RESUMO
L-serine (Ser) and L-glycine (Gly) are critically important for the overall functioning of primary metabolism. We investigated the interaction of the phosphorylated pathway of Ser biosynthesis (PPSB) with the photorespiration-associated glycolate pathway of Ser biosynthesis (GPSB) using Arabidopsis thaliana PPSB-deficient lines, GPSB-deficient mutants, and crosses of PPSB with GPSB mutants. PPSB-deficient lines mainly showed retarded primary root growth. Mutation of the photorespiratory enzyme Ser-hydroxymethyltransferase 1 (SHMT1) in a PPSB-deficient background resumed primary root growth and induced a change in the plant metabolic pattern between roots and shoots. Grafting experiments demonstrated that metabolic changes in shoots were responsible for the changes in double mutant development. PPSB disruption led to a reduction in nitrogen (N) and sulfur (S) contents in shoots and a general transcriptional response to nutrient deficiency. Disruption of SHMT1 boosted the Gly flux out of the photorespiratory cycle, which increased the levels of the one-carbon (1C) metabolite 5,10-methylene-tetrahydrofolate and S-adenosylmethionine. Furthermore, disrupting SHMT1 reverted the transcriptional response to N and S deprivation and increased N and S contents in shoots of PPSB-deficient lines. Our work provides genetic evidence of the biological relevance of the Ser-Gly-1C metabolic network in N and S metabolism and in interorgan metabolic homeostasis.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Serina/metabolismo , Glicina/metabolismo , Carbono/metabolismo , Nitrogênio/metabolismo , Arabidopsis/metabolismo , Redes e Vias Metabólicas/genética , Enxofre/metabolismo , Desenvolvimento VegetalRESUMO
Cutin and suberin are lipid polyesters deposited in specific apoplastic compartments. Their fundamental roles in plant biology include controlling the movement of gases, water and solutes, and conferring pathogen resistance. Both cutin and suberin have been shown to be present in the Arabidopsis seed coat where they regulate seed dormancy and longevity. In this study, we use accelerated and natural ageing seed assays, glutathione redox potential measures, optical and transmission electron microscopy and gas chromatography-mass spectrometry to demonstrate that increasing the accumulation of lipid polyesters in the seed coat is the mechanism by which the AtHB25 transcription factor regulates seed permeability and longevity. Chromatin immunoprecipitation during seed maturation revealed that the lipid polyester biosynthetic gene long-chain acyl-CoA synthetase 2 (LACS2) is a direct AtHB25 binding target. Gene transfer of this transcription factor to wheat and tomato demonstrated the importance of apoplastic lipid polyesters for the maintenance of seed viability. Our work establishes AtHB25 as a trans-species regulator of seed longevity and has identified the deposition of apoplastic lipid barriers as a key parameter to improve seed longevity in multiple plant species.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Genes Homeobox , Sementes/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Unlike animals, plants possess diverse L-serine (Ser) biosynthetic pathways. One of them, the Phosphorylated Pathway of Serine Biosynthesis (PPSB) has been recently described as essential for embryo, pollen and root development, and required for ammonium and sulfur assimilation. The first and rate limiting step of PPSB is the reaction catalyzed by the enzyme phosphoglycerate dehydrogenase (PGDH). In Arabidopsis, the PGDH family consists of three genes, PGDH1, PGDH2 and PGDH3. PGDH1 is characterized as being the essential gene of the family. However, the biological significance of PGDH2 and PGDH3 remains unknown. In this manuscript, we have functionally characterized PGDH2 and PGDH3. Phenotypic, metabolomic and gene expression analysis in PGDH single, double and triple mutants indicate that both PGDH2 and PGDH3 are functional, affecting plant metabolism and development. PGDH2 has a stronger effect on plant growth than PGDH3, having a partial redundant role with PGDH1. PGDH3, however, could have additional functions in photosynthetic cells unrelated to Ser biosynthesis.
Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/metabolismo , Fosfoglicerato Desidrogenase/genética , Fosfoglicerato Desidrogenase/metabolismo , Serina/biossíntese , Serina/genética , Vias Biossintéticas , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genes de PlantasRESUMO
Intracellular acid stress inhibits plant growth by unknown mechanisms and it occurs in acidic soils and as consequence of other stresses. In order to identify mechanisms of acid toxicity, we screened activation-tagging lines of Arabidopsis thaliana for tolerance to intracellular acidification induced by organic acids. A dominant mutant, sbt4.13-1D, was isolated twice and shown to over-express subtilase SBT4.13, a protease secreted into endoplasmic reticulum. Activity measurements and immuno-detection indicate that the mutant contains less plasma membrane H+-ATPase (PMA) than wild type, explaining the small size, electrical depolarization and decreased cytosolic pH of the mutant but not organic acid tolerance. Addition of acetic acid to wild-type plantlets induces production of ROS (Reactive Oxygen Species) measured by dichlorodihydrofluorescein diacetate. Acid-induced ROS production is greatly decreased in sbt4.13-1D and atrboh-D,F mutants. The latter is deficient in two major NADPH oxidases (NOXs) and is tolerant to organic acids. These results suggest that intracellular acidification activates NOXs and the resulting oxidative stress is important for inhibition of growth. The inhibition of acid-activated NOXs in the sbt4.13-1D mutant compensates inhibition of PMA to increase acid tolerance.
Assuntos
Germinação , Estresse Oxidativo , Prótons , Subtilisinas/genética , Arabidopsis , Proteínas de Arabidopsis/genética , Mutação , NADPH Oxidases/genética , ATPases Translocadoras de Prótons/genética , ATPases Translocadoras de Prótons/metabolismo , Subtilisinas/metabolismoRESUMO
Permeability is a crucial trait that affects seed longevity and is regulated by different polymers including proanthocyanidins, suberin, cutin and lignin located in the seed coat. By testing mutants in suberin transport and biosynthesis, we demonstrate the importance of this biopolymer to cope with seed deterioration. Transcriptomic analysis of cog1-2D, a gain-of-function mutant with increased seed longevity, revealed the upregulation of several peroxidase genes. Reverse genetics analysing seed longevity uncovered redundancy within the seed coat peroxidase gene family; however, after controlled deterioration treatment, seeds from the prx2 prx25 double and prx2 prx25 prx71 triple mutant plants presented lower germination than wild-type plants. Transmission electron microscopy analysis of the seed coat of these mutants showed a thinner palisade layer, but no changes were observed in proanthocyanidin accumulation or in the cuticle layer. Spectrophotometric quantification of acetyl bromide-soluble lignin components indicated changes in the amount of total polyphenolics derived from suberin and/or lignin in the mutant seeds. Finally, the increased seed coat permeability to tetrazolium salts observed in the prx2 prx25 and prx2 prx25 prx71 mutant lines suggested that the lower permeability of the seed coats caused by altered polyphenolics is likely to be the main reason explaining their reduced seed longevity.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Peroxidases/metabolismo , Sementes/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Parede Celular/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Germinação/genética , Germinação/fisiologia , Lignina , Metabolismo dos Lipídeos , Lipídeos , Lipídeos de Membrana , Mutação , Peroxidases/genética , Proantocianidinas , Sementes/genéticaRESUMO
Although the plant Phosphorylated Pathway of l-Ser Biosynthesis (PPSB) is essential for embryo and pollen development, and for root growth, its metabolic implications have not been fully investigated. A transcriptomics analysis of Arabidopsis (Arabidopsis thaliana) PPSB-deficient mutants at night, when PPSB activity is thought to be more important, suggested interaction with the sulfate assimilation process. Because sulfate assimilation occurs mainly in the light, we also investigated it in PPSB-deficient lines in the day. Key genes in the sulfate starvation response, such as the adenosine 5'phosphosulfate reductase genes, along with sulfate transporters, especially those involved in sulfate translocation in the plant, were induced in the PPSB-deficient lines. However, sulfate content was not reduced in these lines as compared with wild-type plants; besides the glutathione (GSH) steady-state levels in roots of PPSB-deficient lines were even higher than in wild type. This suggested that PPSB deficiency perturbs the sulfate assimilation process between tissues/organs. Alteration of thiol distribution in leaves from different developmental stages, and between aerial parts and roots in plants with reduced PPSB activity, provided evidence supporting this idea. Diminished PPSB activity caused an enhanced flux of 35S into thiol biosynthesis, especially in roots. GSH turnover also accelerated in the PPSB-deficient lines, supporting the notion that not only biosynthesis, but also transport and allocation, of thiols were perturbed in the PPSB mutants. Our results suggest that PPSB is required for sulfide assimilation in specific heterotrophic tissues and that a lack of PPSB activity perturbs sulfur homeostasis between photosynthetic and nonphotosynthetic tissues.
