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1.
Int J Biol Macromol ; 229: 766-777, 2023 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-36610562

RESUMO

Invertases are ubiquitous enzymes that catalyze the unalterable cleavage of sucrose into glucose and fructose, and are crucially involved in plant growth, development and stress response. In this study, a total of 17 putative invertase genes, including 3 cell wall invertases, 3 vacuolar invertases, and 11 neutral invertases were identified in apple genome. Subcellular localization of MdNINV7 and MdNINV11 indicated that both invertases were located in the cytoplasm. Comprehensive analyses of physicochemical properties, chromosomal localization, genomic characterization, and gene evolution of MdINV family were conducted. Gene duplication revealed that whole-genome or segmental duplication and random duplication might have been the major driving force for MdINVs expansion. Selection index values, ω, showed strong evidence of positive selection signatures among the INV clusters. Gene expression analysis indicated that MdNINV1/3/6/7 members are crucially involved in fruit development and sugar accumulation. Similarly, expression profiles of MdCWINV1, MdVINV1, and MdNINV1/2/7/11 suggested their potential roles in response to cold stress. Furthermore, overexpression of MdNINV11 in apple calli at least in part promoted the expression of MdCBF1-5 and H2O2 detoxification in response to cold. Overall, our results will be useful for understanding the functions of MdINVs in the regulation of apple fruit development and cold stress response.


Assuntos
Malus , beta-Frutofuranosidase , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo , Malus/genética , Malus/metabolismo , Peróxido de Hidrogênio/metabolismo , Família Multigênica , Filogenia , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
2.
BMC Genomics ; 24(1): 18, 2023 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-36639618

RESUMO

BACKGROUND: The importance of uridine 5'-diphosphate glucose (UDP-G) synthesis and degradation on carbon (C) partitioning has been indicated in several studies of plant systems, whereby the kinetic properties and abundance of involved enzymes had a significant effect upon the volume of C moving into the hemicellulose, cellulose and sucrose pools. In this study, the expression of 136 genes belonging to 32 gene families related to UDP-G metabolism was studied in 3 major sugarcane organs (including leaf, internode and root) at 6 different developmental stages in 2 commercial genotypes. RESULTS: Analysis of the genes associated with UDP-G metabolism in leaves indicated low expression of sucrose synthase, but relatively high expression of invertase genes, specifically cell-wall invertase 4 and neutral acid invertase 1-1 and 3 genes. Further, organs that are primarily responsible for sucrose synthesis or bioaccumulation, i.e., in source organs (mature leaves) and storage sink organs (mature internodes), had very low expression of sucrose, cellulose and hemicellulose synthesis genes, specifically sucrose synthase 1 and 2, UDP-G dehydrogenase 5 and several cellulose synthase subunit genes. Gene expression was mostly very low in both leaf and mature internode samples; however, leaves did have a comparatively heightened invertase and sucrose phosphate synthase expression. Major differences were observed in the transcription of several genes between immature sink organs (roots and immature internodes). Gene transcription favoured utilisation of UDP-G toward insoluble and respiratory pools in roots. Whereas, there was comparatively higher expression of sucrose synthetic genes, sucrose phosphate synthase 1 and 4, and comparatively lower expression of many genes associated with C flow to insoluble and respiratory pools including myo-Inositol oxygenase, UDP-G dehydrogenase 4, vacuolar invertase 1, and several cell-wall invertases in immature internodes. CONCLUSION: This study represents the first effort to quantify the expression of gene families associated with UDP-G metabolism in sugarcane. Transcriptional analysis displayed the likelihood that C partitioning in sugarcane is closely related to the transcription of genes associated with the UDP-G metabolism. The data presented may provide an accurate genetic reference for future efforts in altering UDP-G metabolism and in turn C partitioning in sugarcane.


Assuntos
Saccharum , Saccharum/metabolismo , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo , Difosfato de Uridina/metabolismo , Sacarose/metabolismo , Celulose/metabolismo , Glucose/metabolismo , Oxirredutases/metabolismo
3.
BMC Plant Biol ; 22(1): 574, 2022 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-36496357

RESUMO

BACKGROUND: Cassava (Manihot esculenta Crantz) is an important multiuse crop grown for economic and energy purposes. Its vegetative organs are storage roots, in which the main storage material is starch. The accumulation characteristics of starch in cassava roots can directly affect the yield, starch content and maturation of cassava storage roots. In this study, we used a cassava sexual tetraploid (ST), which showed early maturation heterosis in previous work, as the main test material. We analyzed the sucrose metabolism and starch accumulation characteristics of the ST and its parents from the leaf "source" to the storage root "sink" during different developmental stages and explored the regulatory mechanisms of ST storage root early maturation by combining the transcriptome data of the storage roots during the expansion period. RESULTS: The results showed that the trends in sucrose, glucose and fructose contents in the ST leaves were similar to those of the two parents during different stages of development, but the trends in the ST storage roots were significantly different from those of their parents, which showed high sucrose utilization rates during the early stage of development and decreased utilization capacity in the late developmental stage. Transcriptome data showed that the genes that were expressed differentially between ST and its parents were mainly involved in the degradation and utilization of sucrose in the storage roots, and four key enzyme genes were significantly upregulated (Invertase MeNINV8/MeVINV3, Sucrose synthase MeSuSy2, Hexokinase MeHXK2), while the expressions of key enzyme genes involved in starch synthesis were not significantly different. CONCLUSIONS: The results revealed that the pattern of sucrose degradation and utilization in the cassava ST was different from that of its parents and promoted early maturation in its tuberous roots. Starch accumulation in the ST from sucrose mainly occurred during the early expansion stage of the storage roots, and the starch content during this period was higher than that of both parents, mainly due to the regulation of invertase and hexokinase activities during sucrose metabolism. This study provides a basis for further genetic improvements to cassava traits and for breeding varieties that mature early and are adapted well to provide starch supply requirements.


