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Berlin Brandenburg

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  • 1
    Language: English
    In: Plant physiology, July 2013, Vol.162(3), pp.1486-96
    Description: Catharanthus roseus is one of the most studied medicinal plants due to the interest in their dimeric terpenoid indole alkaloids (TIAs) vinblastine and vincristine, which are used in cancer chemotherapy. These TIAs are produced in very low levels in the leaves of the plant from the monomeric precursors vindoline and catharanthine and, although TIA biosynthesis is reasonably well understood, much less is known about TIA membrane transport mechanisms. However, such knowledge is extremely important to understand TIA metabolic fluxes and to develop strategies aimed at increasing TIA production. In this study, the vacuolar transport mechanism of the main TIAs accumulated in C. roseus leaves, vindoline, catharanthine, and α-3',4'-anhydrovinblastine, was characterized using a tonoplast vesicle system. Vindoline uptake was ATP dependent, and this transport activity was strongly inhibited by NH4(+) and carbonyl cyanide m-chlorophenyl hydrazine and was insensitive to the ATP-binding cassette (ABC) transporter inhibitor vanadate. Spectrofluorimetry assays with a pH-sensitive fluorescent probe showed that vindoline and other TIAs indeed were able to dissipate an H(+) gradient preestablished across the tonoplast by either vacuolar H(+)-ATPase or vacuolar H(+)-pyrophosphatase. The initial rates of H(+) gradient dissipation followed Michaelis-Menten kinetics, suggesting the involvement of mediated transport, and this activity was species and alkaloid specific. Altogether, our results strongly support that TIAs are actively taken up by C. roseus mesophyll vacuoles through a specific H(+) antiport system and not by an ion-trap mechanism or ABC transporters.
    Keywords: Catharanthus -- Metabolism ; Indole Alkaloids -- Metabolism
    ISSN: 00320889
    E-ISSN: 1532-2548
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  • 2
    Language: English
    In: 2015, Vol.10(7), p.e0133810
    Description: Pectin is secreted in a highly methylesterified form and partially de-methylesterified in the cell wall by pectin methylesterases (PMEs). PME activity is expressed during plant growth, development and stress responses. PME activity is controlled at the post-transcriptional level by proteins named PME inhibitors (PMEIs). We have identified, expressed and characterized VvPMEI1, a functional PME inhibitor of Vitis vinifera . VvPMEI1 typically affects the activity of plant PMEs and is inactive against microbial PMEs. The kinetics of PMEI-PME interaction, studied by surface plasmon resonance, indicates that the inhibitor strongly interacts with PME at apoplastic pH while the stability of the complex is reduced by increasing the pH. The analysis of VvPMEI1 expression in different grapevine tissues and during grape fruit development suggests that this inhibitor controls PME activity mainly during the earlier phase of berry development. A proteomic analysis performed at this stage indicates a PME isoform as possible target of VvPMEI1.
    Keywords: Research Article
    E-ISSN: 1932-6203
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  • 3
    In: PLoS ONE, 2016, Vol.11(6)
    Description: The present study demonstrates the plant growth promoting (PGP) potential of a bacterial isolate CDP-13 isolated from ‘ Capparis decidua ’ plant, and its ability to protect plants from the deleterious effect of biotic and abiotic stressors. Based on 16S rRNA gene sequence analysis, the isolate was identified as Serratia marcescens . Among the PGP traits, the isolate was found to be positive for ACC deaminase activity, phosphate solubilization, production of siderophore, indole acetic acid production, nitrogen fixation, and ammonia production. CDP-13 showed growth at an increased salt (NaCl) concentration of up to 6%, indicating its potential to survive and associate with plants growing in saline soil. The inoculation of S . marcescens enhanced the growth of wheat plant under salinity stress (150–200 mM). It significantly reduced inhibition of plant growth (15 to 85%) caused by salt stressors. Application of CDP-13 also modulated concentration (20 to 75%) of different osmoprotectants (proline, malondialdehyde, total soluble sugar, total protein content, and indole acetic acid) in plants suggesting its role in enabling plants to tolerate salt stressors. In addition, bacterial inoculation also reduced the disease severity caused by fungal infection, which illustrated its ability to confer induced systemic resistance (ISR) in host plants. Treatment of wheat plants with the test organism caused alteration in anti-oxidative enzymes activities (Superoxide dismutase, Catalase, and Peroxidase) under various salinity levels, and therefore minimizes the salinity-induced oxidative damages to the plants. Colonization efficiency of strain CDP-13 was confirmed by CFU count, epi-fluorescence microscopy, and ERIC-PCR-based DNA fingerprinting approach. Hence, the study indicates that bacterium CDP-13 enhances plant growth, and has potential for the amelioration of salinity stress in wheat plants. Likewise, the results also provide insights into biotechnological approaches to using PGPR as an alternative to chemicals and pesticides.
