RESUMO
Tuta absoluta ("leafminer"), is a major pest of tomato crops worldwide. Controlling this insect is difficult due to its efficient infestation, rapid proliferation, and resilience to changing weather conditions. Furthermore, chemical pesticides have only a short-term effect due to rapid development of T. absoluta strains. Here, we show that a variety of tomato cultivars, treated with external phenylalanine solutions exhibit high resistance to T. absoluta, under both greenhouse and open field conditions, at different locations. A large-scale metabolomic study revealed that tomato leaves absorb and metabolize externally given Phe efficiently, resulting in a change in their volatile profile, and repellence of T. absoluta moths. The change in the volatile profile is due to an increase in three phenylalanine-derived benzenoid phenylpropanoid volatiles (BPVs), benzaldehyde, phenylacetaldehyde, and 2-phenylethanol. This treatment had no effect on terpenes and green leaf volatiles, known to contribute to the fight against insects. Phe-treated plants also increased the resistance of neighboring non-treated plants. RNAseq analysis of the neighboring non-treated plants revealed an exclusive upregulation of genes, with enrichment of genes related to the plant immune response system. Exposure of tomato plants to either benzaldehyde, phenylacetaldehyde, or 2-phenylethanol, resulted in induction of genes related to the plant immune system that were also induced due to neighboring Phe-treated plants. We suggest a novel role of phenylalanine-derived BPVs as mediators of plant-insect interactions, acting as inducers of the plant defense mechanisms.
Assuntos
Fenilalanina , Folhas de Planta , Solanum lycopersicum , Compostos Orgânicos Voláteis , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Solanum lycopersicum/parasitologia , Fenilalanina/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Animais , Folhas de Planta/metabolismo , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/parasitologia , Benzaldeídos/metabolismo , Benzaldeídos/farmacologia , Acetaldeído/análogos & derivados , Acetaldeído/metabolismo , Acetaldeído/farmacologia , Mariposas/fisiologia , Mariposas/efeitos dos fármacos , Doenças das Plantas/parasitologia , Doenças das Plantas/imunologia , Manduca/fisiologiaRESUMO
Plants are sessile and therefore have developed an extraordinary capacity to adapt to external signals. Here, the focus is on the plasticity of the plant cell to respond to new intracellular cues. Ketocarotenoids are high-value natural red pigments with potent antioxidant activity. In the present study, system-level analyses have revealed that the heterologous biosynthesis of ketocarotenoids in tomato initiated a series of cellular and metabolic mechanisms to cope with the formation of metabolites that are non-endogenous to the plant. The broad multilevel changes were linked to, among others, (i) the remodelling of the plastidial membrane, where the synthesis and storage of ketocarotenoids occurs; (ii) the recruiting of core metabolic pathways for the generation of metabolite precursors and energy; and (iii) redox control. The involvement of the metabolites as regulators of cellular processes shown here reinforces their pivotal role suggested in the remodelled 'central dogma' concept. Furthermore, the role of metabolic reprogramming to ensure cellular homeostasis is proposed.
Assuntos
Carotenoides , Solanum lycopersicum , Carotenoides/metabolismo , Solanum lycopersicum/genética , Reprogramação Metabólica , Plantas/metabolismo , HomeostaseRESUMO
Pistacia palaestina Boiss. is a common tree in the Mediterranean maquis. The leaves of this plant accumulate defensive monoterpenes, whose levels greatly increase in galls induced by the aphid Baizongia pistaciae. We previously found a significant chemopolymorphism in monoterpene content among individual trees, but the chirality of these monoterpenes was unknown. Although most plant species specifically accumulate one enantiomeric form of a given compound, P. palaestina individuals display chemopolymorphism in the chirality of the key monoterpenes accumulated. We report here a marked enantiomeric variation for the limonene, α- and ß-pinene, camphene, sabinene, δ-3-carene, and terpene-4-ol content in leaves and galls of nine different naturally growing P. palaestina trees. Interestingly, insect-induced gall monoterpene composition is an augmentation of the specific enantiopolymorphism originally displayed by each individual tree.
