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Marc Boutry - One of the best experts on this subject based on the ideXlab platform.
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activity of the purified plant abc transporter ntpdr1 is stimulated by diterpenes and sesquiterpenes involved in constitutive and induced defenses
Journal of Biological Chemistry, 2017Co-Authors: Baptiste Pierman, Frederic Toussaint, Aurelie Bertin, Nicolas Smargiasso, Daniel Levy, Edwin De Pauw, Marc BoutryAbstract:Abstract Within the plant ATP-binding cassette transporter family, pleiotropic drug resistance (PDR) transporters play essential functions, such as in hormone transport or defense against biotic and abiotic stresses. NtPDR1 from Nicotiana tabacum has been shown to be involved in the constitutive defense against pathogens through the secretion of toxic cyclic diterpenes, such as the antimicrobial substrates cembrene and Sclareol from the leaf hairs (trichomes). However, direct evidence of an interaction between NtPDR1 and terpenes is lacking. Here, we stably expressed NtPDR1 in N. tabacum BY-2 suspension cells. NtPDR1 was purified as an active monomer glycosylated at a single site in the third external loop. NtPDR1 reconstitution in proteoliposomes stimulated its basal ATPase activity from 21 to 38 nmol of Pi·mg−1·min−1, and ATPase activity was further stimulated by the NtPDR1 substrates cembrene and Sclareol, providing direct evidence of an interaction between NtPDR1 and its two substrates. Interestingly, NtPDR1 was also stimulated by capsidiol, a sesquiterpene produced by N. tabacum upon pathogen attack. We also monitored the transcriptional activity from the NtPDR1 promoter in situ with a reporter gene and found that, although NtPDR1 expression was limited to trichomes under normal conditions, addition of methyl jasmonate, a biotic stress hormone, induced expression in all leaf tissues. This finding indicated that NtPDR1 is involved not only in constitutive but also in induced plant defenses. In conclusion, we provide direct evidence of an interaction between the NtPDR1 transporter and its substrates and that NtPDR1 transports compounds involved in both constitutive (diterpenes) and induced (sesquiterpenes) plant defenses.
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nppdr1 a pleiotropic drug resistance type atp binding cassette transporter from nicotiana plumbaginifolia plays a major role in plant pathogen defense
Plant Physiology, 2005Co-Authors: Yvan Stukkens, Alain Bultreys, Sebastien Grec, Tomasz Trombik, Delphine Vanham, Marc BoutryAbstract:Nicotiana plumbaginifolia NpPDR1, a plasma membrane pleiotropic drug resistance-type ATP-binding cassette transporter formerly named NpABC1, has been suggested to transport the diterpene Sclareol, an antifungal compound. However, direct evidence for a role of pleiotropic drug resistance transporters in the plant defense is still lacking. In situ immunolocalization and histochemical analysis using the gusA reporter gene showed that NpPDR1 was constitutively expressed in the whole root, in the leaf glandular trichomes, and in the flower petals. However, NpPDR1 expression was induced in the whole leaf following infection with the fungus Botrytis cinerea, and the bacteria Pseudomonas syringae pv tabaci, Pseudomonas fluorescens, and Pseudomonas marginalis pv marginalis, which do not induce a hypersensitive response in N. plumbaginifolia, whereas a weaker response was observed using P. syringae pv syringae, which does induce a hypersensitive response. Induced NpPDR1 expression was more associated with the jasmonic acid than the salicylic acid signaling pathway. These data suggest that NpPDR1 is involved in both constitutive and jasmonic acid-dependent induced defense. Transgenic plants in which NpPDR1 expression was prevented by RNA interference showed increased sensitivity to Sclareol and reduced resistance to B. cinerea. These data show that NpPDR1 is involved in pathogen resistance and thus demonstrate a new role for the ATP-binding cassette transporter family.