    Keywords: Research Article ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Ecology And Environmental Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Biology And Life Sciences ; Medicine And Health Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Biology And Life Sciences ; Biology And Life Sciences
    E-ISSN: 1932-6203
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  • 4
    In: PLoS ONE, 2018, Vol.13(10)
    Description: Methotrexate (MTX) is a widely used chemotherapeutic agent, immune suppressant and antimalarial drug. It is a substrate of several human ABC proteins that confer multidrug resistance to cancer cells and determine compartmentalization of a wide range of physiological metabolites and endo or xenobiotics, by their primary active transport across biological membranes. The substrate specificity and tissue distribution of these promiscuous human ABC transporters show a high degree of redundancy, providing robustness to these key physiological and pharmacological processes, such as the elimination of toxins, e . g . methotrexate from the body. A similar network of proteins capable of transporting methotrexate has been recently suggested to exist in Drosophila melanogaster . One of the key players of this putative network is Drosophila Multidrug-resistance Associated Protein (DMRP). DMRP has been shown to be a highly active and promiscuous ABC transporter, capable of transporting various organic anions. Here we provide the first direct evidence that DMRP, expressed alone in a heterologous system lacking other, potentially functionally overlapping D . melanogaster organic anion transporters, is indeed able to transport methotrexate. Our in vitro results support the hypothesized but debated role of DMRP in in vivo methotrexate excretion.
    Keywords: Research Article ; Medicine And Health Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Research And Analysis Methods ; Research And Analysis Methods ; Research And Analysis Methods ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Biology And Life Sciences ; Research And Analysis Methods ; Biology And Life Sciences
    E-ISSN: 1932-6203
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  • 5
    Language: English
    In: PLOS ONE, 5/11/2017, Vol.12(5), p.e0177543
    Description: Sphingosine 1-phosphate (S1P) is a bioactive lipid mediator that is thought to be involved in various diseases. Although the main source of S1P in the plasma is erythrocytes, how S1P is exported from erythrocytes has not been elucidated. When we differentiated K562 cells into erythroblast-like cells with sodium butyrate, we observed that the efflux of S1P was increased without increased expression of previously proposed S1P transporters, while the expression levels of Band3 were increased. Therefore, in this study, we investigated the involvement of Band 3, the most characteristic membranous transporter for erythrocytes, in S1P efflux, using 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid, disodium salt (H2DIDS), which is an inhibitor of Band3. First, we treated human washed erythrocytes with H.sub.2 DIDS and found that H.sub.2 DIDS decreased the S1P levels in the supernatant, while it increased the cellular S1P contents. Next, when we injected H.sub.2 DIDS into mice, the plasma S1P level was significantly decreased. Finally, when we overexpressed or suppressed Band3 in K562 cells, S1P efflux was enhanced or decreased, respectively, while the overexpression of Band3 in HEK293 cells did not modulate S1P efflux. These results suggested the possible involvement of Band3 in the transport of S1P, a multi-functional bioactive phospholipid, from erythrocytes.