Assuntos
Monoterpenos , Pistacia , Folhas de Planta , Folhas de Planta/química , Monoterpenos/química , Pistacia/química , Estereoisomerismo , Animais , Afídeos , Tumores de Planta/parasitologiaRESUMO
MAIN CONCLUSION: Ambrosia species differ both in their trichome types and in metabolic profiles of leaf volatiles. The current study provides tools for easier taxonomic identification of ragweed species. The genus Ambrosia (Asteraceae) includes some of the most noxious allergenic invasive weeds in the world. Due to high polymorphism in this genus, identification of species is often difficult. This study focuses on microscopic investigation of foliar features and GC-MS identification of the main leaf volatile components of three Ambrosia species currently found in Israel-invasive species Ambrosia confertiflora and A. tenuifolia, and transient A. grayi. A. confertiflora and A. tenuifolia have three trichome types: non-glandular trichomes, capitate glandular trichomes and linear glandular trichomes. Their non-glandular trichomes and capitate trichomes have distinct structures and can serve as taxonomic characters. A. grayi (the least successful invader) has only very dense covering trichomes. All three Ambrosia species have secretory structures in their leaf midrib. A. confertiflora, the most problematic invasive plant in Israel, had a ten times higher volatiles content than the other two species. In A. confertiflora, the most abundant volatiles were chrysanthenone (25.5%), borneol (18%), germacrene D and (E)-caryophyllene (both around 12%). In A. tenuifolia, the most abundant volatiles were ß-myrcene (32.9%), (2E)-hexenal (13%) and 1,8-cineole (11.7%). In A. grayi, the most abundant volatiles were ß-myrcene (17.9%), germacrene D (17.8%) and limonene (14%). The three examined species have distinct trichome types and metabolic profiles. Non-glandular trichomes show structural diversification between species and are a good descriptive character. Considering the anthropocentric significance of this highly problematic genus, the current study provides tools for easier identification of ragweed species.
Assuntos
Ambrosia , Asteraceae , Asteraceae/metabolismo , Monoterpenos Acíclicos/análise , Monoterpenos Acíclicos/metabolismo , Tricomas/metabolismo , Folhas de Planta/metabolismoRESUMO
Galling insects gain food and shelter by inducing specialized anatomical structures in their plant hosts. Such galls often accumulate plant defensive metabolites protecting the inhabiting insects from predation. We previously found that, despite a marked natural chemopolymorphism in natural populations of Pistacia palaestina, the monoterpene content in Baizongia pistaciae-induced galls is substantially higher than in leaves of their hosts. Here we show a general up-regulation of key structural genes in both the plastidial and cytosolic terpene biosynthetic pathways in galls as compared with non-colonized leaves. Novel prenyltransferases and terpene synthases were functionally expressed in Escherichia coli to reveal their biochemical function. Individual Pistacia trees exhibiting chemopolymorphism in terpene compositions displayed differential up-regulation of selected terpene synthase genes, and the metabolites generated by their gene products in vitro corresponded to the monoterpenes accumulated by each tree. Our results delineate molecular mechanisms responsible for the formation of enhanced monoterpene in galls and the observed intraspecific monoterpene chemodiversity displayed in P. palaestina. We demonstrate that gall-inhabiting aphids transcriptionally reprogram their host terpene pathways by up-regulating tree-specific genes, boosting the accumulation of plant defensive compounds for the protection of colonizing insects.
Assuntos
Afídeos , Pistacia , Animais , Tumores de Planta , Terpenos , Regulação para CimaRESUMO
Flowers are the most vulnerable plant organ to infection by the necrotrophic fungus Botrytis cinerea. Here we show that pre-treatment of chrysanthemum (Chrysanthemum morifolium) flowers with phenylalanine (Phe) significantly reduces their susceptibility to B. cinerea. To comprehend how Phe treatment induces resistance, we monitored the dynamics of metabolites (by GC/LC-MS) and transcriptomes (by RNAseq) in flowers after Phe treatment and B. cinerea infection. Phe treatment resulted in accumulation of 3-phenyllactate and benzaldehyde, and in particular induced the expression of genes related to Ca2+ signaling and receptor kinases, implicating an induction of the defense response. Interestingly, the main effects of Phe treatment were observed in flowers exposed to B. cinerea infection, stabilizing the global fluctuations in the levels of metabolites and transcripts while reducing susceptibility to the fungus. We suggest that Phe-induced resistance is associated to cell priming, enabling rapid and targeted reprogramming of cellular defense responses to resist disease development. After Phe pre-treatment, the levels of the anti-fungal volatiles phenylacetaldehyde and eugenol were maintained and the level of coniferin, a plausible monolignol precursor in cell wall lignification, was strongly increased. In addition, Phe pre-treatment reduced ROS generation, prevented ethylene emission, and caused changes in the expression of a minor number of genes related to cell wall biogenesis, encoding the RLK THESEUS1, or involved in Ca2+ and hormonal signaling processes. Our findings point to Phe pre-treatment as a potential orchestrator of a broad-spectrum defense response which may not only provide an ecologically friendly pest control strategy but also offers a promising way of priming plants to induce defense responses against B. cinerea.