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a plant plasma membrane atp binding cassette type transporter is involved in antifungal terpenoid secretion
The Plant Cell, 2001Co-Authors: Michal Jasinski, Yvan Stukkens, Herve Degand, Benedicte Purnelle, Jacqueline Marchandbrynaert, Marc BoutryAbstract:ATP binding cassette (ABC) transporters, which are found in all species, are known mainly for their ability to confer drug resistance. To date, most of the ABC transporters characterized in plants have been localized in the vacuolar membrane and are considered to be involved in the intracellular sequestration of cytotoxins. Working on the assumption that certain ABC transporters might be involved in defense metabolite secretion and their expression might be regulated by the concentration of these metabolites, we treated a Nicotiana plumbaginifolia cell culture with Sclareolide, a close analog of Sclareol, an antifungal diterpene produced at the leaf surface of Nicotiana spp; this resulted in the appearance of a 160-kD plasma membrane protein, which was partially sequenced. The corresponding cDNA (NpABC1) was cloned and shown to encode an ABC transporter. In vitro and in situ immunodetection showed NpABC1 to be localized in the plasma membrane. Under normal conditions, expression was found in the leaf epidermis. In cell culture and in leaf tissues, NpABC1 expression was strongly enhanced by Sclareolide and Sclareol. In parallel with NpABC1 induction, cells acquired the ability to excrete a labeled synthetic Sclareolide derivative. These data suggest that NpABC1 is involved in the secretion of a secondary metabolite that plays a role in plant defense.
Yvan Stukkens - One of the best experts on this subject based on the ideXlab platform.
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nppdr1 a pleiotropic drug resistance type atp binding cassette transporter from nicotiana plumbaginifolia plays a major role in plant pathogen defense
Plant Physiology, 2005Co-Authors: Yvan Stukkens, Alain Bultreys, Sebastien Grec, Tomasz Trombik, Delphine Vanham, Marc BoutryAbstract:Nicotiana plumbaginifolia NpPDR1, a plasma membrane pleiotropic drug resistance-type ATP-binding cassette transporter formerly named NpABC1, has been suggested to transport the diterpene Sclareol, an antifungal compound. However, direct evidence for a role of pleiotropic drug resistance transporters in the plant defense is still lacking. In situ immunolocalization and histochemical analysis using the gusA reporter gene showed that NpPDR1 was constitutively expressed in the whole root, in the leaf glandular trichomes, and in the flower petals. However, NpPDR1 expression was induced in the whole leaf following infection with the fungus Botrytis cinerea, and the bacteria Pseudomonas syringae pv tabaci, Pseudomonas fluorescens, and Pseudomonas marginalis pv marginalis, which do not induce a hypersensitive response in N. plumbaginifolia, whereas a weaker response was observed using P. syringae pv syringae, which does induce a hypersensitive response. Induced NpPDR1 expression was more associated with the jasmonic acid than the salicylic acid signaling pathway. These data suggest that NpPDR1 is involved in both constitutive and jasmonic acid-dependent induced defense. Transgenic plants in which NpPDR1 expression was prevented by RNA interference showed increased sensitivity to Sclareol and reduced resistance to B. cinerea. These data show that NpPDR1 is involved in pathogen resistance and thus demonstrate a new role for the ATP-binding cassette transporter family.
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a plant plasma membrane atp binding cassette type transporter is involved in antifungal terpenoid secretion
The Plant Cell, 2001Co-Authors: Michal Jasinski, Yvan Stukkens, Herve Degand, Benedicte Purnelle, Jacqueline Marchandbrynaert, Marc BoutryAbstract:ATP binding cassette (ABC) transporters, which are found in all species, are known mainly for their ability to confer drug resistance. To date, most of the ABC transporters characterized in plants have been localized in the vacuolar membrane and are considered to be involved in the intracellular sequestration of cytotoxins. Working on the assumption that certain ABC transporters might be involved in defense metabolite secretion and their expression might be regulated by the concentration of these metabolites, we treated a Nicotiana plumbaginifolia cell culture with Sclareolide, a close analog of Sclareol, an antifungal diterpene produced at the leaf surface of Nicotiana spp; this resulted in the appearance of a 160-kD plasma membrane protein, which was partially sequenced. The corresponding cDNA (NpABC1) was cloned and shown to encode an ABC transporter. In vitro and in situ immunodetection showed NpABC1 to be localized in the plasma membrane. Under normal conditions, expression was found in the leaf epidermis. In cell culture and in leaf tissues, NpABC1 expression was strongly enhanced by Sclareolide and Sclareol. In parallel with NpABC1 induction, cells acquired the ability to excrete a labeled synthetic Sclareolide derivative. These data suggest that NpABC1 is involved in the secretion of a secondary metabolite that plays a role in plant defense.
Jean-louis Magnard - One of the best experts on this subject based on the ideXlab platform.
