RESUMEN
MYB-bHLH-TTG1 (MBW) transcription factor (TF) complexes regulate Arabidopsis seed coat biosynthesis pathways via a multi-tiered regulatory mechanism. The MYB genes include MYB5, MYB23 and TRANSPARENT TESTA2 (TT2), which regulate GLABRA2 (GL2), HOMEODOMAIN GLABROUS2 (HDG2) and TRANSPARENT TESTA GLABRA2 (TTG2). Here, we examine the role of PECTIN METHYLESTERASE INHIBITOR14 (PMEI14) in seed coat mucilage pectin methylesterification and provide evidence in support of multi-tiered regulation of seed coat mucilage biosynthesis genes including PMEI14. The PMEI14 promoter was active in the seed coat and developing embryo. A pmei14 mutant exhibited stronger attachment of the outer layer of seed coat mucilage, increased mucilage homogalacturonan demethylesterification and reduced seed coat radial cell wall thickness, results consistent with decreased PMEI activity giving rise to increased PME activity. Reduced mucilage release from the seeds of myb5, myb23, tt2 and gl2, hdg2, ttg2 triple mutants indicated that HDG2 and MYB23 play minor roles in seed coat mucilage deposition. Chromatin immunoprecipitation analysis found that MYB5, TT8 and seven mucilage pathway structural genes are directly regulated by MYB5. Expression levels of GL2, HDG2, TTG2 and nine mucilage biosynthesis genes including PMEI14 in the combinatorial mutant seeds indicated that these genes are positively regulated by at least two of those six TFs and that TTG1 and TTG2 are major regulators of PMEI14 expression. Our results show that MYB-bHLH-TTG1 complexes regulate mucilage biosynthesis genes, including PMEI14, both directly and indirectly via a three-tiered mechanism involving GL2, HDG2 and TTG2.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Mucílago de Planta , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Mutación , Pectinas/metabolismo , Proteínas de Unión al ADN/metabolismo , Semillas/genética , Semillas/metabolismo , Regulación de la Expresión Génica de las Plantas , Mucílago de Planta/metabolismoRESUMEN
The Arabidopsis seed coat mucilage is a polysaccharide-rich matrix synthesized by the seed coat epidermal cells. It is a specialized cell wall mainly composed of three types of polysaccharides (i. e. pectin, hemicellulose, and cellulose), and represents as an ideal model system for plant cell wall research. A large number of genes responsible for the synthesis and modification of cell wall polysaccharides have been identified using this model system. Moreover, a subset of regulators controlling mucilage production and modification have been characterized, and the underlying transcriptional regulatory mechanisms have been elucidated. This substantially contributes to the understanding of the molecular mechanisms underlying mucilage synthesis and modification. In this review, we concisely summarize the various genes and regulators involved in seed coat cell differentiation, mucilage biosynthesis and modification, and secondary cell wall formation. In particular, we put emphasis on the latest knowledge gained regarding the transcriptional regulation of mucilage production, which is composed of a hierarchal cascade with three-layer transcriptional regulators. Collectively, we propose an updated schematic framework of the genetic regulatory network controlling mucilage production and modification in the Arabidopsis mucilage secretory cells.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Mucílago de Planta , Arabidopsis/metabolismo , Mucílago de Planta/metabolismo , Redes Reguladoras de Genes , Polisacáridos/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pectinas/metabolismo , Semillas/genética , Semillas/metabolismo , Pared Celular/metabolismo , Regulación de la Expresión Génica de las PlantasAsunto(s)
Proteínas de Arabidopsis , Mucílago de Planta , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Pectinas , Mucílago de Planta/genética , Mucílago de Planta/metabolismo , Semillas/genética , Semillas/metabolismoRESUMEN
