Changes In Cell Wall Components Research Articles

The Resistance of Maize to Ustilago maydis Infection Is Correlated with the Degree of Methyl Esterification of Pectin in the Cell Wall.

Common smut caused by Ustilago maydis is one of the dominant fungal diseases in plants. The resistance mechanism to U. maydis infection involving alterations in the cell wall is poorly studied. In this study, the resistant single segment substitution line (SSSL) R445 and its susceptible recurrent parent line Ye478 of maize were infected with U. maydis, and the changes in cell wall components and structure were studied at 0, 2, 4, 8, and 12 days postinfection. In R445 and Ye478, the contents of cellulose, hemicellulose, pectin, and lignin increased by varying degrees, and pectin methylesterase (PME) activity increased. The changes in hemicellulose and pectin in the cell wall after U. maydis infection were analyzed via immunolabeling using monoclonal antibodies against hemicellulsic xylans and high/low-methylated pectin. U. maydis infection altered methyl esterification of pectin, and the degree of methyl esterification was correlated with the resistance of maize to U. maydis. Furthermore, the relationship between methyl esterification of pectin and host resistance was validated using 15 maize inbred lines with different resistance levels. The results revealed that cell wall components, particularly pectin, were important factors affecting the colonization and propagation of U. maydis in maize, and methyl esterification of pectin played a role in the resistance of maize to U. maydis infection.

Comparative physiological and transcriptomic analyses provide insights into fruit softening in Chinese cherry [Cerasus pseudocerasus (Lindl.) G.Don

Fruit softening is a complex, genetically programmed and environmentally regulated process, which undergoes biochemical and physiological changes during fruit development. The molecular mechanisms that determine these changes in Chinese cherry [Cerasus peseudocerasus (Lindl.) G.Don] fruits are still unknown. In the present study, fruits of hard-fleshed 'Hongfei' and soft-fleshed 'Pengzhoubai' varieties of Chinese cherry were selected to illustrate the fruit softening at different developmental stages. We analyzed physiological characteristics and transcriptome profiles to identify key cell wall components and candidate genes related to fruit softening and construct the co-expression networks. The dynamic changes of cell wall components (cellulose, hemicellulose, pectin, and lignin), the degrading enzyme activities, and the microstructure were closely related to the fruit firmness during fruit softening. A total of 6,757 and 3,998 differentially expressed genes (DEGs) were screened between stages and varieties, respectively. Comprehensive functional enrichment analysis supported that cell wall metabolism and plant hormone signal transduction pathways were involved in fruit softening. The majority of structural genes were significantly increased with fruit ripening in both varieties, but mainly down-regulated in Hongfei fruits compared with Pengzhoubai, especially DEGs related to cellulose and hemicellulose metabolism. The expression levels of genes involving lignin biosynthesis were decreased with fruit ripening, while mainly up-regulated in Hongfei fruits at red stage. These obvious differences might delay the cell all degrading and loosening, and enhance the cell wall stiffing in Hongfei fruits, which maintained a higher level of fruit firmness than Pengzhoubai. Co-expressed network analysis showed that the key structural genes were correlated with plant hormone signal genes (such as abscisic acid, auxin, and jasmonic acid) and transcription factors (MADS, bHLH, MYB, ERF, NAC, and WRKY). The RNA-seq results were supported using RT-qPCR by 25 selected DEGs that involved in cell wall metabolism, hormone signal pathways and TF genes. These results provide important basis for the molecular mechanism of fruit softening in Chinese cherry.

QTL analysis of the developmental changes in cell wall components and forage digestibility in maize (Zea mays L.)

