The Experts below are selected from a list of 285 Experts worldwide ranked by ideXlab platform
Birgit Dräger - One of the best experts on this subject based on the ideXlab platform.
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Overexpression of tropinone reductases alters alkaloid composition in Atropa Belladonna root cultures
2013Co-Authors: Ute Richter, Grit Rothe, Anne-katrin Fabian, Bettina Rahfeld, Birgit DrägerAbstract:composition in Atropa Belladonna root culture
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Alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine N‐methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine n methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Calystegines in root cultures of Atropa Belladonna respond to sucrose, not to elicitation.
Plant Science, 2001Co-Authors: Grit Rothe, Uta Garske, Birgit DrägerAbstract:Calystegines are norpseudotropine alkaloids accumulating in root cultures of Atropa Belladonna, together with tropine derivatives, e.g. hyoscyamine. Both alkaloid groups are derived from the tropane alkaloid pathway. For the investigation of the regulation and individual steps of tropane biosynthesis, methods for the induction of the pathway were tested. Elicitation by chitosan, or defence responses to ABA and methyl jasmonate did not enhance calystegine accumulation, but led to a more or less pronounced decrease. By blocking one arm of the diverged tropane pathway, calystegine accumulation can be increased, but total tropane alkaloid formation does not increase considerably. By elevation of sucrose supply, both, total alkaloids and calystegines in particular were increased approximately threefold. The mechanism of the induction of the biosynthesis by sucrose is not known and needs further experiments.
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Tropinone reduction in Atropa Belladonna root cultures
Phytochemistry, 1994Co-Authors: Birgit Dräger, Angela SchaalAbstract:Abstract Transformed root cultures of Atropa Belladonna accumulate the tropane alkaloid hyoscyamine and the precursors tropane-3α-ol (tropine) and tropane-3β-ol (pseudotropine), depending on the culture stage. The latter are stereoisomeric reduction products of tropinone and are formed by two different NADPH-dependent tropinone reductases. Feeding experiments indicate that tropine and pseudotropine do not isomerize and that only tropine is incorporated into hyocyamine. The pseudotropine-forming tropinone reductase was purified, characterized and compared with other tropinone reductases reported so far. It has a molecular weight of 78 500, three identical subunits measured as 27 500 by SDS-PAGE, an IEP at pH 4.5, and a pH optimum at 6.25. The enzyme has a Km of 90,μM for tropinone and 21 μM for NADPH. It catalyses the reduction of a limited number of analogues of tropinone, but with higher Km values. None of the substrate analogues inhibited tropinone reduction. The regulatory role of the pseudotropine-forming tropinone reductase in tropane alkaloid metabolism is discussed.
Yasuyuki Yamada - One of the best experts on this subject based on the ideXlab platform.
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alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine n methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine N‐methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Expression of Atropa Belladonna putrescine N-methyltransferase gene in root pericycle.
Plant and Cell Physiology, 1999Co-Authors: Ken-ichi Suzuki, Yasuyuki Yamada, Takashi HashimotoAbstract:The cDNAs encoding putrescine N-methyltransferase (PMT), which catalyzes the S-adenosylmethionine-dependent N-methylation of putrescine at the first committed step in the biosynthetic pathways of tropane alkaloids, were isolated from Atropa Belladonna and Hyoscyamus niger. These PMTs, however, lacked the N-terminal tandem repeat arrays previously found in Nicotiana PMTs. AbPMT1 RNA was much more abundant in the root of A. Belladonna than was AbPMT2 RNA. The 5'-flanking region of the AbPMT1 gene was fused to the beta-glucuronidase (GUS) reporter gene and transferred to A. Belladonna. Histochemical analysis showed that GUS is expressed specifically in root pericycle cells and that the 0.3-kb 5'-upstream region was sufficient for pericycle-specific expression. Treatment of A. Belladonna roots with methyl jasmonate did not up-regulate the expression of GUS or endogenous AbPMT genes. The regulation of tropane alkaloid biosynthesis is discussed and compared with that of nicotine biosynthesis.
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Metabolic engineering of medicinal plants: transgenic Atropa Belladonna with an improved alkaloid composition
Proceedings of the National Academy of Sciences of the United States of America, 1992Co-Authors: Dae-jin Yun, Takashi Hashimoto, Yasuyuki YamadaAbstract:Abstract The tropane alkaloid scopolamine is a medicinally important anticholinergic drug present in several solanaceous plants. Hyoscyamine 6 beta-hydroxylase (EC 1.14.11.11) catalyzes the oxidative reactions in the biosynthetic pathway leading from hyoscyamine to scopolamine. We introduced the hydroxylase gene from Hyoscyamus niger under the control of the cauliflower mosaic virus 35S promoter into hyoscyamine-rich Atropa Belladonna by the use of an Agrobacterium-mediated transformation system. A transgenic plant that constitutively and strongly expressed the transgene was selected, first by screening for kanamycin resistance and then by immunoscreening leaf samples with an antibody specific for the hydroxylase. In the primary transformant and its selfed progeny that inherited the transgene, the alkaloid contents of the leaf and stem were almost exclusively scopolamine. Such metabolically engineered plants should prove useful as breeding materials for obtaining improved medicinal components.
