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Human milk oligosaccharides (HMOs) have attracted much attention in recent years not only as a prebiotic factor but also in particular as an essential component of infant nutrition in relation to their impact on innate immunity. The backbone structures of complex HMOs generally contain single or repetitive lacto-N-biose (type 1) or lactosamine (type 2) units in either linear or branched chains extending from a lactose core. While all known branched structures originate from the 3,6-substitution of the lactosyl core galactose, we here describe a new class of HMOs that tentatively branch at the terminal galactose of 6'-galactosyllactose. Another novel feature of this class of HMOs was found in linear oligo-galactosyl chains linked to one of the N-acetylglucosamine (GlcNAc) branches. The novel structures exhibit general formulas with hexose versus hexosamine contents of 5/2 to 8/2 and can be designated as high-galactose (HG)-HMOs. In addition, up to three fucosyl residues are linked to the octa- to dodecasaccharides, which were detected in two human milk samples from the Lewis blood-group-defined donors. Structural analyses of methylated glycans and their alditols comprised matrix-assisted laser desorption ionization mass spectrometry, electrospray-(collision-induced dissociation) mass spectrometry and linkage analyses by gas chromatography-mass spectrometry of the derived partially methylated alditol acetates. Enzymatic degradation by the application of β1-3,4-specific galactosidase supported the presence of terminal galactose-linked β1-6 to one of the two GlcNAc branches. The mass spectrometry glycomic data have been deposited at the GlycoPOST archive with the data set identifier GPST000191 (Username: franz.hanisch@uni-koeln.de; Password: Soma1Dita2Carb. Watanabe, Y. GlycoPOST realizes FAIR principles for glycomics mass spectrometry data. Nucleic Acids Res. 2021, 49, D1523-D1528).

Background Increased soil salt concentration decreases productivity and changes the physiological and chemical properties of plants. Various omics technologies have been used to understand the salt response in plants but overall changes in the metabolite profiles of spinach (Spinacia oleracea L.) under salt stress have not been studied. In this article, therefore, the changes in mineral and metabolite profiles of spinach plants cultivated with different NaCl concentrations of 0-200 mmol L-1 in the irrigation water were analyzed to investigate the effect of salt stress on nutritional quality. Results Increasing NaCl concentration decreased plant growth due to mineral imbalance. The amounts of minerals (K+ , Ca2+ , and Fe2+ ) were reduced with increasing NaCl concentration, resulting in altered ratios of Na+ :K+ and Na+ :Ca2+ . The change in the mineral ratios due to NaCl irrigation led to a decrease in the height and an increase in the weight of spinach. Moreover, the profiles of 32 metabolites, including flavonoids, amino acids, acidic compounds, sugars, and lipid-related compounds, were altered by NaCl irrigation; most of them showed decreased levels. In particular, at 200 mmol L-1 NaCl, the levels of sucrose, glutamic acid, hexose sugars, and acidic compounds significantly decreased upon NaCl irrigation. Based on these metabolites, a salt-stress-related spinach metabolomic pathway was proposed. Conclusion Sodium chloride irrigation increased mineral imbalance, resulting in decreased plant growth, and the levels of most metabolites involved in energy production, sensory quality, and health benefits decreased with NaCl irrigation. The results suggest that NaCl irrigation negatively affects the nutritional quality of spinach. © 2020 Society of Chemical Industry.

A reduction in cytosolic invertase in Arabidopsis roots changes the sucrose to hexose ratio, resulting in profound alterations of metabolism, growth, development, and patterns of gene expression., Plant roots depend on sucrose imported from leaves as the substrate for metabolism and growth. Sucrose and hexoses derived from it are also signalling molecules that modulate growth and development, but the importance for signalling of endogenous changes in sugar levels is poorly understood. We report that reduced activity of cytosolic invertase, which converts sucrose to hexoses, leads to pronounced metabolic, growth, and developmental defects in roots of Arabidopsis (Arabidopsis thaliana) seedlings. In addition to altered sugar and downstream metabolite levels, roots of cinv1 cinv2 mutants have reduced elongation rates, cell and meristem size, abnormal meristematic cell division patterns, and altered expression of thousands of genes of diverse functions. Provision of exogenous glucose to mutant roots repairs relatively few of the defects. The extensive transcriptional differences between mutant and wild-type roots have hallmarks of both high sucrose and low hexose signalling. We conclude that the mutant phenotype reflects both low carbon availability for metabolism and growth and complex sugar signals derived from elevated sucrose and depressed hexose levels in the cytosol of mutant roots. Such reciprocal changes in endogenous sucrose and hexose levels potentially provide rich information about sugar status that translates into flexible adjustments of growth and development.

Two new phenolic glucosides, including a new O-glycoside (1) and a new C-glycoside (2), were isolated from a marine-derived fungus Aspergillus sp. The structures of new compounds were elucidated through interpretations of spectroscopic evidence and high-resolution electrospray ionization mass spectrometry. The hexose unit of 1 was identified as β-D-glucose by comparison with an authentic sample via HPLC after acid hydrolysis and derivatization. All compounds were evaluated for their ability to inhibit LPS-induced NO production in RAW264.7 macrophages, but none of them displayed significant activity.

Chinese cabbage (Brassica campestris L. ssp. pekinensis) is a widely cultivated vegetable crop in eastern Asia. The flavor of Chinese cabbage was mostly affected by soluble sugar content. To make clear its accumulation and regulation mechanism, the content of different sugars in the internal blade (IB), the internal midrib (IM), the external blade (EB) and the external midrib (EM) of the leafy head were determined during the leafy head formation. The results showed that fructose was the major sugar accumulated in the internal tissues, followed by glucose. IM was the main tissue of sugar accumulation with the highest contents of total soluble sugar and hexose at harvest. RNA sequencing data of the four tissues at 90 DAS showed that the number of differentially expressed genes (DEGs) in IB and IM was the least (2408), while the number of DEGs between the external and internal tissues ranged from 6037 to 8674. The enzyme genes differentially expressed in ‘starch and sucrose metabolism’ pathway was detected during the leafy head development. The expression of two sucrose synthase (SUS) genes, SUS1a (Bra002332) and SUS1b (Bra006587), were always higher in IM than in the other tissues, and positively correlated with the content of total soluble sugar and hexose, respectively. SUS1 was considered to play an important role in leafy head development and sugar accumulation in Chinese cabbage.

The co-utilization of mixed (pentose/hexose) sugars constitutes a challenge for microbial fermentations. The fungus Ashbya gossypii, which is currently exploited for the industrial production of riboflavin, has been presented as an efficient biocatalyst for the production of biolipids using xylose-rich substrates. However, the utilization of xylose in A. gossypii is hindered by hexose sugars. Three A. gossypii homologs (AFL204C, AFL205C and AFL207C) of the yeast HXT genes that code for hexose transporters have been identified and characterized by gene-targeting approaches. Significant differences in the expression profile of the HXT homologs were found in response to different concentrations of sugars. More importantly, an amino acid replacement (N355V) in AFL205Cp, introduced by CRISPR/Cas9-mediated genomic edition, notably enhanced the utilization of xylose in the presence of glucose. Hence, the introduction of the afl205c-N355V allele in engineered strains of A. gossypii will further benefit the utilization of mixed sugars in this fungus. Electronic supplementary material The online version of this article (10.1007/s10295-020-02320-5) contains supplementary material, which is available to authorized users.

Glucose, fructose and mannose are the preferred carbon/energy sources for the yeast Saccharomyces cerevisiae. Absence of preferred energy sources activates glucose derepression, which is regulated by the kinase Snf1. Snf1 phosphorylates the transcriptional repressor Mig1, which results in its exit from the nucleus and subsequent derepression of genes. In contrast, Snf1 is inactive when preferred carbon sources are available, which leads to dephosphorylation of Mig1 and its translocation to the nucleus where Mig1 acts as a transcription repressor. Here we revisit the role of the three hexose kinases, Hxk1, Hxk2 and Glk1, in glucose de/repression. We demonstrate that all three sugar kinases initially affect Mig1 nuclear localization upon addition of glucose, fructose and mannose. This initial import of Mig1 into the nucleus was temporary; for continuous nucleocytoplasmic shuttling of Mig1, Hxk2 is required in the presence of glucose and mannose and in the presence of fructose Hxk2 or Hxk1 is required. Our data suggest that Mig1 import following exposure to preferred energy sources is controlled via two different pathways, where (1) the initial import is regulated by signals derived from metabolism and (2) continuous shuttling is regulated by the Hxk2 and Hxk1 proteins. Mig1 nucleocytoplasmic shuttling appears to be important for the maintenance of the repressed state in which Hxk1/2 seems to play an essential role. Electronic supplementary material The online version of this article (10.1007/s00438-020-01715-4) contains supplementary material, which is available to authorized users.

