High Levels Of Trehalose Research Articles

The Role of Osmolytes and Membrane Lipids in the Adaptation of Acidophilic Fungi.

Acidophiles maintain near-neutral intracellular pH using proton pumps. We have suggested the protective role of osmolytes and membrane lipids in the adaptation to an acidic environment. Previously we have observed, for the first time, high levels of trehalose in acidophilic basidiomycete Sistotrema brinkmannii. Here, we have studied the composition of both osmolytes and membrane lipids of two more acidophilic fungi. Trehalose and polyols were among the main osmolytes during growth under optimal conditions (pH 4.0) in basidiomycete Phlebiopsis gigantea and ascomycete Mollisia sp. Phosphatidic acids, phosphatidylethanolamines, phosphatidylcholines, and sterols, were predominant membrane lipids in both fungi. P. gigantea had a narrow optimum of growth at pH 4.0, resulting in a sharp decline of growth rate at pH 2.6 and 5.0, accompanied by a decrease in the number of osmolytes and significant changes in the composition of membrane lipids. In contrast, Mollisia sp. had a broad optimal growth range (pH 3.0-5.0), and the number of osmolytes either stayed the same (at pH 6.0) or increased (at pH 2.6), while membrane lipids composition remained unchanged. Thus, the data obtained indicate the participation of osmolytes and membrane lipids in the adaptation of acidophilic fungi.

Effective Parameters for the Stimulation of Trehalose Synthesis in Saccharomyces cerevisiae

Background: The ability of anhydrobiotic to survive long periods of stress conditions is associated with high trehalose levels found in these organisms as a responsive mechanism against stress and adverse conditions. Many methods and factors function as determinant agents of trehalose production and preservation in yeasts. Objectives: This study aimed to investigate the effect of three parameters on trehalose synthesis, including temperature, rate of feed, and time of aeration. Methods: To induce trehalose production in baker's yeast, we used an experimental design method in an industrial fermenter at the Khuzestan Yeast Company. The investigation of temperature, rate of feed, and time of aeration parameters in our recipe design were also performed after the cessation of feeding molasses. Results: The findings showed that temperature had the greatest effect on the synthesis of trehalose. Moreover, the interaction of three parameters was significantly effective for the stimulation of trehalose synthesis. We could also make a gas production surge in the dough by increasing the level of trehalose. Conclusions: We conclude that controlling the temperature and taking advantage of the interaction between temperature, rate of feed, and aeration time can help achieve the most effective trehalose synthesis and ensuing gas production in the dough.

Trehalose-induced changes in the aggregation behavior and structural properties of wheat gluten

Trehalose is widely used in the food and cosmetics industry owing to its unique stabilizing properties. In this study, we aimed to further understand the effect of trehalose on the aggregation and structural properties of gluten. We investigated changes in the aggregation and textural properties, molecular weight distribution, disulfide bond contents, secondary structure, and network structure for weak-, middle-, and strong-gluten when supplemented with trehalose. The results showed that the macromolecular aggregation of gluten proteins was impaired. The hardness, gumminess, and chewiness of dough reduced, whereas adhesiveness increased significantly. Moreover, a looser gluten network structure was obtained with increase in trehalose content. High levels of trehalose (<5%) caused a decrease in the disulfide bond content. Secondary structure analysis of gluten indicated that the α-helix content increased for weak-gluten. In strong-gluten wheat, the β-sheet content was converted to random coil. This suggests that trehalose affected the hydration level of gluten. Trehalose likely competes with gluten for the available water by disrupting the hydrogen bonds between gluten and water molecules and breaking the ‘‘loop and train” model of gluten. Thus, the decreased disulfide bond content and increased α-helix and random coil contents induced by trehalose inhibited gluten aggregation and interrupted the gluten network. The study findings improve our understanding of trehalose functionality in dough of different gluten strength. Furthermore, these data provide valuable insights into the application of trehalose in wheat flour food products.

Physiological and Molecular Characterization of an Oxidative Stress-Resistant Saccharomyces cerevisiae Strain Obtained by Evolutionary Engineering.