Assuntos
Regulação da Expressão Gênica de Plantas , Manihot , Raízes de Plantas/metabolismo , Melhoramento Vegetal , Amido/metabolismo , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo , Tetraploidia , Sacarose/metabolismo
4.
Int J Mol Sci ; 23(23)2022 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-36499311

RESUMO

Rhodotorula dairenensis ß-fructofuranosidase is a highly glycosylated enzyme with broad substrate specificity that catalyzes the synthesis of 6-kestose and a mixture of the three series of fructooligosaccharides (FOS), fructosylating a variety of carbohydrates and other molecules as alditols. We report here its three-dimensional structure, showing the expected bimodular arrangement and also a unique long elongation at its N-terminus containing extensive O-glycosylation sites that form a peculiar arrangement with a protruding loop within the dimer. This region is not required for activity but could provide a molecular tool to target the dimeric protein to its receptor cellular compartment in the yeast. A truncated inactivated form was used to obtain complexes with fructose, sucrose and raffinose, and a Bis-Tris molecule was trapped, mimicking a putative acceptor substrate. The crystal structure of the complexes reveals the major traits of the active site, with Asn387 controlling the substrate binding mode. Relevant residues were selected for mutagenesis, the variants being biochemically characterized through their hydrolytic and transfructosylating activity. All changes decrease the hydrolytic efficiency against sucrose, proving their key role in the activity. Moreover, some of the generated variants exhibit redesigned transfructosylating specificity, which may be used for biotechnological purposes to produce novel fructosyl-derivatives.


Assuntos
Rhodotorula , beta-Frutofuranosidase , beta-Frutofuranosidase/metabolismo , Rhodotorula/genética , Rhodotorula/metabolismo , Oligossacarídeos/química , Especificidade por Substrato , Sacarose/metabolismo
5.
Planta ; 256(6): 107, 2022 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-36342558

RESUMO

MAIN CONCLUSION: VInv gene editing in potato using CRISPR/Cas9 resulted in knockdown of expression and a lower VInv enzymatic activity resulting in a decrease in post-harvest cold-storage sugars formation and sweetening in potatoes. CRISPR-Cas9-mediated knockdown of vacuolar invertase (VInv) gene was carried out using two sgRNAs in local cultivar of potato plants. The transformation efficiency of potatoes was found to be 11.7%. The primary transformants were screened through PCR, Sanger sequencing, digital PCR, and ELISA. The overall editing efficacy was determined to be 25.6% as per TIDE analysis. The amplicon sequencing data showed maximum indel frequency for potato plant T12 (14.3%) resulting in 6.2% gene knockout and 6% frame shift. While for plant B4, the maximum indel frequency of 2.0% was found which resulted in 4.4% knockout and 4% frameshift as analyzed by Geneious. The qRT-PCR data revealed that mRNA expression of VInv gene was reduced 90-99-fold in edited potato plants when compared to the non-edited control potato plant. Following cold storage, chips analysis of potatoes proved B4 and T12 as best lines. Reducing sugars' analysis by titration method determined fivefold reduction in percentage of reducing sugars in tubers of B4 transgenic lines as compared to the control. Physiologically genome-edited potatoes behaved like their conventional counterpart. This is first successful report of knockdown of potato VInv gene in Pakistan that addressed cold-induced sweetening resulting in minimum accumulation of reducing sugars in genome edited tubers.


Assuntos
Solanum tuberosum , beta-Frutofuranosidase , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo , Solanum tuberosum/genética , Solanum tuberosum/metabolismo , Sistemas CRISPR-Cas , Regulação da Expressão Gênica de Plantas , Expressão Gênica , Açúcares/metabolismo
6.
Int J Mol Sci ; 23(20)2022 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-36293272