Assuntos
Arabidopsis/metabolismo , Serina/biossíntese , Transdução de Sinais/genética , Enxofre/metabolismo , Arabidopsis/genética , Oxirredução , Fosforilação , TranscriptomaRESUMO
In plants, phosphoglycerate kinase (PGK) converts 1,3-bisphosphoglycerate into 3-phosphoglycerate in glycolysis but also participates in the reverse reaction in gluconeogenesis and the Calvin-Benson cycle. In the databases, we found three genes that encode putative PGKs. Arabidopsis (Arabidopsis thaliana) PGK1 was localized exclusively in the chloroplasts of photosynthetic tissues, while PGK2 was expressed in the chloroplast/plastid of photosynthetic and nonphotosynthetic cells. PGK3 was expressed ubiquitously in the cytosol of all studied cell types. Measurements of carbohydrate content and photosynthetic activities in PGK mutants and silenced lines corroborated that PGK1 was the photosynthetic isoform, while PGK2 and PGK3 were the plastidial and cytosolic glycolytic isoforms, respectively. The pgk1.1 knockdown mutant displayed reduced growth, lower photosynthetic capacity, and starch content. The pgk3.2 knockout mutant was characterized by reduced growth but higher starch levels than the wild type. The pgk1.1 pgk3.2 double mutant was bigger than pgk3.2 and displayed an intermediate phenotype between the two single mutants in all measured biochemical and physiological parameters. Expression studies in PGK mutants showed that PGK1 and PGK3 were down-regulated in pgk3.2 and pgk1.1, respectively. These results indicate that the down-regulation of photosynthetic activity could be a plant strategy when glycolysis is impaired to achieve metabolic adjustment and optimize growth. The double mutants of PGK3 and the triose-phosphate transporter (pgk3.2 tpt3) displayed a drastic growth phenotype, but they were viable. This implies that other enzymes or nonspecific chloroplast transporters could provide 3-phosphoglycerate to the cytosol. Our results highlight both the complexity and the plasticity of the plant primary metabolic network.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Fosfoglicerato Quinase/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Citosol/metabolismo , Regulação da Expressão Gênica de Plantas , Ácidos Glicéricos/metabolismo , Metabolômica/métodos , Família Multigênica , Mutação , Fosfoglicerato Quinase/genética , Componentes Aéreos da Planta/genética , Componentes Aéreos da Planta/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Plásticos/metabolismoRESUMO
The presence of two glycolytic pathways working in parallel in plastids and cytosol has complicated the understanding of this essential process in plant cells, especially the integration of the plastidial pathway into the metabolism of heterotrophic and autotrophic organs. It is assumed that this integration is achieved by transport systems, which exchange glycolytic intermediates across plastidial membranes. However, it is unknown whether plastidial and cytosolic pools of 3-phosphoglycerate (3-PGA) can equilibrate in non-photosynthetic tissues. To resolve this question, we employed Arabidopsis mutants of the plastidial glycolytic isoforms of glyceraldehyde-3-phosphate dehydrogenase (GAPCp) that express the triose phosphate translocator (TPT) under the control of the 35S (35S:TPT) or the native GAPCp1 (GAPCp1:TPT) promoters. TPT expression under the control of both promoters complemented the vegetative developmental defects and metabolic disorders of the GAPCp double mutants (gapcp1gapcp2). However, as the 35S is poorly expressed in the tapetum, full vegetative and reproductive complementation of gapcp1gapcp2 was achieved only by transforming this mutant with the GAPCp1:TPT construct. Our results indicate that the main function of GAPCp is to supply 3-PGA for anabolic pathways in plastids of heterotrophic cells and suggest that the plastidial glycolysis may contribute to fatty acid biosynthesis in seeds. They also suggest a 3-PGA deficiency in the plastids of gapcp1gapcp2, and that 3-PGA pools between cytosol and plastid do not equilibrate in heterotrophic cells.