Oxidative stress is a major stress type observed in yeast bioprocesses, resulting in a decrease in yeast growth, viability, and productivity. Thus, robust yeast strains with increased resistance to oxidative stress are in highly demand by the industry. In addition, oxidative stress is also associated with aging and age-related complex conditions such as cancer and neurodegenerative diseases. Saccharomyces cerevisiae, as a model eukaryote, has been used to study these complex eukaryotic processes. However, the molecular mechanisms underlying oxidative stress responses and resistance are unclear. In this study, we have employed evolutionary engineering (also known as adaptive laboratory evolution – ALE) strategies to obtain an oxidative stress-resistant and genetically stable S. cerevisiae strain. Comparative physiological, transcriptomic, and genomic analyses of the evolved strain were then performed with respect to the reference strain. The results show that the oxidative stress-resistant evolved strain was also cross-resistant against other types of stressors, including heat, freeze-thaw, ethanol, cobalt, iron, and salt. It was also found to have higher levels of trehalose and glycogen production. Further, comparative transcriptomic analysis showed an upregulation of many genes associated with the stress response, transport, carbohydrate, lipid and cofactor metabolic processes, protein phosphorylation, cell wall organization, and biogenesis. Genes that were downregulated included those related to ribosome and RNA processing, nuclear transport, tRNA, and cell cycle. Whole genome re-sequencing analysis of the evolved strain identified mutations in genes related to the stress response, cell wall organization, carbohydrate metabolism/transport, which are in line with the physiological and transcriptomic results, and may give insight toward the complex molecular mechanisms of oxidative stress resistance.

Membrane lipid and osmolyte readjustment in the alkaliphilic micromycete Sodiomyces tronii under cold, heat and osmotic shocks.

Previously, we showed for the first time that alkaliphilic fungi, in contrast to alkalitolerant fungi, accumulated trehalose under extremely alkaline conditions, and we have proposed its key role in alkaliphilia. We propose that high levels of trehalose in the mycelium of alkaliphiles may promote adaptation not only to alkaline conditions, but also to other stressors. Therefore, we studied changes in the composition of osmolytes, and storage and membrane lipids under the action of cold (CS), heat (HS) and osmotic (OS) shocks in the obligate alkaliphilic micromycete Sodiomyces tronii. During adaptation to CS, an increase in the degree of unsaturation of phospholipids was observed while the composition of osmolytes, membrane and storage lipids remained the same. Under HS conditions, a twofold increase in the level of trehalose and an increase in the proportion of phosphatidylethanolamines were observed against the background of a decrease in the proportion of phosphatidic acids. OS was accompanied by a decrease in the amount of membrane lipids, while their ratio remained unchanged, and an increase in the level of polyols (arabitol and mannitol) in the fungal mycelium, which suggests their role for adaptation to OS. Thus, the observed consistency of the composition of membrane lipids suggests that trehalose can participate in adaptation not only to extremely alkaline conditions, but also to other stressors - HS, CS and OS. Taken together, the data obtained indicate the adaptability of the fungus to the action of various stressors, which can point to polyextremotolerance.

Impact of trehalose on physicochemical stability of β-carotene high loaded microcapsules fabricated by wet-milling coupled with spray drying

Solvent-free β-carotene high loaded microcapsules formulated by octenyl succinic anhydride (OSA)-starch and trehalose were prepared by a combination of wet-milling and spray drying. The influence of different mass ratios of OSA-starch to trehalose on the particle size distribution, storage stability, hygroscopicity and redispersibility of β-carotene microcapsules was investigated. The optimized microcapsules exhibited a small average diameter (228.2 nm) after rehydration with high encapsulation efficiency and loading capacity (99.06% and 10.32%, respectively). The phase separation in the OSA-starch and trehalose composite matrix was observed in the presence of trehalose at a high level, which was favorable for the prominent storage stability of β-carotene (96.16% for 50 days). The half-life of rehydrated β-carotene microcapsules was also extended by 2.23- and 1.25- folds during the thermal- and photo-degradation treatments. The crystallinity of β-carotene microcapsules was largely reduced due to the enhanced hydrogen bonding interaction between OSA-starch and β-carotene as analyzed by FTIR. SEM images showed that the morphology of β-carotene microcapsules was regulated by the level of trehalose. Confocal Raman spectrum microscopy and hygroscopic variation jointly verified the entrapment of β-carotene into a glassy matrix formed at a high level of trehalose. Moreover, a high level of trehalose could improve the spontaneous sinkability and dispersibility of β-carotene microcapsules, which were completely rehydrated in less than 1 min.

