RESUMEN
Pollen fertility is critical for successful fertilization and, accordingly, for crop yield. While sugar unloading affects the growth and development of all types of sink organs, the molecular nature of sugar import to tomato (Solanum lycopersicum) pollen is poorly understood. However, sugar will eventually be exported transporters (SWEETs) have been proposed to be involved in pollen development. Here, reverse transcription-quantitative polymerase chain reaction (PCR) revealed that SlSWEET5b was markedly expressed in flowers when compared to the remaining tomato SlSWEETs, particularly in the stamens of maturing flower buds undergoing mitosis. Distinct accumulation of SlSWEET5b-ß-glucuronidase activities was present in mature flower buds, especially in anther vascular and inner cells, symplasmic isolated microspores (pollen grains), and styles. The demonstration that SlSWEET5b-GFP fusion proteins are located in the plasma membrane supports the idea that the SlSWEET5b carrier functions in apoplasmic sugar translocation during pollen maturation. This is consistent with data from yeast complementation experiments and radiotracer uptake, showing that SlSWEET5b operates as a low-affinity hexose-specific passive facilitator, with a Km of â¼36 mM. Most importantly, RNAi-mediated suppression of SlSWEET5b expression resulted in shrunken nucleus-less pollen cells, impaired germination, and low seed yield. Moreover, stamens from SlSWEET5b-silenced tomato mutants showed significantly lower amounts of sucrose (Suc) and increased invertase activity, indicating reduced carbon supply and perturbed Suc homeostasis in these tissues. Taken together, our findings reveal the essential role of SlSWEET5b in mediating apoplasmic hexose import into phloem unloading cells and into developing pollen cells to support pollen mitosis and maturation in tomato flowers.
Asunto(s)
Solanum lycopersicum , Flores/genética , Flores/metabolismo , Hexosas/metabolismo , Solanum lycopersicum/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Polen , Sacarosa/metabolismoRESUMEN
Tomato (Solanum lycopersium), an important fruit crop worldwide, requires efficient sugar allocation for fruit development. However, molecular mechanisms for sugar import to fruits remain poorly understood. Expression of sugars will eventually be exported transporters (SWEETs) proteins is closely linked to high fructose/glucose ratios in tomato fruits and may be involved in sugar allocation. Here, we discovered that SlSWEET15 is highly expressed in developing fruits compared to vegetative organs. In situ hybridization and ß-glucuronidase fusion analyses revealed SlSWEET15 proteins accumulate in vascular tissues and seed coats, major sites of sucrose unloading in fruits. Localizing SlSWEET15-green fluorescent protein to the plasma membrane supported its putative role in apoplasmic sucrose unloading. The sucrose transport activity of SlSWEET15 was confirmed by complementary growth assays in a yeast (Saccharomyces cerevisiae) mutant. Elimination of SlSWEET15 function by clustered regularly interspaced short palindromic repeats (CRISPRs)/CRISPR-associated protein gene editing significantly decreased average sizes and weights of fruits, with severe defects in seed filling and embryo development. Altogether, our studies suggest a role of SlSWEET15 in mediating sucrose efflux from the releasing phloem cells to the fruit apoplasm and subsequent import into storage parenchyma cells during fruit development. Furthermore, SlSWEET15-mediated sucrose efflux is likely required for sucrose unloading from the seed coat to the developing embryo.
Asunto(s)
Frutas/crecimiento & desarrollo , Proteínas de Transporte de Membrana/metabolismo , Floema/metabolismo , Semillas/crecimiento & desarrollo , Solanum lycopersicum/crecimiento & desarrollo , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Sacarosa/metabolismo , Productos Agrícolas/genética , Productos Agrícolas/crecimiento & desarrollo , Productos Agrícolas/metabolismo , Frutas/genética , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Proteínas de Transporte de Membrana/genética , Floema/genética , Semillas/genéticaRESUMEN
Gastrodia elata, a fully mycoheterotrophic orchid without photosynthetic ability, only grows symbiotically with the fungus Armillaria. The mechanism of carbon distribution in this mycoheterotrophy is unknown. We detected high sucrose concentrations in all stages of Gastrodia tubers, suggesting sucrose may be the major sugar transported between fungus and orchid. Thick symplasm-isolated wall interfaces in colonized and adjacent large cells implied involvement of sucrose importers. Two sucrose transporter (SUT)-like genes, GeSUT4 and GeSUT3, were identified that were highly expressed in young Armillaria-colonized tubers. Yeast complementation and isotope tracer experiments confirmed that GeSUT4 functioned as a high-affinity sucrose-specific proton-dependent importer. Plasma-membrane/tonoplast localization of GeSUT4-GFP fusions and high RNA expression of GeSUT4 in symbiotic and large cells indicated that GeSUT4 likely functions in active sucrose transport for intercellular allocation and intracellular homeostasis. Transgenic Arabidopsis overexpressing GeSUT4 had larger leaves but were sensitive to excess sucrose and roots were colonized with fewer mutualistic Bacillus, supporting the role of GeSUT4 in regulating sugar allocation. This is not only the first documented carbon import system in a mycoheterotrophic interaction but also highlights the evolutionary importance of sucrose transporters for regulation of carbon flow in all types of plant-microbe interactions.
