Lipoxygenase and Its Relationship with Ethylene During Ripening of Genetically Modified Tomato (Solanum lycopersicum).

RESEARCH BACKGROUND: TomloxB is the main isoform of lipoxygenase associated with ripening and senescence of fruits. On the other hand, ethylene, a gaseous hormone, is essential for the regulation of ripening in climacteric fruits like tomatoes. However, the relationship between TomloxB and ethylene production has not been thoroughly studied. Therefore, we aim to assess the effect of exogenous ethylene in transgenic tomatoes that contain a silenced TomloxB gene, and subsequently evaluate lipoxygenase activity, 1-aminocyclopropane-1-carboxylic acid oxidase and ethylene production; as well as to quantify the expression of the genes encoding 1-aminocyclopropane-1-carboxylic acid oxidase and TomloxB. EXPERIMENTAL APPROACH: To investigate the effect of lipoxygenase and 1-aminocyclopropane-1-carboxylic acid oxidase activity, fruits harvested at the stages of break, turning and pink were used. Tomatoes at break stage collected from transgenic and wild type plants were used to determine ethylene production and gene expression. Genetically modified and wild type tomato fruits were exposed to 100 µL/L exogenous ethylene. Lipoxygenase activity was measured spectrophotometrically. Activity of 1-aminocyclopropane-1-carboxylic acid oxidase and ethylene production were determined by gas chromatography. Oligonucleotides for differentially expressed genes: 1-aminocyclopropane-1-carboxylic acid oxidase and TomloxB were used to determine gene expression by real-time PCR. RESULTS AND CONCLUSIONS: The data showed that silencing of TomloxB caused a reduction in lipoxygenase activity and ethylene production in tomato fruits, and also reduced 1-aminocyclopropane-1-carboxylic acid oxidase activity. Hence, the addition of exogenous ethylene increased lipoxygenase activity in all treatments and 1-aminocyclopropane-1-carboxylic acid oxidase activity only in transgenic lines at break stage, consequently there was a positive regulation between TomloxB and ethylene, as increasing the amount of ethylene increased the activity of lipoxygenase. The results suggest that lipoxygenase may be a regulator of 1-aminocyclopropane-1-carboxylic acid oxidase and production of ethylene at break stage. NOVELTY AND SCIENTIFIC CONTRIBUTION: These results lead to a better understanding of the metabolic contribution of TomloxB in fruit ripening and how it is linked to the senescence-related process, which can lead to a longer shelf life of fruits. Understanding this relationship between lipoxygenase and ethylene can be useful for better post-harvest handling of tomatoes.

Isolation and characterization of novel soil- and plant-associated bacteria with multiple phytohormone-degrading activities using a targeted methodology.

Ethylene (ET), salicylic acid (SA) and indole-3-acetic acid (IAA) are important phytohormones regulating plant growth and development, as well as plant-microbe interactions. Plant growth-promoting bacteria (PGPB) naturally associate with plants and facilitate plant growth through a variety of mechanisms, including the ability to modulate the concentrations of these phytohormones in planta. Importantly, the wide presence of phytohormone degradation mechanisms amongst symbiotic and other soil- and plant-associated bacteria indicates that the ability to modulate phytohormone concentrations plays an important role in bacterial colonization and plant-growth promotion abilities. Obtaining phytohormone-degrading bacteria is therefore key for the development of novel solutions aiming to increase plant growth and protection. In this paper, we report an optimized targeted methodology and the consequent isolation of novel soil- and plant-associated bacteria, including rhizospheric, endophytic and phyllospheric strains, with the ability to degrade the phytohormones, SA and IAA, as well as the ET precursor, 1-aminocyclopropane-1-carboxylic acid (ACC). By using an optimized targeted methodology, we rapidly isolated diverse soil- and plant-associated bacteria presenting phytohormone-degrading abilities from several plants, plant tissues and environments, without the need for prior extensive and laborious isolation and maintenance of large numbers of isolates. The developed methodology facilitates PGPB research, especially in developing countries. Here, we also report, for the first time, the isolation of bacterial strains able to concomitantly catabolize three phytohormones (SA, IAA and ACC). Ultimately, the described targeted methodology and the novel phytohormone-degrading bacteria obtained in this work may be useful tools for future plant-microbe interaction studies, and in the development of new inoculant formulations for agriculture and biotechnology.

Germination of salt-stressed seeds as related to the ethylene biosynthesis ability in three Stylosanthes species.

Stylosanthes, a genus of tropical forage legume, is known to exhibit good persistence in saline soils, yet mechanisms for regulation of seed germination under salt stress are poorly understood. This study was carried out to evaluate the mode of action of salt stress on seed germination of Stylosanthes. 1-Aminocyclopropane-1-carboxylic acid (ACC) increased ethylene biosynthesis and germination of NaCl-inhibited seeds in a dose-dependent manner. Contents of ACC and germination of Stylosanthes humilis seeds increased following transfer from NaCl solution to deionised water, but not after transfer to l-α-(2-aminoethoxyvinyl)-glycine (AVG) solution, an inhibitor of ethylene biosynthesis. Ethylene biosynthesis was much larger in NaCl-treated seeds of Stylosanthes guianensis than in seeds of S. humilis and Stylosanthes capitata, a fact which was reflected in higher germination rates. S. guianensis seedlings also displayed higher growth and survival rates than S. humilis and S. capitata under salt stress. Moreover, smaller ACC levels, as well as reduced ethylene biosynthesis of S. capitata seeds were accompanied by lower germination under salt stress. In addition, S. capitata seedlings treated with NaCl solutions exhibited relatively lower growth and survival rates in comparison with S. humilis and S. guianensis. Thus, different abilities to synthesize ethylene by S. guianensis, S. humilis and S. capitata seeds explain the differences in tolerance to salt stress of the three species.