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Stefan Gleissberg - One of the best experts on this subject based on the ideXlab platform.
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Duplicated STM-like KNOX I genes act in floral meristem activity in Eschscholzia Californica (Papaveraceae)
Development Genes and Evolution, 2013Co-Authors: Angelika Stammler, Annette Becker, Sandra S. Meyer, Alastair R. Plant, Brad T. Townsley, Stefan GleissbergAbstract:In angiosperms, the shoot apical meristem is at the origin of leaves and stems and is eventually transformed into the floral meristem. Class I knotted -like homeobox ( KNOX I ) genes are known as crucial regulators of shoot meristem formation and maintenance. KNOX I genes maintain the undifferentiated state of the apical meristem and are locally downregulated upon leaf initiation. In Arabidopsis , KNOX I genes, especially SHOOTMERISTEMLESS ( STM ), have been shown to regulate flower development and the formation of carpels. We investigated the role of STM -like genes in the reproductive development of Eschscholzia Californica , to learn more about the evolution of KNOX I gene function in basal eudicots. We identified two orthologs of STM in Eschscholzia , EcSTM1 and EcSTM2 , which are predominantly expressed in floral tissues. In contrast, a KNAT1 / BP -like and a KNAT2 / 6 -like KNOX I gene are mainly expressed in vegetative organs. Virus-induced gene silencing (VIGS) was used to knockdown gene expression, revealing that both EcSTM genes are required for the formation of reproductive organs. Silencing of EcSTM1 resulted in the loss of the gynoecium and a reduced number of stamens. EcSTM2 -VIGS flowers had reduced and defective gynoecia and a stronger reduction in the number of stamen than observed in EcSTM1 -VIGS. Co-silencing of both genes led to more pronounced phenotypes. In addition, silencing of EcSTM2 alone or together with EcSTM1 resulted in altered patterns of internodal elongation and sometimes in other floral defects. Our data suggest that some aspects of STM function present in Arabidopsis evolved already before the basal eudicots diverged from core eudicots.
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silencing of ecflo a floricaula leafy gene of the california poppy Eschscholzia Californica affects flower specification in a perigynous flower context
International Journal of Plant Sciences, 2013Co-Authors: Sara Wreath, Andrea Scholz, Conny Bartholmes, Oriane Hidalgo, Stefan GleissbergAbstract:Flowers are reproductive shoots produced by determinate floral meristems. The role of FLORICAULA/LEAFY-like genes in the specification of flowers varies between lineages and has not been characterized in a basal eudicot species. Here we report the phenotypic effects of virus-induced silencing of EcFLO, a FLORICAULA/LEAFY homologue in the California poppy Eschscholzia Californica. EcFLO silencing resulted in repeated sepal and petal whorl formation and internodes between calyxes. A subset of silenced flowers showed reduced petal and stamen numbers and mosaic identities of floral organs. We also provide expression data of other floral regulator genes and conclude that EcFLO contributes to certain aspects of flower development, reflecting both functional conservation and differentiation among basal eudicots, core eudicots, and monocot grasses. EcFLO is involved in stamen and petal initiation, floral organ identity delimitation, suppression of internodes, and limitation of the number of perianth whorls. Furth...
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evolution of the yabby gene family with emphasis on the basal eudicot Eschscholzia Californica papaveraceae
Plant Biology, 2011Co-Authors: Conny Bartholmes, Oriane Hidalgo, Stefan GleissbergAbstract:YABBY genes are seed plant-specific transcriptional regulators that are involved in diverse aspects of leaf, shoot and flower development. A series of duplications gave rise to five gene groups found throughout flowering plants. In Arabidopsis and other species, expression of two gene groups, CRABS CLAW and INNER NO OUTER, is restricted to floral organs. In contrast, members of the FILAMENTOUS FLOWER, YABBY2 and YABBY5 gene groups are also expressed in leaves and have been termed 'vegetative YABBYs'. How the five paralogue groups evolved and how their expression and function diversified have remained largely unresolved, precluding a reconstruction of the natural history of this gene family. Here, we report new genes from Eschscholzia Californica (Ranunculales, Papaveraceae) that we use together with currently available database sequences in a comprehensive phylogenetic re-evaluation of the YABBY gene family. Multilayered Bayesian analysis covering seed plants allowed us to locate Eschscholzia YABBY sequences within the gene family phylogeny. We established that vegetative YABBYs do not form a monophyletic clade, and that CRABS CLAW and FILAMENTOUS FLOWER arose from a common ancestor gene. INNER NO OUTER genes are sister to that ancestral gene. We identified several conserved motifs outside of known amino acid domains that define all five angiosperm YABBY gene clades. Further, we inferred the evolution of gene expression and provide evidence for release of purifying constraint in certain branches of the gene family tree. Finally, we report expression patterns for five Eschscholzia YABBY genes consistent with functional conservation between early-diverged and core eudicots.