Cell wall architecture plays a key role in stalk strength and forage digestibility. Lignin, cellulose, and hemicellulose are the three main components of plant cell walls, and they can impact stalk quality by affecting the structure and strength of the cell wall. To explore cell wall development during secondary cell wall lignification in maize stalks, conventional and conditional genetic mapping were used to identify the dynamic quantitative trait loci (QTLs) of the cell wall components and digestibility traits during five growth stages after silking. Acid detergent lignin (ADL), cellulose (CEL), acid detergent fiber (ADF), neutral detergent fiber (NDF), and in vitro dry matter digestibility (IVDMD) were evaluated in a maize recombinant inbred line (RIL) population. ADL, CEL, ADF, and NDF gradually increased from 10 to 40 days after silking (DAS), and then they decreased. IVDMD initially decreased until 40 DAS, and then it increased slightly. Seventy-two QTLs were identified for the five traits, and each accounted for 3.48–24.04% of the phenotypic variation. Six QTL hotspots were found, and they were localized in the 1.08, 2.04, 2.07, 7.03, 8.05, and 9.03 bins of the maize genome. Within the interval of the pleiotropic QTL identified in bin 1.08 of the maize genome, six genes associated with cell wall component biosynthesis were identified as potential candidate genes for stalk strength as well as cell wall-related traits. In addition, 26 conditional QTLs were detected in the five stages for all of the investigated traits. Twenty-two of the 26 conditional QTLs were found at 30 DAS conditioned using the values of 20 DAS, and at 50 DAS conditioned using the values of 40 DAS. These results indicated that cell wall-related traits are regulated by many genes, which are specifically expressed at different stages after silking. Simultaneous improvements in both forage digestibility and lodging resistance could be achieved by pyramiding multiple beneficial QTL alleles identified in this study.

Responses of Cell Wall Components to Low Nitrogen in Rapeseed Roots

Rapeseed (Brassica napus L.) is a major oil crop in China, with the world’s largest planted area and total yield. Rapeseed has a high demand for nitrogen (N), and nitrogen deficiency in soil is an important limiting factor for rapeseed production. However, rapeseed responds to N deprivation by regulating its own morphology, structure, and physiology. We carried out the current experiment by utilizing low N (LN: 0.3 mM NO3−) and normal N (CK: 6.0 mM NO3−) treatments using Brassica napus as the experimental material. The study results showed that low N induced root elongation in rapeseed, and the root length of LN treatment was 2.37 times that of HN treatment. The dry matter of roots also significantly increased due to low N treatment. Meanwhile, low N treatment decreased photosynthetic pigment (including chlorophyll a, chlorophyll b, and carotenoids) contents and dry mass accumulation of leaves. A higher root/shoot ratio and N physiological efficiency were observed under low N treatment. The changes in cell wall components (pectin, cellulose, hemicellulose, and lignin), related enzymes, and genes’ transcription levels in roots were determined and the results suggested that low N promoted the demethylation of ion-bound pectin (ISP) and covalently bound pectin (CSP), the content of CSP and cellulose. The promoted pectin methylesterase (PME) activity, inhibited pectin and cellulose degradation enzymes, and up/downregulation of related genes also confirming the results of cell wall components. The low N-increased demethylation degree of pectin and content of pectin and cellulose in cell walls was conducive to cell wall loosening and cell wall synthesis during cell division and elongation, ultimately promoting root-adaptive elongation. The study revealed a possible mechanism in which the alteration of cell wall component content and structure participates in cell elongation and expansion, which directly induces root elongation under N deficiency. The successful implementation of this research may be conducive to facilitating the development of rapeseed cultivars with high N use efficiency through root-based genetic improvements and improving plant adaptability to low N.

Proteomics Reveals the Mechanism Underlying the Autolysis of Postharvest Coprinus comatus Fruiting Bodies

Autolysis occurs widely in edible mushroom fruiting bodies after harvest, but the mechanism is still unclear. In this study, quantitative proteomics and bioinformatics analyses have been applied for revealing the autolysis mechanism of postharvest Coprinus comatus fruiting bodies. The results indicated that the autolysis mechanism of postharvest C. comatus was complicated. Before pileus opening, the carbohydrate metabolism including cell wall hydrolysis and energy biosynthesis, which were probably regulated by the ribosome, was involved in mushroom autolysis, whereas after pileus opening, the autolysis mechanism was related to the accumulated reactive oxygen species (ROS) and activated mitogen-activated protein kinase (MAPK) signaling pathway based on the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Furthermore, the changes in cell wall components and hydrolases, along with the production of ROS and the activities of oxidoreductase in C. comatus, were also verified to confirm the proteomic analysis results.