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Extracellular Localization of the Diterpene Sclareol in Clary Sage (Salvia sclarea L., Lamiaceae)
PLoS Biology, 2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:Sclareol is a high-value natural product obtained by solid/liquid extraction of clary sage (Salvia sclarea L.) inflorescences. Because processes of excretion and accumulation of this labdane diterpene are unknown, the aim of this work was to gain knowledge on its sites of accumulation in planta. Samples were collected in natura or during different steps of the industrial process of extraction (steam distillation and solid/liquid extraction). Samples were then analysed with a combination of complementary analytical techniques (gas chromatography coupled to a mass spectrometer, polarized light microscopy, environmental scanning electron microscopy, two-photon fluorescence microscopy, second harmonic generation microscopy). According to the literature, it is hypothesized that Sclareol is localized in oil pockets of secretory trichomes. This study demonstrates that this is not the case and that Sclareol accumulates in a crystalline epicuticular form, mostly on calyces.
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discovery and functional characterization of two diterpene synthases for Sclareol biosynthesis in salvia sclarea l and their relevance for perfume manufacture
BMC Plant Biology, 2012Co-Authors: Anne Caniard, Sylvain Legrand, Nadine Valot, Jean-louis Magnard, Philipp Zerbe, Allison Cohade, Jorg BohlmannAbstract:Sclareol is a diterpene natural product of high value for the fragrance industry. Its labdane carbon skeleton and its two hydroxyl groups also make it a valued starting material for semisynthesis of numerous commercial substances, including production of Ambrox® and related ambergris substitutes used in the formulation of high end perfumes. Most of the commercially-produced Sclareol is derived from cultivated clary sage (Salvia sclarea) and extraction of the plant material. In clary sage, Sclareol mainly accumulates in essential oil-producing trichomes that densely cover flower calices. Manool also is a minor diterpene of this species and the main diterpene of related Salvia species. Based on previous general knowledge of diterpene biosynthesis in angiosperms, and based on mining of our recently published transcriptome database obtained by deep 454-sequencing of cDNA from clary sage calices, we cloned and functionally characterized two new diterpene synthase (diTPS) enzymes for the complete biosynthesis of Sclareol in clary sage. A class II diTPS (SsLPPS) produced labda-13-en-8-ol diphosphate as major product from geranylgeranyl diphosphate (GGPP) with some minor quantities of its non-hydroxylated analogue, (9 S, 10 S)-copalyl diphosphate. A class I diTPS (SsSS) then transformed these intermediates into Sclareol and manool, respectively. The production of Sclareol was reconstructed in vitro by combining the two recombinant diTPS enzymes with the GGPP starting substrate and in vivo by co-expression of the two proteins in yeast (Saccharomyces cerevisiae). Tobacco-based transient expression assays of green fluorescent protein-fusion constructs revealed that both enzymes possess an N-terminal signal sequence that actively targets SsLPPS and SsSS to the chloroplast, a major site of GGPP and diterpene production in plants. SsLPPS and SsSS are two monofunctional diTPSs which, together, produce the diterpenoid specialized metabolite Sclareol in a two-step process. They represent two of the first characterized hydroxylating diTPSs in angiosperms and generate the dihydroxylated labdane Sclareol without requirement for additional enzymatic oxidation by activities such as cytochrome P450 monoxygenases. Yeast-based production of Sclareol by co-expresssion of SsLPPS and SsSS was efficient enough to warrant the development and use of such technology for the biotechnological production of scareol and other oxygenated diterpenes.
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Discovery and functional characterization of two diterpene synthases for Sclareol biosynthesis in Salvia sclarea (L.) and their relevance for perfume manufacture
BMC Plant Biology, 2012Co-Authors: Anne Caniard, Sylvain Legrand, Nadine Valot, Jean-louis Magnard, Philipp Zerbe, Allison Cohade, Joerg Bohlmann, Laurent LegendreAbstract:Background: Sclareol is a diterpene natural product of high value for the fragrance industry. Its labdane carbon skeleton and its two hydroxyl groups also make it a valued starting material for semisynthesis of numerous commercial substances, including production of Ambrox (R) and related ambergris substitutes used in the formulation of high end perfumes. Most of the commercially-produced Sclareol is derived from cultivated clary sage (Salvia sclarea) and extraction of the plant material. In clary sage, Sclareol mainly accumulates in essential oil-producing trichomes that densely cover flower calices. Manool also is a minor diterpene of this species and the main diterpene of related Salvia species. Results: Based on previous general knowledge of diterpene biosynthesis in angiosperms, and based on mining of our recently published transcriptome database obtained by deep 454-sequencing of cDNA from clary sage calices, we cloned and functionally characterized two new diterpene synthase (diTPS) enzymes for the complete biosynthesis of Sclareol in clary sage. A class II diTPS (SsLPPS) produced labda-13-en-8-ol diphosphate as major product from geranylgeranyl diphosphate (GGPP) with some minor quantities of its non-hydroxylated analogue, (9 S, 10 S)-copalyl diphosphate. A class I diTPS (SsSS) then transformed these intermediates into Sclareol and manool, respectively. The production of Sclareol was reconstructed in vitro by combining the two recombinant diTPS enzymes with the GGPP starting substrate and in vivo by co-expression of the two proteins in yeast (Saccharomyces cerevisiae). Tobacco-based transient expression assays of green fluorescent protein-fusion constructs revealed that both enzymes possess an N-terminal signal sequence that actively targets SsLPPS and SsSS to the chloroplast, a major site of GGPP and diterpene production in plants. Conclusions: SsLPPS and SsSS are two monofunctional diTPSs which, together, produce the diterpenoid specialized metabolite Sclareol in a two-step process. They represent two of the first characterized hydroxylating diTPSs in angiosperms and generate the dihydroxylated labdane Sclareol without requirement for additional enzymatic oxidation by activities such as cytochrome P450 monoxygenases. Yeast-based production of Sclareol by co-expresssion of SsLPPS and SsSS was efficient enough to warrant the development and use of such technology for the biotechnological production of scareol and other oxygenated diterpenes.