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highly efficient virus induced gene silencing vigs in california poppy Eschscholzia Californica an evaluation of vigs as a strategy to obtain functional data from non model plants
Annals of Botany, 2007Co-Authors: Stefanie Wege, Stefan Gleissberg, Andrea Scholz, Annette BeckerAbstract:†Background and Aims Eschscholzia Californica (California poppy) is an emerging model plant for ‘evo‐devo’ studies from the basal eudicot clade of Papaveraceae. California poppy has a relatively small genome, a short life cycle and, most importantly, it is amenable for transformation. However, since this transformation protocol is time consuming, virus-induced gene silencing (VIGS) was evaluated as a fast method to obtain functional data for California poppy genes. †Methods Commercially available California poppy plants were infiltrated with Agrobacterium tumefaciens carrying the tobacco rattle virus plasmids pTRV1 and pTRV2. pTRV2 contained part of the Eschscholzia Phytoene Desaturase (EcPDS) gene whose loss of function results in photobleaching of the green parts of the plant and in a lack of floral coloration. The degree and duration of these symptoms was evaluated for vegetative rosettes and plants in flower. †Key Results It is shown that VIGS is able to effectively down-regulate the EcPDS gene in Eschscholzia. Various degrees of silencing were observed starting ,2 weeks after infiltration with Agrobacterium tumefaciens in 92 % of the plants. Tissue with silencing symptoms also showed complete or strong reduction of EcPDS transcripts. Strong silencing resulted in almost completely white petals, fruits, shoots and leaves. Plants with a strong degree of silencing will eventually die off; however, others are able to produce EcPDS gene product even after a strong initial silencing and will recover. Silencing was found to be not always systemic, but was often restricted to certain organs or parts of organs. †Conclusions VIGS is an effective, fast and transient method to down-regulate gene expression in Eschscholzia. It serves well to detect prominent phenotypes which may become obvious even if some target gene transcript remains in the plant tissue. However, subtle phenotypes will be more difficult to detect, as extremely strong silencing effects occur in ,10 % of all flowers from infected plants.
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floral and vegetative morphogenesis in california poppy Eschscholzia Californica cham
International Journal of Plant Sciences, 2005Co-Authors: Annette Becker, Stefan Gleissberg, David R SmythAbstract:For studies of the evolution of development in angiosperms, early‐diverging eudicot taxa are of particular interest for comparisons with established core eudicot model plants, such as Arabidopsis. Here we provide a detailed description of shoot and floral development of the basal eudicot California poppy (Eschscholzia Californica). Rosette formation in the vegetative phase is accompanied by increased leaf complexity and shoot apex size. The flowering phase is characterized by internode elongation, formation of dissected cauline leaves, terminal flowers, and basipetal inflorescence branching. For developing flowers and fruits, we have defined 14 stages according to important landmark events, from inflorescence primordium initiation through seed dispersal. Floral organ initiation, morphogenesis, increase in floral meristem size, and the surface structure of mature floral organs are recorded in detail. The duration of the later floral stages, as well as the path of pollen tube growth in the gynoecium, is doc...
Fumihiko Sato - One of the best experts on this subject based on the ideXlab platform.
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genome wide profiling of wrky genes involved in benzylisoquinoline alkaloid biosynthesis in california poppy Eschscholzia Californica
Frontiers in Plant Science, 2021Co-Authors: Yasuyuki Yamada, Nobukazu Shitan, Fumihiko Sato, Shohei NishidaAbstract:Transcription factors of the WRKY family play pivotal roles in plant defense responses, including the biosynthesis of specialized metabolites. Based on the previous findings of WRKY proteins regulating benzylisoquinoline alkaloid (BIA) biosynthesis, such as CjWRKY1-a regulator of berberine biosynthesis in Coptis japonica-and PsWRKY1-a regulator of morphine biosynthesis in Papaver somniferum-we performed genome-wide characterization of the WRKY gene family in Eschscholzia Californica (California poppy), which produces various BIAs. Fifty WRKY genes were identified by homology search and classified into three groups based on phylogenetic, gene structure, and conserved motif analyses. RNA sequencing showed that several EcWRKY genes transiently responded to methyl jasmonate, a known alkaloid inducer, and the expression patterns of these EcWRKY genes were rather similar to those of BIA biosynthetic enzyme genes. Furthermore, tissue expression profiling suggested the involvement of a few subgroup IIc EcWRKYs in the regulation of BIA biosynthesis. Transactivation analysis using luciferase reporter genes harboring the promoters of biosynthetic enzyme genes indicated little activity of subgroup IIc EcWRKYs, suggesting that the transcriptional network of BIA biosynthesis constitutes multiple members. Finally, we investigated the coexpression patterns of EcWRKYs with some transporter genes and discussed the diversified functions of WRKY genes based on a previous finding that CjWRKY1 overexpression in California poppy cells enhanced BIA secretion into the medium.