The mucilage surrounding hydrated Arabidopsis thaliana seeds is a specialized extracellular matrix composed mainly of the pectic polysaccharide rhamnogalacturonan I (RG-I). Although, several genes responsible for RG-I biosynthesis have been identified, the transcriptional regulatory mechanisms controlling RG-I production remain largely unknown. Here we report that the trihelix transcription factor DE1 BINDING FACTOR 1 (DF1) is a key regulator of mucilage RG-I biosynthesis. RG-I biosynthesis is significantly reduced in loss-of-function mutants of DF1. DF1 physically interacts with GLABRA2 (GL2) and both proteins transcriptionally regulate the expression of the RG-I biosynthesis genes MUCILAGE MODIFIED 4 (MUM4) and GALACTURONOSYLTRANSFERASE-LIKE5 (GATL5). Through chromatin immunoprecipitation-quantitative PCR and transcriptional activation assays, we uncover a cooperative mechanism of the DF1-GL2 module in activating MUM4 and GATL5 expression, in which DF1 binds to the promoters of MUM4 and GATL5 through interacting with GL2 and facilitates the transcriptional activity of GL2. The expression of DF1 and GL2 is directly regulated by TRANSPARENT TESTA GLABRA2 (TTG2) and, in turn, DF1 directly represses the expression of TTG2. Taken together, our data reveal that the transcriptional regulation of mucilage RG-I biosynthesis involves a regulatory module, comprising DF1, GL2, and TTG2.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Mucílago de Planta , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Pectinas , Mucílago de Planta/metabolismo , Polisacáridos/metabolismo , Semillas/genética , Semillas/metabolismoRESUMEN
Pectin is one of the major components of plant primary cell wall polysaccharides. The degree of pectin methylesterification (DM) plays an important role in the process of plant growth. However, little is known about the underlying regulatory mechanisms during the process of pectin demethylesterification. Here, we characterized mucilage defect 1 (mud1), a novel Arabidopsis thaliana mutant, which displays increased mucilage adherence resulting from increased activities of pectin methylesterases (PMEs) and decreased degree of pectin methylesterification (DM). MUD1 encodes a nuclear protein with a Really Interesting New Gene (RING)-v domain and is highly expressed in developing seed coat when seed coat mucilage starts to accumulate. We have demonstrated that MUD1 has E3 ubiquitin ligase activity in vitro. The expression of PME-related genes, including MYB52, LUH, SBT1.7, PMEI6, and PMEI14 decreased considerably in mud1. We propose that MUD1 acts as an ubiquitin ligase potentially regulating the DM of pectin by post-transcriptionally removing proteins that normally negatively regulate the level or activity of PMEs in the seed coat mucilage.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Mucílago de Planta , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pared Celular/metabolismo , Regulación de la Expresión Génica de las Plantas , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Semillas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Flax (Linum usitatissimum L.) seed oil, which accumulates in the embryo, and mucilage, which is synthesized in the seed coat, are of great economic importance for food, pharmaceutical as well as chemical industries. Theories on the link between oil and mucilage production in seeds consist in the spatio-temporal competition of both compounds for photosynthates during the very early stages of seed development. In this study, we demonstrate a positive relationship between seed oil production and seed coat mucilage extrusion in the agronomic model, flax. Three recombinant inbred lines were selected for low, medium and high mucilage and seed oil contents. Metabolite and transcript profiling (1H NMR and DNA oligo-microarrays) was performed on the seeds during seed development. These analyses showed main changes in the seed coat transcriptome during the mid-phase of seed development (25 Days Post-Anthesis), once the mucilage biosynthesis and modification processes are thought to be finished. These transcriptome changes comprised genes that are putatively involved in mucilage chemical modification and oil synthesis, as well as gibberellic acid (GA) metabolism. The results of this integrative biology approach suggest that transcriptional regulations of seed oil and fatty acid (FA) metabolism could occur in the seed coat during the mid-stage of seed development, once the seed coat carbon supplies have been used for mucilage biosynthesis and mechanochemical properties of the mucilage secretory cells.