Koichiro Shimomura - One of the best experts on this subject based on the ideXlab platform.
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Salicylic acid carboxyl methyltransferase induced in hairy root cultures of Atropa Belladonna after treatment with exogeneously added salicylic acid.
Plant and Cell Physiology, 2002Co-Authors: Hiroyuki Fukami, Koichiro Shimomura, Hiroshi Hirano, Tomiko Asakura, Keiko Abe, Takashi YamakawaAbstract:;In Atropa Belladonna hairy roots, exogeneously added salicylic acid (SA) is converted to methyl salicylate (MSA) through the reaction, which might be catalysed by Sadenosyl-L-methionine: salicylic acid carboxyl methyltransferase (SAMT). Here we cloned a cDNA for A. Belladonna SAMT (AbSAMT1), which consisted of 357 aa residues. It was expressed in E. coli, and the recombinant AbSAMT1 showed SAMT activity. When A. Belladonna hairy roots were exposed to a high concentration of SA, AbSAMT1 mRNA begins to be expressed 12 h after the exposure, and steady expression continued over 144 h.
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Responses of Transformed Root Culture of Atropa Belladonna to Salicylic Acid Stress
Journal of Bioscience and Bioengineering, 2002Co-Authors: Kung-ta Lee, Hiroshi Hirano, Takashi Yamakawa, Tohru Kodama, Yasuo Igarashi, Koichiro ShimomuraAbstract:The effect of salicylic acid (SA) on tropane alkaloid production and the responses to SA stress of transformed root cultures of Atropa Belladonna (Belladonna) were investigated. Treatment of A. Belladonna transformed roots with 0.2 mM SA did not have any effect on tropane alkaloid production, but two compounds were produced in the medium. These were identified as the SA derivatives methylsalicylate and methyl-o-methoxybenzoate by high-resolution mass spectrometry and UV spectrometry. In contrast, treatment with 2 mM salicylic acid stimulated tropane alkaloid release from the transformed roots into the medium by up to 35% of the total alkaloids after 24 h, and the SA derivatives were not observed in the medium. These results revealed that transformed root of A. Belladonna exhibits distinct by different responses to SA stress depending on the SA concentration.
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Genetic Transformation of Atropa Belladonna
Transgenic Medicinal Plants, 1999Co-Authors: Mondher El Jaziri, Kayo Yoshimatsu, Koichiro ShimomuraAbstract:Leaves of Atropa Belladonna L. (Solanaceae) are mainly used as a source for the tropane alkaloids, hyoscyamine and scopolamine, which are not easily produced by chemical synthesis. Thus because of its high morphogenetic potential, the in vitro culture of A. Belladonna has been actively investigated as an alternative to field cultivation and as a possible source for tropane alkaloid production (for review see Bajaj and Simola 1991) Several studies have confirmed that the product levels in unorganized A. Belladonna callus and suspension cultures are generally very low and that root morphology is required for synthesis of tropane alkaloid in vitro. However, molecular genetic technology is now available to achieve transformation and regeneration of transgenic A. Belladonna plants in which morphological traits and/or particular metabolic step(s) are artificially modified. The hairy root system is demonstrated to be much more genetically stable than dedifferentiated normal (callus) or transformed (crown gall) plant cells. In this chapter some important recent studies related to the genetic transformation of A. Belladonna are discussed.
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Traits of transgenic Atropa Belladonna doubly transformed with different Agrobacterium rhizogenes strains
Plant Cell Tissue and Organ Culture, 1994Co-Authors: Mondher El Jaziri, Kayo Yoshimatsu, Jacques Homes, Koichiro ShimomuraAbstract:Hairy root cultures of Atropa Belladonna L. were established by infection either with Agrobacterium rhizogenes ATCC 15834 or MAFF 03-01724, and transgenic plants were obtained from both hairy root cultures. Doubly transformed roots were induced by re-infection of the leaf segments of transgenic Atropa Belladonna plants (A. rhizogenes 15834) with MAFF 03-01724. Shoots and viviparous leaves were regenerated from the doubly transformed roots. The genetic transformation was determined by the opine assay (agropine, mannopine and/or mikimopine) and polymerase chain reaction. Physiological changes and tropane alkaloid biosynthesis in the hairy roots (singly and doubly transformed) were investigated. The alkaloid content in the doubly transformed root strain was intermediate as compared to the root strains which were singly transformed. On the other hand endogenous IAA levels in doubly transformed roots were significantly decreased compared to both singly transformed roots.
Grit Rothe - One of the best experts on this subject based on the ideXlab platform.
