• Journal of Life Science
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• v.17 no.8 s.88
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• pp.1157-1165
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• 2007

VvMSA, a grapevine ASR which is highly inducible by sugar and abscisic acid signals was previously shown to be a transcription factor for a hexose transporter gene VvHT1. We isolated a cDNA clone, VlASR which is regulated temporally during the grape berry development by ACP RT-PCR (annealing control primer reverse transcriptase-polymerase chain reaction) and it proved identical to VvMSA. RT-PCR and real-time PCR analyses revealed that the VlASR gene was expressed in berries at fruit set and that its expression increased as berries aged but decreased at the late ripening stage. In order to understand the regulatory mechanism of the asr gene, a genomic fragment was cloned from grapevine. The genomic DNA was 1375 bp long and a sugar box (sucrose box 3 and sucrose responsive element 1) was identified in the 611 bp upstream region of the open reading frame. Analysis of the VlASR promoter::reporter gene fusion demonstrated that this promoter was expressed in transgenic Arabidopsis even without sucrose treatment. This result suggests that the ASR/VvHT1-mediated sugar/ABA signaling, previously reported in grapevine, may not function in Arabidopsis which has no ASR homologue.

• Journal of Ginseng Research
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• v.44 no.1
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• pp.161-167
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• 2020

Background: The ascomycete fungi Cylindrocarpon destructans (Cd) and Fusarium solani (Fs) cause ginseng root rot and significantly reduce the quality and yield of ginseng. Cd produces the secondary metabolite radicicol, which targets the molecular chaperone Hsp90. Fs is resistant to radicicol, whereas other fungal genera associated with ginseng disease are sensitive to it. Radicicol resistance mechanisms have not yet been elucidated. Methods: Transcriptome analyses of Fs and Cd mycelia treated with or without radicicol were conducted using RNA-seq. All of the differentially expressed genes (DEGs) were functionally annotated using the Fusarium graminearum transcript database. In addition, deletions of two transporter genes identified by RNA-seq were created to confirm their contributions to radicicol resistance. Results: Treatment with radicicol resulted in upregulation of chitin synthase and cell wall integrity genes in Fs and upregulation of nicotinamide adenine dinucleotide dehydrogenase and sugar transporter genes in Cd. Genes encoding an ATP-binding cassette transporter, an aflatoxin efflux pump, ammonium permease 1 (mep1), and nitrilase were differentially expressed in both Fs and Cd. Among these four genes, only the ABC transporter was upregulated in both Fs and Cd. The aflatoxin efflux pump and mep1 were upregulated in Cd, but downregulated in Fs, whereas nitrilase was downregulated in both Fs and Cd. Conclusion: The transcriptome analyses suggested radicicol resistance pathways, and deletions of the transporter genes indicated that they contribute to radicicol resistance.

• Biomolecules & Therapeutics
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• v.12 no.2
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• pp.74-78
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• 2004

Paecilomyces tenuipes, a caterpillar fungus, contains many health-promoting ingredients. Recent reports indicate that consumption of P. tenuipes helps reducing blood sugar content for diabetes. Mechanism for reduction in the circulatory sugar content, however, still remains least understood. Methanolic extraction of P. tenuipes (MPT) was prepared and acetoxyscirpendiol (ASD) was subsequently purified limn MPT. Glucose transporter-1 (GLUT-1) was expressed in the Xenopus oocytes and the effect of MPT or ASD on the expressed GLUT-1 was analyzed according to the uptake of 2-dideoxy-D-glucose (2-DOG). MPT was shown to inhibit GLUT-1 activity significant1y compared to the non-treated control. In the presence of ASD and its derivatives, GLUT-1 activity was greatly inhibited in a dose-dependent manner. Among ASD and its derivatives, AS-1 showed most significant inhibition. Taken together, these results strongly indicate that ASD in P. tenuipes may serve as a functional substance in lowering blood sugar in the circulatory system. ASD and its derivatives can be utilized as inhibitors of GLUT-1.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2005.11a
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• pp.9-21
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• 2005