Asunto(s)
Lino/crecimiento & desarrollo , Lino/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Mucílago de Planta/metabolismo , Semillas/crecimiento & desarrollo , Semillas/genética , Transcripción Genética , Pared Celular/metabolismo , Endospermo/metabolismo , Ácidos Grasos/metabolismo , Lino/ultraestructura , Giberelinas/metabolismo , Glucosa/metabolismo , Endogamia , Cinética , Metabolómica , Fenotipo , Mucílago de Planta/ultraestructura , Aceites de Plantas/metabolismo , Análisis de Componente Principal , Recombinación Genética/genética , Semillas/ultraestructura , Almidón/metabolismo , Sacarosa/metabolismo , Transcriptoma/genéticaRESUMEN
The cell wall is essential for plant survival. Determining the relationship between cell wall structure and function using mutant analysis or overexpressing cell wall-modifying enzymes has been challenging due to the complexity of the cell wall and the appearance of secondary, compensatory effects when individual polymers are modified. In addition, viability of the plants can be severely impacted by wall modification. A useful model system for studying structure-function relationships among extracellular matrix components is the seed coat epidermal cells of Arabidopsis thaliana. These cells synthesize relatively simple, easily accessible, pectin-rich mucilage that is not essential for plant viability. In this study, we expressed enzymes predicted to modify polysaccharide components of mucilage in the apoplast of seed coat epidermal cells and explored their impacts on mucilage. The seed coat epidermal-specific promoter TESTA ABUNDANT2 (TBA2) was used to drive expression of these enzymes to avoid adverse effects in other parts of the plant. Mature transgenic seeds expressing Rhamnogalacturonate lyase A (RglA) or Rhamnogalacturonate lyase B (RglB) that degrade the pectin rhamnogalacturonan-I (RG-I), a major component of mucilage, had greatly reduced mucilage capsules surrounding the seeds and concomitant decreases in the monosaccharides that comprise the RG-I backbone. Degradation of the minor mucilage component homogalacturonan (HG) using the HG-degrading enzymes Pectin lyase A (PLA) or ARABIDOPSIS DEHISCENCE ZONE POLYGALACTURONASE2 (ADPG2) resulted in developing seed coat epidermal cells with disrupted cell-cell adhesion and signs of early cell death. These results demonstrate the feasibility of manipulating the seed coat epidermal cell extracellular matrix using a targeted genetic engineering approach.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Arabidopsis/enzimología , Proteínas de Arabidopsis/metabolismo , Semillas/químicaRESUMEN
Homogalacturonan (HG), a component of pectin, is synthesized in the Golgi apparatus in its fully methylesterified form. It is then secreted into the apoplast where it is typically de-methylesterified by pectin methylesterases (PME). Secretion and de-esterification are critical for normal pectin function, yet the underlying transcriptional regulation mechanisms remain largely unknown. Here, we uncovered a mechanism that fine-tunes the degree of HG de-methylesterification (DM) in the mucilage that surrounds Arabidopsis thaliana seeds. We demonstrate that the APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) transcription factor (TF) ERF4 is a transcriptional repressor that positively regulates HG DM. ERF4 expression is confined to epidermal cells in the early stages of seed coat development. The adhesiveness of the erf4 mutant mucilage was decreased as a result of an increased DM caused by a decrease in PME activity. Molecular and genetic analyses revealed that ERF4 positively regulates HG DM by suppressing the expression of three PME INHIBITOR genes (PMEIs) and SUBTILISIN-LIKE SERINE PROTEASE 1.7 (SBT1.7). ERF4 shares common targets with the TF MYB52, which also regulates pectin DM. Nevertheless, the erf4-2 myb52 double mutant seeds have a wild-type mucilage phenotype. We provide evidence that ERF4 and MYB52 regulate downstream gene expression in an opposite manner by antagonizing each other's DNA-binding ability through a physical interaction. Together, our findings reveal that pectin DM in the seed coat is fine-tuned by an ERF4-MYB52 transcriptional complex.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de la Membrana/metabolismo , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Proteínas Represoras/metabolismo , Semillas/metabolismo , Factores Generales de Transcripción/metabolismo , Adhesividad , Arabidopsis/embriología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Calcio/metabolismo , Hidrolasas de Éster Carboxílico/metabolismo , Reactivos de Enlaces Cruzados/química , Esterificación , Genes de Plantas , Mutación/genética , Motivos de Nucleótidos/genética , Fenotipo , Epidermis de la Planta/citología , Epidermis de la Planta/metabolismo , Unión Proteica , Proteínas Represoras/genéticaRESUMEN
Basil seeds are widely cultivated throughout the world because of their extensive applications in various fields of life. The Basil seeds mucilage (BSM) exhibits remarkable physical and chemical properties like high water absorbing capacity, emulsifying, and stabilizing properties. The extraction of this mucilage from the seed surface has always been done by physical and chemical methods, which has certain drawbacks. Here, we report for the first time a chemical method for the effective extraction of this mucilage using ionic liquids (ILs); the green solvents. Pyridinium chloride based ILs were investigated for the effective extraction of mucilage and the process was optimized for various variables i.e. time, temperature, basil seed loading, co-solvents, anti-solvents. The extraction yield (up to 25% w/w of mucilage per basil seeds dry weight) was obtained at optimum conditions. Extracted mucilage was characterized by analytical techniques. The extracted BSM was used to prepare AuNps/BSM nanocomposite by stabilizing the gold nanoparticles. The AuNps/BSM nanocomposite was applied for the catalytic degradation of dyes (congo red; 12 min, methyl orange; 4 min, whereas 4-nitrophenol; 6 min).