Boron-toxicity induced changes in cell wall components, boron forms, and antioxidant defense system in rice seedlings

Boron (B) is an indispensable micronutrient that ensures the optimal growth and productivity of the plant. However, excessive use of B fertilizers results in B toxicity which is relatively difficult to correct as compared to B deficiency. Moreover, underlying mechanisms of B toxicity induced changes in cell wall components and the association of B forms in the appearance of toxicity symptoms in rice seedlings are lacking. Therefore, the present investigation was carried out on rice seedlings by employing different concentrations of B (CK, B1; 100 µM, B2; 300 µM, and B3; 400 µM). The results showed that a high concentration of B caused inhibition of root and shoot growth with noticeable signs of stress on leaves in terms of chlorophyll contents. In addition, B toxicity caused oxidative stress and lipid oxidation of membranes. The higher concentrations of B were accumulated in the leaves than roots. In the roots and leaves, more than 80% B was adsorbed on the cell wall. In the treatment of B3, the free form of B was higher than the bound-B. Fourier Transform Infrared Spectrometer (FTIR) results showed that higher concentrations led to variation in functional groups of cell walls of leaves. The results of this investigation showed that B stress-induced inhibition of growth might be linked with higher B uptake in the upper parts, oxidative damages, and forms of B may play important role in the chlorosis. The findings of the study may help to understand the mechanisms of B stress-induced growth inhibition in rice seedlings.

Changes in cell wall components and hygroscopic properties of Pinus radiata caused by heat treatment

After thermal modification at 210 °C using the Thermowood method, changes in Pinus radiata D. Don wood cell wall components and hygroscopicity were studied for subsequent comparison with recently felled wood of the same species in samples from the same region of provenance (Basque Country, Spain). To this end, samples were characterised by sorption isotherms at 15, 35 and 50 °C fitted to the Guggenheim–Anderson–de Boer (GAB) model, the chemical composition was obtained by high performance liquid chromatography (HPLC) and infrared spectroscopy (FTIR), and crystallinity and structural organisation of cellulose (crystal orientation) were determined using powder and 2D X-ray diffraction (XRD). Heat treatment caused the following changes in the wood: a decrease in the equilibrium moisture content (EMC); a smaller hysteresis area and therefore more hygroscopically stable wood; a decrease in the hemicellulose content; an increase in the relative percentage of cellulose, lignin and extractives; and a higher degree of crystallinity and crystal orientation of the cellulose. The reorganisation of cellulose could be explained by epitaxial growth of cellulose starting from the highly oriented crystalline regions during the recrystallisation process. All these chemical and structural changes induced by heating could explain the reduction in hygroscopic properties of wood, as well as its stability.

Accelerating thermal decomposition of wood cell wall with glycerol

Thermal modification reduces the hygroscopicity of timber potentially reducing susceptibility to biological degradation, but it is also energetically intensive. Identifying methods for accelerating the process could reduce these inputs. The potential effects of glycerol pretreatment on thermal modification was investigated by pre-treating poplar wood (Populus tomentosa Carr.) with 60% glycerol followed by thermal modification at temperatures between 120 and 200 °C for 4 h. Potential changes in the wood were analyzed using high performance liquid chromatography, infrared spectroscopy, X-ray diffraction, scanning electron microscopy and fluorescence microscopy to detect the changes in cell wall components, crystallinity, the average size of the crystalline region and cell wall thickness. Glycerol accelerated lignin decomposition over the entire temperature range. Thermal modification altered the degree of cellulose crystallinity as well as the average size of the crystalline region and the effects were more noticeable with the glycerol pre-treatment. The cell wall thickness of thermally treated wood decreased with increasing temperature and glycerol accelerated this process. Glycerol appears to enhance the modification process with increasing treatment temperature.

Changes in cell wall components and polysaccharide-degrading enzymes in relation to differences in texture during sweetpotato storage root growth