The aim of this work was to evaluate the general role of the vacuolar pyrophosphatase proton pump (V-PPase) in sucrose accumulation in citrus species. First, three citrus V-PPase genes, designated CsVPP-1, CsVPP-2, and CsVPP-4, were identified in the citrus genome. CsVPP-1 and CsVPP-2 belonging to citrus type I V-PPase genes are targeted to the tonoplast, and CsVPP-4 belonging to citrus type II V-PPase genes is located in the Golgi bodies. Moreover, there was a significantly positive correlation between transcript levels of type I V-PPase genes and sucrose, rather than hexose, content in fruits of seven citrus cultivars. Drought and abscisic acid treatments significantly induced the CsVPP-1 and CsVPP-2 transcript levels, as well as the sucrose content. The overexpression of type I V-PPase genes significantly increased PPase activity, decreased pyrophosphate contents, and increased sucrose contents, whereas V-PPase inhibition produced the opposite effect in both citrus fruits and leaves. Furthermore, altering the expression levels of type I V-PPase genes significantly influenced the transcript levels of sucrose transporter genes. Taken together, this study demonstrated that CsVPP-1 and CsVPP-2 play key roles in sucrose storage in the vacuole by regulating pyrophosphate homeostasis, ultimately the sucrose biosynthesis and transcript levels of sucrose transport genes, providing a novel lead for engineering or breeding modified taste in citrus and other fruits.

To clarify the complex regulatory relationship between changes in sugar content and leaf senescence during the grain-filling stage of rice, genotype-dependent differences in sugar content and the temporal transcriptional patterns of genes involved in sugar signaling pathways were determined in mutant rice exhibiting early leaf senescence and its wild type Zhefu 142. The effects of exogenous glucose or sucrose on the senescence of detached leaves under dark conditions were also investigated. Chlorophyll, soluble sugar, sucrose and fructose contents decreased, whereas electrolytic leakage and malondialdehyde levels increased in mutant leaves at the grain-filling stage. These results suggested that sugar starvation is positively correlated with the early leaf senescence of mutant plants. Detached leaf segments incubated in exogenous sugar solutions under dark conditions exhibited delayed senescence. The high expression of Hxk1 in leaves of mutant plants at the initial grain-filling stage suggested that Hxk1 is involved in the hexose-sensing process at the early stage of leaf senescence. The low expression levels of Hxk2 and Frk1 in the senescing leaves of mutant rice during the grain-filling stage are indicative of weakened hexose phosphorylation. In addition, the high expression levels of SuSy1, SuSy2 and SuSy4 in leaves of mutant plants at the initial grain-filling stage are accompanied by the high transcript levels of SUT1, which favor sucrose translocation and remobilization from the early senescing leaves of mutant rice. The relatively reduced transcript levels of chFBP, cyFBP, SPS1, SPS2 and SPS6 indicated that during the grain-filling stage, sucrose biosynthesis is weakened in the senescing leaves of mutant rice. Keywords: rice, sugar starvation, leaf senescence, signaling pathway, detached leaf segment, grain-filling stage

Shading and drought stresses during the flowering stage of maize (Zea mays L.) production cause seed losses, making them two of the major limitations to yield output. The mechanisms by which the two stresses reduce seed numbers are still unclear. Here, different durations of shading and different degrees of drought were applied, accompanied by synchronous pollination, and the early development of fertilized apical kernels was suppressed, with decreases in sucrose and hexose levels and the promotion of ethylene emission. Consequently, the two distinct stresses led to a common consequence: up to 20–30% of the fertilized kernels were aborted at maturity, which accounted for the yield losses under both scenarios. Notably, by removing the shading net or the preventing pollination of the basal kernels, the apical kernels, were partial or fully restored from abortion, with an increased sugar status and suppressed ethylene emission, even under stress conditions. We found that dry matter accumulation within the early stage was positively correlated with sucrose and hexose contents but negatively correlated with ethylene emission. By altering the sugar status within developing kernels, we verified the antagonistic relationships between endogenous ethylene emission and soluble sugars, including sucrose and hexoses. Collectively, these findings suggest that the equilibrium between sugars and ethylene may determine the development or abortion of early developing kernels exposed to environmental stresses.

Background Biotrophic fungi make intimate contact with host cells to access nutrients. Sugar is considered as the main carbon sources absorbed from host cells by pathogens. Partition, exchanges and competition for sugar at plant-pathogen interfaces are controlled by sugar transporters. Previous studies have indicated that the leaf rust resistance (Lr) gene Lr67, a natural mutation of TaSTP13 encoding a wheat sugar transport protein, confers partial resistance to all three wheat rust species and powdery mildew possibly due to weakened sugar transport activity of TaSTP13 by heterodimerization. However, one major problem that remains unresolved concerns whether TaSTP13 participates in wheat susceptibility to rust and mildew. Results In this study, expression of TaSTP13 was highly induced in wheat leaves challenged by Puccinia striiformis f. sp. tritici (Pst) and certain abiotic treatments. TaSTP13 was localized in the plasma membrane and functioned as homooligomers. In addition, a functional domain for its transport activity was identified in yeast. Suppression of TaSTP13 reduced wheat susceptibility to Pst by barley stripe mosaic virus-induced gene silencing (VIGS). While overexpression of TaSTP13 promoted Arabidopsis susceptibility to powdery mildew and led to increased glucose accumulation in the leaves. Conclusions These results indicate that TaSTP13 is transcriptionally induced and contributes to wheat susceptibility to stripe rust, possibly by promoting cytoplasmic hexose accumulation for fungal sugar acquisition in wheat-Pst interactions.

Rapid pollen tube growth requires uptake of Suc or its hydrolytic products, hexoses, from the apoplast of surrounding tissues in the style. Due to species-specific sugar requirements, reliance of pollen germination and tube growth on cell wall invertase and Suc or hexose transporters varies between species, but it is not known if plants have a sugar transporter that mediates the uptake of both hexose and Suc for pollen tube growth. Here, we show that a sugar transporter protein in apple (Malus domestica), MdSTP13a, takes up both hexose and Suc when expressed in yeast, and is essential for pollen tube growth on Glc and Suc but not on maltose. MdSTP13a-mediated direct uptake of Suc is primarily responsible for apple pollen tube growth on Suc medium. Sorbitol, a major photosynthate and transport carbohydrate in apple, modulates pollen tube growth via the MYB transcription factor MdMYB39L, which binds to the promoter of MdSTP13a to activate its expression. Antisense repression of MdSTP13a blocks sorbitol-modulated pollen tube growth. These findings demonstrate that MdSTP13a takes up both hexose and Suc for sorbitol-modulated pollen tube growth in apple, revealing a situation where acquisition of sugars for pollen tube growth is regulated by a sugar alcohol.

Nostoc microscopicum an effective extracellular polymers (EPs) synthesizer among cyanobacteria was isolated, and its elementary morphological features were defined with the aid of light microscope and CLSM. Bioseparation of EPs from 44 days-aged culture gave 0.90 g/L of the dry powdered extract. Chemical quantification of EPs showed the presence of 550 mg/g of carbohydrate and 395 mg/g of protein. HPLC results depicted the presence of mannose (monomer sugar), fucose (hexose deoxy sugar), mannitol (sugar alcohol) and N- acetylglucosamine (glycoprotein) in the EPs. The vibration-based spectrum produced by FT-IR proves the β-Sheet structure of EPs glycoprotein and the size as 45 kDa by performing SDS-PAGE. Bactericidal activity evaluation of EPs on Pseudomonas aeruginosa and Staphylococcus aureus co-expressed the MIC value as 125 μg/mL, while zone of inhibition was 12 mm for Gram-negative and 8 mm for Gram-positive bacteria. Biofilm inhibition assay was effective in 1.0 mg/mL concentration of EPs in both bacterial strains with a mean rate of 60 percentages which was further confirmed using confocal laser scanning microscopic imaging. This natural polymeric extract of Nostoc microscopicum indicates its possible applications in bactericidal and biofilm inhibition.