RESUMO

The reconfiguration of the primary metabolism is essential in plant-pathogen interactions. We compared the local metabolic responses of cucumber leaves inoculated with Pseudomonas syringae pv lachrymans (Psl) with those in non-inoculated systemic leaves, by examining the changes in the nicotinamide adenine dinucleotides pools, the concentration of soluble carbohydrates and activities/gene expression of carbohydrate metabolism-related enzymes, the expression of photosynthesis-related genes, and the tricarboxylic acid cycle-linked metabolite contents and enzyme activities. In the infected leaves, Psl induced a metabolic signature with an altered [NAD(P)H]/[NAD(P)+] ratio; decreased glucose and sucrose contents, along with a changed invertase gene expression; and increased glucose turnover and accumulation of raffinose, trehalose, and myo-inositol. The accumulation of oxaloacetic and malic acids, enhanced activities, and gene expression of fumarase and l-malate dehydrogenase, as well as the increased respiration rate in the infected leaves, indicated that Psl induced the tricarboxylic acid cycle. The changes in gene expression of ribulose-l,5-bis-phosphate carboxylase/oxygenase large unit, phosphoenolpyruvate carboxylase and chloroplast glyceraldehyde-3-phosphate dehydrogenase were compatible with a net photosynthesis decline described earlier. Psl triggered metabolic changes common to the infected and non-infected leaves, the dynamics of which differed quantitatively (e.g., malic acid content and metabolism, glucose-6-phosphate accumulation, and glucose-6-phosphate dehydrogenase activity) and those specifically related to the local or systemic response (e.g., changes in the sugar content and turnover). Therefore, metabolic changes in the systemic leaves may be part of the global effects of local infection on the whole-plant metabolism and also represent a specific acclimation response contributing to balancing growth and defense.


Assuntos
Carbono-Nitrogênio Ligases , Cucumis sativus , Pseudomonas syringae/fisiologia , Cucumis sativus/genética , Cucumis sativus/metabolismo , Carbono/metabolismo , Fosfoenolpiruvato Carboxilase/genética , beta-Frutofuranosidase/metabolismo , Malato Desidrogenase/metabolismo , Rafinose/metabolismo , Trealose/metabolismo , NAD/metabolismo , Fumarato Hidratase , Glucose-6-Fosfato/metabolismo , Glucosefosfato Desidrogenase/metabolismo , Folhas de Planta/metabolismo , Fotossíntese/fisiologia , Metabolismo dos Carboidratos , Sacarose/metabolismo , Fosfatos/metabolismo , Oxigenases/metabolismo , Inositol/metabolismo , Carbono-Nitrogênio Ligases/metabolismo , Niacinamida/metabolismo , Adenina/metabolismo , Glucose/metabolismo
7.
Plant Physiol Biochem ; 190: 101-108, 2022 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-36108354

RESUMO

At the end of the summer season, grapevine buds (Vitis vinifera L) grown in temperate climates enter a state of winter recess or endodormancy (ED), which is induced by the shortening of the photoperiod, and during this period, the buds accumulate sucrose. In this study, we investigated whether the shortening of the photoperiod regulates the accumulation of sucrose in the buds in the same way as it regulates its entry into the ED. Because sucrose accumulation is regulated by genes that control its transport and degradation, the effect of the SD photoperiod and the transition of buds from paradormancy (PD) to ED on the expression of sucrose transporter (VvSUTs) and invertase genes (VvINVs) was studied. To analyze the possible role of sucrose during ED development, its effect on bud swelling and sprouting was studied on dormant and nondormant buds under forced growth conditions. The results showed that the SD photoperiod upregulates the expression of the VvSUT genes and downregulates that of the VvINV genes in grapevine buds. Additionally, during the transition of buds from PD to ED, the sucrose content increased, the expression of the VvINV genes decreased, and the expression of the VvSUT genes did not change significantly. Sucrose delayed bud swelling and sprouting when applied to dormant buds but had no effect when applied to nondormant buds. Therefore, we concluded that ED development and sucrose accumulation were synchronized events induced by the SD photoperiod and that a sucrose peak marks the end of ED development in grapevine buds.


Assuntos
Fotoperíodo , Vitis , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sacarose/metabolismo , Vitis/metabolismo , beta-Frutofuranosidase/metabolismo
8.
J Environ Manage ; 323: 116197, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36126591

RESUMO

Baker's yeast industries generate highly polluted effluents, especially the cell free broth (i.e., vinasse) characterized by high chemical oxygen demand, nitrogen, and salts. In this work, it was found that the residual by-products (i.e., ethanol and acetic acid) and salts in the vinasse severely inhibited the cell growth, which hindered the reuse of the vinasse for the production of Saccharomyces cerevisiae. Through optimizing a suitable control strategy, the productions of ethanol and acetic acid were eliminated. Then, a nanofiltration membrane (i.e., NF5) was preferred for preliminarily and simultaneously separating and concentrating valuable molecules (i.e., invertase, food grade proteins and pigments) in the vinasse, and the main fouling mechanism was cake layer formation. Subsequently, a reverse osmosis membrane (RO) was suitable to separate and concentrate salts in the NF5 permeate, where the membrane fouling was negligible. Finally, the RO permeate was successfully reused for the production of S. cerevisiae. In addition, without calculating the benefit from the recovery of the valuable molecules, the cost of the integrated process can be decreased by 59.8% compared with the sole triple effect evaporation. Meanwhile, the volume of the fresh water used in the fermentation process can be decreased by 68.8%. Thus, it is a sustainable process for the cleaner production of baker's yeast using the integrated fermentation and membrane separation process.