Assuntos
Botrytis , Chrysanthemum/fisiologia , Flores/fisiologia , Fenilalanina/fisiologia , Doenças das Plantas/imunologia , Chrysanthemum/imunologia , Chrysanthemum/microbiologia , Etilenos/metabolismo , Flores/imunologia , Fenilalanina/metabolismo , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas/metabolismo , Espécies Reativas de OxigênioRESUMO
Lisianthus (Eustoma grandiflorum), a leading plant in the cut flower industry, is scentless. Here we show that lisianthus flowers have potential to produce several fragrant benzenoid-phenylpropanoids when substrate availability is not limited. To enable hyperaccumulation of substrates for the production of volatile benzenoid-phenylpropanoids, lisianthus commercial hybrid "Excalibur Pink" was transformed via floral dipping with a feedback-insensitive Escherichia coli DAHP synthase (AroG*) and Clarkia breweri benzyl alcohol acetyltransferase (BEAT), under constitutive promoters. The T1 progeny of "Excalibur Pink" plants segregated into four visual phenotypes, with pink or white colored petals and multiple or single petal layers. Interestingly, transformation with AroG* and BEAT caused no significant effect in the pigment composition among phenotypes, but did increase the levels of down-stream fragrant volatile benzenoids. All the transgenic lines exclusively accumulated methyl benzoate, a fragrant benzenoid, either in their petals or leaves. Furthermore, feeding with benzyl alcohol resulted in the accumulation of two novel benzenoids, benzyl acetate (the product of BEAT) and benzoate, as well as a dramatic increase in the concentrations of additional benzenoid-phenylpropanoid volatiles. Presumably, the degree of benzaldehyde overproduction after benzyl alcohol feeding in both leaves and flowers revealed their reverse conversion in lisianthus plants. These findings demonstrate the concealed capability of lisianthus plants to produce a wide array of fragrant benzenoid-phenylpropanoids, given high substrate concentrations, which could in turn open opportunities for future scent engineering.
Assuntos
Flores , Odorantes , Pigmentação , Folhas de Planta , PlantasRESUMO
Combined quantitative trait loci (QTL) and expression-QTL (eQTL) mapping analysis was performed to identify genetic factors affecting melon (Cucumis melo) fruit quality, by linking genotypic, metabolic and transcriptomic data from a melon recombinant inbred line (RIL) population. RNA sequencing (RNA-Seq) of fruit from 96 RILs yielded a highly saturated collection of >â 58â 000 single-nucleotide polymorphisms, identifying 6636 recombination events that separated the genome into 3663 genomic bins. Bin-based QTL analysis of 79 RILs and 129 fruit-quality traits affecting taste, aroma and color resulted in the mapping of 241 QTL. Thiol acyltransferase (CmThAT1) gene was identified within the QTL interval of its product, S-methyl-thioacetate, a key component of melon fruit aroma. Metabolic activity of CmThAT1-encoded protein was validated in bacteria and in vitro. QTL analysis of flesh color intensity identified a candidate white-flesh gene (CmPPR1), one of two major loci determining fruit flesh color in melon. CmPPR1 encodes a member of the pentatricopeptide protein family, involved in processing of RNA in plastids, where carotenoid and chlorophyll pigments accumulate. Network analysis of >â 12â 000 eQTL mapped for >â 8000 differentially expressed fruit genes supported the role of CmPPR1 in determining the expression level of plastid targeted genes. We highlight the potential of RNA-Seq-based QTL analysis of small to moderate size, advanced RIL populations for precise marker-assisted breeding and gene discovery. We provide the following resources: a RIL population genotyped with a unique set of SNP markers, confined genomic segments that harbor QTL governing 129 traits and a saturated set of melon eQTLs.