To study the biochemical and physiological role of the plastidic glucose transporter (pGlcT) in carbohydrate metabolism, we characterized transgenic plants with mutations in the pGlcT gene (GT), gt-1 and gt-2, as well double mutants of GT and the maltose transporter (MEX1) and GT and the triose phosphate/phosphate translocator (TPT), GT and the cytosolic fructose-1,6-bisphosphatase gene (cFBP), and MEX1 and TPT, gt-1/mex2, gt-1/tpt-2, gt-1/cfbp-1, mex1-1/tpt-2, respectively. Compared to the wild type, all mutants except the gt-1/cfbp-1 mutant lines displayed higher starch accumulation and higher levels of maltose. Starch accumulation is due to a decrease in starch turnover, leading to an imbalance between the rates of synthesis and degradation. Sucrose levels of gt alleles were higher than those in wild-type plants during the light period, suggesting possible nightly supplementation via the maltose transport pathway to maintain proper carbohydrate partitioning in the plant leaves. The gt plants displayed less growth retardation than mex1-1 mutant and gt-1/mex2 double mutant displayed accumulativesevere growth retardation as compared to individual gt-1 and mex1-1 mutants, implying that the maltose transporter-mediated pathway is a major route for carbohydrate partitioning at night. The gt-1/tpt-2, mex1-1/tpt-2 and gt-1/cfbp-1 double mutants had retarded growth and low chlorophyll content to differing degrees, indicating that photosynthetic capacity had diminished. Interestingly, the gt-1/tpt-2 line displayed a glucose-insensitive phenotype and higher germination rates than wild type, suggesting its involvement not only in carbon partitioning, but also in the sugar signaling network of the pGlcT and TPT.

• BMB Reports
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• v.38 no.2
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• pp.211-217
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• 2005

Cordyceps, an entomopathogenic fungus, contains many health-promoting ingredients. Recent reports indicate that the consumption of cordyceps helps reduce blood-sugar content in diabetics. However, the mechanism underlying this reduction in circulatory sugar content is not fully understood. Methanolic extracts were prepared from the fruiting bodies of Paecilomyces tenuipes, and 4-beta acetoxyscirpendiol (4-ASD) was eventually isolated and purified. $Na^+$/Glucose transporter-1 (SGLT-1) was expressed in Xenopus oocytes, and the effect of 4-ASD on SGLT-1 was analyzed utilizing a voltage clamp and by performing 2-deoxy-D-glucose (2-DOG) uptake studies. 4-ASD was shown to significantly inhibit SGLT-1 activity compared to the non-treated control in a dose-dependent manner. In the presence of the derivatives of 4-ASD (diacetoxyscirpenol or 15-acetoxyscirpendiol), SGLT-1 activity was greatly inhibited in an 4-ASD-like manner. Of these derivatives, 15-acetoxyscirepenol inhibited SGLT-1 as well as 4-ASD, whereas diacetoxyscirpenol was slightly less effective. Taken together, these results strongly indicate that 4-ASD in P. tenuipes may lower blood sugar levels in the circulatory system. We conclude that 4-ASD and its derivatives are effective SGLT-1 inhibitors.

• Biomolecules & Therapeutics
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• v.12 no.4
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• pp.250-256
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• 2004

Cordyceps possesses numerous health-promoting ingredients including hypoglycemic agents. The mechanism for the reduction of circulatory sugar content, however, is still not fully understand. In this study, 4-beta acetoxyscirpendiol (ASD) was purified from the methanolic extracts from fruiting bodies of Paecilomyces tenuipes. Na+/Glucose transporter-1 (SGLT-1) was expressed in the Xenopus oocytes. The effect of ASD on the oocyte expressed SGLT-1 was analyzed utilizing the voltage clamp and 2-deoxy-D-glucose (2-DOG) uptake studies. ASD was shown to significantly inhibit SGLT-1 activity compared to the non-treated control in a dose- dependent manner. In the presense of its two derivatives (diacetoxyscirpenol or 15-acetoxyscirpendiol), SGLT-1 activity was greatly inhibited similarly as ASD. Between ASD derivatives, 15-acetoxyscirepenol showed inhibition equivalent to that of ASD while diacetoxyscirpenol did less degree of inhibition. Insummary , these results strongly indicate that ASD in P. tenuipes may serve as a functional substance in lowering blood sugar in the circulatory system. ASD and its derivatives can be utilized as inhibitors of SGLT-1.