Peach PpSnRK1α interacts with bZIP11 and maintains trehalose balance in plants

The nonreducing disaccharide trehalose is widespread in nature. It plays a very important role in plant growth and development. In plants, trehalose is present in trace amounts. High concentration of trehalose disrupts energy balance and inhibits normal growth and development. Studies have shown that high levels of trehalose and trehalose-6-phosphate (T6P), the metabolic precursor of trehalose, inhibit sucrose non-fermenting-1-related protein kinase1 (SnRK1) activity, which affect plant growth and development. However, the role of SnRK1, the energy balance center, in the regulation of trehalose metabolism in plants is unknown. In this study, exogenous trehalose at higher concentrations inhibited the expression of SnRK1 genes, especially PpSnRK1α in peach (Prunus persica) seedlings. This change in gene expression was dependent on trehalose concentration. Furthermore, overexpression of peach PpSnRK1α in Arabidopsis thaliana significantly promoted trehalase activity, reduced T6P content, and suppressed the trehalose synthesis related genes (TPSs, TPPB) expression, promoted the trehalose metabolism of gene expression (TRE1), in addition the transgenic plants alleviated photosynthetic product distribution imbalance (aboveground and underground parts), and enhanced root growth. Yeast two-hybrid and bimolecular fluorescence assays revealed the interaction between PpSnRK1α and peach basic domain leucine zipper transcription factor 11 (PpbZIP11), a key transcription factor of trehalose metabolism, in the nucleus. To summarize, PpSnRK1α overexpression improved bZIP11 transcriptional activity and regulated trehalose metabolism to protect the plants against trehalose-induced damage. This study preliminarily explained the mechanism of SnRK1 regulating trehalose metabolism balance in plants, which laid a foundation for further understanding of energy metabolism and function of SnRK1 in plants.

Physiological and biochemical role of proline, trehalose, and compost on enhancing salinity tolerance of quinoa plant

Background and objectiveResearches on compost introduced the evidence of its benefits to plant productivity and soil fertility. These advantages are noticed in forms of improving soil water holding capacity and nutrient availability for plants. These changes can also improve plants’ capability to overcome salinity stress conditions. The application of osmo-protectant materials (proline and trehalose) and/or compost addition enhances plant antioxidative defense system against stress conditions. This experiment conducted to study the effect of spraying quinoa plants with proline and trehalose with and without soil compost addition under salinity stress on some morphological and physiological aspects.Materials and methodsQuinoa plant was grown with or without compost in the soil and foliar sprayed with proline or trehalose under salt irrigation. Plant samples were taken after 60 days from sowing and at the end of the experiment for growth, yield, and biochemical measurements.ResultsGrowth and yield measurements were decreased with salinity stress. High levels of both proline and trehalose recorded the highest values of total soluble sugars, proline, and free amino acids in both unstressed or salinity stressed plants with or without compost addition. The use of compost in soil for cultivating quinoa plants with either proline or trehalose treatments increased growth parameters, photosynthetic pigments, and yield attributes. In addition, these treatments improved the accumulation of some organic solutes in leaves and promoted antioxidant enzyme activities.ConclusionCompost addition to soil with spraying proline or trehalose improved quinoa growth and yield and produced seed nutritional value.