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Microscopic imaging of crystal-like structures of clary sage and Sclareol crystals.
2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:(A) Polarized light micrography of Sclareol crystals. (B, C, D) Polarized light micrography of crystal-like structures on the cuticle of calyces showing a cluster on C and different shapes on D. (E) Differential interference contrast micrography of crystal-like structures on the cuticle of calyces. (F) TPF micrography of the same structures as in E. (G) Imaging of SHG signal (green canal) merge with photo F. (H) TPF micrography of Sclareol crystals. (G) Imaging of SHG signal (green canal) merge with photo H. Legends: Full arrowheads, epicuticular crystal-like structures. Scale bars: 10 µm (D), 15 µm (B, C, E, F, G), 30 µm (A, H, I).
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Observation and GC-MS analysis of epicuticular crystals.
2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:(A) Light micrography of cuticular crystals just before the industrial process of Sclareol extraction. (B) Light micrography of cuticular crystals after steam distillation. (C) Light micrography of the cuticle after solid/liquid extraction with hexane. (D) Polarized micrography of crystals observed in the pellet after centrifugation (note the cuticle scraps). Trichomes were photographed because they allowed the best imaging after the treatments. (E) GC-MS analysis of essential oil obtained by steam distillation of straw and diluted 10-fold in hexane (% of total peak area: 16% peak 1, 62% peak 2, 4% peak 3, 6% peak 4, 2% peak 5). (F) GC-MS analysis of an hexane wash of calyces dissected after steam distillation of the straw (% of total peak area: 97% peak 5). (G) GC-MS analysis of pellets collected after centrifugation of calyces in water and phase extracted with hexane (% of total peak area: 11% peak 2, 84% peak 5). (H) GC-MS analysis of commercial Sclareol (purity >2%, 2 g/L in hexane; total peak area, 96% peak 5). Peak identifications are based on retention time, mass spectrum and comparison with authentic standards. Legends: 1, linalool; 2, linalyl acetate; 3, β-caryophyllene; 4, germacrene D; 5, Sclareol; Arrows, cuticle; Arrow head, crystals; TIC, total ion count. Scale bars: 30 µm.
Nadine Valot - One of the best experts on this subject based on the ideXlab platform.
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Extracellular Localization of the Diterpene Sclareol in Clary Sage (Salvia sclarea L., Lamiaceae)
PLoS Biology, 2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:Sclareol is a high-value natural product obtained by solid/liquid extraction of clary sage (Salvia sclarea L.) inflorescences. Because processes of excretion and accumulation of this labdane diterpene are unknown, the aim of this work was to gain knowledge on its sites of accumulation in planta. Samples were collected in natura or during different steps of the industrial process of extraction (steam distillation and solid/liquid extraction). Samples were then analysed with a combination of complementary analytical techniques (gas chromatography coupled to a mass spectrometer, polarized light microscopy, environmental scanning electron microscopy, two-photon fluorescence microscopy, second harmonic generation microscopy). According to the literature, it is hypothesized that Sclareol is localized in oil pockets of secretory trichomes. This study demonstrates that this is not the case and that Sclareol accumulates in a crystalline epicuticular form, mostly on calyces.