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comparative analysis using the draft genome sequence of california poppy Eschscholzia Californica for exploring the candidate genes involved in benzylisoquinoline alkaloid biosynthesis
Bioscience Biotechnology and Biochemistry, 2021Co-Authors: Yasuyuki Yamada, Kentaro Hori, Hideki Hirakawa, Yohei Minakuchi, Atsushi Toyoda, Nobukazu Shitan, Fumihiko SatoAbstract:Genome characterization of California poppy (Eschscholzia Californica cv. "Hitoezaki"), which produces pharmaceutically important benzylisoquinoline alkaloids (BIAs), was carried out using the draft genome sequence. The numbers of tRNA and rRNA genes were close to those of the other plant species tested, whereas the frequency of repetitive sequences was distinct from those species. Comparison of the predicted genes with those of Amborella trichopoda, Nelumbo nucifera, Solanum lycopersicum, and Arabidopsis thaliana, and analyses of gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway indicated that the enzyme genes involved in BIA biosynthesis were highly enriched in the California poppy genome. Further comparative analysis using the genome information of Papaver somniferum and Aquilegia coerulea, both BIA-producing plants, revealed that many genes encoding BIA biosynthetic enzymes, transcription factors, transporters, and candidate proteins, possibly related to BIA biosynthesis, were specifically distributed in these plant species.
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genome wide identification of ap2 erf transcription factor encoding genes in california poppy Eschscholzia Californica and their expression profiles in response to methyl jasmonate
Scientific Reports, 2020Co-Authors: Yasuyuki Yamada, Nobukazu Shitan, Fumihiko Sato, Shohei NishidaAbstract:With respect to the biosynthesis of plant alkaloids, that of benzylisoquinoline alkaloids (BIAs) has been the most investigated at the molecular level. Previous investigations have shown that the biosynthesis of BIAs is comprehensively regulated by WRKY and bHLH transcription factors, while promoter analyses of biosynthesis enzyme-encoding genes have also implicated the involvement of members of the APETALA2/ethylene responsive factor (AP2/ERF) superfamily. To investigate the physiological roles of AP2/ERF transcription factors in BIA biosynthesis, 134 AP2/ERF genes were annotated using the draft genome sequence data of Eschscholzia Californica (California poppy) together with transcriptomic data. Phylogenetic analysis revealed that these genes could be classified into 20 AP2, 5 RAV, 47 DREB, 60 ERF and 2 Soloist family members. Gene structure, conserved motif and orthologous analyses were also carried out. Gene expression profiling via RNA sequencing in response to methyl jasmonate (MeJA) indicated that approximately 20 EcAP2/ERF genes, including 10 group IX genes, were upregulated by MeJA, with an increase in the expression of the transcription factor-encoding gene EcbHLH1 and the biosynthesis enzyme-encoding genes Ec6OMT and EcCYP719A5. Further quantitative RT-PCR confirmed the MeJA responsiveness of the EcAP2/ERF genes, i.e., the increased expression of 9 group IX, 2 group X and 2 group III ERF subfamily genes. Transactivation activity of group IX EcAP2/ERFs was also confirmed by a luciferase reporter assay in conjunction with the promoters of the Ec6OMT and EcCYP719A5 genes. The physiological roles of AP2/ERF genes in BIA biosynthesis and their evolution in the regulation of alkaloid biosynthesis are discussed.
Ramiro O Bustamante - One of the best experts on this subject based on the ideXlab platform.