Assuntos
Mapeamento Cromossômico , Cucurbitaceae/genética , Frutas/genética , Locos de Características Quantitativas/genética , Cucurbitaceae/metabolismo , Qualidade dos Alimentos , Frutas/metabolismo , Genes de Plantas/genética , Genes de Plantas/fisiologia , Ligação Genética , Estudo de Associação Genômica Ampla , Polimorfismo de Nucleotídeo Único/genética , Análise de Sequência de RNARESUMO
Isoprenoids consist of a large class of compounds that are present in all living organisms. They are derived from the 5C building blocks isopentenyl diphosphate (IDP) and its isomer dimethylallyl diphosphate (DMADP). In plants, IDP is synthesized in the cytoplasm from mevalonic acid via the MVA pathway, and in plastids from 2-C-methyl-d-erythritol-4-phosphate through the MEP pathway. The enzyme IDP isomerase (IDI) catalyzes the interconversion between IDP and DMADP. Most plants contain two IDI enzymes, the functions of which are characteristically compartmentalized in the cells. Carotenoids are isoprenoids that play essential roles in photosynthesis and provide colors to flowers and fruits. They are synthesized in the plastids via the MEP pathway. Fruits of Solanum lycopersicum (tomato) accumulate high levels of the red carotene lycopene. We have identified mutations in tomato that reduce overall carotenoid accumulation in fruits. Four alleles of a locus named FRUIT CAROTENOID DEFICIENT 1 (fcd1) were characterized. Map-based cloning of fcd1 indicated that this gene encodes the plastidial enzyme IDI1. Lack of IDI1 reduced the concentration of carotenoids in fruits, flowers and cotyledons, but not in mature leaves. These results indicate that the plastidial IDI plays an important function in carotenoid biosynthesis, thus highlighting its role in optimizing the ratio between IDP and DMADP as precursors for different downstream isoprenoid pathways.
Assuntos
Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Carotenoides/biossíntese , Frutas/metabolismo , Solanum lycopersicum/metabolismo , Isomerases de Ligação Dupla Carbono-Carbono/genética , Frutas/genética , Hemiterpenos/metabolismo , Solanum lycopersicum/genética , Mutação , Compostos Organofosforados/metabolismoRESUMO
Certain insect species can induce gall formation on numerous plants species. Although the mechanism of gall development is largely unknown, it is clear that insects manipulate their hosts' anatomy, physiology, and chemistry for their own benefit. It is well known that insect-induced galls often contain vast amounts of plant defensive compounds as compared to non-colonized tissues, but it is not clear if defensive compounds can be produced in situ in the galled tissues. To answer this question, we analyzed terpene accumulation patterns and possible independent biosynthetic potential of galls induced by the aphid Baizongia pistaciae L. on the terminal buds of Pistacia palaestina Boiss. We compared monoterpene levels and monoterpene synthase enzyme activity in galls and healthy leaves from individual trees growing in a natural setting. At all developmental stages, monoterpene content and monoterpene synthase activity were consistently (up to 10 fold on a fresh weight basis) higher in galls than in intact non-colonized leaves. A remarkable tree to tree variation in the products produced in vitro from the substrate geranyl diphosphate by soluble protein extracts derived from individual trees was observed. Furthermore, galls and leaves from the same trees displayed enhanced and often distinct biosynthetic capabilities. Our results clearly indicate that galls possess independent metabolic capacities to produce and accumulate monoterpenes as compared to leaves. Our study indicates that galling aphids manipulate the enzymatic machinery of their host plant, intensifying their own defenses against natural enemies.