Glomalin, one of the glycoproteins generated in the spores and hyphae of arbuscular mycorrhizal (AM) fungi, has multiple functions in plants and soil, while the role of foliar spray of easily extractable glomalin-related soil proteins (EE-GRSP) in citrus fruits is not well defined. Our study aimed to use referenced transcriptome sequencing to uncover the mechanism and the role of exogenous EE-GRSP in two late-ripening varieties of sweet orange (Citrus sinensis) fruits including Navel Lane Late (LW) and Rohde Red Valencia (XC). The 1804 and 1861 differentially expressed genes were identified in fruits of LW and XC, respectively, following foliar spray of EE-GRSP. Photosynthesis ranked second in the Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolism in the LW variety, and carbon fixation in photosynthetic organizations ranked first in KEGG metabolism in the XC variety. The response to foliar spray of EE-GRSP affected the fruit starch and sucrose metabolism of KEGG, with 15 (10 up-regulated and 5 down-regulated) and 13 (2 up-regulated and 11 down-regulated) differentially expressed genes identified in the LW and XC variety, respectively. Cs5g19060 (sucrose phosphate synthase 4) was activated and reduced by EE-GRSP on XC and LW, respectively. Cs1g18220 (β-fructofuranosidase) and Cs2g12180 (glycosyl hydrolase family 9) genes were up-regulated and down-regulated in LW and XC, respectively. These results established the involvement of molecular signaling in response to foliar spray of EE-GRSP activating fruit sugar metabolism is dependent on citrus varieties.

International audience; The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica, a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).

Simultaneous co-fermentation of glucose and xylose is a key desired trait of engineered Saccharomyces cerevisiae for efficient and rapid production of biofuels and chemicals. However, glucose strongly inhibits xylose transport by endogenous hexose transporters of S. cerevisiae. We identified structurally distant sugar transporters (Lipomyces starkeyi LST1_205437 and Arabidopsis thaliana AtSWEET7) capable of co-transporting glucose and xylose from previously unexplored oleaginous yeasts and plants. Kinetic analysis showed that LST1_205437 had lenient glucose inhibition on xylose transport and AtSWEET7 transported glucose and xylose simultaneously with no inhibition. Modelling studies of LST1_205437 revealed that Ala335 residue at sugar binding site can accommodates both glucose and xylose. Docking studies with AtSWEET7 revealed that Trp59, Trp183, Asn145 and Asn179 residues stabilized the sugars, allowing both xylose and glucose to be co-transported. In addition, we altered sugar preference of LST1_205437 by single amino acid mutation at Asn365. Our findings provide a new mechanistic insight on glucose and xylose transport mechanism of sugar transporters and the identified sugar transporters can be employed to develop engineered yeast strains for producing cellulosic biofuels and chemicals. This article is protected by copyright. All rights reserved.

Cucumber (Cucumis sativus L.), an important vegetable plant species, is susceptible to low temperature stress especially during the seedling stage. Vacuolar invertase (VI) plays important roles in plant responses to abiotic stress. However, the molecular and biochemical mechanisms of VI function in cucumber, have not yet been completely understood and VI responses to low temperature stress and it functions in cold tolerance in cucumber seedlings are also in need of exploration. The present study found that hexose accumulation in the roots of cucumber seedlings under low temperature stress is closely related to the observed enhancement of invertase activity. Our genome-wide search for the vacuolar invertase (VI) genes in cucumber identified the candidate VI-encoding gene CsVI1. Expression profiling of CsVI1 showed that it was mainly expressed in the young roots of cucumber seedlings. In addition, transcriptional analysis indicated that CsVI1 expression could respond to low temperature stress. Recombinant CsVI1 proteins purified from Pichia pastoris and Nicotiana benthamiana leaves could hydrolyze sucrose into hexoses. Further, overexpression of CsVI1 in cucumber plants could increase their hexose contents and improve their low temperature tolerance. Lastly, a putative cucumber invertase inhibitor was found could form a complex with CsVI1. In summary, these results confirmed that CsVI1 functions as an acid invertase involved in hexose accumulation and responds to low temperature stress in cucumber seedlings.

Sugars, especially glucose and fructose, contribute to the taste and quality of tomato fruits. These compounds are translocated from the leaves to the fruits and then unloaded into the fruits by various sugar transporters at the plasma membrane. SWEETs, are sugar transporters that regulate sugar efflux independently of energy or pH. To date, the role of SWEETs in tomato has received very little attention. In this study, we performed functional analysis of SlSWEET7a and SlSWEET14 to gain insight into the regulation of sugar transport and storage in tomato fruits. SlSWEET7a and SlSWEET14 were mainly expressed in peduncles, vascular bundles, and seeds. Both SlSWEET7a and SlSWEET14 are plasma membrane-localized proteins that transport fructose, glucose, and sucrose. Apart from the resulting increase in mature fruit sugar content, silencing SlSWEET7a or SlSWEET14 resulted in taller plants and larger fruits (in SlSWEET7a-silenced lines). We also found that invertase activity and gene expression of some SlSWEET members increased, which was consistent with the increased availability of sucrose and hexose in the fruits. Overall, our results demonstrate that suppressing SlSWEET7a and SlSWEET14 could be a potential strategy for enhancing the sugar content of tomato fruits.

Hungateiclostridium thermocellum ATCC 27405 is a promising bacterium for consolidated bioprocessing with a robust ability to degrade lignocellulosic biomass through a multienzyme cellulosomal complex. The bacterium uses the released cellodextrins, glucose polymers of different lengths, as its primary carbon source and energy. In contrast, the bacterium exhibits poor growth on monosaccharides such as fructose and glucose. This phenomenon raises many important questions concerning its glycolytic pathways and sugar transport systems. Until now, the detailed mechanisms of H. thermocellum adaptation to growth on hexose sugars have been relatively poorly explored. In this study, adaptive laboratory evolution was applied to train the bacterium in hexose sugars-based media, and genome resequencing was used to detect the genes that got mutated during adaptation period. RNA-seq data of the first culture growing on either fructose or glucose revealed that several glycolytic genes in the Embden–Mayerhof–Parnas pathway were expressed at lower levels in these cells than in cellobiose-grown cells. After seven consecutive transfer events on fructose and glucose (~42 generations for fructose-adapted cells and ~40 generations for glucose-adapted cells), several genes in the EMP glycolysis of the evolved strains increased the levels of mRNA expression, accompanied by a faster growth, a greater biomass yield, a higher ethanol titer than those in their parent strains. Genomic screening also revealed several mutation events in the genomes of the evolved strains, especially in those responsible for sugar transport and central carbon metabolism. Consequently, these genes could be applied as potential targets for further metabolic engineering to improve this bacterium for bio-industrial usage.