Sweetpotato has special texture characteristics, which directly affect the eating quality and post-production processing quality of sweetpotato. To investigate the texture change mechanism of sweetpotato during the growth process, this study selected two varieties with significant differences in texture from 35 varieties. The storage roots were sampled at 50, 80, 110, and 140 days after planting. Measure the texture parameters, the cell wall composition content, cell wall-related enzyme activities and the expression of expansin genes of sweetpotato storage roots. The results show that the hardness, adhesiveness and chewiness parameters of ‘Yushu No 10′ were significantly lower than those of ‘Mianfen No 1′, they have significantly different texture properties. In terms of cell wall composition, the soluble pectin content of ‘Yushu No 10′ was more than twice that of ‘Mianfen No 1′, whereas the insoluble pectin content was lower than that of ‘Mianfen No 1′, with the cellulose content of ‘Yushu No 10′ being significantly higher than that of ‘Mianfen No 1′. In terms of cell wall-related enzymes, ‘Yushu No 10′ hardness gumminess and chewiness had a significant correlation with hemicellulose activity, and ‘Mianfen No 1′ had insignificant correlation with four cell wall-related enzymes. Expansin genes were also expressed differently during the various stages of root tubers expansin. The expressions of IbEXP1, IbEXP2 and IbEXPL1 were significantly correlated with the changes in cell wall component content, and were related to the qualitative structure changes. The research conclusion shows that the texture changes during the growth of sweetpotato are related to cell wall composition, cell wall-related enzyme activity changes, and the expression of expansin genes. This study provides theoretical guidance for in-depth study of texture changes of sweetpotato, post-harvest processing and utilization and quality improvement of storage roots.

Changes of cell wall components during embryogenesis of Castanea mollissima.

The Chinese chestnut (Castanea mollissima Blume) 'Huaihuang' was chosen as the experimental material to observe embryogenesis and the dynamic changes of cell wall components during this process. Various developmental stages of embryos, including globular embryos, heart embryos, torpedo embryos and cotyledon embryos, were observed. The results showed that during embryogenesis, cellulose increased, and callose rapidly degraded. In the cell walls of developing embryos, pectic homogalacturonan (HG), especially low-esterified HG, was abundant, suggesting rapid synthesis and de-methyl-esterification of HG. Extensin and galactan increased with the development of the embryos. In contrast, the arabinan epitopes decreased in developing embryos but were more abundant than galactan epitopes at all stages. Xylan epitopes showed explicit boundaries between the outer epidermal wall and the rest of the inner tissues, and the fluorescence intensity of the outer epidermal wall was significantly higher than that of the inner tissues. Furthermore, the results indicated that the outer epidermal wall contained high amounts of cellulose, HG pectin and hemicellulose, especially arabinan and xylan. These results suggested the presence of rapid pectin metabolism, cellulose synthesis, rapid degradation of callose, different distributive patterns and dynamic changes of hemicellulose (galactan, arabinan and xylan) and extensin during embryogenesis. Various cell wall components exist in different tissues of the embryo, and dynamic changes in cell wall components are involved in the embryonic development process.

Changes in cell wall components of Pinus sylvestris L. wood after 300 years in contact with salt (NaCl)

Changes were evaluated in cell wall components, hygroscopicity, and thermodynamic properties of Pinus sylvestris L. wood that had been in contact with salt (NaCl) for 300 years in a former saltworks storehouse in Gerri de la Sal (Lérida, Spain), comparing it with recently felled wood (new wood) of the same species. The wood in contact with salt (salt-covered wood) had higher equilibrium moisture content values except in the first interval of the adsorption isotherm, probably because of mechanical blocking of polar sites by salt crystals. The high fibre saturation point values of this wood are caused by the salt dissolved in water above water activity of about 0.75. More energy is invested in the desorption process, and the values are higher in the salt-covered wood.

Transcriptomic analysis of boron hyperaccumulation mechanisms in Puccinellia distans

Puccinellia distans, common alkali grass, is found throughout the world and can survive in soils with boron concentrations that are lethal for other plant species. Indeed, P. distans accumulates very high levels of this element. Despite these interesting features, very little research has been performed to elucidate the boron tolerance mechanism in this species. In this study, P. distans samples were treated for three weeks with normal (0.5 mg L−1) and elevated (500 mg L−1) boron levels in hydroponic solution. Expressed sequence tags (ESTs) derived from shoot tissue were analyzed by RNA sequencing to identify genes up and down-regulated under boron stress. In this way, 3312 differentially expressed transcripts were detected, 67.7% of which were up-regulated and 32.3% of which were down-regulated in boron-treated plants. To partially confirm the RNA sequencing results, 32 randomly selected transcripts were analyzed for their expression levels in boron-treated plants. The results agreed with the expected direction of change (up or down-regulation). A total of 1652 transcripts had homologs in A. thaliana and/or O. sativa and mapped to 1107 different proteins. Functional annotation of these proteins indicated that the boron tolerance and hyperaccumulation mechanisms of P. distans involve many transcriptomic changes including: alterations in the malate pathway, changes in cell wall components that may allow sequestration of excess boron without toxic effects, and increased expression of at least one putative boron transporter and two putative aquaporins. Elucidation of the boron accumulation mechanism is important in developing approaches for bioremediation of boron contaminated soils.