• Proceedings of the Korean Society of Potoscience Conference
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• 2005.06a
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• pp.115-115
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• 2005

• Biomedical Science Letters
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• v.16 no.3
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• pp.201-206
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• 2010

In order to develop procedures for the rapid isolation of recombinant sugar transporter in functional form from away from the endogenous insect cell transporter, gene fusion techniques were exploited. Briefly, BamH1-digested human HepG2 type glucose transport protein cDNA was first cloned into a transfer vector pBlueBacHis, containing a tract of six histidine residues. Recombinant baculoviruses including the human cDNA were then generated by allelic exchange following transfection of insect cells with wild-type BaculoGold virus DNA and the recombinant transfer vector. Plaque assay was then performed to obtain and purify recombinant viruses expressing the human transport protein. All the cell samples that had been infected with viruses from the several blue plaques exhibited a positive reaction in the immnuassay, demonstrating expression of the glucose transport protein. In contrast, no color development in the immunoassay was observed for cells infected with the wild-type virus or no virus. Immunoblot analysis showed that a major immunoreactive band of apparent Mr 43,000~44,000 was evident in the lysate from cells infected with the recombinant baculovirus. Following expression of the recombinant fusion protein with the metal-binding domain and enterokinase cleavage site, the fusion protein was recovered by competition with imidizole using immobilized metal charged resin. The leader peptide was then removed from the fusion protein by cleavage with porcine enterokinase. Final separation of the recombinant protein of the interest was achieved by passage over $Ni^{2+}$-charged resin under binding conditions. The expressed transport protein bound cytochalasin B and demonstrated a functional similarity to its human counterpart.

• Journal of Plant Biotechnology
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• v.44 no.3
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• pp.296-302
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• 2017

Rice (Oryza sativa) cultivars show an impairment of growth and development in response to abiotic stresses such as drought, salinity, heat and cold at the early seedling stage. The tolerance to heat stress in plants has been genetically modulated by the overexpression of heat shock transcription factor genes or proteins. In addition to a high temperature-tolerance that has also been altered by elevating levels of osmolytes, increasing levels of cell detoxification enzymes and through altering membrane fluidity. To examine the heat tolerance in transgenic rice plants, three OsOPT10 overexpressing lines were characterized through a physiological analysis, which examined factors such as the electrolyte leakage (EL), soluble sugar and proline contents. We further functionally characterized the OsOPT10 gene and found that heat induced the expression of OsOPT10 and P5CS gene related proline biosynthesis. It has been suggested that the expression of OsOPT10 led to elevated heat tolerance in transgenic lines.

• Biomedical Science Letters
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• v.12 no.1
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• pp.17-22
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• 2006

The baculovirus expression system is a powerful method for producing large amounts of the human erythrocyte-type glucose transport protein, heterologously. Characterization of the expressed protein is expected to show its ability to transport sugars directly. To achieve this, it is a prerequisite to know the properties of the endogenous sugar transport system in Spodoptera frugiperda Clone 21 (Sf21) cells, which are commonly employed as a host permissive cell line to support the baculovirus replication. The Sf21 cells can grow well on TC-100 medium that contains 0.1% D-glucose as the major carbon source, strongly suggesting the presence of endogenous glucose transport system. However, unlike the human glucose transport protein that has a broad substrate and inhibitor specificity, very little is known about the nature of the endogenous sugar transport system in Sf21 cells. In order to characterize further the inhibitor recognition properties of the Sf21 cell transporter, the ability of phloretin, cytochalasin B and D-fructose to inhibit 2-deoxy-D-glucose (2dGlc) transport was examined by measuring inhibition constants $(K_i)$. The $K_i's$ for reversible inhibitors were determined from plots of uptake versus inhibitor concentration. The 2dGlc transport in the Sf21 cells was very potently inhibited by phloretin, the aglucone of phlorizin with a $K_i$ similar to the value of about $2{\mu}M$ reported for inhibition of glucose transport in human erythrocytes. However, the Sf21 cell transport system was found to differ from the human transport protein in being much less sensitive to inhibition by cytochalasin B (apparent $K_i$ approximately $10\;{\mu}M$). In contrast, It is reported that the inhibitor binds the human erythrocyte counterpart with a $K_d$ of approximately $0.12\;{\mu}M$. Interestingly, the Sf21 glucose transport system also appeared to have high affinity for D-fructose with a $K_i$ of approximately 5mM, contrasting the reported $K_m$ of the human erythrocyte transport protein for the ketose of 1.5M.