Background Cellulose is not only a common component in vascular plants, but also has great economic benefits for paper, wood, and industrial products. In addition, its biosynthesis is highly regulated by carbohydrate metabolism and allocation in plant. MdFRK2, which encodes a key fructokinase (FRK) in apple, showed especially high affinity to fructose and regulated carbohydrate metabolism. Results It was observed that overexpression of MdFRK2 in apple decreased sucrose (Suc) and fructose (Fru) with augmented FRK activity in stems, and caused the alterations of many phenotypic traits that include increased cellulose content and an increase in thickness of the phloem region. To further investigate the involved mechanisms, we generated FRK2-OE poplar lines OE#1, OE#4 and OE#9 and discovered (1) that overexpression of MdFRK2 resulted in the huge increased cellulose level by shifting the fructose 6-phosphate or glucose 6-phsophate towards UDPG formation, (2) a direct metabolic pathway for the biosynthesis of cellulose is that increased cleavage of Suc into UDP-glucose (UDPG) for cellulose synthesis via the increased sucrose synthase (SUSY) activity and transcript levels of PtrSUSY1, (3) that the increased FRK activity increases the sink strength overall so there is more carbohydrate available to fuel increased cambial activity and that resulted in more secondary phloem. These results demonstrated that MdFRK2 overexpression would significantly changes the photosynthetic carbon flux from sucrose and hexose to UDPG for increased cellulose synthesis. Conclusions The present data indicated that MdFRK2 overexpression in apple and poplar changes the photosynthetic carbon flux from sucrose and hexose to UDPG for stem cellulose synthesis. A strategy is proposed to increase cellulose production by regulating sugar metabolism as a whole.

Human milk oligosaccharides (HMOs) have attracted much attention in recent years not only as a prebiotic factor, but in particular as an essential component in infant nutrition related to their impact in innate immunity. The backbone structures of complex HMOs generally contain single or repetitive lacto-N-biose (type 1) or lactosamine (type 2) units in either linear or branched chains extending from a lactose core. While all known branched structures originate from 3,6-substitution of the lactosyl core galactose, we here describe a new class of HMOs that tentatively branch at terminal galactose of 6’-galactosyllactose. Another novel feature of this class of HMOs was found in linear oligo-galactosyl chains linked to one of the N-acetylglucosamine (GlcNAc) branches. The novel structures exhibit general formulas with hexose vs. hexosamine contents of 5/2 to 8/2 and can be designated as high-galactose (HG)-HMOs. In addition, up to three fucosyl residues are linked to the octa- to dodecasaccharides, which were detected in two human milk samples from Lewis blood group defined donors. Structural analyses of methylated glycans and their alditols comprised MALDI mass spectrometry, ESI-(CID)MS and linkage analyses by GC-MS of the derived partially methylated alditol acetates. Enzymatic degradation by application of β1-3,4-specific galactosidase supported the presence of terminal galactose linked β1−6 to one of the two GlcNAc branches.

Summary • Stomatal opening is caused by guard cell swelling due to an accumulation of osmotica. We investigated the release of carbon from guard cell chloroplasts as a source for the production of organic osmotica. • Photosynthetically active chloroplasts were isolated from guard cell protoplasts of Vicia faba. Export of metabolites into the surrounding medium was analyzed by silicone oil filtering centrifugation and spectrophotometrically by coupled metabolite assays. Effects of external oxaloacetate and 3-phosphoglycerate on photosynthetic electron transport were examined by recording chlorophyll fluorescence. • In the light, guard cell chloroplasts exported triose phosphates, glucose, maltose and hexose phosphates. The presence of phosphate in the medium was essential for the release of phosphorylated compounds and also strongly enhanced the export of glucose and maltose. Total efflux of carbon from illuminated guard cell chloroplasts was on average 486 µatom C (mg Chl)−1 h−1, which was significant with respect to the carbon requirement for stomatal opening. • Metabolites released by illuminated guard cell chloroplasts originated predominantly from starch breakdown. Photosynthetic electron transport provided redox power for the reduction of oxaloacetate and 3-phosphoglycerate.

Insufficient delivery of oligonucleotides to muscle and heart remains a barrier for clinical implementation of antisense oligonucleotide (AO)-mediated exon-skipping therapeutics in Duchenne muscular dystrophy (DMD), a lethal monogenic disorder caused by frame-disrupting mutations in the DMD gene. We previously demonstrated that hexose, particularly an equal mix of glucose:fructose (GF), significantly enhanced oligonucleotide delivery and exon-skipping activity in peripheral muscles of mdx mice; however, its efficacy in the heart remains limited. Here we show that co-administration of GF with peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO, namely, BMSP-PMO) induced an approximately 2-fold higher level of dystrophin expression in cardiac muscles of adult mdx mice compared to BMSP-PMO in saline at a single injection of 20 mg/kg, resulting in evident phenotypic improvement in dystrophic mdx hearts without any detectable toxicity. Dystrophin expression in peripheral muscles also increased. However, GF failed to potentiate BMSP-PMO efficiency in aged mdx mice. These findings demonstrate that GF is applicable to both PMO and PPMO. Furthermore, GF potentiates oligonucleotide activity in mdx mice in an age-dependent manner, and, thus, it has important implications for its clinical deployment for the treatment of DMD and other muscular disorders. Keywords: hexose, peptide-conjugated morpholino oligomer, Duchenne muscular dystrophy, exon skipping, cardiac functions

Background Candida albicans is an opportunistic commensal microorganism, often associated with severe infections in immunosuppressed individuals. C. albicans has hexose transporters that may favor the intracellular accumulation of photosensitizer (PS). the aims of this study were to investigate the influence of glucose load on photodynamic antimicrobial chemotherapy (PACT); and the role that membrane transport system plays on this therapy in the presence of glucose. Material and methods Strains of C. albicans were selected: ATCC 10231, YEM 12, YEM 13, YEM 14 and YEM 15. All strains were grown aerobically on Sabouraud agar and incubated at 30 °C for 24 h. The strains were treated with and without glucose, and divided into Control (no treatment), LED light (660 nm, 166 mW/cm2), Photosensitizer (100 μM methylene blue) and PACT at 1, 3 and 6 min of irradiation groups. The colony forming units were counted and data submitted to statistical analysis (ANOVA) and Tukey's test. The concentration of methylene blue (MB) outside the yeast was measured by fluorescence spectroscopy. Results PACT inactivate C. albicans and the presence of glucose did not affect the killing effect for most strains. Only YEM12 was partially affected by its presence. Regarding efflux systems, ABC overexpressing strain showed a protective effect on the yeast cells. We observed that yeast with overexpression of major facilitator superfamily (MFS) membrane pore tended to accumulate more MB in its cytoplasm, whereas strains that overexpressed ABC pumps (ATP-binding cassette transporters) tended to decrease MB uptake and survive the photodynamic challenge. Conclusion Presence of glucose showed a small effect on PACT . The accumulation of MB on yeast induces more photodynamic inactivation; however, the photodynamic efficacy depends on the type and characteristics of the microbial strain.

Summary Sugar transporters are necessary to transfer hexose from cell wall spaces into parenchyma cells to boost hexose accumulation to high concentrations in fruit. Here, we have identified an apple hexose transporter (HTs), MdHT2.2, located in the plasma membrane, which is highly expressed in mature fruit. In a yeast system, the MdHT2.2 protein exhibited high 14C‐fructose and 14C‐glucose transport activity. In transgenic tomato heterologously expressing MdHT2.2, the levels of both fructose and glucose increased significantly in mature fruit, with sugar being unloaded via the apoplastic pathway, but the level of sucrose decreased significantly. Analysis of enzyme activity and the expression of genes related to sugar metabolism and transport revealed greatly up‐regulated expression of SlLIN5, a key gene encoding cell wall invertase (CWINV), as well as increased CWINV activity in tomatoes transformed with MdHT2.2. Moreover, the levels of fructose, glucose and sucrose recovered nearly to those of the wild type in the sllin5‐edited mutant of the MdHT2.2‐expressing lines. However, the overexpression of MdHT2.2 decreased hexose levels and increased sucrose levels in mature leaves and young fruit, suggesting that the response pathway for the apoplastic hexose signal differs among tomato tissues. The present study identifies a new HTs in apple that is able to take up fructose and glucose into cells and confirms that the apoplastic hexose levels regulated by HT controls CWINV activity to alter carbohydrate partitioning and sugar content.

