered H₂O₂-induced increases in cell expansion and root diameter by inhibiting pectin synthesis, PME activity and pectin demethylesterification. H₂O₂ plays a novel role in the regulation of pectin synthesis, PME activity and pectin demethylesterification. H₂O₂ increases cell expansion and root diameter by increasing pectin content and demethylesterification.

The effects of bruising and temperature on enzyme activity and textural qualities of tomato juice.

During harvest and transportation, processing tomatoes are exposed to elevated temperatures, compression and vibration in the harvester and truck, making them prone to bruising. The objective of this study was to determine how bruising and exposure to high temperatures affect pectin methylesterase (PME) activation and the textural quality of tomato juice. Tomatoes were both hand and mechanically harvested using current harvest practices. Mechanically harvested fruits were significantly softer, had greater PME activity and greater juice consistency than hand harvested fruits. In a controlled bruising study, whole tomatoes were exposed to various compressive forces at 21 or 40 °C and held for 0 or 4 h. Greater bruising force and higher temperature resulted in a decrease in firmness and an increase in PME activity. Consistency of tomato juice improved when tomatoes were exposed to 40 °C. Tomatoes subjected to a temperature range from 21 to 65 °C had activated PME at 40 °C and increased activity as temperature increased. Consistency increased at 35 °C but decreased with increasing temperature. Tomatoes harvested using current mechanical techniques are likely to be less firm and have increased PME activity; however, increased consistency of processed juice is observed. Tomatoes harvested at higher temperatures are also likely to have better consistency when processed.

Biochemical Characterization of Pectin Methylesterase from Musa acuminata Referring to Delayed Ripening

Several cell-wall degrading enzymes have been studied and their action involved in the softening concomitant with ripening of various fruits. However, studies pertaining to pectin methylesterase (PME) that catalyzes the demethylesterification of cell wall polygalacturonans have been very limited especially in fruit tissue. The present study covers the biochemical characterization of pectin methylesterase from Musa acuminata in context to delayed ripening phenomenon, principally dealing withidentification of a PME gene, namely, PME 1 from Grand Naineveriety of banana. The expression of this gene was well correlated with the ripening. After Ethylene treatment the expression of PME1 gene was peaked on day 3 while in control fruit, the progression of ripening was rather slow. The GA content was observed to augment significantly during the course of ripening of banana fruit. The overall data obtained provide new insights into correlation of the expression of PME1 with increased PME1 with increased PME activity as well as pectin content.

边缘细胞对荞麦根尖铝毒的防护效应和对细胞壁多糖的影响

Root border cells(RBCs) are special cells that surround the root apices and are physically separate from the root cap epidermis.The mRNAs and proteins produced by RBCs differ from those produced by the root cap cells.RBCs and the mucilage they secrete are thought to have roles in protecting root tips from biotic and abiotic stresses,such as aluminum toxicity.In this research,we aimed to elucidate the role of RBCs in protecting root apices of buckwheat(Fygopyrum esculentum Moench) from Al toxicity,and to determine their effect on polysaccharide contents in the cell wall of root tips.We used two cultivars of buckwheat: the Al-tolerant genotype 'Jiangxi' and the Al-sensitive genotype 'Neimeng'.Seedlings were aeroponically cultured in 0.5 mmol/L CaCl2 solution,pH 4.5,containing 0,25,50 or 200 μmol/L AlCl3.All of the primary roots were sprayed with AlCl3 solutions,and one half of the roots was washed with a stream of distilled water every 4 h to remove the RBCs that continuously formed during the experimental period.We evaluated root growth,Al accumulation in root tips,phosphatase(APA) and pectin methylesterase(PME) activities,and polysaccharide contents in the roots.In roots subjected to Al treatment,elongation was inhibited and there was increased Al accumulation in the tips.Physical removal of RBCs from root tips resulted in more severe Al-induced inhibition of root elongation and greater Al accumulation in the root tips.For both buckwheat genotypes,removal of RBCs from the roots resulted in higher APA activity in the root tips compared with that in root tips with RBCs.This effect was more pronounced in the Al-tolerant genotype(Jiangxi) than in the Al-sensitive genotype(Neimeng).Physical removal of RBCs from root tips also significantly increased PME activity and the contents of pectin,hemicellulose 1,and hemicellulose 2 in the cell walls of the root tips.These results suggested that RBCs adhered to the root tips tend to exclude Al from the rice root apex,decrease the cell-wall polysaccharide content,and promote synthesis of APA,thereby promoting Al tolerance in buckwheat.