Comparative metabolic profiling of Ophiocordyceps sinensis and its cultured mycelia using GC–MS

Ophiocordyceps sinensis, one of the well-known traditional Chinese medicine, has multiple health-promoting effects. It is used as herbal remedy and health food in Asian countries, together with its cultured mycelia used as a substitute of natural O. sinensis. In the present study, natural O. sinensis collected from three geographical regions and its cultured mycelia derived from three strains were analyzed by gas chromatography-mass spectrometry (GC–MS) combined with chemometrics. A total of 72 metabolites were identified from all samples with different metabolic profiles observed between natural O. sinensis and cultured mycelia. Among them, 50 metabolites showed significant differences between natural O. sinensis and cultured mycelia. Higher levels of trehalose, glycerol and citric acid in natural O. sinensis were found compared to those in cultured mycelia, while myo-inositol and some amino acids were more abundant in cultured mycelia. In addition, chemical compositions of natural O. sinensis varied depending on the geographical regions. Natural O. sinensis from three locations were clearly differentiated by the concentrations of meso-erythritol, D-mannitol, glucose and organic acids. The current study provides a comprehensive metabolic profiles of natural O. sinensis and its cultured mycelia, which is potentially important for understanding the metabolism of O. sinensis and facilitating the application of cultured mycelia as a supplement of natural O. sinensis.

Molecular markers of biomass burning and primary biological aerosols in urban Beijing: size distribution and seasonal variation

Abstract. Biomass burning and primary biological aerosol particles account for an important part of urban aerosols. Floods of studies have been conducted on the chemical compositions of fine aerosols (PM2.5) in megacities where the haze pollution is one of the severe environmental issues in China. However, little is known about their size distributions in atmospheric aerosols in the urban boundary layer. Here, size-segregated aerosol samples were collected in Beijing during haze and clear days from April 2017 to January 2018. Three anhydrosugars, six primary saccharides and four sugar alcohols in these samples were identified and quantified by gas chromatography/mass spectrometry (GC/MS). Higher concentrations of a biomass burning tracer, levoglucosan, were detected in autumn and winter than in other seasons. Sucrose, glucose, fructose, mannitol and arabitol were more abundant in the bloom and glowing seasons. A particularly high level of trehalose was found in spring, which was largely associated with the Asian dust outflows. Anhydrosugars, xylose, maltose, inositol and erythritol are mainly present in the fine mode (&lt;2.1 µm), while the others showed the coarse-mode preference. The concentrations of measured tracers of biomass burning particles and primary biological particles in the haze events were higher than those in the non-hazy days, with enrichment factors of 2–10. Geometric mean diameters (GMDs) of molecular markers of biomass burning and primary biological aerosols showed that there was no significant difference in the coarse mode (&gt;2.1 µm) between the haze and non-haze samples, while a size shift towards large particles and large GMDs in the fine fraction (&lt;2.1 µm) was detected during the hazy days, which highlights that the stable meteorological conditions with high relative humidity in urban Beijing may favor the condensation of organics onto coarse particles.The contributions of reconstructed primary organic carbon (POC) by tracer-based methods from plant debris, fungal spores and biomass burning to aerosol OC in the total-mode particles were in the ranges of 0.09 %–0.30 % (on average 0.21 %), 0.13 %–1.0 % (0.38 %) and 1.2 %–7.5 % (4.5 %), respectively. This study demonstrates that the contribution of biomass burning was significant in Beijing throughout the whole year with the predominance in the fine mode, while the contributions of plant debris and fungal spores dominated in spring and summer in the coarse mode, especially in sizes &gt;5.8 µm. Our observations demonstrate that the sources, abundance and chemical composition of urban aerosol particles are strongly size dependent in Beijing, which is important to better understand the environmental and health effects of urban aerosols and should be considered in air quality and climate models.