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negative photoblastism in the invasive specie Eschscholzia Californica cham papaveraceae patterns of altitudinal variation in native and invasive range
Gayana Botánica; Vol. 70 Núm. 2 (2013); 330-336, 2021Co-Authors: Maria L C Castillo, Ramiro O Bustamante, Francisco T Penagomez, Victor L Gutierrez, Claudio A Reyes, Alicia Arredondonunez, Marilyn MareyAbstract:Negative photoblastism is defined as the inhibitory effect of light on seed germination. This effect can be modulated by abiotic variables, such as temperature, light condition and water potential. This conditions change notoriously at higher altitudes, could promote differentiations in the photoblastic response among populations. Also, this physiological attribute poses an interesting conflict for plant regeneration, because prevents seed germination on the soil surface, however, it can also to reduce the mortality of seeds germinants in unsuitable conditions and consequently, seedling mortality. Eschscholzia Californica is one of the few species with negative photoblastism and it is invasive in Chile, growing primarily in open and disturbed places. Some invasive species have the potential to adapt their morphological and ecophysiological characteristics faced to new environmental conditions. In this study, we examined variation in negative photoblastism between populations from central Chile (invasive) and California (native) located at the extremes of their altitudinal distribution. We used common garden experiments where seeds from different origins were germinated under controlled lab condition in Chile. If negative photoblastism is conserved, we would see similar responses among seeds original from this climatic analogues regions. Also, we evaluated whether seed burial increases seed germination of this species as a mechanism for escaping the intense luminosity at the soil surface, by planting seeds at different soil depths. For this experiment, we expected an increase of germination at higher soil depth. The results indicate no differences in negative photoblastism between Chilean and Californian populations. A significant variation across altitudinal range in California suggests the existence of genetic differentiation in the native region, however, the absence of differences across the altitudinal range in Chile suggests trait conservatism at local scale. Seed germination was zero at the soil surface and increased when seeds were experimentally buried, suggesting that negative photoblastism is inhibited. Three possible explanations are given to explain the pattern of establishment of E. Californica despite having negative photoblastism. This is a fairly specialist trait, related with Mediterranean climates and does not explain by itself the invasiveness described for this specie. In any case, is clear that more studies are necessary to disentangle the adaptive value of this physiological trait.
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seed mass seed number and evolutionary trade off across geographic distribution do they explain invasiveness in Eschscholzia Californica central chile
Plant Ecology & Diversity, 2017Co-Authors: Daniel Zamorano, Ramiro O BustamanteAbstract:Background: Seed mass is a life history trait that is related to invasiveness. Under limiting conditions, a trade-off is observed whereby an increase in seed mass occurs at the expense of seed numbers; if the above trade-off holds across climatic gradients it can provide an opportunity to assess the fitness/invasive potential of exotic plants.Aims: To examine the variation in the life history traits of Eschscholzia Californica populations across climatic gradients and to relate these traits to observed invasiveness.Methods: We examined 19 populations in Chile. For each population we related seed mass, seed number, the slope of trade-off between seed mass and number and plant density with annual precipitation and mean annual temperature.Results: Seed number and the coefficient of variation in seed mass were positively correlated with climatic variables. Trade-off was detected in 26% of the populations and no relationship was detected with climatic gradient. Plant density was negatively associated with prec...
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Climatic Niche Conservatism and Biogeographical Non-Equilibrium in Eschscholzia Californica (Papaveraceae), an Invasive Plant in the Chilean Mediterranean Region
2016Co-Authors: Francisco T. Peña-gómez, Pablo C. Guerrero, Gustavo Bizama, Ramiro O BustamanteAbstract:Species climate requirements are useful for predicting their geographic distribution. It is often assumed that the niche requirements for invasive plants are conserved during invasion, especially when the invaded regions share similar climate conditions. California and central Chile have a remarkable degree of convergence in their vegetation structure, and a similar Mediterranean climate. Such similarities make these geographic areas an interesting natural experiment for testing climatic niche dynamics and the equilibrium of invasive species in a new environment. We tested to see if the climatic niche of Eschscholzia Californica is conserved in the invaded range (central Chile), and we assessed whether the invasion process has reached a biogeographical equilibrium, i.e., occupy all the suitable geographic locations that have suitable conditions under native niche requirements. We compared the climatic niche in the native and invaded ranges as well as the projected potential geographic distribution in the invaded range. In order to compare climatic niches, we conducted a Principal Component Analysis (PCA) and Species Distribution Models (SDMs), to estimate E. Californica’s potential geographic distribution. We also used SDMs to predict altitudinal distribution limits in central Chile. Our results indicated that the climatic niche occupied by E. Californica in the invaded range is firmly conserved, occupying a subset of the native climatic niche but leaving a substantial fraction of it unfilled. Comparisons of projected SDMs for central Chile indicate a similarity, yet the projection from native range predicted a larger geographic distribution in central Chile compared to the predictio