Softening and cell wall metabolism in late-season peach in response to controlled atmosphere and 1-MCP treatment

SummarySoftening of peach (Prunus persica L. Batsch) fruit is generally rapid at ambient temperature, which limits considerably the commercial life of the produce. Modifications in the cell wall are believed to underlie changes in fruit firmness and texture. In this work, the effects of controlled atmosphere (CA) storage and 1-methylcyclopropene (1-MCP) on cell wall composition and on the activities of some cell wall-modifying enzymes were assessed in ‘Tardibelle’ fruit, a late-season, melting-flesh peach cultivar. CA storage and 1-MCP treatment applied separately were effective in delaying the softening of ‘Tardibelle’ peach during shelf-life at 20ºC following cold storage, apparently through different mechanisms. CA storage partially inhibited the activities of some pectolytic enzymes and increased the content of pectic polysaccharides, possibly related to delayed softening. 1-MCP treatment caused increased pectin methylesterase activity, but other pectolytic enzymes were inhibited, resulting in an enrichment of chelate-soluble pectin in the cell walls. In contrast, a combined treatment (i.e., CA plus 1-MCP) was unsatisfactory for retaining fruit firmness, apparently due to the extensive degradation of pectin.

Sequences Variation and Classification of B-Hordein Genes in Hull-Less Barley from the Qinghai-Tibet Plateau

Pectin methylesterase (PME), cell wall enzyme, is known as a factor of plant growth and morphogenesis. Recently, PME participation in gene silencing, plant virus reproduction and transgenesis was revealed. Here, the role of PME in antivirus resistance was studied. It has been shown that increasing PME enzymatic activity induced by additional PME gene and even empty vector resulted in suppression of tobacco mosaic virus (TMV) reproduction including short- and long-distance virus movement in plant. We revealed increased PME activity in stably-transformed intact plants. For example, transgenic tobacco and N. benthamiana plants expressing TMV movement protein gene and GFP appeared increased PME activity. The tomato plants co-suppressing polygalacturonase gene have increased PME activity as well. Moreover, light-induced psbO gene activation was accompanied by transcription of PME gene. The introduction of foreign gene in plant cell or excess transcription of own genome resulted in increasing PME activity leading to transcriptome status quo returning.

Cell wall polysaccharides are specifically involved in the exclusion of aluminum from the rice root apex.