Assuntos
Saccharomyces cerevisiae , Gerenciamento de Resíduos , Ácido Acético/metabolismo , Etanol/metabolismo , Fermentação , Nitrogênio/metabolismo , Saccharomyces cerevisiae/metabolismo , Sais/metabolismo , beta-Frutofuranosidase/metabolismo
9.
Ying Yong Sheng Tai Xue Bao ; 33(9): 2431-2440, 2022 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-36131659

RESUMO

To understand the development mechanism of the epiphyllous bud of waterlily, we examined the morphological anatomy of the leaf-navel epiphyllous bud by paraffin section technique at four stages, and compared the differences of carbohydrate metabolism between viviparous and non-viviparous waterlily leaves. Three tropical waterlily cultivars of Brachyceras were used, including two viviparous cultivars Nymphaea 'Margaret Mary', Nymphaea 'Ruby', and a non-viviparous cultivar Nymphaea 'Pink Star'. The results showed that parenchyma cells below the epidermis of leaf-navel divided and grew continuously after the leaf unfolded, forming a closely arranged cell cluster in viviparous waterlily and raised upward to a spherical shape. In contrast, no change was observed in leaf-navel of non-viviparous waterlily with the expansion of leaves. With the development of leaves, the contents of all physiological variables except sucrose and enzyme activities in the leaves of viviparous waterlily showed a first increase and then a decrease, which was significantly higher than those of non-viviparous waterlily. The carbohydrate contents in different parts showed the order of leaf > leaf-navel > petiole (except for starch content, which was highest in the leaf-navel). The activities of sucrose synthase (SS) and acid invertase (AI) were higher than those of sucrose phosphate synthase (SPS) and neutral invertase (NI). The activities of SPS and NI in different tissues of viviparous waterlily were significantly higher than those in non-viviparous one, but SS and AI did not show pronounced cultivar advantage in viviparous cultivars. AI activity varied greatly among cultivars, whereas NI activity varied less and was at a low level in different tissues. The sucrose of Nymphaea 'Ruby' leaves was positively correlated with the SPS and AI, and significantly associated with NI. The accumulation of sucrose content increased the activities of SS and NI of waterlily leaves, which was conducive to promoting the formation of epiphyllous buds.


Assuntos
Nymphaea , beta-Frutofuranosidase , Metabolismo dos Carboidratos , Nymphaea/metabolismo , Parafina/metabolismo , Folhas de Planta/metabolismo , Amido/metabolismo , Sacarose/metabolismo , beta-Frutofuranosidase/metabolismo
10.
Plant Physiol Biochem ; 189: 1-13, 2022 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-36030618

RESUMO

Sugar synthesis from photosynthesis and its utilization through sugar metabolism jointly determine leaf sugar content, and in contrast, excess sugar represses leaf photosynthesis. Although plant photosynthesis is affected by leaf sugar metabolism, the relationship between sugar metabolism and photosynthetic capacity of different grape genotypes remains unclear. In this study, two grape (Vitis vinifera L.) genotypes 'Riesling' (RI, high sugar content in leaf) and 'Petit Manseng' (PM, low sugar content in leaf) were used to evaluate the relationship between sugar metabolism and photosynthesis. Sugar content, chlorophyll content, photosynthetic parameters, enzyme activity, and gene expression related to sucrose metabolism in leaves were measured, and the correlations between photosynthesis and sugar metabolism were assessed. The contents of sucrose and glucose were significantly higher in RI leaves than in PM leaves, while the fructose content pattern was reversed. Cell wall invertase activity for sucrose hydrolysis and the transcript levels of VvCWINV, VvHTs, VvTMT1, VvFKs, and VvHXK2 were also higher in RI leaves than in PM leaves, whereas that of VvHXK1 mediating glucose phosphorylation, was lower in RI leaves than in PM leaves. Net photosynthetic rate, stomatal conductance, transpiration rate, and chlorophyll content were lower in RI leaves than in PM leaves and negatively correlated with glucose content, and the transcript levels of VvCWINV, VvHTs, VvTMT1, and VvHXK2. In conclusion, this study indicates that leaf sugar metabolism and transport are related to photosynthesis in Vitis vinifera L., which provides a theoretical basis for improving grape photosynthesis.