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Plastídeos/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Gliceraldeído-3-Fosfato Desidrogenases/genética , Ácidos Glicéricos/metabolismo , Glicólise/genética , Glicólise/fisiologia , Plastídeos/genéticaRESUMO
Light is a major regulator of plant growth and development by antagonizing gibberellins (GA), and we provide evidence for a role of light perception and GA in seed coat formation and seed tolerance to deterioration. We have identified two activation-tagging mutants of Arabidopsis thaliana, cog1-2D and cdf4-1D, with improved seed tolerance to deterioration linked to increased expression of COG1/DOF1.5 and CDF4/DOF2.3, respectively. These encode two homologous DOF transcription factors, with COG1 most highly expressed in seeds. Improved tolerance to seed deterioration was reproduced in transgenic plants overexpressing these genes, and loss of function from RNA interference resulted in opposite phenotypes. Overexpressions of COG1 and CDF4 have been described to attenuate various light responses mediated by phytochromes. Accordingly, we found that phyA and phyB mutants exhibit increased seed tolerance to deterioration. The phenotype of tolerance to deterioration conferred by gain of function of COG1 and by loss of function of phytochromes is of maternal origin, is also observed under natural aging conditions and correlates with a seed coat with increased suberin and reduced permeability. In developing siliques of the cog1-2D mutant the expression of the GA biosynthetic gene GA3OX3 and levels of GA1 are higher than in the wild type. These results explain the antagonism between phytochromes and COG1 in terms of the inhibition and the activation, respectively, of GA action.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Giberelinas/metabolismo , Sementes/fisiologia , Fatores de Transcrição/metabolismo , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Luz , Lipídeos/genética , Mutação , Fitocromo/genética , Fitocromo/metabolismo , Plantas Geneticamente Modificadas , Fatores de Transcrição/genéticaRESUMO
The cellular compartmentalization of metabolic processes is an important feature in plants where the same pathways could be simultaneously active in different compartments. Plant glycolysis occurs in the cytosol and plastids of green and non-green cells in which the requirements of energy and precursors may be completely different. Because of this, the relevance of plastidial glycolysis could be very different depending on the cell type. In the associated study, we investigated the function of plastidial glycolysis in photosynthetic and heterotrophic cells by specifically driving the expression of plastidial glyceraldehyde-3-phosphate dehydrogenase (GAPCp) in a glyceraldehyde-3-phosphate dehydrogenase double mutant background (gapcp1gapcp2). We showed that GAPCp is not functionally significant in photosynthetic cells, while it plays a crucial function in heterotrophic cells. We also showed that (i) GAPCp activity expression in root tips is necessary for primary root growth, (ii) its expression in heterotrophic cells of aerial parts and roots is necessary for plant growth and development, and (iii) GAPCp is an important metabolic connector of carbon and nitrogen metabolism through the phosphorylated pathway of serine biosynthesis (PPSB). We discuss here the role that this pathway could play in the control of plant growth and development.