Rice (Oryza sativa) is the most aluminum (Al)-resistant crop species among the small-grain cereals, but the mechanisms responsible for this trait are still unclear. Using two rice cultivars differing in Al resistance, rice sp. japonica 'Nipponbare' (an Al-resistant cultivar) and rice sp. indica 'Zhefu802' (an Al-sensitive cultivar), it was found that Al content in the root apex (0-10 mm) was significantly lower in Al-resistant 'Nipponbare' than in sensitive 'Zhefu802', with more of the Al localized to cell walls in 'Zhefu802', indicating that an Al exclusion mechanism is operating in 'Nipponbare'. However, neither organic acid efflux nor changes in rhizosphere pH appear to be responsible for the Al exclusion. Interestingly, cell wall polysaccharides (pectin, hemicellulose 1, and hemicellulose 2) in the root apex were found to be significantly higher in 'Zhefu802' than in 'Nipponbare' in the absence of Al, and Al exposure increased root apex hemicellulose content more significantly in 'Zhefu802'. Root tip cell wall pectin methylesterase (PME) activity was constitutively higher in 'Zhefu802' than in 'Nipponbare', although Al treatment resulted in increased PME activity in both cultivars. Immunolocalization of pectins showed a higher proportion of demethylated pectins in 'Zhefu802', indicating a higher proportion of free pectic acid residues in the cell walls of 'Zhefu802' root tips. Al adsorption and desorption kinetics of root tip cell walls also indicated that more Al was adsorbed and bound Al was retained more tightly in 'Zhefu802', which was consistent with Al content, PME activity, and pectin demethylesterification results. These responses were specific to Al compared with other metals (CdCl(2), LaCl(3), and CuCl(2)), and the ability of the cell wall to adsorb these metals was also not related to levels of cell wall pectins. All of these results suggest that cell wall polysaccharides may play an important role in excluding Al specifically from the rice root apex.

Improvement in texture using calcium lactate and heat-shock treatments for stored ready-to-eat carrots

The use of calcium lactate solutions has been shown to be a healthy alternative to chlorine washing in order to maintain the shelf-life of fresh-cut products. The aim of this research was to analyse the effects of calcium lactate (15 g L −1) treatment at 25 °C and 50 °C (heat-shock) on the textural properties of sliced carrots and to compare those with the chlorine treatment (120 mg L −1) widely used in industry. Several direct and indirect markers of textural changes in carrots during storage were used: Instron textural analysis, Cryo-SEM and optical microscopic, sensory analysis, pectin methylesterase (PME) activity, calcium content and water activity. Samples treated with calcium lactate maintained texture significantly ( p < 0.05) better than samples treated with chlorine throughout storage. Calcium lactate treatment produced a reduction in the water activity in sliced carrots and a higher firmness (Instron analysis) than chlorine treatment. In addition, combined use of heat-shock and calcium lactate treatment increased PME activity significantly when compared to the other treatments, results that were confirmed by sensory analysis. Cryo-SEM analyses showed that combined heat-shock and calcium lactate treatment was more effective in maintaining the turgor of cortex tissue cells and reduced the extent of lignification at cutting-edge areas. The use of calcium lactate combined with heat-shock is a promising washing method for fresh-cut carrots in order to preserve their texture and improve their nutritional value, avoiding the use of chlorine washing.

Chilling‐Associated Softening of Tomato Fruit is Related to Increased Pectinmethylesterase Activity

ABSTRACTMature‐green tomatoes chilled 15 days (5°C; RH &gt;85%) were softer than nonchilled during subsequent ripening (22°C) by both whole fruit and pericarp tissue puncture (p&lt;0.05), but not by flat‐plate compression. No differences in total polygalactnronase (PG) or PG isozyme activity were evident although total activity was greater in nonchilled after 10 days ripening. Softening of nonchilled fruit correlated (p&lt;0.05) with extracted PGI activity, while chilling‐associated softening correlated (p&lt;0.05) with higher initial extracted pectinmethylesterase (PME) activity. Extracted peroxidase remained constant throughout ripening but was greater (p&lt;0.05) in pre‐chilled fruit consistent with chill‐induced membrane dysfunction. Transmission electron microscopy showed the middle lamella from pre‐chilled tomatoes was swollen and less defined. Loss of turgor from translocation of water to the PME‐modified cell wall was suggested to be responsible for softening as a consequence of chilling.

Association of the degree of methylation of intercellular pectin with plant resistance to aphids and with induction of aphid biotypes

Increased methylation of middle lamellar pectin in plants hinders aphids in penetrating host-plant tissue. In sorghum, a new aphid biotype has overcome this host-plant barrier by having increased pectin methylesterase activity. Results suggest that resistance in crop-plants against sap-feeding insects may possibly be manipulated by altering middle lamellar chemistry either through breeding or use of certain plant-growth regulators.