Asunto(s)
Nanocompuestos/química , Ocimum basilicum/química , Mucílago de Planta/aislamiento & purificación , Catálisis , Colorantes , Emulsiones/química , Oro , Líquidos Iónicos/química , Nanopartículas del Metal/química , Extractos Vegetales/química , Mucílago de Planta/metabolismo , Polisacáridos/análisis , Semillas/química , Temperatura , Agua/químicaRESUMEN
Following exposure to water, mature Arabidopsis seeds are surrounded by a gelatinous capsule, termed mucilage. The mucilage consists of pectin-rich polysaccharides, which are produced in epidermal cells of the seed coat. Although pectin is a major component of plant cell walls, its biosynthesis and biological functions are not fully understood. Previously, we reported that a transmembrane RING E3 ubiquitin ligase, FLYING SAUCER 1 (FLY1) regulates the degree of pectin methyl esterification for mucilage capsule formation. The Arabidopsis thaliana genome has a single FLY1 homolog, FLY2. In this study, we show that the FLY2 protein functions in mucilage modification together with FLY1. FLY2 was expressed in seed coat epidermal cells during mucilage synthesis, but its expression level was much lower than that of FLY1. While fly2 showed no obvious difference in mucilage capsule formation from wild type, the fly1 fly2 double mutants showed more severe defects in mucilage than fly1 alone. FLY2-EYFP that was expressed under the control of the FLY1 promoter rescued fly1 mucilage, showing that FLY2 has the same molecular function as FLY1. FLY2-EYFP colocalized with marker proteins of Golgi apparatus (sialyltransferase-mRFP) and late endosome (mRFP-ARA7), indicating that as FLY1, FLY2 controls pectin modification by functioning in these endomembrane organelles. Furthermore, phylogenetic analysis suggests that FLY1 and FLY2 originated from a common ancestral gene by gene duplication prior to the emergence of Brassicaceae. Taken together, our findings suggest that FLY2 functions in the Golgi apparatus and/or the late endosome of seed coat epidermal cells in a manner similar to FLY1.
Asunto(s)
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Mucílago de Planta/genética , Mucílago de Planta/metabolismo , Semillas/metabolismo , Arabidopsis/genética , Pared Celular/metabolismo , Endosomas/metabolismo , Células Epidérmicas , Esterificación , Regulación de la Expresión Génica de las Plantas , Aparato de Golgi/metabolismo , Pectinas/metabolismo , Filogenia , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas , Semillas/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Flaxseed protein (FP) and mucilage (FM) complex bioparticles as sustainable ingredients were assembled by electrostatic interaction for plant-based Pickering stabilization of flaxseed oil (FO)-in-water emulsions. The effect of FO content (1-5 wt%) on droplet size and accelerated creaming stability of the emulsions was evaluated, from which it was found that 2.5 wt% FO emulsion had the smallest initial droplet size (i.e. D[4,3] = 8 µm) and creaming velocity (2.9 µm/s). The microstructure of the emulsions was observed using Cryo-SEM, confocal and optical microscopy, showing a thick layer of the particles on the oil surface responsible for the stabilization. The physical stability of FO emulsions stabilized by complex bioparticles against various environmental stress conditions (pH, salt and temperature) was higher compared to plain FP- and polysorbate 80-stabilized emulsions. Thus, the droplet size of FP-stabilized emulsions (pH 3) increased from 21 to 38 µm after thermal treatment (80 °C), whereas the size distribution of particle-stabilized emulsions hardly changed. The latter emulsions also remained stable during 28 days of storage and displayed good stability against a wide range of pH conditions (2-9) and salt concentrations (0-500 mM) with no sign of oiling-off. The complex particles as Pickering emulsifiers were successful to depress the FO oxidation at 4 °C and 50 °C. This study could open a promising pathway for producing natural and surfactant-free emulsions through Pickering stabilization using plant-based biopolymer particles for protecting lipophilic bioactive components.