Evaluating Changes in Cell-Wall Components Associated with Clubroot Resistance Using Fourier Transform Infrared Spectroscopy and RT-PCR.

Clubroot disease is a serious threat to canola production in western Canada and many parts of the world. Rcr1 is a clubroot resistance (CR) gene identified recently and its molecular mechanisms in mediating CR have been studied using several omics approaches. The current study aimed to characterize the biochemical changes in the cell wall of canola roots connecting to key molecular mechanisms of this CR gene identified in prior studies using Fourier transform infrared (FTIR) spectroscopy. The expression of nine genes involved in phenylpropanoid metabolism was also studied using qPCR. Between susceptible (S) and resistance (R) samples, the most notable biochemical changes were related to an increased biosynthesis of lignin and phenolics. These results were supported by the transcription data on higher expression of BrPAL1. The up-regulation of PAL is indicative of an inducible defence response conferred by Rcr1; the activation of this basal defence gene via the phenylpropanoid pathway may contribute to clubroot resistance conferred by Rcr1. The data indicate that several cell-wall components, including lignin and pectin, may play a role in defence responses against clubroot. Principal components analysis of FTIR data separated non-inoculated samples from inoculated samples, but not so much between inoculated S and inoculated R samples. It is also shown that FTIR spectroscopy can be a useful tool in studying plant-pathogen interaction at cellular levels.

JoVE Monthly Highlights: June 2017

Here's a look at what's coming up in the June 2017 issue of JoVE: The World's Premier Video Journal A handy method to test the efficacy of sanitizing techniques is our first feature this month, from JoVE Immunology and Infection. Here, our Authors first spike the gloved hands of volunteers with a test organism, and then employ a hand-washing technique of interest. Then, the remaining organisms are collected, and unwashed and test samples are compared. These results allow researchers to provide recommendations for best cleanliness practices during emerging infectious disease outbreaks. We can all hi-five with confidence to that! As spring is in full bloom so are allergies, and these are the subjects of our next video highlight. In JoVE Medicine, our Authors describe a novel diagnostic approach for determining specific types of grass pollen allergy using a cohort of patients from Southern China - an area with previously limited data on local allergens. Here, the researchers employed component-resolved diagnostics involving recombinant or purified allergens, to detect sensitization to specific types of grass pollen in test and control individuals. This allows for more targeted treatment of allergic airway diseases in patients with allergic rhinitis or asthma, and that's nothing to sneeze at! From grassy gametes to adult plant cells, next we showcase a method for visualizing plant cell wall components, from JoVE Biochemistry. Here, our Authors demonstrate a technique for accurately distinguishing and quantifying cell wall constituents - including those with similar chemical structures such as lignin, cellulose and hemicellulose - using a combination of Raman imaging and multivariate analysis. This method overcomes the barriers to studies of dynamic changes in cell wall components during chemical, physical or biological treatments. Finally this month we move from the sessile to the mobile and examine motor neuron development with JoVE Neuroscience. Studying late stage-16 Drosophila embryos, our Author details an immunohistochemistry method to analyze axonal motor neuron projections. By positioning fixed embryos in a filleted arrangement and staining with FasII antibody, the resulting preparations are ideal for the characterization of genes involved in motor axon path-finding and target recognition. It's a neat little technique, with lots of muscle. You've just had a sneak peek of the June 2017 issue of JoVE. Visit the website to see the full-length articles, plus many more, in JoVE: The World's Premier Video Journal.