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discovery and functional characterization of two diterpene synthases for Sclareol biosynthesis in salvia sclarea l and their relevance for perfume manufacture
BMC Plant Biology, 2012Co-Authors: Anne Caniard, Sylvain Legrand, Nadine Valot, Jean-louis Magnard, Philipp Zerbe, Allison Cohade, Jorg BohlmannAbstract:Sclareol is a diterpene natural product of high value for the fragrance industry. Its labdane carbon skeleton and its two hydroxyl groups also make it a valued starting material for semisynthesis of numerous commercial substances, including production of Ambrox® and related ambergris substitutes used in the formulation of high end perfumes. Most of the commercially-produced Sclareol is derived from cultivated clary sage (Salvia sclarea) and extraction of the plant material. In clary sage, Sclareol mainly accumulates in essential oil-producing trichomes that densely cover flower calices. Manool also is a minor diterpene of this species and the main diterpene of related Salvia species. Based on previous general knowledge of diterpene biosynthesis in angiosperms, and based on mining of our recently published transcriptome database obtained by deep 454-sequencing of cDNA from clary sage calices, we cloned and functionally characterized two new diterpene synthase (diTPS) enzymes for the complete biosynthesis of Sclareol in clary sage. A class II diTPS (SsLPPS) produced labda-13-en-8-ol diphosphate as major product from geranylgeranyl diphosphate (GGPP) with some minor quantities of its non-hydroxylated analogue, (9 S, 10 S)-copalyl diphosphate. A class I diTPS (SsSS) then transformed these intermediates into Sclareol and manool, respectively. The production of Sclareol was reconstructed in vitro by combining the two recombinant diTPS enzymes with the GGPP starting substrate and in vivo by co-expression of the two proteins in yeast (Saccharomyces cerevisiae). Tobacco-based transient expression assays of green fluorescent protein-fusion constructs revealed that both enzymes possess an N-terminal signal sequence that actively targets SsLPPS and SsSS to the chloroplast, a major site of GGPP and diterpene production in plants. SsLPPS and SsSS are two monofunctional diTPSs which, together, produce the diterpenoid specialized metabolite Sclareol in a two-step process. They represent two of the first characterized hydroxylating diTPSs in angiosperms and generate the dihydroxylated labdane Sclareol without requirement for additional enzymatic oxidation by activities such as cytochrome P450 monoxygenases. Yeast-based production of Sclareol by co-expresssion of SsLPPS and SsSS was efficient enough to warrant the development and use of such technology for the biotechnological production of scareol and other oxygenated diterpenes.
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Discovery and functional characterization of two diterpene synthases for Sclareol biosynthesis in Salvia sclarea (L.) and their relevance for perfume manufacture
BMC Plant Biology, 2012Co-Authors: Anne Caniard, Sylvain Legrand, Nadine Valot, Jean-louis Magnard, Philipp Zerbe, Allison Cohade, Joerg Bohlmann, Laurent LegendreAbstract:Background: Sclareol is a diterpene natural product of high value for the fragrance industry. Its labdane carbon skeleton and its two hydroxyl groups also make it a valued starting material for semisynthesis of numerous commercial substances, including production of Ambrox (R) and related ambergris substitutes used in the formulation of high end perfumes. Most of the commercially-produced Sclareol is derived from cultivated clary sage (Salvia sclarea) and extraction of the plant material. In clary sage, Sclareol mainly accumulates in essential oil-producing trichomes that densely cover flower calices. Manool also is a minor diterpene of this species and the main diterpene of related Salvia species. Results: Based on previous general knowledge of diterpene biosynthesis in angiosperms, and based on mining of our recently published transcriptome database obtained by deep 454-sequencing of cDNA from clary sage calices, we cloned and functionally characterized two new diterpene synthase (diTPS) enzymes for the complete biosynthesis of Sclareol in clary sage. A class II diTPS (SsLPPS) produced labda-13-en-8-ol diphosphate as major product from geranylgeranyl diphosphate (GGPP) with some minor quantities of its non-hydroxylated analogue, (9 S, 10 S)-copalyl diphosphate. A class I diTPS (SsSS) then transformed these intermediates into Sclareol and manool, respectively. The production of Sclareol was reconstructed in vitro by combining the two recombinant diTPS enzymes with the GGPP starting substrate and in vivo by co-expression of the two proteins in yeast (Saccharomyces cerevisiae). Tobacco-based transient expression assays of green fluorescent protein-fusion constructs revealed that both enzymes possess an N-terminal signal sequence that actively targets SsLPPS and SsSS to the chloroplast, a major site of GGPP and diterpene production in plants. Conclusions: SsLPPS and SsSS are two monofunctional diTPSs which, together, produce the diterpenoid specialized metabolite Sclareol in a two-step process. They represent two of the first characterized hydroxylating diTPSs in angiosperms and generate the dihydroxylated labdane Sclareol without requirement for additional enzymatic oxidation by activities such as cytochrome P450 monoxygenases. Yeast-based production of Sclareol by co-expresssion of SsLPPS and SsSS was efficient enough to warrant the development and use of such technology for the biotechnological production of scareol and other oxygenated diterpenes.