Asunto(s)
Emulsiones/química , Lino/química , Aceite de Linaza/química , Mucílago de Planta/química , Proteínas de Plantas/química , Tensoactivos/química , Productos Biológicos/química , Productos Biológicos/metabolismo , Microscopía por Crioelectrón , Estabilidad de Medicamentos , Lino/metabolismo , Concentración de Iones de Hidrógeno , Microscopía Confocal , Oxidación-Reducción , Tamaño de la Partícula , Mucílago de Planta/metabolismo , Proteínas de Plantas/metabolismo , Polisorbatos/química , Tensoactivos/síntesis química , Temperatura , Agua/químicaRESUMEN
The current study aimed to investigate the impacts of different concentrations of GO/PANI nanocomposites (25, 50 and 100â¯mgâ¯L-1), in comparison with GO and PANI, on seed germination behaviors, morpho-physiological and biochemical traits in intact (mucilaginous) and demucilaged seeds, and young seedlings of the medicinal plant Salvia mirzayanii. Upon exposure to GO, seed germination was delayed and reduced, and growth attributes (root and shoot length, shoot fresh weight, and total chlorophyll content) declined, all of which could be attributed to the reductions in water uptake and oxidative stress particularly in demucilaged seeds. A hormetic dose-dependent response was observed for the growth traits in both intact and demucilaged seedlings upon exposure to GO/PANI concentrations, i.e. low-concentration stimulation and high-concentration repression. Elevated levels of H2O2 in shoot tissue of the seedlings exposed to GO and high concentration of GO/PANI, in comparison with those exposed to low levels of GO/PANI and control, were linked with the activities of the antioxidant enzymes SOD, CAT, POD, and total phenolics. Overall, the results showed high toxicity of GO on germination and early growth of S. mirzayani that was more evident in demucilaged seedlings, whereas GO/PANI stimulated germination, and the effects on seedling growth were stimulatory or inhibitory depending on the application dose and presence of mucilage. Furthermore, the capacity of GO/PANI nanocomposites to improve germination and cause a regular porosity pattern in roots accompanied by improved water uptake and early establishment of S. mirzayanii propose potential implications of GO/PANI nanocomposites for seeds/plants in drought-prone ecosystems.
Asunto(s)
Compuestos de Anilina/toxicidad , Antioxidantes/farmacología , Germinación/efectos de los fármacos , Grafito/toxicidad , Salvia/metabolismo , Plantones/crecimiento & desarrollo , Semillas/efectos de los fármacos , Catalasa/metabolismo , Clorofila , Peróxido de Hidrógeno/farmacología , Nanocompuestos , Estrés Oxidativo/efectos de los fármacos , Mucílago de Planta/metabolismo , Superóxido Dismutasa/metabolismo , AguaRESUMEN
KEY MESSAGE: A possible transcription factor TLP2 was identified to be involved in the regulation of HG biosynthesis in Arabidopsis seed mucilage. TLP2 can translocate into nucleus from plasma membrane by interacting with NF-YC3. The discovery of TLP2 gene function can further fulfill the regulatory network of pectin biosynthesis in Arabidopsis thaliana. Arabidopsis seed coat mucilage is an excellent model system to study the biosynthesis, function and regulation of pectin. Rhamnogalacturonan I (RG-I) and homogalacturonan (HG) are the major polysaccharides constituent of the Arabidopsis seed coat mucilage. Here, we identified a Tubby-like gene, Tubby-like protein 2 (TLP2), which was up-regulated in developing siliques when mucilage began to be produced. Ruthenium red (RR) staining of the seeds showed defective mucilage of tlp2-1 mutant after vigorous shaking compared to wild type (WT). Monosaccharide composition analysis revealed that the amount of total sugars and galacturonic acid (GalA) decreased significantly in the adherent mucilage (AM) of tlp2-1 mutant. Immunolabelling and dot immunoblotting analysis showed that unesterified HG decreased in the tlp2-1 mutant. Furthermore, TLP2 can translocate into nucleus by interacting with Nuclear Factor Y subunit C3 (NF-YC3) to function as a transcription factor. RNA-sequence and transactivation assays revealed that TLP2 could activate UDP-glucose 4-epimerase 1 (UGE1). In all, it is concluded that TLP2 could regulate the biosynthesis of HG possibly through the positive activation of UGE1.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Pectinas/biosíntesis , Mucílago de Planta/metabolismo , Semillas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Regulación de la Expresión Génica de las Plantas , Ácidos Hexurónicos , Mutación , Fenotipo , Plantas Modificadas Genéticamente , Polisacáridos , Semillas/crecimiento & desarrollo , Análisis de Secuencia de ARN , Factores de Transcripción , Activación Transcripcional , Uridina Difosfato Glucosa Deshidrogenasa/metabolismoRESUMEN