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number of conspecifics and reproduction in the invasive plant Eschscholzia Californica papaveraceae is there a pollinator mediated allee effect
Plant Biology, 2015Co-Authors: V Anic, Carolina A Henriquez, Sebastian Abades, Ramiro O BustamanteAbstract:The component Allee effect has been defined as 'a positive relationship between any measure of individual fitness and the number or density of conspecifics'. Larger plant populations or large patches have shown a higher pollinator visitation rate, which may give rise to an Allee effect in reproduction of the plants. We experimentally tested the effect of number of conspecifics on reproduction and pollinator visitation in Eschscholzia Californica Cham., an invasive plant in Chile. We then built patches with two, eight and 16 flowering individuals of E. Californica (11 replicates per treatment) in an area characterised by dominance of the study species. We found that E. Californica exhibits a component Allee effect, as the number of individuals of this species has a positive effect on individual seed set. However, individual fruit production was not affected by the number of plants examined. Pollinator visitation rate was also independent of the number of plants, so this factor would not explain the Allee effect. This rate was positively correlated with the total number of flowers in the patches. We also found that the number of plants did not affect the seed mass or proportion of germinated seeds in the patches. Higher pollen availability in patches with 16 plants and pollination by wind could explain the Allee effect. The component Allee effect identified could lead to a weak demographic Allee effect that might reduce the rate of spread of E. Californica. Knowledge of this would be useful for management of this invasive plant in Chile.
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climatic niche conservatism and biogeographical non equilibrium in Eschscholzia Californica papaveraceae an invasive plant in the chilean mediterranean region
PLOS ONE, 2014Co-Authors: Francisco T Penagomez, Gustavo Bizama, Pablo Guerrero, Milen Duarte, Ramiro O BustamanteAbstract:Species climate requirements are useful for predicting their geographic distribution. It is often assumed that the niche requirements for invasive plants are conserved during invasion, especially when the invaded regions share similar climate conditions. California and central Chile have a remarkable degree of convergence in their vegetation structure, and a similar Mediterranean climate. Such similarities make these geographic areas an interesting natural experiment for testing climatic niche dynamics and the equilibrium of invasive species in a new environment. We tested to see if the climatic niche of Eschscholzia Californica is conserved in the invaded range (central Chile), and we assessed whether the invasion process has reached a biogeographical equilibrium, i.e., occupy all the suitable geographic locations that have suitable conditions under native niche requirements. We compared the climatic niche in the native and invaded ranges as well as the projected potential geographic distribution in the invaded range. In order to compare climatic niches, we conducted a Principal Component Analysis (PCA) and Species Distribution Models (SDMs), to estimate E. Californica's potential geographic distribution. We also used SDMs to predict altitudinal distribution limits in central Chile. Our results indicated that the climatic niche occupied by E. Californica in the invaded range is firmly conserved, occupying a subset of the native climatic niche but leaving a substantial fraction of it unfilled. Comparisons of projected SDMs for central Chile indicate a similarity, yet the projection from native range predicted a larger geographic distribution in central Chile compared to the prediction of the model constructed for central Chile. The projected niche occupancy profile from California predicted a higher mean elevation than that projected from central Chile. We concluded that the invasion process of E. Californica in central Chile is consistent with climatic niche conservatism but there is potential for further expansion in Chile.
Yasuyuki Yamada - One of the best experts on this subject based on the ideXlab platform.
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genome wide profiling of wrky genes involved in benzylisoquinoline alkaloid biosynthesis in california poppy Eschscholzia Californica
Frontiers in Plant Science, 2021Co-Authors: Yasuyuki Yamada, Nobukazu Shitan, Fumihiko Sato, Shohei NishidaAbstract:Transcription factors of the WRKY family play pivotal roles in plant defense responses, including the biosynthesis of specialized metabolites. Based on the previous findings of WRKY proteins regulating benzylisoquinoline alkaloid (BIA) biosynthesis, such as CjWRKY1-a regulator of berberine biosynthesis in Coptis japonica-and PsWRKY1-a regulator of morphine biosynthesis in Papaver somniferum-we performed genome-wide characterization of the WRKY gene family in Eschscholzia Californica (California poppy), which produces various BIAs. Fifty WRKY genes were identified by homology search and classified into three groups based on phylogenetic, gene structure, and conserved motif analyses. RNA sequencing showed that several EcWRKY genes transiently responded to methyl jasmonate, a known alkaloid inducer, and the expression patterns of these EcWRKY genes were rather similar to those of BIA biosynthetic enzyme genes. Furthermore, tissue expression profiling suggested the involvement of a few subgroup IIc EcWRKYs in the regulation of BIA biosynthesis. Transactivation analysis using luciferase reporter genes harboring the promoters of biosynthetic enzyme genes indicated little activity of subgroup IIc EcWRKYs, suggesting that the transcriptional network of BIA biosynthesis constitutes multiple members. Finally, we investigated the coexpression patterns of EcWRKYs with some transporter genes and discussed the diversified functions of WRKY genes based on a previous finding that CjWRKY1 overexpression in California poppy cells enhanced BIA secretion into the medium.