Proteomic analysis of stipe explants reveals differentially expressed proteins involved in early direct somatic embryogenesis of the tree fern Cyathea delgadii Sternb

Using cyto-morphological analysis of somatic embryogenesis (SE) in the tree fern Cyathea delgadii as a guide, we performed a comparative proteomic analysis in stipe explants undergoing direct SE. Plant material was cultured on hormone-free medium supplemented with 2% sucrose. Phenol extracted proteins were separated using two-dimensional gel electrophoresis (2-DE) and mass spectrometry was performed for protein identification. A total number of 114 differentially regulated proteins was identified during early SE, i.e. when the first cell divisions started and several-cell pro-embryos were formed. Proteins were assigned to seven functional categories: carbohydrate metabolism, protein metabolism, cell organization, defense and stress responses, amino acid metabolism, purine metabolism, and fatty acid metabolism. Carbohydrate and protein metabolism were found to be the most sensitive SE functions with the greatest number of alterations in the intensity of spots in gel. Differences, especially in non-enzymatic and structural protein abundance, are indicative for cell organization, including cytoskeleton rearrangement and changes in cell wall components. The highest induced changes concern those enzymes related to fatty acid metabolism. Global analysis of the proteome reveals several proteins that can represent markers for the first 16days of SE induction and expression in fern. The findings of this research improve the understanding of molecular processes involved in direct SE in C. delgadii.

Immunolocalization of pectic polysaccharides during abscission in pea seeds (Pisum sativum L.) and in abscission less def pea mutant seeds.

BackgroundIn pea seeds (Pisum sativum L.), the presence of the Def locus determines abscission event between its funicle and the seed coat. Cell wall remodeling is a necessary condition for abscission of pea seed. The changes in cell wall components in wild type (WT) pea seed with Def loci showing seed abscission and in abscission less def mutant peas were studied to identify the factors determining abscission and non-abscission event.MethodsChanges in pectic polysaccharides components were investigated in WT and def mutant pea seeds using immunolabeling techniques. Pectic monoclonal antibodies (1 → 4)-β-d-galactan (LM5), (1 → 5)-α-l-arabinan(LM6), partially de-methyl esterified homogalacturonan (HG) (JIM5) and methyl esterified HG (JIM7) were used for this study.ResultsPrior to abscission zone (AZ) development, galactan and arabinan reduced in the predestined AZ of the pea seed and disappeared during the abscission process. The AZ cells had partially de-methyl esterified HG while other areas had highly methyl esterified HG. A strong JIM5 labeling in the def mutant may be related to cell wall rigidity in the mature def mutants. In addition, the appearance of pectic epitopes in two F3 populations resulting from cross between WT and def mutant parents was studied. As a result, we identified that homozygous dominant lines (Def/Def) showing abscission and homozygous recessive lines (def/def) showing non-abscission had similar immunolabeling pattern to their parents. However, the heterogeneous lines (Def/def) showed various immunolabeling pattern and the segregation pattern of the Def locus.ConclusionsThrough the study of the complexity and variability of pectins in plant cell walls as well as understanding the segregation patterns of the Def locus using immunolabeling techniques, we conclude that cell wall remodeling occurs in the abscission process and de-methyl esterification may play a role in the non-abscission event in def mutant. Overall, this study contributes new insights into understanding the structural and architectural organization of the cell walls during abscission.

A Cotton Annexin Affects Fiber Elongation and Secondary Cell Wall Biosynthesis Associated with Ca2+ Influx, ROS Homeostasis, and Actin Filament Reorganization

Annexins play pivotal roles in a variety of cellular processes as well as in fiber development; however, the functional mechanisms of their activities are unclear. Here, an annexin gene that is preferentially expressed in fibers, GhFAnnxA, was found to be significantly associated with various cotton (Gossypium hirsutum) fiber traits. Transgenic analysis demonstrated that GhFAnnxA affected cotton fiber elongation and was involved in secondary cell wall (SCW) biosynthesis. Functional studies demonstrated that GhFAnnxA may act as a Ca(2+) conductance regulator and that reactive oxygen species (ROS) produced by Rbohs in a Ca(2+)-dependent manner may determine fiber elongation caused by elevated intracellular turgor and cell wall loosening. However, excessive hydrogen peroxide (H2O2) inhibited cotton fiber elongation in vitro. We speculate that a positive feedback loop involving ROS and Ca(2+) is regulated by GhCDPK1 and regulates fiber cell elongation. Furthermore, the convergence of actin filaments is altered by their interaction with GhFAnnxA, and this also may contribute to fiber elongation. Moreover, GhFAnnxA may affect SCW biosynthesis through changes in cell wall components caused by an increase in H2O2 levels. These results not only provide new insights into the signaling pathways of GhFAnnxA in fiber development but also clarify the role of ROS in fiber development.