The differentiation of the seed coat epidermal (SCE) cells in Arabidopsis thaliana leads to the production of a large amount of pectin-rich mucilage and a thick cellulosic secondary cell wall. The mechanisms by which cortical microtubules are involved in the formation of these pectinaceous and cellulosic cell walls are still largely unknown. Using a reverse genetic approach, we found that TONNEAU1 (TON1) recruiting motif 4 (TRM4) is implicated in cortical microtubule organization in SCE cells, and functions as a novel player in the establishment of mucilage structure. TRM4 is preferentially accumulated in the SCE cells at the stage of mucilage biosynthesis. The loss of TRM4 results in compact seed mucilage capsules, aberrant mucilage cellulosic structure, short cellulosic rays and disorganized cellulose microfibrils in mucilage. The defects could be rescued by transgene complementation of trm4 alleles. Probably, this is a consequence of a disrupted organization of cortical microtubules, observed using fluorescently tagged tubulin proteins in trm4 SCE cells. Furthermore, TRM4 proteins co-aligned with microtubules and interacted directly with CELLULOSE SYNTHASE 3 in two independent assays. Together, the results indicate that TRM4 is essential for microtubule array organization and therefore correct cellulose orientation in the SCE cells, as well as the establishment of the subsequent mucilage architecture.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Celulosa/metabolismo , Glucosiltransferasas/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Alelos , Arabidopsis/fisiología , Arabidopsis/ultraestructura , Proteínas de Arabidopsis/genética , Pared Celular/metabolismo , Pared Celular/ultraestructura , Glucosiltransferasas/genética , Proteínas Asociadas a Microtúbulos/genética , Microtúbulos/metabolismo , Microtúbulos/ultraestructura , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Semillas/genética , Semillas/fisiología , Semillas/ultraestructuraRESUMEN
Seedling emergence is a critical stage in the establishment of desert plants. Soil microbes participate in plant growth and development, but information is lacking with regard to the role of microbes on seedling emergence. We applied the biocides (captan and streptomycin) to assess how seed mucilage interacts with soil microbial community and physiochemical processes to affect seedling emergence of Artemisia sphaerocephala on the desert sand dune. Fungal and bacterial community composition and diversity and fungal-bacterial interactions were changed by both captan and streptomycin. Mucilage increased soil enzyme activities and fungal-bacterial interactions. Highest seedling emergence occurred under streptomycin and mucilage treatment. Members of the phyla Firmicutes and Glomeromycota were the keystone species that improved A. sphaerocephala seedling emergence, by increasing resistance of young seedlings to drought and pathogen. Seed mucilage directly improved seedling emergence and indirectly interacted with the soil microbial community through strengthening fungal-bacterial interactions and providing favourable environment for soil enzymes to affect seedling emergence. Our study provides a comprehensive understanding of the regulatory mechanisms by which soil microbial community and seed mucilage interactively promote successful establishment of populations of desert plants on the barren and stressful sand dune.
Asunto(s)
Interacciones Microbiota-Huesped , Mucílago de Planta/fisiología , Plantones/crecimiento & desarrollo , Semillas/fisiología , Microbiología del Suelo , Antiinfecciosos/farmacología , Artemisia/crecimiento & desarrollo , Artemisia/metabolismo , Artemisia/microbiología , Captano/farmacología , Clima Desértico , Secuenciación de Nucleótidos de Alto Rendimiento , Interacciones Microbiota-Huesped/fisiología , Mucílago de Planta/metabolismo , ARN Ribosómico 16S/genética , Plantones/metabolismo , Plantones/microbiología , Semillas/metabolismo , Semillas/microbiología , Estreptomicina/farmacologíaRESUMEN
Pectin is a vital component of the plant cell wall and provides the molecular glue that maintains cell-cell adhesion, among other functions. As the most complex wall polysaccharide, pectin is composed of several covalently linked domains, such as homogalacturonan (HG) and rhamnogalacturonan I (RG I). Pectin has widespread uses in the food industry and has emerging biomedical applications, but its synthesis remains poorly understood. For instance, the enzymes that catalyze RG I elongation remain unknown. Recently, a coexpression- and sequence-based MUCILAGE-RELATED (MUCI) reverse genetic screen uncovered hemicellulose biosynthetic enzymes in the Arabidopsis (Arabidopsis thaliana) seed coat. Here, we use an extension of this strategy to identify MUCI70 as the founding member of a glycosyltransferase family essential for the accumulation of seed mucilage, a gelatinous wall rich in unbranched RG I. Detailed biochemical and histological characterization of two muci70 mutants and two galacturonosyltransferase11 (gaut11) mutants identified MUCI70 and GAUT11 as required for two distinct RG I domains in seed mucilage. We demonstrate that, unlike MUCI70, GAUT11 catalyzes HG elongation in vitro and, thus, likely is required for the synthesis of an HG region important for RG I elongation. Analysis of a muci70 gaut11 double mutant confirmed that MUCI70 and GAUT11 are indispensable for the production and release of the bulk of mucilage RG I and for shaping the surface morphology of seeds. In addition, we uncover relationships between pectin and hemicelluloses and show that xylan is essential for the elongation of at least one RG I domain.