Assuntos
Vitis , Carboidratos/farmacologia , Clorofila/metabolismo , Frutose/metabolismo , Glucose/metabolismo , Hidrólise , Fotossíntese , Folhas de Planta/metabolismo , Sacarose/metabolismo , Açúcares/metabolismo , Vitis/metabolismo , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo
11.
Plant J ; 112(1): 115-134, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35942603

RESUMO

Vegetative propagation (VP) is an important practice for production in many horticultural plants. Sugar supply constitutes the basis of VP in bulb flowers, but the underlying molecular basis remains elusive. By performing a combined sequencing technologies coupled with ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry approach for metabolic analyses, we compared two Lycoris species with contrasting regeneration rates: high-regeneration Lycoris sprengeri and low-regeneration Lycoris aurea. A comprehensive multi-omics analyses identified both expected processes involving carbohydrate metabolism and transcription factor networks, as well as the metabolic characteristics for each developmental stage. A higher abundance of the differentially expressed genes including those encoding ethylene responsive factors was detected at bulblet initiation stage compared to the late stage of bulblet development. High hexose-to-sucrose ratio correlated to bulblet formation across all the species examined, indicating its role in the VP process in Lycoris bulb. Importantly, a clear difference between cell wall invertase (CWIN)-catalyzed sucrose unloading in high-regeneration species and the sucrose synthase-catalyzed pathway in low-regeneration species was observed at the bulblet initiation stage, which was supported by findings from carboxyfluorescein tracing and quantitative real-time PCR analyses. Collectively, the findings indicate a sugar-mediated model of the regulation of VP in which high CWIN expression or activity may promote bulblet initiation via enhancing apoplasmic unloading of sucrose or sugar signals, whereas the subsequent high ratio of hexose-to-sucrose likely supports cell division characterized in the next phase of bulblet formation.


Assuntos
Lycoris , Transcriptoma , Metabolismo dos Carboidratos/genética , Etilenos , Lycoris/genética , Lycoris/metabolismo , Metaboloma , Sacarose/metabolismo , Fatores de Transcrição/metabolismo , beta-Frutofuranosidase/metabolismo
12.
Plant Cell Physiol ; 63(10): 1510-1525, 2022 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-35946132

RESUMO

Phloem unloading plays an important role in photoassimilate partitioning and grain yield improvements in cereal crops. The phloem unloading strategy and its effects on photoassimilate translocation and yield formation remain unclear in rice. In this study, plasmodesmata were observed at the interface between the sieve elements (SEs) and companion cells (CCs), and between the SE-CC complex and surrounding parenchyma cells (PCs) in phloem of the dorsal vascular bundle in developing caryopses. Carboxyfluorescein (CF) signal was detected in the phloem of caryopses, which showed that CF was unloaded into caryopses. These results indicated that the SE-CC complex was symplasmically connected with adjacent PCs by plasmodesmata. Gene expression for sucrose transporter (SUT) and cell wall invertase (CWI), and OsSUT1 and OsCIN1 proteins were detected in developing caryopses, indicating that rice plants might actively unload sucrose into caryopses by the apoplasmic pathway. Among three rice recombinant inbred lines, R201 exhibited lower plasmodesmal densities at the boundaries between cell types (SE-CC, SE-PC and CC-PC) in developing caryopses than R91 and R156. R201 also had lower expression of SUT and CWI genes and lower protein levels of OsSUT1 and OsCIN1, as well as CWI activity, than R91 and R156. These data agreed with stem non-structural carbohydrate (NSC) translocation and grain yields for the three lines. The nitrogen application rate had no significant effect on plasmodesmal densities at the interfaces between different cells types, and did not affect CF unloading in the phloem of developing caryopses. Low nitrogen treatment enhanced expression levels of OsSUT and OsCIN genes in the three lines. These results suggested that nitrogen application had no substantial effect on symplasmic unloading but affected apoplasmic unloading. Therefore, we concluded that poor symplasmic and apoplasmic unloading in developing caryopses might result in low stem NSC translocation and poor grain yield formation of R201.


Assuntos
Oryza , Floema , Floema/metabolismo , Oryza/genética , Oryza/metabolismo , Grão Comestível/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , beta-Frutofuranosidase/metabolismo , Sacarose/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Nitrogênio/metabolismo , Transporte Biológico
13.
New Phytol ; 235(6): 2331-2349, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35695205

RESUMO

Invertase (INV)-mediated sucrose (Suc) hydrolysis, leading to the irreversible production of glucose (Glc) and fructose (Frc), plays an essential role in abiotic stress tolerance of plants. However, the regulatory network associated with the Suc catabolism in response to cold environment remains largely elusive. Herein, the cold-induced alkaline/neutral INV gene PtrA/NINV7 of trifoliate orange (Poncirus trifoliata (L.) Raf.) was shown to function in cold tolerance via mediating the Suc hydrolysis. Meanwhile, a nuclear matrix-associated region containing A/T-rich sequences within its promoter was indispensable for the cold induction of PtrA/NINV7. Two AT-Hook Motif Containing Nuclear Localized (AHL) proteins, PtrAHL14 and PtrAHL17, were identified as upstream transcriptional activators of PtrA/NINV7 by interacting with the A/T-rich motifs. PtrAHL14 and PtrAHL17 function positively in the cold tolerance by modulating PtrA/NINV7-mediated Suc catabolism. Furthermore, both PtrAHL14 and PtrAHL17 could form homo- and heterodimers between each other, and interacted with two histone acetyltransferases (HATs), GCN5 and TAF1, leading to elevated histone3 acetylation level under the cold stress. Taken together, our findings unraveled a new cold-responsive signaling module (AHL14/17-HATs-A/NINV7) for orchestration of Suc catabolism and cold tolerance, which shed light on the molecular mechanisms underlying Suc catabolism catalyzed by A/NINVs under cold stress.