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Overexpression of tropinone reductases alters alkaloid composition in Atropa Belladonna root cultures
2013Co-Authors: Ute Richter, Grit Rothe, Anne-katrin Fabian, Bettina Rahfeld, Birgit DrägerAbstract:composition in Atropa Belladonna root culture
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Alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine N‐methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine n methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Calystegines in root cultures of Atropa Belladonna respond to sucrose, not to elicitation.
Plant Science, 2001Co-Authors: Grit Rothe, Uta Garske, Birgit DrägerAbstract:Calystegines are norpseudotropine alkaloids accumulating in root cultures of Atropa Belladonna, together with tropine derivatives, e.g. hyoscyamine. Both alkaloid groups are derived from the tropane alkaloid pathway. For the investigation of the regulation and individual steps of tropane biosynthesis, methods for the induction of the pathway were tested. Elicitation by chitosan, or defence responses to ABA and methyl jasmonate did not enhance calystegine accumulation, but led to a more or less pronounced decrease. By blocking one arm of the diverged tropane pathway, calystegine accumulation can be increased, but total tropane alkaloid formation does not increase considerably. By elevation of sucrose supply, both, total alkaloids and calystegines in particular were increased approximately threefold. The mechanism of the induction of the biosynthesis by sucrose is not known and needs further experiments.
Takashi Hashimoto - One of the best experts on this subject based on the ideXlab platform.
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alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine n methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Alkaloids in plants and root cultures of Atropa Belladonna overexpressing putrescine N‐methyltransferase
Journal of Experimental Botany, 2003Co-Authors: Grit Rothe, Yasuyuki Yamada, Takashi Hashimoto, Akira Hachiya, Birgit DrägerAbstract:Putrescine N-methyltransferase (PMT) is the first alkaloid-specific enzyme for nicotine and tropane alkaloid formation. The pmt gene from Nicotiana tabacum was fused to the CaMV 35S promoter and integrated into the Atropa Belladonna genome. Transgenic plants and derived root cultures were analysed for gene expression and for levels of alkaloids and their precursors. Scopolamine, hyoscyamine, tropine, pseudotropine, tropinone, and calystegines were found unaltered or somewhat decreased in pmt-overexpressing lines compared to controls. When root cultures were treated with 5% sucrose, calystegine levels were elevated in control roots, but were not affected in pmt-overexpressing roots. 1 microM auxin reduced calystegine levels in control roots, while in pmt-overexpressing roots all alkaloids remained unaltered. Expression level of pmt alone is apparently not limiting for tropane alkaloid formation in A. Belladonna.
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Expression of Atropa Belladonna putrescine N-methyltransferase gene in root pericycle.
Plant and Cell Physiology, 1999Co-Authors: Ken-ichi Suzuki, Yasuyuki Yamada, Takashi HashimotoAbstract:The cDNAs encoding putrescine N-methyltransferase (PMT), which catalyzes the S-adenosylmethionine-dependent N-methylation of putrescine at the first committed step in the biosynthetic pathways of tropane alkaloids, were isolated from Atropa Belladonna and Hyoscyamus niger. These PMTs, however, lacked the N-terminal tandem repeat arrays previously found in Nicotiana PMTs. AbPMT1 RNA was much more abundant in the root of A. Belladonna than was AbPMT2 RNA. The 5'-flanking region of the AbPMT1 gene was fused to the beta-glucuronidase (GUS) reporter gene and transferred to A. Belladonna. Histochemical analysis showed that GUS is expressed specifically in root pericycle cells and that the 0.3-kb 5'-upstream region was sufficient for pericycle-specific expression. Treatment of A. Belladonna roots with methyl jasmonate did not up-regulate the expression of GUS or endogenous AbPMT genes. The regulation of tropane alkaloid biosynthesis is discussed and compared with that of nicotine biosynthesis.
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Metabolic engineering of medicinal plants: transgenic Atropa Belladonna with an improved alkaloid composition
Proceedings of the National Academy of Sciences of the United States of America, 1992Co-Authors: Dae-jin Yun, Takashi Hashimoto, Yasuyuki YamadaAbstract:Abstract The tropane alkaloid scopolamine is a medicinally important anticholinergic drug present in several solanaceous plants. Hyoscyamine 6 beta-hydroxylase (EC 1.14.11.11) catalyzes the oxidative reactions in the biosynthetic pathway leading from hyoscyamine to scopolamine. We introduced the hydroxylase gene from Hyoscyamus niger under the control of the cauliflower mosaic virus 35S promoter into hyoscyamine-rich Atropa Belladonna by the use of an Agrobacterium-mediated transformation system. A transgenic plant that constitutively and strongly expressed the transgene was selected, first by screening for kanamycin resistance and then by immunoscreening leaf samples with an antibody specific for the hydroxylase. In the primary transformant and its selfed progeny that inherited the transgene, the alkaloid contents of the leaf and stem were almost exclusively scopolamine. Such metabolically engineered plants should prove useful as breeding materials for obtaining improved medicinal components.