Asunto(s)
Proteínas de Arabidopsis/fisiología , Arabidopsis/enzimología , Glucuronosiltransferasa/metabolismo , Hidrolasas/fisiología , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Semillas/enzimología , Arabidopsis/genética , Arabidopsis/ultraestructura , Proteínas de Arabidopsis/genética , Pared Celular/metabolismo , Pared Celular/ultraestructura , Glucuronosiltransferasa/genética , Glicosiltransferasas/genética , Glicosiltransferasas/metabolismo , Hidrolasas/genética , Microscopía Electrónica de Rastreo , Filogenia , Mucílago de Planta/química , Mucílago de Planta/ultraestructura , Polisacáridos/metabolismo , Semillas/genética , Semillas/ultraestructuraRESUMEN
Pectin, which is a major component of the plant primary cell walls, is synthesized and methyl-esterified in the Golgi apparatus and then demethylesterified by pectin methylesterases (PMEs) located in the cell wall. The degree of methylesterification affects the functional properties of pectin, and thereby influences plant growth, development and defense. However, little is known about the mechanisms that regulate pectin demethylesterification. Here, we show that in Arabidopsis (Arabidopsis thaliana) seed coat mucilage, the absence of the MYB52 transcription factor is correlated with an increase in PME activity and a decrease in the degree of pectin methylesterification. Decreased methylesterification in the myb52 mutant is also correlated with an increase in the calcium content of the seed mucilage. Chromatin immunoprecipitation analysis and molecular genetic studies suggest that MYB52 transcriptionally activates PECTIN METHYLESTERASE INHIBITOR6 (PMEI6), PMEI14, and SUBTILISIN-LIKE SER PROTEASE1.7 (SBT1.7) by binding to their promoters. PMEI6 and SBT1.7 have previously been shown to be involved in seed coat mucilage demethylesterification. Our characterization of two PMEI14 mutants suggests that PMEI14 has a role in seed coat mucilage demethylesterification, although its activity may be confined to the seed coat in contrast to PMEI6, which functions in the whole seed. Our demonstration that MYB52 negatively regulates pectin demethylesterification in seed coat mucilage, and the identification of components of the molecular network involved, provides new insight into the regulatory mechanism controlling pectin demethylesterification and increases our understanding of the transcriptional regulation network involved in seed coat mucilage formation.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Pectinas/metabolismo , Mucílago de Planta/metabolismo , Semillas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Calcio/metabolismo , Hidrolasas de Éster Carboxílico/genética , Hidrolasas de Éster Carboxílico/metabolismo , Pared Celular/enzimología , Pared Celular/genética , Esterificación , Regulación de la Expresión Génica de las Plantas , Redes Reguladoras de Genes , Mutación , Regiones Promotoras Genéticas/genética , Unión Proteica , Semillas/genéticaRESUMEN
A comparative study of mucilage (locular tissue) and pulp polysaccharides from ripe tamarillo fruits (Solanum betaceum Cav.) was carried out. After aqueous and alkaline extractions and various purification steps (freeze-thaw and α-amylase - EC 3.2.1.1 treatments, Fehling precipitation and ultrafiltration through 50 kDa cut-off membrane), the obtained fractions from mucilage were analyzed by sugar composition, HPSEC, and NMR spectroscopy analyses. The results showed that the mucilage of tamarillo contains a highly methoxylated homogalacturonans mixed with type I arabinogalactans, a linear (1 â 5)-linked α-L-arabinan, and a linear (1 â 4)-ß-D-xylan. A comparison with polysaccharides extracted from the pulp revealed that differences were observed in the yield and in the ratio of extracted polysaccharides. Moreover, structural differences between pulp and mucilage polysaccharides were also observed, such as in the length of side chains of the pectins, and in the degree of branching of the xylans.