Assuntos
Afídeos/fisiologia , Interações Hospedeiro-Parasita , Monoterpenos/metabolismo , Pistacia/parasitologia , Folhas de Planta/parasitologia , Tumores de Planta/parasitologia , Animais , Monoterpenos/análise , Pistacia/química , Pistacia/metabolismo , Folhas de Planta/química , Folhas de Planta/metabolismoRESUMO
Bay laurel (Laurus nobilis) is an agriculturally and economically important dioecious tree in the basal dicot family Lauraceae used in food and drugs and in the cosmetics industry. Bay leaves, with their abundant monoterpenes and sesquiterpenes, are used to impart flavor and aroma to food, and have also drawn attention in recent years because of their potential pharmaceutical applications. To identify terpene synthases (TPSs) involved in the production of these volatile terpenes, we performed RNA sequencing to profile the transcriptome of L. nobilis leaves. Bioinformatic analysis led to the identification of eight TPS complementary DNAs. We characterized the enzymes encoded by three of these complementary DNAs: a monoterpene synthase that belongs to the TPS-b clade catalyzes the formation of mostly 1,8-cineole; a sesquiterpene synthase belonging to the TPS-a clade catalyzes the formation of mainly cadinenes; and a diterpene synthase of the TPS-e/f clade catalyzes the formation of geranyllinalool. Comparison of the sequences of these three TPSs indicated that the TPS-a and TPS-b clades of the TPS gene family evolved early in the evolution of the angiosperm lineage, and that geranyllinalool synthase activity is the likely ancestral function in angiosperms of genes belonging to an ancient TPS-e/f subclade that diverged from the kaurene synthase gene lineages before the split of angiosperms and gymnosperms.
Assuntos
Alquil e Aril Transferases/genética , Laurus/enzimologia , Terpenos/metabolismo , Alquil e Aril Transferases/metabolismo , Sequência de Bases , Cicloexanóis/metabolismo , DNA Complementar/genética , Eucaliptol , Evolução Molecular , Genes Reporter , Liases Intramoleculares/genética , Liases Intramoleculares/metabolismo , Laurus/química , Laurus/genética , Modelos Moleculares , Dados de Sequência Molecular , Monoterpenos/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , RNA de Plantas/química , RNA de Plantas/genética , Proteínas Recombinantes , Análise de Sequência de RNARESUMO
BACKGROUND: Melon (Cucumis melo) fruits exhibit phenotypic diversity in several key quality determinants such as taste, color and aroma. Sucrose, carotenoids and volatiles are recognized as the key compounds shaping the above corresponding traits yet the full network of biochemical events underlying their synthesis have not been comprehensively described. To delineate the cellular processes shaping fruit quality phenotypes, a population of recombinant inbred lines (RIL) was used as a source of phenotypic and genotypic variations. In parallel, ripe fruits were analyzed for both the quantified level of 77 metabolic traits directly associated with fruit quality and for RNA-seq based expression profiles generated for 27,000 unigenes. First, we explored inter-metabolite association patterns; then, we described metabolites versus gene association patterns; finally, we used the correlation-based associations for predicting uncharacterized synthesis pathways. RESULTS: Based on metabolite versus metabolite and metabolite versus gene association patterns, we divided metabolites into two key groups: a group including ethylene and aroma determining volatiles whose accumulation patterns are correlated with the expression of genes involved in the glycolysis and TCA cycle pathways; and a group including sucrose and color determining carotenoids whose accumulation levels are correlated with the expression of genes associated with plastid formation. CONCLUSIONS: The study integrates multiple processes into a genome scale perspective of cellular activity. This lays a foundation for deciphering the role of gene markers associated with the determination of fruit quality traits.
Assuntos
Cor , Cucurbitaceae/metabolismo , Odorantes , Paladar , Cucurbitaceae/genética , Expressão Gênica , Genes de PlantasRESUMO
Purple Petunia × hybrida V26 plants accumulate fragrant benzenoid-phenylpropanoid molecules and anthocyanin pigments in their petals. These specialized metabolites are synthesized mainly from the aromatic amino acids phenylalanine. Here, we studied the profile of secondary metabolites of petunia plants, expressing a feedback-insensitive bacterial form of 3-deoxy-di-arabino-heptulosonate 7-phosphate synthase enzyme (AroG*) of the shikimate pathway, as a tool to stimulate the conversion of primary to secondary metabolism via the aromatic amino acids. We focused on specialized metabolites contributing to flower showy traits. The presence of AroG* protein led to increased aromatic amino acid levels in the leaves and high phenylalanine levels in the petals. In addition, the AroG* petals accumulated significantly higher levels of fragrant benzenoid-phenylpropanoid volatiles, without affecting the flowers' lifetime. In contrast, AroG* abundance had no effect on flavonoids and anthocyanins levels. The metabolic profile of all five AroG* lines was comparable, even though two lines produced the transgene in the leaves, but not in the petals. This implies that phenylalanine produced in leaves can be transported through the stem to the flowers and serve as a precursor for formation of fragrant metabolites. Dipping cut petunia stems in labelled phenylalanine solution resulted in production of labelled fragrant volatiles in the flowers. This study emphasizes further the potential of this metabolic engineering approach to stimulate the production of specialized metabolites and enhance the quality of various plant organs. Furthermore, transformation of vegetative tissues with AroG* is sufficient for induced production of specialized metabolites in organs such as the flowers.