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comparative analysis using the draft genome sequence of california poppy Eschscholzia Californica for exploring the candidate genes involved in benzylisoquinoline alkaloid biosynthesis
Bioscience Biotechnology and Biochemistry, 2021Co-Authors: Yasuyuki Yamada, Kentaro Hori, Hideki Hirakawa, Yohei Minakuchi, Atsushi Toyoda, Nobukazu Shitan, Fumihiko SatoAbstract:Genome characterization of California poppy (Eschscholzia Californica cv. "Hitoezaki"), which produces pharmaceutically important benzylisoquinoline alkaloids (BIAs), was carried out using the draft genome sequence. The numbers of tRNA and rRNA genes were close to those of the other plant species tested, whereas the frequency of repetitive sequences was distinct from those species. Comparison of the predicted genes with those of Amborella trichopoda, Nelumbo nucifera, Solanum lycopersicum, and Arabidopsis thaliana, and analyses of gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway indicated that the enzyme genes involved in BIA biosynthesis were highly enriched in the California poppy genome. Further comparative analysis using the genome information of Papaver somniferum and Aquilegia coerulea, both BIA-producing plants, revealed that many genes encoding BIA biosynthetic enzymes, transcription factors, transporters, and candidate proteins, possibly related to BIA biosynthesis, were specifically distributed in these plant species.
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genome wide identification of ap2 erf transcription factor encoding genes in california poppy Eschscholzia Californica and their expression profiles in response to methyl jasmonate
Scientific Reports, 2020Co-Authors: Yasuyuki Yamada, Nobukazu Shitan, Fumihiko Sato, Shohei NishidaAbstract:With respect to the biosynthesis of plant alkaloids, that of benzylisoquinoline alkaloids (BIAs) has been the most investigated at the molecular level. Previous investigations have shown that the biosynthesis of BIAs is comprehensively regulated by WRKY and bHLH transcription factors, while promoter analyses of biosynthesis enzyme-encoding genes have also implicated the involvement of members of the APETALA2/ethylene responsive factor (AP2/ERF) superfamily. To investigate the physiological roles of AP2/ERF transcription factors in BIA biosynthesis, 134 AP2/ERF genes were annotated using the draft genome sequence data of Eschscholzia Californica (California poppy) together with transcriptomic data. Phylogenetic analysis revealed that these genes could be classified into 20 AP2, 5 RAV, 47 DREB, 60 ERF and 2 Soloist family members. Gene structure, conserved motif and orthologous analyses were also carried out. Gene expression profiling via RNA sequencing in response to methyl jasmonate (MeJA) indicated that approximately 20 EcAP2/ERF genes, including 10 group IX genes, were upregulated by MeJA, with an increase in the expression of the transcription factor-encoding gene EcbHLH1 and the biosynthesis enzyme-encoding genes Ec6OMT and EcCYP719A5. Further quantitative RT-PCR confirmed the MeJA responsiveness of the EcAP2/ERF genes, i.e., the increased expression of 9 group IX, 2 group X and 2 group III ERF subfamily genes. Transactivation activity of group IX EcAP2/ERFs was also confirmed by a luciferase reporter assay in conjunction with the promoters of the Ec6OMT and EcCYP719A5 genes. The physiological roles of AP2/ERF genes in BIA biosynthesis and their evolution in the regulation of alkaloid biosynthesis are discussed.