Assuntos
Citrus , Poncirus , Citrus/genética , Temperatura Baixa , Resposta ao Choque Frio/fisiologia , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Poncirus/genética , Poncirus/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sacarose/metabolismo , beta-Frutofuranosidase/metabolismo
14.
J Exp Bot ; 73(14): 4908-4922, 2022 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-35552692

RESUMO

Fructans such as inulin and levan accumulate in certain taxonomic groups of plants and are a reserve carbohydrate alternative to starch. Onion (Allium cepa L.) is a typical plant species that accumulates fructans, and it synthesizes inulin-type and inulin neoseries-type fructans in the bulb. Although genes for fructan biosynthesis in onion have been identified so far, no genes for fructan degradation had been found. In this study, phylogenetic analysis predicted that we isolated a putative vacuolar invertase gene (AcpVI1), but our functional analyses demonstrated that it encoded a fructan 1-exohydrolase (1-FEH) instead. Assessments of recombinant proteins and purified native protein showed that the protein had 1-FEH activity, hydrolyzing the ß-(2,1)-fructosyl linkage in inulin-type fructans. Interestingly, AcpVI1 had an amino acid sequence close to those of vacuolar invertases and fructosyltransferases, unlike all other FEHs previously found in plants. We showed that AcpVI1 was localized in the vacuole, as are onion fructosyltransferases Ac1-SST and Ac6G-FFT. These results indicate that fructan-synthesizing and -degrading enzymes are both localized in the vacuole. In contrast to previously reported FEHs, our data suggest that onion 1-FEH evolved from a vacuolar invertase and not from a cell wall invertase. This demonstrates that classic phylogenetic analysis on its own is insufficient to discriminate between invertases and FEHs, highlighting the importance of functional markers in the nearby active site residues.


Assuntos
Cebolas , beta-Frutofuranosidase , Frutanos/metabolismo , Glicosídeo Hidrolases/metabolismo , Inulina , Cebolas/genética , Cebolas/metabolismo , Filogenia , Vacúolos/metabolismo , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo
15.
Ecotoxicol Environ Saf ; 240: 113701, 2022 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-35636237

RESUMO

In this study, six different treatments involving extracellular polymeric substances (EPS) from Enterobacter sp. FM-1 (FM-1) (no EPS (control), original bacterial cells (FM-1), FM-1 cells with EPS artificially removed (EPS-free cells, EPS-R), different forms of EPS (soluble EPS (S-EPS), loosely bound EPS (LB-EPS) and tightly bound EPS (TB-EPS)) obtained from FM-1) and three types of soils (non-contaminated soil (NC soil), high-contamination soil (HC soil) and low-contamination soil (LC soil)) were used to investigate the impact of different EPS treatments on soil microbial community composition and their potential role in the remediation of heavy metal (HM)-contaminated soil. The results indicate that the EPS secreted by FM-1 played a vital role in changing soil pH and helped increase soil bio- HMs. In addition, EPS secretion by FM-1 helped increase the soil EPS-polysaccharide and EPS-nucleic acid contents; even in HC soil, where the HM content was relatively high, LB-EPS addition still increased the EPS-polysaccharide and EPS-nucleic acid contents in the soil by 1.18- and 15.54-fold, respectively. FM-1, LB-EPS and TB-EPS addition increased the soil invertase, urease and alkaline phosphatase activities and increased the soil organic matter (SOM), NH4+-N and available phosphorus (AP) contents, which helped regulate soil nutrient reserves. Moreover, the addition of different EPS fractions modified the soil microbial community composition to help microbes adapt to an HM-contaminated environment. In the HC and LC soils, where the HM content was relatively high, the soil bacteria were dominated by Protobacteria, while fungi in the soil were dominated by Ascomycota. Among the soil physicochemical properties, the soil SOM and NH4+-N contents and invertase activity significantly impacted the diversity and community composition of both bacteria and fungi in the soil.


Assuntos
Metais Pesados , Microbiota , Ácidos Nucleicos , Bactérias/metabolismo , Biodegradação Ambiental , Matriz Extracelular de Substâncias Poliméricas/química , Fungos , Metais Pesados/análise , Ácidos Nucleicos/metabolismo , Solo , Microbiologia do Solo , beta-Frutofuranosidase/metabolismo
16.
Sci Total Environ ; 833: 155163, 2022 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-35413342