Assuntos
Aminoácidos Aromáticos/metabolismo , Cruzamentos Genéticos , Flores/crescimento & desenvolvimento , Odorantes , Petunia/crescimento & desenvolvimento , Pigmentação , Transporte Biológico , Isótopos de Carbono , Flores/genética , Genes Bacterianos , Metabolômica , Fenilalanina/metabolismo , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Propanóis/metabolismo , Transformação Genética , Transgenes , Compostos Orgânicos Voláteis/análiseRESUMO
Sulfur-containing aroma volatiles are important contributors to the distinctive aroma of melon and other fruits. Melon cultivars and accessions differ in the content of sulfur-containing and other volatiles. L-methionine has been postulated to serve as a precursor of these volatiles. Incubation of melon fruit cubes with ¹³C- and ²H-labeled L-methionine revealed two distinct catabolic routes into volatiles. One route apparently involves the action of an L-methionine aminotransferase and preserves the main carbon skeleton of L-methionine. The second route apparently involves the action of an L-methionine-γ-lyase activity, releasing methanethiol, a backbone for formation of thiol-derived aroma volatiles. Exogenous L-methionine also generated non-sulfur volatiles by further metabolism of α-ketobutyrate, a product of L-methionine-γ-lyase activity. α-Ketobutyrate was further metabolized into L-isoleucine and other important melon volatiles, including non-sulfur branched and straight-chain esters. Cell-free extracts derived from ripe melon fruit exhibited L-methionine-γ-lyase enzymatic activity. A melon gene (CmMGL) ectopically expressed in Escherichia coli, was shown to encode a protein possessing L-methionine-γ-lyase enzymatic activity. Expression of CmMGL was relatively low in early stages of melon fruit development, but increased in the flesh of ripe fruits, depending on the cultivar tested. Moreover, the levels of expression of CmMGL in recombinant inbred lines co-segregated with the levels of sulfur-containing aroma volatiles enriched with +1 m/z unit and postulated to be produced via this route. Our results indicate that L-methionine is a precursor of both sulfur and non-sulfur aroma volatiles in melon fruit.
Assuntos
Cucumis melo/enzimologia , Frutas/metabolismo , Metionina/metabolismo , Enxofre/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Liases de Carbono-Enxofre/metabolismo , Cucumis melo/genética , Cucumis melo/crescimento & desenvolvimento , Ativação Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Frutas/genética , Frutas/crescimento & desenvolvimento , Genes de Plantas , Isoleucina/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Solubilidade , Especificidade da Espécie , Transaminases/metabolismoRESUMO
Pistacia palaestina Boiss. (Anacardiaceae), a sibling species of P. terebinthus also known as turpentine tree or terebinth tree, is common in the Levant region. The aphid Baizongia pistaciae L. manipulates the leaves of the plant to form large galls, which provide both food and protection for its developing offspring. We analyzed the levels and composition of mono-and sesquiterpenes in both leaves and galls of ten naturally growing trees. Our results show that monoterpene hydrocarbons are the main constituents of P. palaestina leaves and galls, but terpene levels and composition vary among trees. Despite this inter-tree variation, terpene levels and compositions in galls from different trees resemble each other more than the patterns displayed by leaves from the same trees. Generally, galls contain 10 to 60 fold higher total terpene amounts than leaves, especially of the monoterpenes α-pinene and limonene. Conversely, the leaves generally accumulate more sesquiterpenes, in particular E-caryophyllene, germacrene D and δ-cadinene, in comparison to galls. Our results clearly show that the terpene pattern in the galls is not a simple reflection of that of the leaves and suggest that aphids have a strong impact on the metabolism of their host plant, possibly for their own defense.