Although hexoses glucose and fructose serve as important energy sources of food, up to now, there is little information about hexose transporters in birds’ intestinal epithelium during their first postnatal week. The aim of the investigation was to carry out an immunohistochemical study of integral membrane proteins glucose transporter-2 and -5 (GLUT-2 and GLUT-5) on intestinal epithelial cells of ostrich chicks during their first postnatal week. The material from duodenum and ileum was collected from 9 female ostriches (Struthio camelus var. Domesticus) divided into three age groups, three birds in each group: chicks immediately after hatching, 3-day-old ostriches and 7-day-old chicks. The material was fixed in 10% formalin, embedded into paraffin, slices 7μm thick were cut followed by immunohistochemical staining with polyclonal primary antibodies Rabbit anti- GLUT-2 and Rabbit anti-GLUT-5, carried out according to the manufacturer’s guidelines (IHC kit, Abcam, UK). Immunohistochemical localization of GLUT-2 and -5 in the intestinal epithelial cells in ostriches of different age groups was determined. In the groups of chicks after hatching and 3-day-old ostriches, enterocytes in duodenal epithelium were mostly unstained and goblet cells stained weakly for both antibodies. Weak staining of enterocytes and goblet cells was also noted in the ileal epithelium of the chick after hatching. Moderate staining of goblet cells was noted in the 3-day-old chicks’ ileal epithelium. In 7-day-old ostriches, the expression of both antibodies was weak in duodenal but moderate in ileal epithelial cells. The pattern of immunohistochemical expression of GLUT-2 and GLUT-5 in ostriches’ intestinal epithelial cells confirms our hypothesis that the intestinal tract of ostriches after hatching is not yet entirely capable of transportation of hexoses and showed that it is completing gradually during the first postnatal week.

Hexokinase (HXK) catalyses hexose phosphorylation and senses glucose signalling to control carbon partition in plants. In sugarcane, HXK is involved in regulating source/sink carbon balance. However, the characteristics of sugarcane HXK genes remain unclear. This study revealed that the full-length cDNA sequence of the HXK gene ShHXK8 cloned from sugarcane mature leaves contains a 1560 bp open reading frame and encodes a putative protein of 519 amino acids. Amongst HXKs from monocot and dicot subjected to phylogenetic analysis, ShHXK8 was clustered into a monocot-specific group with OsHXK7, SbHXK8 and ZmHXK1. Gene expression analysis in Saccharum spp. hybrid ROC22 plants showed that ShHXK8 was expressed in leaves and stalks and expressed highly in mature leaves and young internodes. The difference in the expression of ShHXK8 was compared in high-sugar-content (HSC) and low-sugar-content (LSC) Saccharum officinarum plants. The expression of ShHXK8 in the leaves and internodes 3–24 of HSC plants was lower than that of LSC plants. The expression of ShHXK8 in the internodes 1–2 of HSC plants was higher than that of LSC plants. ShHXK8–GFP fusion was located in the cytosol by transiently expressing the corresponding vectors in rice mesophyll protoplasts. ShHXK8 might function as a cytoplasmic hexokinase involved in hexose metabolism in sugarcane mature leaves and young internodes.

Silencing expression of SlSWEET1a substantially reduced hexose accumulation in young leaves of tomato plants, revealing the key role of SlSWEET1a in glucose import to sink leaves., Sugar allocation from source to sink (young) leaves, critical for plant development, relies on activities of plasma membrane sugar transporters. However, the key sugar unloading mechanism to sink leaves remains elusive. SWEET transporters mediate sugar efflux into reproductive sinks; therefore, they are promising candidates for sugar unloading during leaf growth. Transcripts of SlSWEET1a, belonging to clade I of the SWEET family, were markedly more abundant than those of all other 30 SlSWEET genes in young leaves of tomatoes. High expression of SlSWEET1a was also detected in reproductive sinks, such as flowers. SlSWEET1a was dominantly expressed in leaf unloading veins, and the green fluorescent protein (GFP) fusion protein was localized to the plasma membrane using Arabidopsis protoplasts, further implicating this carrier in sugar unloading. In addition, yeast growth assays and radiotracer uptake analyses further demonstrated that SlSWEET1a acted as a low-affinity (Km ~100 mM) glucose-specific carrier with a passive diffusion manner. Finally, virus-induced gene silencing of SlSWEET1a expression reduced hexose accumulation to ~50% in young leaves, with a parallel 2-fold increase in mature leaves. Thus, we propose a novel function for SlSWEET1a in the uptake of glucose into unloading cells as part of the sugar unloading mechanism in sink leaves of tomato.

In this study, a novel inoculation method to mitigate the inhibition of 5-hydroxymethylfurfural (5-HMF) is proposed. Acid algae hydrolysate containing 1.5 g 5-HMF/L and 15 g hexose/L hexose was fed to a continuous fixed bed reactor (C-FBR) partially packed with hybrid-immobilized beads. The inoculation method enabled a high rate of H2 production, due to the reduction of 5-HMF inhibition and enhanced biofilm formation. Maximum hydrogen production was achieved at a hydraulic retention time of 6 h with a hydrogen production rate (HPR) of 20.0 ± 3.3 L H2/L-d and a hydrogen yield (HY) of 2.3 ± 0.4 mol H2/mol hexose added. Butyrate and acetate were the major soluble metabolic products released during fermentation. Quantitative real-time polymerase chain reaction analysis revealed that Clostridium butyricum comprised 94.3% of the total bacteria, which was attributed to the high rate of biohydrogen production.

Dark fermentative biohydrogen production mainly relies on strains that follow the butyric acid fermentation pathway and substrate that could be easily fermented, i.e., hexose. In this paper, biohyd...

Saccharomyces paradoxus, a native microorganism of the aguamiel, was used successfully for endoinulinase synthesis for agave fructooligasaccharide (FOS) production. We optimized the fermentation parameters to maximize the enzyme synthesis, and we performed enzyme kinetics studies to achieve agave fructans hydrolysis. The results showed that under constant operating conditions (pH 7.7, 40 °C, 175 rpm of agitation, and 0.005 VVM of aeration) results in the production of an enzymatic extract with 49.57 mg/L. This enzymatic extract, when mixed with an agave fructans solution containing 37.8 g/L, allowed us to obtain products with 18% more FOS content the original concentration. The mass spectrum plot shows that the hydrolyzed product contains FOS with a degree of polymerization from 5 to 9 hexose units. These results are promising because they show FOS production from agave and confirm that importance of using native strains in the design of directed fermentation processes.

Background: Cadmium (Cd) pollution is of serious concern due to its toxic effects in both humans and animals. The study investigates the protective effect of Tinospora cordifolia stem methanolic extract (TCME) on Cd induced hepatotoxicity. Objective(s): The objective of the study was to explore the hepatoprotective effects of T. cordifolia extract. Materials and methods: Rats were administered orally with Cd (5 mg/kg) and TCME (100 mg/kg) for 28 days. At the end of the treatment period, serum and liver tissues homogenates were subjected to biochemical analysis. Results: Cd treated rats showed increased activities of the serum marker enzymes of liver damage such as AST and ALT along with increased levels of LPO and protein carbonyl content in liver tissues. Cd treatment also leads to decreased activities of endogenous antioxidants (SOD, CAT, GSH, GPx and GST), membrane ATPases (Na+K+ATPase, Ca2+ATPase and Mg2+K+ATPase) and the tissue glycoprotein levels (hexose, fucose, hexosamine and sialic acid). Histological analysis revealed vacuolar degeneration of hepatocytes with focal necrosis upon Cd administration. TCME co-treatment restored the biochemical and histological alterations caused by Cd intoxication to near normal levels. Conclusion: The results of the present investigation reveal the hepatoprotective nature of T. cordifolia against Cd induced hepatotoxicity. Keywords: Cadmium, Oxidative stress, Tinospora cordifolia, Hepatotoxicity, Antioxidant

Present knowledge on the quantitative aerobic physiology of the yeastSaccharomyces cerevisiaeduring growth on sucrose as sole carbon and energy source is limited to either adapted cells or to the model laboratory strain CEN.PK113-7D. To broaden our understanding of this matter and open novel opportunities for sucrose-based biotechnological processes, we characterized three strains, with distinct backgrounds, during aerobic batch bioreactor cultivations. Our results reveal that sucrose metabolism inS. cerevisiaeis a strain-specific trait. Each strain displayed a distinct extracellular hexose concentration and invertase activity profiles. Especially, the inferior maximum specific growth rate (0.21 h−1) of the CEN.PK113-7D strain, with respect to that of strains UFMG-CM-Y259 (0.37 h−1) and JP1 (0.32 h−1), could be associated to its low invertase activity (0.04 to 0.09 U mgDM−1). Moreover, comparative experiments with glucose or fructose alone, or in combination, suggest mixed mechanisms of sucrose utilization by the industrial strain JP1, and points out the remarkable ability of the wild isolate UFMG-CM-259 to grow faster on sucrose than on glucose in a well-controlled cultivation system. This work hints to a series of metabolic traits that can be exploited to increase sucrose catabolic rates and bioprocess efficiency.Abstract Figure