Assuntos
Arabidopsis/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/fisiologia , Glicólise , Plastídeos/metabolismo , Arabidopsis/citologia , Arabidopsis/fisiologia , Carbono/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/genética , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Mutação , Nitrogênio/metabolismo , Fosforilação , Fotossíntese , Raízes de Plantas/citologia , Raízes de Plantas/metabolismo , Raízes de Plantas/fisiologia , Serina/biossínteseRESUMO
This study functionally characterizes the Arabidopsis (Arabidopsis thaliana) plastidial glycolytic isoforms of glyceraldehyde-3-phosphate dehydrogenase (GAPCp) in photosynthetic and heterotrophic cells. We expressed the enzyme in gapcp double mutants (gapcp1gapcp2) under the control of photosynthetic (Rubisco small subunit RBCS2B [RBCS]) or heterotrophic (phosphate transporter PHT1.2 [PHT]) cell-specific promoters. Expression of GAPCp1 under the control of RBCS in gapcp1gapcp2 had no significant effect on the metabolite profile or growth in the aerial part (AP). GAPCp1 expression under the control of the PHT promoter clearly affected Arabidopsis development by increasing the number of lateral roots and having a major effect on AP growth and metabolite profile. Our results indicate that GAPCp1 is not functionally important in photosynthetic cells but plays a fundamental role in roots and in heterotrophic cells of the AP. Specifically, GAPCp activity may be required in root meristems and the root cap for normal primary root growth. Transcriptomic and metabolomic analyses indicate that the lack of GAPCp activity affects nitrogen and carbon metabolism as well as mineral nutrition and that glycerate and glutamine are the main metabolites responding to GAPCp activity. Thus, GAPCp could be an important metabolic connector of glycolysis with other pathways, such as the phosphorylated pathway of serine biosynthesis, the ammonium assimilation pathway, or the metabolism of γ-aminobutyrate, which in turn affect plant development.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Carbono/metabolismo , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/metabolismo , Nitrogênio/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Clonagem Molecular , Regulação Enzimológica da Expressão Gênica/fisiologia , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Isoenzimas , Regiões Promotoras GenéticasRESUMO
MAIN CONCLUSION: A fungal gene encoding a transcription factor is expressed from its own promoter in Arabidopsis phloem and improves drought tolerance by reducing transpiration and increasing osmotic potential. Horizontal gene transfer from unrelated organisms has occurred in the course of plant evolution, suggesting that some foreign genes may be useful to plants. The CtHSR1 gene, previously isolated from the halophytic yeast Candida tropicalis, encodes a heat-shock transcription factor-related protein. CtHSR1, with expression driven by its own promoter or by the Arabidopsis UBQ10 promoter, was introduced into the model plant Arabidopsis thaliana by Agrobacterium tumefaciens-mediated transformation and the resulting transgenic plants were more tolerant to drought than controls. Fusions of the CtHSR1 promoter with ß-glucuronidase reporter gene indicated that this fungal promoter drives expression to phloem tissues. A chimera of CtHSR1 and green fluorescence protein is localized at the cell nucleus. The physiological mechanism of drought tolerance in transgenic plants is based on reduced transpiration (which correlates with decreased opening of stomata and increased levels of jasmonic acid) and increased osmotic potential (which correlates with increased proline accumulation). Transcriptomic analysis indicates that the CtHSR1 transgenic plants overexpressed a hundred of genes, including many relevant to stress defense such as LOX4 (involved in jasmonic acid synthesis) and P5CS1 (involved in proline biosynthesis). The promoters of the induced genes were enriched in upstream activating sequences for water stress induction. These results demonstrate that genes from unrelated organisms can have functional expression in plants from its own promoter and expand the possibilities of useful transgenes for plant biotechnology.