Assuntos
Pistacia/química , Folhas de Planta/química , Sesquiterpenos/análise , Animais , Afídeos , Monoterpenos Bicíclicos , Cicloexenos/análise , Israel , Limoneno , Monoterpenos/análise , Folhas de Planta/metabolismo , Sesquiterpenos Policíclicos , Sesquiterpenos/química , Sesquiterpenos de Germacrano/análise , Terpenos/análiseRESUMO
In order to broaden the available genetic variation of melon, we developed an ethyl methanesulfonate mutation library in an orange-flesh 'Charentais' type melon line that accumulates ß-carotene. One mutagenized M2 family segregated for a novel recessive trait, a yellow-orange fruit flesh ('yofI'). HPLC analysis revealed that 'yofI' accumulates pro-lycopene (tetra-cis-lycopene) as its major fruit pigment. The altered carotenoid composition of 'yofI' is associated with a significant change of the fruit aroma since cleavage of ß-carotene yields different apocarotenoids than the cleavage of pro-lycopene. Normally, pro-lycopene is further isomerized by CRTISO (carotenoid isomerase) to yield all-trans-lycopene, which is further cyclized to ß-carotene in melon fruit. Cloning and sequencing of 'yofI' CRTISO identified two mRNA sequences which lead to truncated forms of CRTISO. Sequencing of the genomic CRTISO identified an A-T transversion in 'yofI' which leads to a premature STOP codon. The early carotenoid pathway genes were up regulated in yofI fruit causing accumulation of other intermediates such as phytoene and ζ-carotene. Total carotenoid levels are only slightly increased in the mutant. Mutants accumulating pro-lycopene have been reported in both tomato and watermelon fruits, however, this is the first report of a non-lycopene accumulating fruit showing this phenomenon.
Assuntos
Cucumis melo/genética , Metanossulfonato de Etila/química , Mutagênese , beta Caroteno/metabolismo , cis-trans-Isomerases/genética , Vias Biossintéticas/genética , Carotenoides/genética , Cromatografia Líquida de Alta Pressão , Cucumis melo/química , Cucumis melo/crescimento & desenvolvimento , Licopeno , beta Caroteno/química , beta Caroteno/genética , cis-trans-Isomerases/químicaRESUMO
Pepper fruit is typically red, but green, orange and yellow cultivars are gaining consumer acceptance. This color variation is mainly due to variations in carotenoid composition. Orange color in pepper can result from a number of carotenoid profiles, but its genetic basis is only partly known. We identified an EMS-induced orange-fruited mutant using the wild-type blocky red-fruited cultivar 'Maor' as progenitor. This mutant accumulates mainly ß-carotene in its fruit, instead of the complex pattern of red and yellow carotenoids in 'Maor'. We identified an A(709) to G transition in the cDNA of ß-CAROTENE HYDROXYLASE2 in the orange pepper and complete co-segregation of this single-nucleotide polymorphism with the mutated phenotype. We therefore hypothesized that ß-CAROTENE HYDROXYLASE2 controls the orange mutation in pepper. Interestingly, the expression of ß-CAROTENE HYDROXYLASE2 and additional carotenogenesis genes was elevated in the orange fruit compared with the red fruit, indicating possible feedback regulation of genes in the pathway. Because carotenoids serve as precursors for volatile compounds, we compared the volatile profiles of the two parents. The orange pepper contained more volatile compounds than 'Maor', with predominant elevation of norisoprenoids derived from ß-carotene degradation, while sesquiterpenes predominated in the red fruit. Because of the importance of ß-carotene as a provitamin A precursor in the human diet, the orange-fruited mutant might serve as a natural source for pepper fruit biofortification. Moreover, the change in volatile profile may result in a fruit flavor that differs from other pepper cultivars.