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Microscopic imaging of crystal-like structures of clary sage and Sclareol crystals.
2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:(A) Polarized light micrography of Sclareol crystals. (B, C, D) Polarized light micrography of crystal-like structures on the cuticle of calyces showing a cluster on C and different shapes on D. (E) Differential interference contrast micrography of crystal-like structures on the cuticle of calyces. (F) TPF micrography of the same structures as in E. (G) Imaging of SHG signal (green canal) merge with photo F. (H) TPF micrography of Sclareol crystals. (G) Imaging of SHG signal (green canal) merge with photo H. Legends: Full arrowheads, epicuticular crystal-like structures. Scale bars: 10 µm (D), 15 µm (B, C, E, F, G), 30 µm (A, H, I).
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Observation and GC-MS analysis of epicuticular crystals.
2012Co-Authors: Jean-claude Caissard, Nadine Valot, Thomas Olivier, Claire Delbecque, Sabine Palle, Pierre-philippe Garry, Arthur Audran, Sandrine Moja, Florence Nicolè, Jean-louis MagnardAbstract:(A) Light micrography of cuticular crystals just before the industrial process of Sclareol extraction. (B) Light micrography of cuticular crystals after steam distillation. (C) Light micrography of the cuticle after solid/liquid extraction with hexane. (D) Polarized micrography of crystals observed in the pellet after centrifugation (note the cuticle scraps). Trichomes were photographed because they allowed the best imaging after the treatments. (E) GC-MS analysis of essential oil obtained by steam distillation of straw and diluted 10-fold in hexane (% of total peak area: 16% peak 1, 62% peak 2, 4% peak 3, 6% peak 4, 2% peak 5). (F) GC-MS analysis of an hexane wash of calyces dissected after steam distillation of the straw (% of total peak area: 97% peak 5). (G) GC-MS analysis of pellets collected after centrifugation of calyces in water and phase extracted with hexane (% of total peak area: 11% peak 2, 84% peak 5). (H) GC-MS analysis of commercial Sclareol (purity >2%, 2 g/L in hexane; total peak area, 96% peak 5). Peak identifications are based on retention time, mass spectrum and comparison with authentic standards. Legends: 1, linalool; 2, linalyl acetate; 3, β-caryophyllene; 4, germacrene D; 5, Sclareol; Arrows, cuticle; Arrow head, crystals; TIC, total ion count. Scale bars: 30 µm.
Michal Jasinski - One of the best experts on this subject based on the ideXlab platform.
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a plant plasma membrane atp binding cassette type transporter is involved in antifungal terpenoid secretion
The Plant Cell, 2001Co-Authors: Michal Jasinski, Yvan Stukkens, Herve Degand, Benedicte Purnelle, Jacqueline Marchandbrynaert, Marc BoutryAbstract:ATP binding cassette (ABC) transporters, which are found in all species, are known mainly for their ability to confer drug resistance. To date, most of the ABC transporters characterized in plants have been localized in the vacuolar membrane and are considered to be involved in the intracellular sequestration of cytotoxins. Working on the assumption that certain ABC transporters might be involved in defense metabolite secretion and their expression might be regulated by the concentration of these metabolites, we treated a Nicotiana plumbaginifolia cell culture with Sclareolide, a close analog of Sclareol, an antifungal diterpene produced at the leaf surface of Nicotiana spp; this resulted in the appearance of a 160-kD plasma membrane protein, which was partially sequenced. The corresponding cDNA (NpABC1) was cloned and shown to encode an ABC transporter. In vitro and in situ immunodetection showed NpABC1 to be localized in the plasma membrane. Under normal conditions, expression was found in the leaf epidermis. In cell culture and in leaf tissues, NpABC1 expression was strongly enhanced by Sclareolide and Sclareol. In parallel with NpABC1 induction, cells acquired the ability to excrete a labeled synthetic Sclareolide derivative. These data suggest that NpABC1 is involved in the secretion of a secondary metabolite that plays a role in plant defense.