Quantitative traits such as human height are measurable phenotypes that show continuous variation over a wide phenotypic range. Enormous effort has recently been put into determining the genetic influences on a variety of quantitative traits, including human genetic diseases, with mixed success. We identified a quantitative trait in a tractable model system, the GAL pathway in yeast, which controls the uptake and metabolism of the sugar galactose. GAL pathway activation depends both on galactose concentration and on the concentrations of competing, preferred sugars such as glucose. Natural yeast isolates show substantial variation in the behavior of the pathway. All studied yeast strains exhibit bimodal responses relative to external galactose concentration, i.e. a set of galactose concentrations existed at which both GAL-induced and GAL-repressed subpopulations were observed. However, these concentrations differed in different strains. We built a mechanistic model of the GAL pathway and identified parameters that are plausible candidates for capturing the phenotypic features of a set of strains including standard lab strains, natural variants, and mutants. In silico perturbation of these parameters identified variation in the intracellular galactose sensor, Gal3p, the negative feedback node within the GAL regulatory network, Gal80p, and the hexose transporters, HXT, as the main sources of the bimodal range variation. We were able to switch the phenotype of individual yeast strains in silico by tuning parameters related to these three elements. Determining the basis for these behavioral differences may give insight into how the GAL pathway processes information, and into the evolution of nutrient metabolism preferences in different strains. More generally, our method of identifying the key parameters that explain phenotypic variation in this system should be generally applicable to other quantitative traits.Author summaryMicrobes adopt elaborate strategies for the preferred uptake and use of nutrients to cope with complex and fluctuating environments. As a result, yeast strains originating from different ecological niches show significant variation in the way they induce genes in the galactose metabolism (GAL) pathway in response to nutrient signals. To identify the mechanistic sources of this variation, we built a mathematical model to simulate the dynamics of the galactose metabolic regulation network, and studied how parameters with different biological implications contributed to the natural variation. We found that variations in the behavior of the galactose sensor Gal3p, the negative feedback node Gal80p, and the hexose transporters HXT were critical elements in the GAL pathway response. Tuning single parameters in silico was sufficient to achieve phenotype switching between different yeast strains. Our computational approach should be generally useful to help pinpoint the genetic and molecular bases of natural variation in other systems.

The aim of this study was to investigate the effect of 20 mg/L 2,4-D on the quality during ripening of on-tree longan fruits. We investigated external and internal properties of the on-tree longan fruits. The results showed that 2,4-D treatment promotes the growth based on the fruit size and weight. The respiration rate, contents of TSS, total soluble sugar, sucrose, glucose, fructose, and hexose revealed an increasing tendency with advancing the maturity and reached the high status during 110-126DPA. By contrast, the relative electric conductivity and TA content displayed a declining trend during the ripening stage, increase with the senescence. All these pieces of information indicated that 2,4-D treatment could effectively promote the sensory quality of on-tree longan fruit, prolong the harvest time to 118DPA, while CK should harvest before 110DPA.

During recent years we have advanced our understanding of plant molecular responses to ultraviolet radiation (UV, 280-400 nm); however, how plants respond to UV radiation under different spectral light qualities is poorly understood. In this study, cucumber plants (Cucumis sativus ‘Lausanna RZ F1’) were grown under monochromatic blue, green, red and broadband white light in combination with UV radiation. The effects of light quality and UV radiation on acclimatory responses were assessed by measuring transcript accumulation of ELONGATED HYPOCOTYL 5 (HY5), CHALCONE SYNTHASE 2 (CHS2) and LIGHT HARVESTING COMPLEX II (LHCII), and the accumulation of flavonoids and hydroxycinnamic acids in the leaves. The growth light backgrounds differentially regulated gene expression and metabolite accumulation. While HY5 and CHS2 transcripts were induced by blue and white light, LHCII was induced by white and red light. Furthermore, UV radiation antagonized the effects of blue, red, green, and white light on transcript accumulation in a gene dependent manner. Plants grown under blue light with supplementary UV radiation increased phenylalanine, flavonol disaccharide I and caffeic acid contents compared to those exposed only to blue light. UV radiation also induced the accumulation of flavonol disaccharide I and II, ferulic acid hexose and coumaric acid hexose in plants grown under green light. Our findings provide further understanding of plant responses to UV radiation in combination with different light spectra and contribute to the design of light recipes for horticultural practices that aim to modify plant metabolism and ultimately improve crop quality., Funding agencies:GUDP (Danish Ministry of Food, Agriculture and Fisheries)Faculty for Business, Science and Technology at Örebro UniversityÖrebro University

Hungateiclostridium thermocellum ATCC 27405 is a promising bacterium for consolidated bioprocessing with a robust ability to degrade lignocellulosic biomass through a multienzyme cellulosomal complex. The bacterium uses the released cellodextrins, glucose polymers of different lengths, as its primary carbon source and energy. In contrast, the bacterium exhibits poor growth on monosaccharides such as fructose and glucose. This phenomenon raises many important questions concerning its glycolytic pathways and sugar transport systems. Until now, the detailed mechanisms of H. thermocellum adaptation to growth on hexose sugars have been relatively poorly explored. In this study, adaptive laboratory evolution was applied to train the bacterium in hexose sugars-based media, and genome resequencing was used to detect the genes that got mutated during adaptation period. RNA-seq data of the first culture growing on either fructose or glucose revealed that several glycolytic genes in the Embden–Mayerhof–Parnas pathway were expressed at lower levels in these cells than in cellobiose-grown cells. After seven consecutive transfer events on fructose and glucose (~42 generations for fructose-adapted cells and ~40 generations for glucose-adapted cells), several genes in the EMP glycolysis of the evolved strains increased the levels of mRNA expression, accompanied by a faster growth, a greater biomass yield, a higher ethanol titer than those in their parent strains. Genomic screening also revealed several mutation events in the genomes of the evolved strains, especially in those responsible for sugar transport and central carbon metabolism. Consequently, these genes could be applied as potential targets for further metabolic engineering to improve this bacterium for bio-industrial usage.

Background The members of the genus Caldicellulosiruptor have the potential for future integration into a biorefinery system due to their capacity to generate hydrogen close to the theoretical limit of 4 mol H2/mol hexose, use a wide range of sugars and can grow on numerous lignocellulose hydrolysates. However, members of this genus are unable to survive in high sugar concentrations, limiting their ability to grow on more concentrated hydrolysates, thus impeding their industrial applicability. In this study five members of this genus, C.owensensis, C. kronotskyensis, C.bescii, C.acetigenus and C.kristjanssonii, were developed to tolerate higher sugar concentrations through an adaptive laboratory evolution (ALE) process. The developed mixed population C.owensensis CO80 was further studied and accompanied by the development of a kinetic model based on Monod kinetics to quantitatively compare it with the parental strain. Results Mixed populations of Caldicellulosiruptor tolerant to higher glucose concentrations were obtained with C.owensensis adapted to grow up to 80 g/L glucose; other strains in particular C. kristjanssonii demonstrated a greater restriction to adaptation. The C.owensensis CO80 mixed population was further studied and demonstrated the ability to grow in glucose concentrations up to 80 g/L glucose, but with reduced volumetric hydrogen productivities ($$Q_{{{\text{H}}_{2} }}$$ Q H 2 ) and incomplete sugar conversion at elevated glucose concentrations. In addition, the carbon yield decreased with elevated concentrations of glucose. The ability of the mixed population C.owensensis CO80 to grow in high glucose concentrations was further described with a kinetic growth model, which revealed that the critical sugar concentration of the cells increased fourfold when cultivated at higher concentrations. When co-cultured with the adapted C.saccharolyticus G5 mixed culture at a hydraulic retention time (HRT) of 20 h, C.owensensis constituted only 0.09–1.58% of the population in suspension. Conclusions The adaptation of members of the Caldicellulosiruptor genus to higher sugar concentrations established that the ability to develop improved strains via ALE is species dependent, with C.owensensis adapted to grow on 80 g/L, whereas C.kristjanssonii could only be adapted to 30 g/L glucose. Although C.owensensis CO80 was adapted to a higher sugar concentration, this mixed population demonstrated reduced $$Q_{{{\text{H}}_{2} }}$$ Q H 2 with elevated glucose concentrations. This would indicate that while ALE permits adaptation to elevated sugar concentrations, this approach does not result in improved fermentation performances at these higher sugar concentrations. Moreover, the observation that planktonic mixed culture of CO80 was outcompeted by an adapted C.saccharolyticus, when co-cultivated in continuous mode, indicates that the robustness of CO80 mixed culture should be improved for industrial application.