Functional Characterization of Class I Trehalose Biosynthesis Genes in Physcomitrella patens

The function of trehalose metabolism in plants during growth and development has been extensively studied, mostly in the eudicot Arabidopsis thaliana. So far, however, not much is known about trehalose metabolism in the moss Physcomitrella patens. Here, we show that in P. patens, two active trehalose-6-phosphate synthase enzymes exist, PpTPS1 and PpTPS2. Expression of both enzymes in Saccharomyces cerevisiae can complement the glucose-growth defect of the yeast tps1∆ mutant. Truncation of N-terminal extension in PpTPS1 and PpTPS2 resulted in higher TPS activity and high trehalose levels, upon expression in yeast. Physcomitrella knockout plants were generated and analyzed in various conditions to functionally characterize these proteins. tps1∆ and tps2∆ knockouts displayed a lower amount of caulonema filaments and were significantly reduced in size of gametophores as compared to the wild type. These phenotypes were more pronounced in the tps1∆ tps2∆ mutant. Caulonema formation is induced by factors such as high energy and auxins. Only high amounts of supplied energy were able to induce caulonema filaments in the tps1∆ tps2∆ mutant. Furthermore, this mutant was less sensitive to auxins as NAA-induced caulonema development was arrested in the tps1∆ tps2∆ mutant. In contrast, formation of caulonema filaments is repressed by cytokinins. This effect was more severe in the tps1∆ and tps1∆ tps2∆ mutants. Our results demonstrate that PpTPS1 and PpTPS2 are essential for sensing and signaling sugars and plant hormones to monitor the balance between caulonema and chloronema development.

Evolutionary Engineering of an Iron-Resistant Saccharomyces cerevisiae Mutant and Its Physiological and Molecular Characterization.

Iron plays an essential role in all organisms and is involved in the structure of many biomolecules. It also regulates the Fenton reaction where highly reactive hydroxyl radicals occur. Iron is also important for microbial biodiversity, health and nutrition. Excessive iron levels can cause oxidative damage in cells. Saccharomyces cerevisiae evolved mechanisms to regulate its iron levels. To study the iron stress resistance in S. cerevisiae, evolutionary engineering was employed. The evolved iron stress-resistant mutant “M8FE” was analysed physiologically, transcriptomically and by whole genome re-sequencing. M8FE showed cross-resistance to other transition metals: cobalt, chromium and nickel and seemed to cope with the iron stress by both avoidance and sequestration strategies. PHO84, encoding the high-affinity phosphate transporter, was the most down-regulated gene in the mutant, and may be crucial in iron-resistance. M8FE had upregulated many oxidative stress response, reserve carbohydrate metabolism and mitophagy genes, while ribosome biogenesis genes were downregulated. As a possible result of the induced oxidative stress response genes, lower intracellular oxidation levels were observed. M8FE also had high trehalose and glycerol production levels. Genome re-sequencing analyses revealed several mutations associated with diverse cellular and metabolic processes, like cell division, phosphate-mediated signalling, cell wall integrity and multidrug transporters.

The Effect of the Viscosity of a Trehalose Solution on ATP Hydrolysis by Chloroplast F1-ATPase

The effect of trehalose solution viscosity on the kinetics of ATP hydrolysis by CF1-ATPase (pea chloroplast F1-ATPase) was studied. Trehalose added to the reaction mixture had a double effect: the Mg-dependent enzyme activity was stimulated at trehalose levels below 20 wt % and suppressed at higher trehalose levels. The Ca-dependent activity of CF1 decreases monotonically with the increasing trehalose level. It was shown that stimulation of the Mg-dependent enzyme activity was a result of diminished Mg-ADP-dependent enzyme inactivation. In the case of negligible inactivation, the elevated trehalose content caused a decrease in the maximum rate of ATP hydrolysis and the apparent Michaelis constant increased. The changes in the values of the Michaelis constant and enzyme activity indicate that the delivery of reaction substrates and the conformational changes in CF1-ATPase accompanying the hydrolysis are impeded by the viscosity of the medium. In accordance with Cramer’s rule, the energy loss due to interaction with the medium leads to the conclusion that the efficiency of energy conversion by CF1-ATPase never reaches 100%.