Assuntos
Adaptação Fisiológica , Arabidopsis/fisiologia , Candida/genética , Secas , Proteínas Fúngicas/genética , Regiões Promotoras Genéticas , Fatores de Transcrição/genética , Arabidopsis/genética , Núcleo Celular/metabolismo , Proteínas Fúngicas/metabolismo , Regulação da Expressão Gênica de Plantas , Genes Fúngicos , Proteínas de Fluorescência Verde/metabolismo , Motivos de Nucleotídeos/genética , Floema/genética , Fotossíntese , Estômatos de Plantas/fisiologia , Plantas Geneticamente Modificadas , Prolina/metabolismo , Nicotiana/metabolismo , Fatores de Transcrição/metabolismo , Transcriptoma/genéticaRESUMO
Membrane-delimited events play a crucial role for ABA signaling and PYR/PYL/RCAR ABA receptors, clade A PP2Cs and SnRK2/CPK kinases modulate the activity of different plasma membrane components involved in ABA action. Therefore, the turnover of PYR/PYL/RCARs in the proximity of plasma membrane might be a step that affects receptor function and downstream signaling. In this study we describe a single-subunit RING-type E3 ubiquitin ligase RSL1 that interacts with the PYL4 and PYR1 ABA receptors at the plasma membrane. Overexpression of RSL1 reduces ABA sensitivity and rsl1 RNAi lines that impair expression of several members of the RSL1/RFA gene family show enhanced sensitivity to ABA. RSL1 bears a C-terminal transmembrane domain that targets the E3 ligase to plasma membrane. Accordingly, bimolecular fluorescent complementation (BiFC) studies showed the RSL1-PYL4 and RSL1-PYR1 interaction is localized to plasma membrane. RSL1 promoted PYL4 and PYR1 degradation in vivo and mediated in vitro ubiquitylation of the receptors. Taken together, these results suggest ubiquitylation of ABA receptors at plasma membrane is a process that might affect their function via effect on their half-life, protein interactions or trafficking.
Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Receptores de Superfície Celular/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Membrana Celular/metabolismo , Regulação da Expressão Gênica de Plantas , Meia-Vida , Proteínas de Membrana Transportadoras/genética , Receptores de Superfície Celular/genética , Transdução de Sinais , Ubiquitina-Proteína Ligases/genética , UbiquitinaçãoRESUMO
Spike lavender (Lavandula latifolia) is an economically important aromatic plant producing essential oils, whose components (mostly monoterpenes) are mainly synthesized through the plastidial methylerythritol 4-phosphate (MEP) pathway. 1-Deoxy-D-xylulose-5-phosphate (DXP) synthase (DXS), that catalyzes the first step of the MEP pathway, plays a crucial role in monoterpene precursors biosynthesis in spike lavender. To date, however, it is not known whether the DXP reductoisomerase (DXR), that catalyzes the conversion of DXP into MEP, is also a rate-limiting enzyme for the biosynthesis of monoterpenes in spike lavender. To investigate it, we generated transgenic spike lavender plants constitutively expressing the Arabidopsis thaliana DXR gene. Although two out of the seven transgenic T0 plants analyzed accumulated more essential oils than the controls, this is hardly imputable to the DXR transgene effect since a clear correlation between transcript accumulation and monoterpene production could not be established. Furthermore, these increased essential oil phenotypes were not maintained in their respective T1 progenies. Similar results were obtained when total chlorophyll and carotenoid content in both T0 transgenic plants and their progenies were analyzed. Our results then demonstrate that DXR enzyme does not play a crucial role in the synthesis of plastidial monoterpene precursors, suggesting that the control flux of the MEP pathway in spike lavender is primarily exerted by the DXS enzyme.
Assuntos
Aldose-Cetose Isomerases/metabolismo , Lavandula/enzimologia , Óleos Voláteis/metabolismo , Óleos de Plantas/metabolismo , Transferases/metabolismo , Aldose-Cetose Isomerases/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Carotenoides/metabolismo , Clorofila/metabolismo , Eritritol/análogos & derivados , Eritritol/metabolismo , Flores/química , Flores/enzimologia , Flores/genética , Expressão Gênica , Lavandula/química , Lavandula/genética , Monoterpenos/metabolismo , Fenótipo , Folhas de Planta/química , Folhas de Planta/enzimologia , Folhas de Planta/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Fosfatos Açúcares/metabolismo , Transferases/genéticaRESUMO
Serine (Ser) has a fundamental role in metabolism and signaling in living organisms. In plants, the existence of different pathways of Ser biosynthesis has complicated our understanding of this amino acid homeostasis. The photorespiratory glycolate pathway has been considered to be of major importance, whereas the nonphotorespiratory phosphorylated pathway has been relatively neglected. Recent advances indicate that the phosphorylated pathway has an important function in plant metabolism and development. Plants deficient in this pathway display developmental defects in embryos, male gametophytes, and roots. We propose that the phosphorylated pathway is more important than was initially thought because it is the only Ser source for specific cell types involved in developmental events. Here, we discuss its importance as a link between metabolism and development in plants.