Tomato, an important fruit crop worldwide, requires efficient sugar allocation for fruit development. However, molecular mechanisms for sugar import to fruits remain poorly understood. Expression of SWEET (Sugars Will Eventually be Exported Transporters) proteins is closely linked with hexose ratio in tomato fruits and may be involved in sugar allocation. Here, using quantitative PCR, we discovered that SlSWEET15 was highly expressed in developing fruits compared to vegetative organs. Based on in situ hybridization and GUS fusion analyses, SlSWEET15 proteins accumulated in vascular tissues and seed coats, major sites of sucrose unloading in fruits. Localizing SlSWEET15-GFP to the plasma membrane supported its putative role in apoplasmic sucrose unloading. The sucrose transport activity of SlSWEET15 was confirmed by complementary growth assays in a yeast mutant. Elimination of the SlSWEET15 function by CRISPR/cas9 gene editing significantly decreased average sizes and weights of fruits, with severe defects in seed filling and embryo development. Together, we confirmed the role of SlSWEET15 in mediating sucrose efflux from the releasing phloem to the fruit apoplasm and subsequent import into parenchyma cells during fruit development. Furthermore, SlSWEET15-mediated sucrose efflux was also required for sucrose unloading from the seed coat to the developing embryo.One-sentence SummarySlSWEET15, a specific sucrose uniporter in tomato, mediates apoplasmic sucrose unloading from releasing phloem cells and seed coat for carbon supply during fruit expansion and seed filling.

We hypothesized that maternal nutrition during the first 50 d of gestation would influence the abundance of hexose transporters, SLC2A1, SLC2A3, and SLC2A5, and cationic amino acid transporters, SLC7A1 and SLC7A2, in heifer uteroplacental tissues. Angus-cross heifers (n = 43) were estrus synchronized, bred via artificial insemination, and assigned at breeding to 1 of 2 dietary intake groups (CON = 100% of requirements to achieve 0.45 kg/d of BW gain or RES = 60% of CON intake) and ovariohysterectomized on day 16, 34, or 50 of gestation (n = 6 to 9/d) in a completely randomized design with a 2 × 3 factorial arrangement of treatments. Uterine cross-sections were collected from the horn ipsilateral to the corpus luteum, fixed in 10% neutral buffered formalin, sectioned at 5 µm, and stained via immunofluorescence for transporters. For each image, areas of fetal membrane (FM; chorioallantois), luminal epithelium (ENDO), superficial glands (SG), deep glands (DG), and myometrium (MYO) were analyzed separately for relative intensity of fluorescence as an indicator of transporter abundance. Analysis of FM was only conducted for days 34 and 50. No transporters in target areas were influenced by a day × treatment interaction (P ≥ 0.06). In ENDO, all transporters were differentially abundant from days 16 to 50 of gestation (P ≤ 0.04), and SLC7A2 was greater (P = 0.05) for RES vs. CON. In SG, SLC7A1 and SLC7A2 were greater (P ≤ 0.04) at day 34 vs. day 16. In DG, SLC2A3 and SLC7A1 were greater (P ≤ 0.05) for CON vs. RES heifers; furthermore, SLC7A1 was greater (P < 0.01) at day 50 vs. days 16 and 34 of gestation. In MYO, SLC7A1 was greater (P < 0.01) for CON vs. RES and was greater (P = 0.02) at days 34 and 50 vs. day 16. There were no differences in FM (P ≥ 0.06). Analysis of all uterine tissues at day 16 determined that SLC2A1, SLC2A3, and SLC7A2 were all differentially abundant across uterine tissue type (P < 0.01), and SLC7A1 was greater (P = 0.02) for CON vs. RES. Analysis of all uteroplacental tissues at days 34 and 50 demonstrated that all transporters differed (P < 0.01) across uteroplacental tissues, and SLC7A1 was greater (P < 0.01) for CON vs. RES. These data are interpreted to imply that transporters are differentially affected by day of gestation, and that hexose and cationic amino acid transporters are differentially abundant across utero-placental tissue types, and that SLC7A1 is responsive to maternal nutritional treatment.

International audience; LC-HRMS Q-TOF method was used to identify and quantify metabolites ofFoleyola billotiiMaire plant. A series of 15 compounds were tentatively identified: three glucosinolates, 2 aliphatic (progoitrin and gluconapin) and 1 indolic (glucobrassicin); six chlorogenic acids, four coumaroyl quinic acid (CoQA) isomers and two feruloyl quinic acid (FQA) isomers; two diglycosylated flavonols, one quercetin-derived and one kaempferol-derived; three sinapic acid derivatives, sinapoyl threonic acid, hexose sinapate and a third derivative supposedly attributed to a feruloyl quinic acid sinapate; and finally, a glycosylatedtrans-cinnamic acid which could correspond to feruloylthreonic acid (FTA). Glucosinolates content was 55 times higher in cultivated plants; chlorogenic acids and flavonols increased too by 5 and 3.5 times, respectively. In contrast, sinapates decreased by 3 times in cultivated plants, this last result can be explained by reduced UV-B intensity in the north of Algeria. The change inF. billotiicompound profile was more quantitative rather than qualitative.

We previously reported the precise structure of acidic free-glycans in human urine. In the present study, structural analysis of neutral free-glycans in urine was performed in fine detail. Urine samples were collected from 21 healthy volunteers and free-glycans extracted from the creatinine-adjusted urine and then fluorescently labeled with 2-aminopyridine. Neutral glycan profiling was achieved by a combination of high-performance liquid chromatography, mass spectrometry, enzymatic digestion, and periodate cleavage. A total of 79 glycans were identified. Because the ABO-blood group antigen containing urinary neutral glycans are major components, profiling patterns were similar between individuals of the same ABO-group. The neutral glycans were composed of lactose-core (Galβ1-4Glc) glycans, type-II N-acetyllactosamine-core (GlcNAcβ1-4Glc) glycans, hexose oligomers, N-glycans and to our surprise β1-3 galactosylglucose-core (Galβ1-3Glc) glycans. Although glycans with a β1-3 galactosylglucose-core were identified as major components in urine, comprising structurally simple isomers of a lactose-core, the core structure has not previously been reported. The major β1-3 galactosylglucose-core glycans were Fucα1-2Galβ1-3(Fucα1-4)Glc, GalNAcα1-3(Fucα1-2)Galβ1-3(Fucα1-4)Glc and Galα1-3(Fucα1-2)Galβ1-3(Fucα1-4)Glc, corresponding to H-, A-, and B-blood group antigens, respectively. Three lactosamine extended β1-3 galactosylglucose-core glycans were also detected as minor components. Elucidating the biosynthesis of β1-3 galactosylglucose will be crucial for understanding the in vivo function of these glycans.