Adult Paederus fuscipes (Coleoptera: Staphylinidae) Beetles Overcome Water Loss With Increased Total Body Water Content, Energy Metabolite Storage, and Reduced Cuticular Permeability: Age, Sex-Specific, and Mating Status Effects on Desiccation.

The ability of Paederus beetles to resist desiccation stress is vital to their adaptability in various ecological niches. How water relations and their response to desiccation vary among adult beetles of different age, sex, and mating status is unclear. We examined the water relations of adult Paederus fuscipes Curtis and the mechanisms used to reduce desiccation stress. One-day-old beetles had an exceptionally high percent total body water (%TBW) content and tolerated a high level of %TBW loss. Newly emerged beetles contained a high level of trehalose and 40 to 60% lipid content of their total dry mass, which allowed them to endure desiccation. Beetles that were 10 wk old and older exhibited reduced cuticular permeability. Glucose, glycogen, and lipid contents were crucial throughout most of the adult life span, as they helped compensate for water loss via increased water vapor absorption and metabolic water. In particular, the accumulation of lipid after mating was significant and may further confer tolerance to water loss. The effect of melanization on the desiccation tolerance of beetles was not significant. Females had better tolerance in response to desiccation stress compared with males. We suggest that the observed differences between sexes likely were a function of water relations and an effect of energy metabolite reserves. However, the mortality of females at 24-h postdesiccating stage was marginally significant compared with males. These results demonstrate that P. fuscipes adults prevent dehydration using multiple mechanisms that collectively reduce desiccation stress and increase dehydration tolerance.

Roots of the Resurrection Plant Tripogon loliiformis Survive Desiccation Without the Activation of Autophagy Pathways by Maintaining Energy Reserves.

Being sessile, plants must regulate energy balance, potentially via source-sink relations, to compromise growth with survival in stressful conditions. Crops are sensitive, possibly because they allocate their energy resources toward growth and yield rather than stress tolerance. In contrast, resurrection plants tightly regulate sugar metabolism and use a series of physiological adaptations to suppress cell death in their vegetative tissue to regain full metabolic capacity from a desiccated state within 72 h of watering. Previously, we showed that shoots of the resurrection plant Tripogon loliiformis, initiate autophagy upon dehydration as one strategy to reinstate homeostasis and suppress cell death. Here, we describe the relationship between energy status, sugar metabolism, trehalose-mediated activation of autophagy pathways and investigate whether shoots and roots utilize similar desiccation tolerance strategies. We show that despite containing high levels of trehalose, dehydrated Tripogon roots do not display elevated activation of autophagy pathways. Using targeted and non-targeted metabolomics, transmission electron microscopy (TEM) and transcriptomics we show that T. loliiformis engages a strategy similar to the long-term drought responses of sensitive plants and continues to use the roots as a sink even during sustained stress. Dehydrating T. loliiformis roots contained more sucrose and trehalose-6-phosphate compared to shoots at an equivalent water content. The increased resources in the roots provides sufficient energy to cope with stress and thus autophagy is not required. These results were confirmed by the absence of autophagosomes in roots by TEM. Upregulation of sweet genes in both shoots and roots show transcriptional regulation of sucrose translocation from leaves to roots and within roots during dehydration. Differences in the cell’s metabolic status caused starkly different cell death responses between shoots and roots. These findings show how shoots and roots utilize different stress response strategies and may provide candidate targets that can be used as tools for the improvement of stress tolerance in crops.

Concomitant changes in radiation resistance and trehalose levels during life stages of Drosophila melanogaster suggest radio-protective function of trehalose