    Keywords: Phosphates -- Research ; Sphingosine -- Research ; Red Blood Cells -- Research ; Gene Expression -- Research;
    ISSN: PLOS ONE
    E-ISSN: 1932-6203
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  • 6
    In: PLoS ONE, 2016, Vol.11(12)
    Description: Analyzing variations in silicon (Si) isotopes can help elucidate the biogeochemical Si cycle and Si accumulation processes of higher plants. Importantly, the composition of Si isotopes in higher plants has yet to be studied comprehensively and our knowledge of the distribution of Si isotopes in higher plants lags behind that of Si isotopes in marine organisms, such as diatoms. In the present study, we investigated the isotope fractionation that occurs during the uptake and transport of Si in rice, using a series of hydroponic experiments with different external concentrations of Si. We found that an active mechanism was responsible for the majority of Si uptake and transport at lower Si levels and that the uptake of Si by rice roots was significantly suppressed by both low temperature and metabolic inhibitors. In addition, light Si isotopes ( 28 Si) entered roots more readily than heavy Si isotopes ( 30 Si) when the active mechanism was inhibited. Therefore, we conclude that biologically mediated isotope fractionation occurs during the uptake of Si by rice roots. In addition, both active and passive Si uptake components co-exist in rice, and the fractionation effect is enhanced when more Si is absorbed by plants.
    Keywords: Research Article ; Physical Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Research And Analysis Methods ; Research And Analysis Methods ; Biology And Life Sciences ; Biology And Life Sciences ; Medicine And Health Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Earth Sciences ; Biology And Life Sciences ; Earth Sciences
    E-ISSN: 1932-6203
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  • 7
    In: PLoS ONE, 2015, Vol.10(9)
    Description: Equilibrative nucleoside transporters (ENTs) are integral membrane proteins, which reside in plasma membranes of all eukaryotic cells and mediate thermodynamically downhill transport of nucleosides. This process is essential for nucleoside recycling, and also plays a key role in terminating adenosine-mediated cellular signaling. Furthermore, ENTs mediate the uptake of many drugs, including anticancer and antiviral nucleoside analogues. The structure and mechanism, by which ENTs catalyze trans-membrane transport of their substrates, remain unknown. To identify the core of the transporter needed for stability, activity, and for its correct trafficking to the plasma membrane, we have expressed human ENT deletion mutants in Xenopus laevis oocytes and determined their localization, transport properties and susceptibility to inhibition. We found that the carboxyl terminal trans-membrane segments are essential for correct protein folding and trafficking. In contrast, the soluble extracellular and intracellular loops appear to be dispensable, and must be involved in the fine-tuning of transport regulation.
    Keywords: Research Article
    E-ISSN: 1932-6203
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  • 8
    In: PLoS ONE, 2016, Vol.11(5)
    Description: Bacterial sugar symporters in the Major Facilitator Superfamily (MFS) use the H + (and in a few cases Na + ) electrochemical gradients to achieve active transport of sugar into the cell. Because a number of structures of MFS sugar symporters have been solved recently, molecular insight into the transport mechanism is possible from detailed functional analysis. We present here a comparative electrophysiological study of the lactose permease (LacY), the fucose permease (FucP) and the xylose permease (XylE), which reveals common mechanistic principles and differences. In all three symporters energetically downhill electrogenic sugar/H + symport is observed. Comparison of the pH dependence of symport at symmetrical pH exhibits broad bell-shaped pH profiles extending over 3 to 6 pH units and a decrease at extremely alkaline pH ≥ 9.4 and at acidic to neutral pH = 4.6–7.5. The pH dependence can be described by an acidic to neutral apparent pK (pK app ) and an alkaline pK app . Experimental evidence suggests that the alkaline pK app is due to H + depletion at the protonation site, while the acidic pK app is due to inhibition of deprotonation. Since previous studies suggest that a single carboxyl group in LacY (Glu325) may be the only side chain directly involved in H + translocation and a carboxyl side chain with similar properties has been identified in FucP (Asp46) and XylE (Asp27), the present results imply that the pK of this residue is switched during H + /sugar symport in all three symporters.