Assuntos
Regulação da Expressão Gênica de Plantas/fisiologia , Plantas/metabolismo , Serina/biossíntese , Glicólise/fisiologia , Fosforilação , Fotossíntese/fisiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas , Sementes/metabolismo , Serina/metabolismo , Estresse FisiológicoRESUMO
Transgenic Lavandula latifolia plants overexpressing the linalool synthase (LIS) gene from Clarkia breweri, encoding the LIS enzyme that catalyzes the synthesis of linalool were generated. Most of these plants increased significantly their linalool content as compared to controls, especially in the youngest leaves, where a linalool increase up to a 1000% was observed. The phenotype of increased linalool content observed in young leaves was maintained in those T1 progenies that inherit the LIS transgene, although this phenotype was less evident in the flower essential oil. Cross-pollination of transgenic spike lavender plants allowed the generation of double transgenic plants containing the DXS (1-deoxy-d-xylulose-5-P synthase), coding for the first enzyme of the methyl-d-erythritol-4-phosphate pathway, and LIS genes. Both essential oil yield and linalool content in double DXS-LIS transgenic plants were lower than that of their parentals, which could be due to co-suppression effects linked to the structures of the constructs used.
Assuntos
Lavandula , Monoterpenos/metabolismo , Folhas de Planta , Plantas Geneticamente Modificadas , Monoterpenos Acíclicos , Clarkia/enzimologia , Clarkia/genética , Eritritol/análogos & derivados , Eritritol/genética , Eritritol/metabolismo , Hidroliases/biossíntese , Hidroliases/genética , Lavandula/genética , Lavandula/metabolismo , Folhas de Planta/genética , Folhas de Planta/metabolismo , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Fosfatos Açúcares/genética , Fosfatos Açúcares/metabolismo , TransgenesRESUMO
Seed longevity is important to preserve crops and wild plants and it is limited by progressive cellular damage (aging) during storage. The induction of cellular stress defenses and the formation of the seed coat are crucial protecting events during seed development, a process mediated in Arabidopsis thaliana by the transcription factors LEC1, LEC2, FUS3 and the abscisic acid-activated ABI3. In order to identify novel determinants of seed longevity we have screened an activation-tagging mutant collection of Arabidopsis and isolated a dominant mutant with increased seed longevity under both natural and accelerated aging conditions. Molecular characterization indicates that the mutant phenotype is caused by over-expression of the At2g26130 gene encoding a RING-type zinc finger putative ubiquitin ligase. Loss of function of this gene in a T-DNA insertion mutant resulted in decreased seed longevity. We named this important gene for seed longevity RSL1 (from Ring finger of Seed Longevity1) and we could demonstrate ubiquitin ligase activity with the recombinant protein. Morphological alterations in shoot tissues of the RSL1 over-expressing plants and analysis of gibberellins levels suggest that RSL1 may increase gibberellins responses by some unknown mechanism. These results validate the forward genetic approach to seed longevity and anticipate the identification of many novel determinants of this important trait.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Senescência Celular/genética , Genes de Plantas , Domínios RING Finger/genética , Sementes/fisiologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina/genética , Ácido Abscísico/metabolismo , Arabidopsis/enzimologia , Arabidopsis/fisiologia , Proteínas de Arabidopsis/metabolismo , DNA Bacteriano , Expressão Gênica , Giberelinas/genética , Giberelinas/metabolismo , Humanos , Longevidade , Mutagênese Insercional , Fenótipo , Brotos de Planta/metabolismo , Sementes/enzimologia , Sementes/metabolismo , Estresse Fisiológico/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Seed longevity is crucial for agriculture and plant genetic diversity, but it is limited by cellular damage during storage. Seeds are protected against aging by cellular defenses and by structures such as the seed coat. We have screened an activation-tagging mutant collection of Arabidopsis (Arabidopsis thaliana) and selected four dominant