Purpose: During development, various life stages of Drosophila melanogaster (D. melanogaster) show different levels of resistance to gamma irradiation, with the early pupal stage being the most radiation sensitive. This provides us an opportunity to explore the biochemical basis of such variations. The present study was carried out to understand the mechanisms underlying radiation resistance during life stages of D. melanogaster.Materials and methods: Homogenates from all the life stages of D. melanogaster were prepared at stipulated age. These homogenates were used for the determination of (1) enzymatic antioxidants: superoxide dismutase (SOD), catalase, D. melanogaster glutathione peroxidase (DmGPx), and glutathione S-transferase (GST); (2) reducing non-enzymatic antioxidants: total antioxidant capacity (TAC), reduced glutathione (GSH) and non-reducing non-enzymatic antioxidant trehalose; and (3) levels of protein carbonyl (PC) content. Age-dependent changes in radiation resistance and associated biochemical changes were also studied in young (2 d) and old (20 and 30 d) flies.Results: TAC and GSH were found high in the early pupal stage, whereas catalase and DmGPx were found to increase in the early pupal stage. The non-feeding third instar (NFTI) larvae were found to have high levels of SOD and GST, besides NFTI larvae showed high levels of trehalose. A remarkable decrease was observed in radiation resistance and trehalose levels during the early pupal stage. The PC level was the highest during early pupal stage and was the lowest in NFTI larvae. Older flies showed high level of PC compared with young flies.Conclusion: In vitro increments in trehalose concentration correspond to reduced formation of PCs, suggesting a protective role of trehalose against free radicals. A strong correlation between levels of trehalose and PC formation suggests amelioration of proteome damage due to ionizing radiation (IR). Stages with high trehalose levels showed protected proteome and high radiation resistance, suggesting a significant role for this disaccharide in radiation resistance.

Supercooling capacity along with up-regulation of glycerol content in an overwintering butterfly, Parnassius bremeri

Eggs of an endangered butterfly, Parnassius bremeri, hatch during December and young larvae grow during winter and early spring. Cold tolerance is required for survival and growth of the young larvae. Supercooling points (SCPs) were below −20°C for the young larvae compared to −9.8°C for the last fifth instar (L5) larvae, which developed during May. To investigate the depression of SCPs in young third instar (L3) larvae, polyols and free amino acids in hemolymph were quantified using a BioLC. Compared to L5 larvae, L3 larvae had higher polyols (glycerol and mannitol). However, L5 larvae had much higher levels of trehalose than L3 larvae in hemolymph. Increase of glycerol in L3 larvae was explained by enhanced expression of genes associated with glycerol metabolism. RNA-Seq analysis indicated that expression level of glycerol-3-phosphate dehydrogenase in L3 larvae was higher than that of L5 larvae and more number of glycerol kinase contigs was detected in L3 larvae. In contrast, trehalose-6-phosphate dehydrogenase was highly expressed in L5 larvae by more than 3.2 folds compared to L3 larvae. These results suggest that deep supercooling along with glycerol and other small cryoprotectants is associated with cold tolerance of young larvae of P. bremeri.

Preliminary characterization of a Moroccan honey with a predominance of Bupleurum spinosum pollen

Honey with Bupleurum spinosum (zandaz) as a main pollen source has not been the subject of previous detailed study. Therefore, twelve Moroccan samples of this honey were subjected to melissopalynological, physicochemical and microbiological quality characterization, as well as antioxidant activity assessment. From a quality point of view, almost all samples were within the limits established by Codex Alimentarius, and/or the European legislation. All samples presented predominance of B. spinosum pollen (more than 48%). Relatively high levels of trehalose (1.3–4.0 g/100 g) and melezitose (1.5–2.8 g/100 g) were detected. Those sugars, not common in monofloral honeys, could be used as an important factor to discriminate zandaz honey. Flavonoid content correlated positively with the honey color, melanoidin and polyphenol content, and negatively with the IC50 values of scavenging ABTS (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) free radicals, while proline amount correlated negatively with IC50 values of nitric oxide scavenging activity and chelating power. This correlation supports the use of anti-oxidant activities as important variables for PCA (principal component analysis). Both components explained 70% from the given data, and showed certain homogeneity upon analyzed samples independent of the region, suggesting the importance of B. spinosum nectar in the resulting honey characteristics.