RESUMO

Nitrogen (N) and phosphorus (P) control biogeochemical cycling in terrestrial ecosystems. However, N and P addition effects on litter decomposition, especially biological pathways in subtropical forests, remain unclear. Here, a two-year field litterbag experiment was employed in a subtropical forest in southwestern China to examine N and P addition effects on litter biological decomposition with nine treatments: low and high N- and P-only addition (LN, HN, LP, and HP), NP coaddition (LNLP, LNHP, HNLP, and HNHP), and a control (CK). The results showed that the decomposition coefficient (k) was higher in NP coaddition treatments (P < 0.05), and lower in N- and P-only addition treatments than in CK (P < 0.05). The highest k was observed with LNLP (P < 0.05). The N- and P-only addition treatments decreased the losses of litter mass, lignin, cellulose, and condensed tannins, litter microbial biomass carbon (MBC), litter cellulase, and soil pH (P < 0.05). The NP coaddition treatments increased the losses of litter mass, lignin, and cellulose, MBC concentration, litter invertase, urease, cellulase, and catalase activities, soil arthropod diversity (S) in litterbags, and soil pH (P < 0.05). Litter acid phosphatase activity and N:P ratio were lower in N-only addition treatments but higher in P-only addition and NP coaddition treatments than in CK (P < 0.05). Structural equation model showed that litter MBC, S, cellulase, acid phosphatase, and polyphenol oxidase contributed to the loss of litter mass (P < 0.05). The litter N:P ratio was negatively logarithmically correlated with mass loss (P < 0.01). In conclusion, the negative effect of N addition on litter decomposition was reversed when P was added by increasing decomposed litter soil arthropod diversity, MBC concentration, and invertase and cellulase activities. Finally, the results highlighted the important role of the N:P ratio in litter decomposition.


Assuntos
Celulases , Nitrogênio , Fosfatase Ácida/metabolismo , Carbono/análise , Celulases/análise , Celulases/metabolismo , China , Ecossistema , Florestas , Lignina/metabolismo , Nitrogênio/análise , Fósforo/análise , Folhas de Planta/química , Solo/química , beta-Frutofuranosidase/análise , beta-Frutofuranosidase/metabolismo
17.
Physiol Plant ; 174(2): e13673, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35307852

RESUMO

Invertases are key enzymes for carbon metabolism, cleaving sucrose into energy-rich and signaling metabolites, glucose and fructose. Invertases play pivotal roles in development and stress response, determining yield and quality of seed production. In this context, the repertoire of invertase gene families is critically scarce in legumes. Here, we performed a systematic search for invertase families in 16 Fabaceae genomes. For instance, we identified 19 invertase genes in the model plant Medicago and 17 accessions in the agronomic crop Pisum sativum. Our comprehensive phylogenetic analysis sets a milestone for the scientific community as we propose a new nomenclature to correctly name plant invertases. Thus, neutral invertases were classified into four clades of cytosolic invertase (CINV). Acid invertases were classified into two cell wall invertase clades (CWINV) and two vacuolar invertase clades (VINV). Then, we explored transcriptional regulation of the pea invertase family, focusing on seed development and water stress. Invertase expression decreased sharply from embryogenesis to seed-filling stages, consistent with higher sucrose and lower monosaccharide contents. The vacuolar invertase PsVINV1.1 clearly marked the transition between both developmental stages. We hypothesize that the predominantly expressed cell wall invertase, PsCWINV1.2, may drive sucrose unloading towards developing seeds. The same candidates, PsVINV1.1 and PsCWINV1.2, were also regulated by water deficit during embryonic stage. We suggest that PsVINV1.1 along with vacuolar sugar transporters maintain cellular osmotic pressure and PsCWINV1.2 control hexose provision, thereby ensuring embryo survival in drought conditions. Altogether, our findings provide novel insights into the regulation of plant carbon metabolism in a challenging environment.


Assuntos
Fabaceae , beta-Frutofuranosidase , Carbono/metabolismo , Secas , Fabaceae/genética , Fabaceae/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Ervilhas/genética , Ervilhas/metabolismo , Filogenia , Sementes/genética , Sementes/metabolismo , Sacarose/metabolismo , beta-Frutofuranosidase/genética , beta-Frutofuranosidase/metabolismo
18.
Int J Phytoremediation ; 24(14): 1505-1517, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35266855

RESUMO

To understand the plant (Vigna unguiculata) and plant-growth promoting bacteria (PGPB) (Microcococcus luteus WN01) interactions in crude oil contaminated soil, experiments were conducted based on the newly designed rhizobox system. The rhizobox was divided into three main compartments namely the rhizosphere zone, the mid-zone, and the bulk soil zone, in accordance with the distance from the plant. Plants were grown in these three-chambered pots for 30 days under natural conditions. The plant root exudates were determined by analyzing for carbohydrates, amino acids, and phenolic compounds. The degradation of alkane, polycyclic aromatic hydrocarbons (PAHs), and total petroleum hydrocarbons (TPHs) were quantified by GC-FID. Soil catalase, dehydrogenase, and invertase activities were determined. The microbial community structure was assessed using denaturing gradient gel electrophoresis (DGGE). Results showed that the inoculation of M. luteus WN01 significantly enhanced cowpea root biomass and exudates, especially the phenolic compounds. Bioaugmented phytoremediation by cowpea and M. luteus promoted rhizodegradation of TPH. Cowpea stimulated microbial growth, soil dehydrogenase, and invertase activities and enhanced bacterial community diversity in oil contaminated soil. The rhizosphere zone of cowpea inoculated with M. luteus showed the highest removal efficiency, microbial activities, microbial population, and bacterial community diversity indicating the strong synergic interactions between M. luteus and cowpea.