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metabolic engineering of plant alkaloid biosynthesis
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Takashi Hashimoto, Kum Boo Choi, Takashi Morishige, Kenichi Tamura, Akira Hachiya, Hideki Fujimoto, Yasuyuki YamadaAbstract:Plant alkaloids, one of the largest groups of natural products, provide many pharmacologically active compounds. Several genes in the biosynthetic pathways for scopolamine, nicotine, and berberine have been cloned, making the metabolic engineering of these alkaloids possible. Expression of two branching-point enzymes was engineered: putrescine N-methyltransferase (PMT) in transgenic plants of Atropa belladonna and Nicotiana sylvestris and (S)-scoulerine 9-O-methyltransferase (SMT) in cultured cells of Coptis japonica and Eschscholzia Californica. Overexpression of PMT increased the nicotine content in N. sylvestris, whereas suppression of endogenous PMT activity severely decreased the nicotine content and induced abnormal morphologies. Ectopic expression of SMT caused the accumulation of benzylisoquinoline alkaloids in E. Californica. The prospects and limitations of engineering plant alkaloid metabolism are discussed.
Peter J. Facchini - One of the best experts on this subject based on the ideXlab platform.
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Khat unigenes representing enzymes putatively involved in ephedrine alkaloid biosynthesis.
2015Co-Authors: Ryan A. Groves, Jillian M. Hagel, Ye Zhang, Korey Kilpatrick, Asaf Levy, Frédéric Marsolais, Efraim Lewinsohn, Christoph W. Sensen, Peter J. FacchiniAbstract:Each unigene is assigned an identifier, which corresponds to a database ID in the CED-Trinity library. Percent amino acid identity between unigenes and queries is provided. Abbreviations: AAO4, aromatic aldehyde oxidase 4; Am, Antirrhinum majus; AHAS, acetohydroxyacid synthase; ArAT, aromatic amino acid transaminase; At, Arabidopsis thaliana; BALDH, benzaldehyde dehydrogenase; BDH, benzaldehyde dehydrogenase; BL, benzoate CoA-ligase; BZO, benzoyloxyglucosinolate; CHD, cinnamoyl-CoA hydratase-dehydrogenase; 4CL, 4-coumaroyl-CoA ligase; Ce, Catha edulis; Cm, Cucumis melo; COR, codeinone reductase; Ds, Datura stramonium; Ec, Eschscholzia Californica; Es, Ephedra sinica; KAT, 3-ketoacyl-CoA thiolasae; Ps, Papaver somniferum; PAL, L-phenylalanine ammonia lyase; PDC, pyruvate decarboxylase; Ph, Petunia x hybrida; PPA-AT, prephenate aminotransferase; RED, reductase; SanR, sanguinarine reductase; TA, transaminase; ThDPC, thiamin diphosphate-dependent carboligase; TR, tropinone reducase.Khat unigenes representing enzymes putatively involved in ephedrine alkaloid biosynthesis.
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methods for regeneration and transformation in Eschscholzia Californica
Methods of Molecular Biology, 2006Co-Authors: Benjamin P Macleod, Peter J. FacchiniAbstract:Eschscholzia Californica Cham. (California poppy) is a plant species that accumulates pharmacologically active alkaloids biosynthetically related to the morphinan alkaloids of Papaver somniferum. This, in combination with the relative ease with which it is propagated in vitro, makes it a key model for benzylisoquinoline biosynthesis. Transformation techniques are an important tool for these studies and for metabolic engineering attempts. Agrobacterium mediated transformation techniques for this model species have been developed in our lab and used for modulation of transcript levels relevant to the biosynthesis of these alkaloids. Here we describe the techniques used in our lab for production of transgenic callus, hairy root cultures, and whole plants.
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Somatic embryogenesis from embryogenic cell suspension cultures of california poppy, Eschscholzia Californica Cham
In Vitro Cellular & Developmental Biology - Plant, 2001Co-Authors: S U Park, Peter J. FacchiniAbstract:A somatic embryogenesis protocol was developed for Eschscholzia Californica Chan. (California poppy) using embryogenic cell suspensions and optimized media conditions. Rapidly-growing, finely-dispersed embryogenic cell suspension cultures were established from embryogenic callus and maintained in B5 liquid media supplemented with 0.5 mg 1^−1 (2.26 μ M ) 2,4-dichlorophenoxyacetic acid. Culture conditions were optimized by investigating the effect of basal media composition, gyratory shaker speed, various carbon sources, different cytokinins, and AgNO_3 on the efficiency of somatic embryogenesis. After 40 d in culture, the somatic embryos that formed were counted and their overall growth expressed as pecked cell volume. The selected media consisted of either Gamborg (B5) or Murashige and Skoog (MS) salts and vitamins supplemented with 40 g 1^−1 (117 m M ) sucrose, 0.05 mg 1^−1 (0.22 μ M ) 6-benzylaminopurine, and 10 mg l^−1 (58.8 μ M ) AgNO_3. Somatic embryo production was substantially reduced at shaker speeds above 40 rpm. Glucose and snerose were the most effective carbon sources, whereas fructose, galactose, and maltose resulted in a reduced yield and growth of somatic embryos. The development of somatic embryos was promoted by AgNO_3 at concentrations below 10 mg l^−1 (58.8 μ M ). A semi-solid medium containing 1.5 g l^−1 Gel-rite produced the highest frequency of somatic embryo conversion, and promoted the efficient growth of plantlets. Using the reported protocol, over 500 viable somatic embryos were produced per 25 ml of embryogenic cell suspension culture.