Quantitative proteomic analysis of the Saccharomyces cerevisiae industrial strains CAT-1 and PE-2

Brazilian ethanol fermentation process commonly uses baker's yeast as inoculum. In recent years, wild type yeast strains have been widely adopted. The two more successful examples are PE-2 and CAT-1, currently employed in Brazilian distilleries. In the present study, we analyzed how these strains compete for nutrients in the same environment and compared the potential characteristics which affect their performance by applying quantitative proteomics methods. Through the use of isobaric tagging, it was possible to compare protein abundances between both strains during the fermentation process. Our results revealed a better fermentation performance and robustness of CAT-1 strain. The proteomic results demonstrated many possible features that may be linked to the improved fermentation traits of the CAT-1. Proteins involved in response to oxidative stress (Sod1 and Trx1) and trehalose synthesis (Tps3) were more abundant in CAT-1 than in PE-2 after a fermentation batch. Tolerance to oxidative stress and trehalose accumulation were subsequently demonstrated to be enhanced for CAT-1, corroborating the comparative proteomic results. The importance of trehalose and the antioxidant system was confirmed by using mutant stains deleted in Sod1, Trx1 or Tps3. These deletions impaired fermentation performance, strengthening the idea that the abilities of accumulating high levels of trehalose and coping with oxidative stress are crucial for improving fermentation. SignificanceThe importance of the present works emerges from the necessity to better understand the peculiar biological features from two important bioethanol industrial strains of Saccharomyces cerevisiae during batch fermentation. We applied an iTRAQ-based quantitative proteomics analysis to compare these two important strains during batch fermentation and identified possible features involved in the fermentation performance. The results provided by this work will serve as an initial framework for future investigations on the biology of both strains.

Interspecies and Intraspecies Analysis of Trehalose Contents and the Biosynthesis Pathway Gene Family Reveals Crucial Roles of Trehalose in Osmotic-Stress Tolerance in Cassava.

Trehalose is a nonreducing α,α-1,1-disaccharide in a wide range of organisms, and has diverse biological functions that range from serving as an energy source to acting as a protective/signal sugar. However, significant amounts of trehalose have rarely been detected in higher plants, and the function of trehalose in the drought-tolerant crop cassava (Manihot esculenta Crantz) is unclear. We measured soluble sugar concentrations of nine plant species with differing levels of drought tolerance and 41 cassava varieties using high-performance liquid chromatography with evaporative light-scattering detector (HPLC-ELSD). Significantly high amounts of trehalose were identified in drought-tolerant crops cassava, Jatropha curcas, and castor bean (Ricinus communis). All cassava varieties tested contained high amounts of trehalose, although their concentrations varied from 0.23 to 1.29 mg·g−1 fresh weight (FW), and the trehalose level was highly correlated with dehydration stress tolerance of detached leaves of the varieties. Moreover, the trehalose concentrations in cassava leaves increased 2.3–5.5 folds in response to osmotic stress simulated by 20% PEG 6000. Through database mining, 24 trehalose pathway genes, including 12 trehalose-6-phosphate synthases (TPS), 10 trehalose-6-phosphate phosphatases (TPP), and two trehalases were identified in cassava. Phylogenetic analysis indicated that there were four cassava TPS genes (MeTPS1–4) that were orthologous to the solely active TPS gene (AtTPS1 and OsTPS1) in Arabidopsis and rice, and a new TPP subfamily was identified in cassava, suggesting that the trehalose biosynthesis activities in cassava had potentially been enhanced in evolutionary history. RNA-seq analysis indicated that MeTPS1 was expressed at constitutionally high level before and after osmotic stress, while other trehalose pathway genes were either up-regulated or down-regulated, which may explain why cassava accumulated high level of trehalose under normal conditions. MeTPS1 was then transformed into tobacco (Nicotiana benthamiana). Results indicated that transgenic tobacco lines accumulated significant level of trehalose and possessed improved drought stress tolerance. In conclusion, cassava accumulated significantly high amount of trehalose under normal conditions due to multiplied trehalose biosynthesis gene families and constant expression of the active MeTPS1 gene. High levels of trehalose subsequently contributed to high drought stress tolerance.