This is the first study to characterize the rhizosphere effect of cowpea on microbial activities, population, and community structure in crude oil contaminated soil in the presence and absence of PGPB, M. luteus WN01. The rhizosphere of cowpea was found to be a degradation hotspot where microbial abundance and metabolic activities were most active. Cowpea-M. luteus association can be a good candidate that can be implemented in real field sites.


Assuntos
Microbiota , Petróleo , Poluentes do Solo , Biodegradação Ambiental , Petróleo/metabolismo , Rizosfera , Solo/química , beta-Frutofuranosidase/metabolismo , Poluentes do Solo/metabolismo , Microbiologia do Solo , Bactérias/metabolismo , Oxirredutases/metabolismo
19.
Appl Microbiol Biotechnol ; 106(7): 2455-2470, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35267055

RESUMO

Fructooligosaccharide is a mixture of mostly the trisaccharide 1-kestose (GF2), tetrasaccharide nystose (GF3), and fructosyl nystose (GF4). Enzymes that hydrolyze GF3 may be useful for preparing GF2 from the fructooligosaccharide mixture. A ß-fructofuranosidase belonging to glycoside hydrolase family 32 (GH32) from the honeybee gut bacterium Frischella perrara (FperFFase) was expressed in Escherichia coli and purified. The time course of the hydrolysis of 60 mM sucrose, GF2, and GF3 by FperFFase was analyzed, showing that the hydrolytic activity of FperFFase for trisaccharide GF2 was lower than those for disaccharide sucrose and tetrasaccharide GF3. The crystal structure of FperFFase and its structure in complex with fructose were determined. FperFFase was found to be structurally homologous to bifidobacterial ß-fructofuranosidases even though bifidobacterial enzymes preferably hydrolyze GF2 and the amino acid residues interacting with fructose at subsite - 1 are mostly conserved between them. A proline residue was inserted between Asp298 and Ser299 using site-directed mutagenesis, and the activity of the variant 298P299 was measured. The ratio of activities for 60 mM GF2/GF3 by wild-type FperFFase was 35.5%, while that of 298P299 was 23.6%, indicating that the structure of the loop comprising Trp297-Asp298-Ser299 correlated with the substrate preference of FperFFase. The crystal structure also shows that a loop consisting of residues 117-127 is likely to contribute to the substrate binding of FperFFase. The results obtained herein suggest that FperFFase is potentially useful for the manufacture of GF2. KEY POINTS: • Frischella ß-fructofuranosidase hydrolyzed nystose more efficiently than 1-kestose. • Trp297-Asp298-Ser299 was shown to be correlated with the substrate preference. • Loop consisting of residues 117-127 appears to contribute to the substrate binding.


Assuntos
Oligossacarídeos , beta-Frutofuranosidase , Animais , Abelhas , Frutose , Gammaproteobacteria , Oligossacarídeos/metabolismo , Sacarose , Trissacarídeos/metabolismo , beta-Frutofuranosidase/metabolismo
20.
Glycobiology ; 32(6): 540-549, 2022 05 23.
Artigo em Inglês | MEDLINE | ID: mdl-35138388

RESUMO

Bifidobacterium pseudocatenulatum grows well in the early stages of cultivation in medium containing sucrose (Suc), whereas its growth in medium containing the analogue disaccharide N-acetylsucrosamine (SucNAc) tends to exhibit a considerable delay. To elucidate the cause of this phenomenon, we investigated the proliferation pattern of B. pseudocatenulatum in medium containing D-glucose (Glc) and SucNAc and identified the enzyme that degrades this disaccharide. We found that B. pseudocatenulatum initially proliferates by assimilating Glc, with subsequent growth based on SucNAc assimilation depending on production of the ß-fructofuranosidase, which can hydrolyze SucNAc, after Glc is completely consumed. Thus, B. pseudocatenulatum exhibited a diauxic growth pattern in medium containing Glc and SucNAc. In contrast, when cultured in medium containing Glc and Suc, B. pseudocatenulatum initially grew by degrading Suc via the phosphorolysis activity of Suc phosphorylase, which did not react to SucNAc. These observations indicate that B. pseudocatenulatum proliferates by assimilating Suc and SucNAc via different pathways. The ß-fructofuranosidase of B. pseudocatenulatum exhibited higher hydrolytic activity against several naturally occurring Suc-based tri- or tetrasaccharides than against Suc, suggesting that this enzyme actively catabolizes oligosaccharides other than Suc.


Assuntos
Bifidobacterium pseudocatenulatum , Bifidobacterium pseudocatenulatum/metabolismo , Dissacarídeos/metabolismo , Oligossacarídeos/metabolismo , Sacarose/metabolismo , beta-Frutofuranosidase/metabolismo
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