Asunto(s)
Gastrodia/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Proteínas de Plantas/metabolismo , Sacarosa/metabolismo , Simbiosis , Arabidopsis , Armillaria/metabolismo , Armillaria/fisiología , Gastrodia/microbiología , Gastrodia/fisiología , Hibridación in Situ , Proteínas de Transporte de Membrana/fisiología , Microscopía Electrónica de Transmisión , Micorrizas/metabolismo , Micorrizas/ultraestructura , Proteínas de Plantas/fisiología , Tubérculos de la Planta/metabolismo , Tubérculos de la Planta/microbiología , Tubérculos de la Planta/ultraestructura , Plantas Modificadas GenéticamenteRESUMEN
Sugar allocation from source to sink (young) leaves, critical for plant development, relies on activities of plasma membrane sugar transporters. However, the key sugar unloading mechanism to sink leaves remains elusive. SWEET transporters mediate sugar efflux into reproductive sinks; therefore, they are promising candidates for sugar unloading during leaf growth. Transcripts of SlSWEET1a, belonging to clade I of the SWEET family, were markedly more abundant than those of all other 30 SlSWEET genes in young leaves of tomatoes. High expression of SlSWEET1a was also detected in reproductive sinks, such as flowers. SlSWEET1a was dominantly expressed in leaf unloading veins, and the green fluorescent protein (GFP) fusion protein was localized to the plasma membrane using Arabidopsis protoplasts, further implicating this carrier in sugar unloading. In addition, yeast growth assays and radiotracer uptake analyses further demonstrated that SlSWEET1a acted as a low-affinity (Km ~100 mM) glucose-specific carrier with a passive diffusion manner. Finally, virus-induced gene silencing of SlSWEET1a expression reduced hexose accumulation to ~50% in young leaves, with a parallel 2-fold increase in mature leaves. Thus, we propose a novel function for SlSWEET1a in the uptake of glucose into unloading cells as part of the sugar unloading mechanism in sink leaves of tomato.
Asunto(s)
Glucosa/metabolismo , Proteínas de Plantas/genética , Solanum lycopersicum/genética , Transporte Biológico , Solanum lycopersicum/metabolismo , Hojas de la Planta/metabolismo , Proteínas de Plantas/metabolismoRESUMEN
Plant roots secrete a significant portion of their assimilated carbon into the rhizosphere. The putative sugar transporter SWEET2 is highly expressed in Arabidopsis roots. Expression patterns of SWEET2-ß-glucuronidase fusions confirmed that SWEET2 accumulates highly in root cells and thus may contribute to sugar secretion, specifically from epidermal cells of the root apex. SWEET2-green fluorescent protein fusions localized to the tonoplast, which engulfs the major sugar storage compartment. Functional analysis of SWEET2 activity in yeast showed low uptake activity for the glucose analog 2-deoxyglucose, consistent with a role in the transport of glucose across the tonoplast. Loss-of-function sweet2 mutants showed reduced tolerance to excess glucose, lower glucose accumulation in leaves, and 15-25% higher glucose-derived carbon efflux from roots, suggesting that SWEET2 has a role in preventing the loss of sugar from root tissue. SWEET2 root expression was induced more than 10-fold during Pythium infection. Importantly, sweet2 mutants were more susceptible to the oomycete, showing impaired growth after infection. We propose that root-expressed vacuolar SWEET2 modulates sugar secretion, possibly by reducing the availability of glucose sequestered in the vacuole, thereby limiting carbon loss to the rhizosphere. Moreover, the reduced availability of sugar in the rhizosphere due to SWEET2 activity contributes to resistance to Pythium.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Proteínas de Transporte de Monosacáridos/metabolismo , Enfermedades de las Plantas/microbiología , Raíces de Plantas/metabolismo , Pythium/patogenicidad , Arabidopsis/microbiología , Proteínas de Arabidopsis/genética , Secuestro de Carbono , Regulación de la Expresión Génica de las Plantas , Glucosa/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Interacciones Huésped-Patógeno , Proteínas de Transporte de Monosacáridos/genética , Hojas de la Planta/metabolismo , Raíces de Plantas/microbiología , Plantas Modificadas Genéticamente , Vacuolas/metabolismoRESUMEN
Potassium (K) is essential for the processes critical for plant performance, including photosynthesis, carbon assimilation, and response to stress. K also influences translocation of sugars in the phloem and regulates sucrose metabolism. Several plant species synthesize polyols and transport these sugar alcohols from source to sink tissues. Limited knowledge exists about the involvement of K in the above processes in polyol-translocating plants. We, therefore, studied K effects in Plantago major, a species that accumulates the polyol sorbitol to high concentrations. We grew P. major plants on soil substrate adjusted to low-, medium-, or high-potassium conditions. We found that biomass, seed yield, and leaf tissue K contents increased in a soil K-dependent manner. K gradually increased the photosynthetic efficiency and decreased the non-photochemical quenching. Concomitantly, sorbitol levels and sorbitol to sucrose ratio in leaves and phloem sap increased in a K-dependent manner. K supply also fostered plant cold acclimation. High soil K levels mitigated loss of water from leaves in the cold and supported cold-dependent sugar and sorbitol accumulation. We hypothesize that with increased K nutrition, P. major preferentially channels photosynthesis-derived electrons into sorbitol biosynthesis and that this increased sorbitol is supportive for sink development and as a protective solute, during abiotic stress.