mutants with improved seed longevity (isl1-1D to isl4-1D) under both natural and accelerated aging conditions. In the isl1-1D mutant, characterized in this work, overexpression of the transcription factor ARABIDOPSIS THALIANA HOMEOBOX25 (ATHB25; At5g65410) increases the expression of GIBBERELLIC ACID3-OXIDASE2, encoding a gibberellin (GA) biosynthetic enzyme, and the levels of GA1 and GA4 are higher (3.2- and 1.4-fold, respectively) in the mutant than in the wild type. The morphological and seed longevity phenotypes of the athb25-1D mutant were recapitulated in transgenic plants with moderate (4- to 6-fold) overexpression of ATHB25. Simultaneous knockdown of ATHB25, ATHB22, and ATHB31 expression decreases seed longevity, as does loss of ATHB25 and ATHB22 function in a double mutant line. Seeds from wild-type plants treated with GA and from a quintuple DELLA mutant (with constitutive GA signaling) are more tolerant to aging, providing additional evidence for a role of GA in seed longevity. A correlation was observed in several genotypes between seed longevity and mucilage formation at the seed surface, suggesting that GA may act by reinforcing the seed coat. This mechanism was supported by the observation of a maternal effect in reciprocal crosses between the wild type and the athb25-1D mutant.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Giberelinas/farmacologia , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas/genética , Genótipo , Germinação/efeitos dos fármacos , Mutação/genética , Motivos de Nucleotídeos/genética , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Mucilagem Vegetal/metabolismo , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas/genética , Sementes/efeitos dos fármacos , Coloração e Rotulagem , Triazóis/farmacologiaRESUMO
In plants, 3 different pathways of serine biosynthesis have been described: the Glycolate pathway, which is associated with photorespiration, and 2 non-photorespiratory pathways, the Glycerate and the Phosphorylated pathways. The Phosphorylated Pathway of Serine Biosynthesis (PPSB) has been known since the 1950s, but has been studied relatively little, probably because it was considered of minor significance as compared with the Glycolate pathway. In the associated study (1), we described for the first time in plants the in vivo functional characterization of the PPSB, by targeting the phosphoserine phosphatase (PSP1), the last enzyme of the pathway. Following a gain- and loss-of-function approach in Arabidopsis, we provided genetic and molecular evidence for the essential role of PSP1 for embryo and pollen development, and for proper root growth. A metabolomics study indicated that the PPSB affects glycolysis, the Krebs cycle, and the biosynthesis of several amino acids, which suggests that this pathway is an important link connecting metabolism and development. The mechanisms underlying the essential functions of PSP1 are discussed.
Assuntos
Arabidopsis/metabolismo , Vias Biossintéticas , Serina/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Fluorescência Verde/metabolismo , Mutação/genética , Monoéster Fosfórico Hidrolases/genética , FosforilaçãoRESUMO
Three different pathways of serine (Ser) biosynthesis have been described in plants: the Glycolate pathway, which is part of the Photorespiratory pathway, and 2 non-Photorespiratory pathways, the Glycerate and the Phosphorylated pathways. The Phosphorylated Pathway of Ser Biosynthesis (PPSB) has been known to exist since the 1950s, but its biological relevance was not revealed until quite recently when the last enzyme of the pathway, the Phosphoserine Phosphatase, was functionally characterized. In the associated study (1), we characterized a family of genes coding for putatite phosphoglycerate dehydrogenases (PGDH, 3-PGDH, and EDA9), the first enzyme of the PPSB. A metabolomics study using overexpressing plants indicated that all PGDH family genes were able to regulate Ser homeostasis but only lacking of EDA9 expression caused drastic developmental defects. We provided genetic and molecular evidence for the essential role of EDA9 for embryo and pollen development. Here, some new insights into the physiological/molecular function of PPSB and Ser are presented and discussed.