Assuntos
Capsicum/genética , Frutas/química , Regulação da Expressão Gênica de Plantas , Oxigenases de Função Mista/genética , beta Caroteno/biossíntese , Capsicum/química , Capsicum/enzimologia , Frutas/enzimologia , Cromatografia Gasosa-Espectrometria de Massas , Genes de Plantas , Marcadores Genéticos , Oxigenases de Função Mista/metabolismo , Mutação , Norisoprenoides/análise , Norisoprenoides/metabolismo , Fenótipo , Pigmentação , Análise de Sequência de DNA , Compostos Orgânicos Voláteis/análiseRESUMO
This paper presents a protocol for the convenient and high-throughput isolation and enrichment of glandular capitate stalked and sessile trichomes from Cannabis sativa. The biosynthetic pathways for cannabinoid and volatile terpene metabolism are localized primarily in the Cannabis trichomes, and isolated trichomes are beneficial for transcriptome analysis. The existing protocols for isolating glandular trichomes for transcriptomic characterization are inconvenient and deliver compromised trichome heads and a relatively low amount of isolated trichomes. Furthermore, they rely on expensive apparatus and isolation media containing protein inhibitors to avoid RNA degradation. The present protocol suggests combining three individual modifications to obtain a large amount of isolated glandular capitate stalked and sessile trichomes from C. sativa mature female inflorescences and fan leaves, respectively. The first modification involves substituting liquid nitrogen for the conventional isolation medium to facilitate the passage of trichomes through the micro-sieves. The second modification involves using dry ice to detach the trichomes from the plant source. The third modification involves passing the plant material consecutively through five micro-sieves of diminishing pore sizes. Microscopic imaging demonstrated the effectiveness of the isolation technique for both trichome types. In addition, the quality of RNA extracted from the isolated trichomes was appropriate for downstream transcriptomic analysis.
Assuntos
Canabinoides , Cannabis , Cannabis/genética , Cannabis/metabolismo , Tricomas/genética , Tricomas/metabolismo , Canabinoides/metabolismo , Folhas de Planta/metabolismo , Extremidade SuperiorRESUMO
Prenylation primarily by geranylgeranylation is required for membrane attachment and function of type I Rho of Plants (ROPs) and Gγ proteins, while type II ROPs are attached to the plasma membrane by S-acylation. Yet, it is not known how prenylation affects ROP membrane interaction dynamics and what are the functional redundancy and specificity of type I and type II ROPs. Here, we have used the expression of ROPs in mammalian cells together with geranylgeranylation and CaaX prenylation-deficient mutants to answer these questions. Our results show that the mechanism of type II ROP S-acylation and membrane attachment is unique to plants and likely responsible for the viability of plants in the absence of CaaX prenylation activity. The prenylation of ROPs determines their steady-state distribution between the plasma membrane and the cytosol but has little effect on membrane interaction dynamics. In addition, the prenyl group type has only minor effects on ROP function. Phenotypic analysis of the CaaX prenylation-deficient pluripetala mutant epidermal cells revealed that type I ROPs affect cell structure primarily on the adaxial side, while type II ROPs are functional and induce a novel cell division phenotype in this genetic background. Taken together, our studies show how prenyl and S-acyl lipid modifications affect ROP subcellular distribution, membrane interaction dynamics, and function.
Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/metabolismo , Proteínas de Membrana/química , Proteínas Monoméricas de Ligação ao GTP/química , Prenilação de Proteína , Acilação , Animais , Arabidopsis/genética , Linhagem Celular , Membrana Celular/química , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Insetos/citologia , Camundongos , Mutação , Células NIH 3T3 , Fenótipo , Epiderme Vegetal/citologiaRESUMO
Khat (Catha edulis Forsk.) is a perennial shrub whose young leaves are chewed for their psychostimulating and anorectic properties. The main active principles of khat are believed to be the phenylpropylamino alkaloids, primarily (-)-cathinone [(S)-α-aminopropiophenone], (+)-cathine [(1S)(2S)-norpseudoephedrine], and (-)-norephedrine [(1R)(2S)-norephedrine]. GC-MS analyses of young leaf extracts indicated the presence of two oxazolidine derivatives, 2,4-dimethyl-5-phenyloxazolidine and 4-methyl-2-(trans-1-pentenyl)-5-phenyloxazolidine. To ascertain the chemical identity of these compounds, we synthesized the putative compounds by condensation of norephedrine and acetaldehyde or trans-2-hexenal, respectively. Spectroscopic analyses (GC-MS, NMR) of the structures of these synthetic compounds showed them to have identical retention indexes and mass spectra characteristic to 2,4-dimethyl-5-phenyloxazolidine and 4-methyl-2-(trans-1-pentenyl)-5-phenyloxazolidine. Marked differences in the ratios between each of these two norephedrine oxazolidine derivatives and total phenylpropylamino alkaloids were found among thirteen different khat accessions further indicating polymorphism in alkaloid ratios and content in C. edulis.