Asunto(s)
Frutas/metabolismo , Mucílago de Planta/metabolismo , Polisacáridos/metabolismo , Solanum/metabolismo , Espectroscopía de Resonancia Magnética con Carbono-13 , Fraccionamiento Químico , Cromatografía en Gel , Monosacáridos/análisis , Pectinas/aislamiento & purificación , Polisacáridos/aislamiento & purificación , Solubilidad , Agua/químicaRESUMEN
Arabidopsis (Arabidopsis thaliana) seed coat epidermal cells produce large amounts of mucilage that is released upon imbibition. This mucilage is structured into two domains: an outer diffuse layer that can be easily removed by agitation and an inner layer that remains attached to the outer seed coat. Both layers are composed primarily of pectic rhamnogalacturonan I (RG-I), the inner layer also containing rays of cellulose that extend from the top of each columella. Perturbation in cellulosic ray formation has systematically been associated with a redistribution of pectic mucilage from the inner to the outer layer, in agreement with cellulose-pectin interactions, the nature of which remained unknown. Here, by analyzing the outer layer composition of a series of mutant alleles, a tight proportionality of xylose, galacturonic acid, and rhamnose was evidenced, except for mucilage modified5-1 (mum5-1; a mutant showing a redistribution of mucilage pectin from the inner adherent layer to the outer soluble one), for which the rhamnose-xylose ratio was increased drastically. Biochemical and in vitro binding assay data demonstrated that xylan chains are attached to RG-I chains and mediate the adsorption of mucilage to cellulose microfibrils. mum5-1 mucilage exhibited very weak adsorption to cellulose. MUM5 was identified as a putative xylosyl transferase recently characterized as MUCI21. Together, these findings suggest that the binding affinity of xylose ramifications on RG-I to a cellulose scaffold is one of the factors involved in the formation of the adherent mucilage layer.
Asunto(s)
Arabidopsis/metabolismo , Pared Celular/metabolismo , Regulación de la Expresión Génica de las Plantas , Mucílago de Planta/genética , Mucílago de Planta/metabolismo , Semillas/metabolismo , Xilanos/metabolismo , Arabidopsis/enzimología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pared Celular/química , Celulosa/metabolismo , Análisis por Conglomerados , Genes de Plantas , Ligamiento Genético , Glucosiltransferasas/genética , Glucosiltransferasas/metabolismo , Ácidos Hexurónicos/metabolismo , Mutación , Pectinas/química , Pectinas/metabolismo , Extractos Vegetales/química , Mucílago de Planta/química , Ramnosa/metabolismo , Semillas/enzimología , Análisis de Secuencia de ADN , Coloración y Etiquetado , Xilanos/química , Xilosa/metabolismoRESUMEN
Pectins are major components of plant primary cell walls. They include homogalacturonans (HGs), which are the most abundant pectin and can be the target of apoplastic enzymes like pectin methylesterases (PMEs) that control their methylesterification level. Several PMEs are expressed in the seed coat of Arabidopsis thaliana, particularly in mucilage secretory cells (MSCs). On the basis of public transcriptomic data, seven PME genes were selected and checked for their seed-specific expression by quantitative reverse transcription PCR. Of these, PME58 presented the highest level of expression and was specifically expressed in MSCs at the early stages of seed development. pme58 mutants presented two discrete phenotypes: (i) their adherent mucilage was less stained by ruthenium red when compared to wild-type seeds, but only in the presence of EDTA, a Ca(2+)chelator; and (ii) the MSC surface area was decreased. These phenotypes are the consequence of an increase in the degree of HG methylesterification connected to a decrease in PME activity. Analysis of the sugar composition of soluble and adherent mucilage showed that, in the presence of EDTA, sugars of adherent mucilage were more readily extracted in pme58 mutants. Immunolabelling with LM19, an antibody that preferentially recognizes unesterified HGs, also showed that molecular interactions with HGs were modified in the adherent mucilage of pme58 mutants, suggesting a role of PME58 in mucilage structure and organization. In conclusion, PME58 is the first PME identified to play a direct role in seed mucilage structure.