In the yeast Saccharomyces cerevisiae, microbial fuels and chemicals production on lignocellulosic hydrolysates is constrained by poor sugar transport. For biotechnological applications, it is desirable to source transporters with novel or enhanced function from nonconventional organisms in complement to engineering known transporters. Here, we identified and functionally screened genes from three strains of early-branching anaerobic fungi (Neocallimastigomycota) that encode sugar transporters from the recently discovered Sugars Will Eventually be Exported Transporter (SWEET) superfamily in Saccharomyces cerevisiae. A novel fungal SWEET, NcSWEET1, was identified that localized to the plasma membrane and complemented growth in a hexose transporter-deficient yeast strain. Single cross-over chimeras were constructed from a leading NcSWEET1 expression-enabling domain paired with all other candidate SWEETs to broadly scan the sequence and functional space for enhanced variants. This led to the identification of a chimera, NcSW1/PfSW2:TM5-7, that enhanced the growth rate significantly on glucose, fructose, and mannose. Additional chimeras with varied cross-over junctions identified residues in TM1 that affect substrate selectivity. Furthermore, we demonstrate that NcSWEET1 and the enhanced NcSW1/PfSW2:TM5-7 variant facilitated novel co-consumption of glucose and xylose in S. cerevisiae. NcSWEET1 utilized 40.1% of both sugars, exceeding the 17.3% utilization demonstrated by the control HXT7(F79S) strain. Our results suggest that SWEETs from anaerobic fungi are beneficial tools for enhancing glucose and xylose co-utilization and offers a promising step towards biotechnological application of SWEETs in S. cerevisiae.

International audience; To examine the physiological role of hexose transporters in determining the sink strength of individual fruits, the regulation of hexose transporters gene expression was studied when the sink/source ratio was artificially altered under the greenhouse condition; this was done in two cultivars of tomato, i.e. Grandella and Isabella. The sink/source ratio treatments included: saving one fruit per truss (1F), two fruits per truss (2F), three fruits per truss (3F) and no fruit pruning (control). The results showed that fruit thinning could increase starch, sucrose, and hexose contents in the fruits; it could also modulate the activity of the key enzymes and the expression of tomato hexose transporter genes (LeHTs). Based on the relative transcript levels, all examined LeHTs were unregulated at the end of cell division and the cell expansion stage of fruit development, but the strongest expression level observed at the onset of ripening was related to LeHT1 and LeHT2. Given the concomitancy of cell wall invertase (EC 3.2.1.26) activity and the LeHTs relative expression cell wall, invertase activity seemed to be involved in the expression level of LeHTs. The increased trends of the LeHTs expression with the decrease of the sink/source ratio confirmed the role of hexose transporters in determining the sink strength of the tomato fruits.

This study examined the feasibility of dark fermentative biohydrogen production from food waste using hybrid immobilization in mesophilic condition. Among four different organic loading rates (OLRs), the highest average hydrogen production rate (HPR) of 9.82 ± 0.30 L/L-d was found at an OLR of 74.7 g hexose/L-d, which was higher than reported values from particulate feedstock in mesophilic condition. The average hydrogen yield (HY) at the condition was 1.25 ± 0.04 mol H2/mol hexoseconsumed. Whereas the average HPR and HY at an OLR 80 g hexose/L-d were 5.82 ± 0.12 L/L-d and 0.64 ± 0.02 mol H2/mol hexoseconsumed, respectively. Metabolic flux analysis showed the low HY was concurrent with the highest propionic acid and homoacetogenis. Bacterial population was shift from Clostridium sp. to non-hydrogen producers including Bifidobacterium, Bacteriodes, Olsenella, Dysgonomonas, and Dialister sp.

The Crabtree effect occurs under high-glucose concentrations and is characterized by the increase of the growth and a decrease in mitochondrial respiration of yeasts. Regulation of the Crabtree effect could enhance ethanol production with biotechnological purposes and a better understanding of the etiology of cancer due to its similitude with the Warburg effect. Nonetheless, the conclusive molecular mechanism of the Crabtree effect is still on debate. The pathway Snf1p/Hxk2p/Mig1p has been linked with the transcriptional regulation of the hexose transporters and has also been identified in the modulation of phenotypes related to the Crabtree effect. Nevertheless, it has not been directly identified the genetic regulation of the hexose transporters with modulation of the Crabtree effect phenotypes by Snf1p/Hxk2p/Mig1p pathway. In this sense, we provide evidence that the deletion of the SNF1 and HXK2 genes affects the exponential growth, mitochondrial respiration, and the transcription of hexose transporters in a glucose-dependent manner in Saccharomyces cerevisiae. The Vmax of the main hexose transporters transcribed showed a positive correlation with the exponential growth and a negative correlation with the mitochondrial respiration. Transcription of the gene HXT2 was the most affected by the deletion of the pathway SNF1/HXK2/MIG1. Deletion of the orthologous genes SNF1 and HXK2 in the Crabtree negative yeast, K. marxianus, has a differential effect in exponential growth and mitochondrial respiration in comparison with S. cerevisiae. Overall, these results indicate that the SNF1/HXK2/MIG1 pathway transcriptionally regulates the hexose transporters having an influence in the exponential growth and mitochondrial respiration in a glucose-dependent manner.

Diurnal and seasonal changes of abiotic environmental factors shape plant performance and distribution. Changes of growth temperature and light intensity may vary significantly on a diurnal, but also on a weekly or seasonal scale. Hence, acclimation to a changing temperature and light regime is essential for plant survival and propagation. In the present study, we analyzed photosynthetic CO2 assimilation and metabolic regulation of the central carbohydrate metabolism in two natural accessions of Arabidopsis thaliana that originate from north western Russia and south Italy during exposure to heat and a combination of heat and high light. Our findings indicate that it is hardly possible to predict photosynthetic capacities under combined stress from single stress experiments. Further, capacities of hexose phosphorylation were found to be significantly lower in the Italian than in the Russian accession, which could explain an inverted sucrose-to-hexose ratio. Together with the finding of significantly stronger accumulation of anthocyanins under heat/high light, these observations indicate a central role of hexokinase activity in the stabilization of photosynthesis and carbohydrate metabolism during environmental changes.

The sugar content of grape berries is affected by many factors. To explore the hexose content in different cultivars, the photosynthesis, vegetative, and reproductive biomass, as well as the enzyme activities and expression levels of genes related to sugar metabolism and sugar contents were measured. Samples were collected 70-110 days after anthesis (DAA), from Riesling (RI), Petit Manseng (PM), and Cabernet Sauvignon (CS) berries cultivated in the field. The results indicated that high expression levels of VvSWEET15 and VvSS3 and a high activity of sucrose synthase (SS) are associated with a higher hexose content in the berries of PM than in the berries of the other two cultivars. These genes promoted hexose accumulation in the berries by regulating sugar hydrolysis and transport. The results of this study indicate that active sugar hydrolysis and transport increase the hexose content of PM berries, which provides insights for grape berry quality improvement and breeding projects in wine production. Main Conclusion: The active VvSS3, sucrose synthase (SS), and VvSWEET15 increases the hexose content in Petit Manseng berries, which are associated with sugar hydrolysis and transport.

The mechanisms underlying nitrogen (N)-regulated plant-fungi interactions are not well understood. N application modulates plant carbohydrate (C) sinks and is involved in the overall plant-fungal association. We hypothesized that N regulates plant-fungi interactions by influencing the carbohydrate metabolism. The mutualistic fungus Phomopsis liquidambaris was found to prioritize host hexose resources through in vitro culture assays and in planta inoculation. Rice-Ph. liquidambaris systems were exposed to N gradients ranging from N-deficient to N-abundant conditions to study whether and how the sugar composition was involved in the dynamics of N-mediated fungal colonization. We found that root soluble acid invertases were activated, resulting in increased hexose fluxes in inoculated roots. These fluxes positively influenced fungal colonization, especially under N-deficient conditions. Further experiments manipulating the carbohydrate composition and root invertase activity through sugar feeding, chemical treatments and the use of different soil types revealed that the external disturbance of root invertase could reduce endophytic colonization and eliminate endophyte-induced host benefits under N-deficient conditions. Collectively, these results suggest that the activation of root invertase is related to N deficiency-enhanced endophytic colonization via increased hexose generation. Certain combinations of farmland ecosystems with suitable N inputs could be implemented to maximize the benefits of plant-fungi associations.