Characterization of the Probiotic Yeast Saccharomyces boulardii in the Healthy Mucosal Immune System.

The probiotic yeast Saccharomyces boulardii has been shown to ameliorate disease severity in the context of many infectious and inflammatory conditions. However, use of S. boulardii as a prophylactic agent or therapeutic delivery vector would require delivery of S. boulardii to a healthy, uninflamed intestine. In contrast to inflamed mucosal tissue, the diverse microbiota, intact epithelial barrier, and fewer inflammatory immune cells within the healthy intestine may all limit the degree to which S. boulardii contacts and influences the host mucosal immune system. Understanding the nature of these interactions is crucial for application of S. boulardii as a prophylactic agent or therapeutic delivery vehicle. In this study, we explore both intrinsic and immunomodulatory properties of S. boulardii in the healthy mucosal immune system. Genomic sequencing and morphological analysis of S. boulardii reveals changes in cell wall components compared to non-probiotic S. cerevisiae that may partially account for probiotic functions of S. boulardii. Flow cytometry and immunohistochemistry demonstrate limited S. boulardii association with murine Peyer’s patches. We also show that although S. boulardii induces a systemic humoral immune response, this response is small in magnitude and not directed against S. boulardii itself. RNA-seq of the draining mesenteric lymph nodes indicates that even repeated administration of S. boulardii induces few transcriptional changes in the healthy intestine. Together these data strongly suggest that interaction between S. boulardii and the mucosal immune system in the healthy intestine is limited, with important implications for future work examining S. boulardii as a prophylactic agent and therapeutic delivery vehicle.

Antisense expression of Gossypium hirsutum UDP-glucuronate decarboxylase in Arabidopsis leads to changes in cell wall components.

UDP-glucuronate decarboxylase (UDP-xylose synthase; UXS, EC 4.1.1.35) is an essential enzyme of the non-cellulosic polysaccharide biosynthetic pathway. In the present study, using transient expression of fluorescently labeled Gossypium hirsutum UXS (GhUXS3) protein in onion epidermal cells, we observed that this protein was distributed in the cytoplasm. The GhUXS3 cDNA of cotton was expressed in an antisense orientation in Arabidopsis thaliana by Agrobacterium tumefaciens-mediated transformation. Homozygous plants showing down-regulation of UXS were analyzed with northern blots. Compared to the untransformed control, transgenic plant showed shorter roots, earlier blossom formation, and delayed senescence. Biochemical analysis indicated that levels of rhamnose, mannose, galactose, glucose, xylose, and cellulose were reduced in some of the down-regulated antisense plants. These results suggest that GhUXS3 regulates the conversion of non-cellulosic polysaccharides and modulates their composition in plant cell walls. We also discuss a possible cellular function for GhUXS in determining the quality of cotton fibers.

The Influence of Hot Water and Calcium Chloride on the Changes in Cell Wall Composition and the Activities of Cell Wall Hydrolases during Storage inAgaricus bisporus

Mushrooms show significant losses in firmness due mainly to the changes in cell walls after harvest. This study was performed to determine the effects of CaCl2 and hot water dips on the changes in firmness, cell wall composition and hydrolase activities of mushrooms during storage. Mushrooms were treated with various concentrations of CaCl2 and hot water at various degrees and stored for 12 days at 10C. The results showed that the firmness of mushrooms decreased significantly and their cell walls have undergone significant changes during 12 days storage period after harvest. Polysaccharide content decreased significantly during storage. This decrease was mainly due to the decrease in the strong alkaline-soluble polysaccharides. Xyloglucanase and beta glucanase activities increased significantly during storage, possibly contributing significantly to the polysaccharide losses. CaCl2 and hot water treatments significantly reduced the changes in cell wall components and cell wall hydrolase activities. The results suggest that CaCl2 and hot water treatments could be used to reduce the changes in firmness and cell walls after harvest and extend the shelf life of mushroom. Practical Applications Mushrooms show rapid deterioration after harvest due mainly to changes in cell walls and have a very short shelf life. This significantly reduces their marketing period. The results demonstrate that both hot water and CaCl2 significantly delay cell wall degradation and prolong the shelf life of mushrooms. The results of the study have the potential to be used to decide on what treatments should be used to extend the shelf life and marketing period of mushrooms.