    Keywords: Research Article ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Physical Sciences ; Physical Sciences ; Research And Analysis Methods ; Biology And Life Sciences ; Biology And Life Sciences ; Biology And Life Sciences ; Medicine And Health Sciences
    E-ISSN: 1932-6203
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  • 9
    In: PLoS ONE, 2017, Vol.12(12)
    Description: Carbohydrates from plant cell walls are often found as heteropolysaccharides intertwined with each other. For competitive advantage against other microorganisms, and ability to fully exploit available carbon and energy sources, Bacillus subtilis possesses a high number of proteins dedicated to the uptake of mono- and oligosaccharides. Here, we characterize transporter complexes, belonging to the ATP-binding cassette (ABC) superfamily, involved in the uptake of oligosaccharides commonly found in pectin. The uptake of these carbohydrates is shown to be MsmX-dependent, assigning a key role in pectin mobilization for MsmX, a multipurpose ATPase serving several distinct ABC-type I sugar importers. Mutagenesis analysis of the transmembrane domains of the AraNPQ MsmX-dependent importer revealed putative residues for MsmX interaction. Interestingly however, although MsmX is shown to be essential for energizing various ABC transporters we found that a second B . subtilis ATPase, YurJ, is able to complement its function when placed in trans at a different locus of the chromosome.
    Keywords: Research Article ; Biology And Life Sciences ; Biology And Life Sciences ; Medicine And Health Sciences ; Research And Analysis Methods ; Biology And Life Sciences ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Biology And Life Sciences ; Physical Sciences ; Physical Sciences ; Research And Analysis Methods ; Biology And Life Sciences ; Research And Analysis Methods
    E-ISSN: 1932-6203
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  • 10
    Language: English
    In: PLOS ONE, 9/21/2017, Vol.12(9), p.e0184247
    Description: We investigated soluble carbohydrate transport in trees that differed in their phloem loading strategies in order to better understand the transport of photosynthetic products into the roots and the rhizosphere as this knowledge is needed to better understand the respiratory processes in the rhizosphere. We compared beech, which is suggested to use mainly passive loading of transport sugars along a concentration gradient into the phloem, with ash that uses active loading and polymer trapping of raffinose family oligosaccharides (RFOs). We pulse-labeled 20 four-year old European beech and 20 four-year old ash trees with .sup.13 CO.sub.2 and tracked the fate of the label within different plant compartments. We extracted soluble carbohydrates from leaves, bark of stems and branches, and fine roots, measured their amount and isotopic content and calculated their turnover times. In beech one part of the sucrose was rapidly transported into sink tissues without major exchange with storage pools whereas another part of sucrose was strongly exchanged with unlabeled possibly stored sucrose. In contrast the storage and allocation patterns in ash depended on the identity of the transported sugars. RFO were the most important transport sugars that had highest turnover in all shoot compartments. However, the turnover of RFOs in the roots was uncoupled from the shoot. The only significant relation between sugars in the stem base and in the roots of ash was found for the amount (r.sup.2 = 0.50; p = 0.001) and isotopic content (r.sup.2 = 0.47; p = 0.01) of sucrose. The negative relation of the amounts suggested an active transport of sucrose into the roots of ash. Sucrose concentration in the root also best explained the concentration of RFOs in the roots suggesting that RFO in the roots of ash may be resynthesized from sucrose. Our results interestingly suggest that in both tree species only sucrose directly entered the fine root system and that in ash RFOs are transported indirectly into the fine roots only. The direct transport of sucrose might be passive in beech but active in ash (sustained active up- and unloading to co-cells), which would correspond to the phloem loading strategies. Our results give first hints that the transport of carbohydrates between shoot and root is not necessarily continuous and involves passive (beech) and active (ash) transport processes, which may be controlled by the phloem unloading.
    Keywords: Photosynthesis;
    ISSN: PLOS ONE
    E-ISSN: 1932-6203
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