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agrobacterium mediated genetic transformation of california poppy Eschscholzia Californica cham via somatic embryogenesis
Plant Cell Reports, 2000Co-Authors: S U Park, Peter J. FacchiniAbstract:An efficient Agrobacterium-mediated protocol for the stable genetic transformation of Eschscholzia Californica Cham. (California poppy) via somatic embryogenesis is reported. Excised cotyledons were co-cultivated with A. tumefaciens strain GV3101 carrying the pBI121 binary vector. Except for the co-cultivation medium, all formulations included 50 mg l−1 paromomycin as the selective agent and 200 mg l−1 timentin to eliminate the Agrobacterium. Four to five weeks after infection, paromomycin-resistant calli grew on 80% of explants in the presence of 2.0 mg l−1 1-naphthaleneacetic acid (NAA) and 0.1 mg l−1 6-benzylaminopurine (BAP). Calli were cultured on somatic embryogenesis induction medium containing 1.0 mg l−1 NAA and 0.5 mg l−1 BAP, and somatic embryos were visible on 30% of the paromomycin-resistant calli within 3–4 weeks. Three to four weeks after the somatic embryos were transferred to phytohormone-free plant regeneration medium, 32% converted to paromomycin-resistant plants. Detection of the neomycin phosphotransferase gene and high levels of β-glucuronidase (GUS) mRNA and enzyme activity, and the cytohistochemical localization of GUS activity in all plant tissues confirmed the integrative transformation of the regenerated plants. The normal alkaloid profile of California poppy was unaffected by the transformation process; thus, the reported protocol could serve as a valuable tool to investigate the molecular and metabolic regulation of the benzophenanthridine alkaloid pathway.
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agrobacterium rhizogenes mediated transformation of opium poppy papaver somniferum l and california poppy Eschscholzia Californica cham root cultures
Journal of Experimental Botany, 2000Co-Authors: S U Park, Peter J. FacchiniAbstract:An efficient protocol for the establishment of transgenic opium poppy (Papaver somniferum L.) and California poppy (Eschscholzia Californica Cham.) root cultures using A. grobacterium rhizogenes is reported. Five strains of A. rhizogenes were tested for their ability to produce hairy roots on wounded opium poppy seedlings and California poppy embryogenic calli. Three of the strains induced hairy root formation on both species, whereas two others either caused the growth of tumorigenic calli or produced no response. To characterize the putative transgenic roots further, explant tissues were co-cultivated with the most effective A: rhizogenes strain (R1000) carrying the pBI121 binary vector. Except for the co-cultivation medium, all formulations included 50 mg l(-1) paromomycin to select for transformants and 200 mg l(-1) timentin to eliminate the Agrobacterium. Four weeks after infection, paromomycin-resistant roots appeared on 92-98% of explants maintained on hormone-free medium. Isolated hairy roots were propagated in liquid medium containing 1.0 mg l(-1) indole-3-acetic acid to promote rapid growth. Detection of the neomycin phosphotransferase gene, high levels of beta-glucuronidase (GUS) transcripts and enzyme activity, and GUS histochemical localization confirmed the integrative transformation of root cultures. Transgenic roots grew faster than wild-type roots, and California poppy roots grew more rapidly than those of opium poppy. With the exception of a less compact arrangement of epidermal cells and more root hairs, transformed roots of both species displayed anatomical features and benzylisoquinoline alkaloid profiles that were virtually identical to those of wild-type roots. Transgenic root cultures of opium poppy and California poppy are a simple, reliable and well-defined model system to investigate the molecular and metabolic regulation of benzylisoquinoline alkaloid biosynthesis, and to evaluate the genetic engineering potential of these important medicinal plants.