Metabolismo de metilglioxal en macroalgas durante desecación

Indexación: Scielo; Web of Science.In primary producers, diverse stressors cause an over-production of methylglyoxal (MG), which is principally detoxified by glyoxalase I (GLO1) activity. A recent proteomic study found that GLO1 was up-regulated during natural desiccation in the red seaweed Pyropia orbicularis, which inhabits the upper intertidal rocky zone and compared to other species, is highly tolerant to air exposure. To better understand and determine differential responses to desiccation stress, this study evaluated MG concentration and GLO1 activity in two species with contrasted vertical distribution, P. orbicularis and Lessonia spicata (lower distribution). Results showed that P. orbicularis successfully scavenges MG via increased GLO1 activity during desiccation. In contrast, GLO1 activity in L. spicata did not increase during desiccation, resulting in MG overproduction. This study is the first to quantify MG and GLO1 levels in seaweeds during natural stress, and partly explain the mechanisms by which P. orbicularis is dominant in the upper rocky intertidal zone.http://ref.scielo.org/tp4sn

Expresión diferencial de genes en Pyropia columbina (Bangiales, Rhodophyta) bajo hidratación y desecación natural

Indexación: Web of Science; Scielo.RESUMEN. En Zonas Costeras rocosas, la desecación es gatillada porción Cambios Diarios en los Niveles de marea, y la Evidencia indica experimental de Me Distribución de las algas en la zona intermareal no está Relacionada estafa do palabra capacidad, párr tolerar la desecación. En Este Contexto, la Presencia de Pyropia columbina en la zona alta del intermareal sí Explica Por Su excepcional tolerancia fisiológica a la desecación. Este Estudio explora las Vías Metabólicas involucradas en la tolerancia a la desecación en P. columbina, un Través de la Caracterización de do transcriptoma Bajo Condiciones de hidratación contrastantes. Se obtuvó 1410 TER provenientes de dos Librerías de substracción de ADNc de frondas Naturalmente hidratadas y desecadas. Los transcriptomas de emba Librerías contienen transcritos de Diversas Rutas Metabólicas Relacionadas a la tolerancia. Entre el los transcritos expresados ​​15% estan involucrados en la Síntesis de Proteínas, do Procesamiento y degradacion, 14,4% Asociados un Fotosíntesis y cloroplasto, el 13,1% una mitocondrial Respiración and function, 10,6% al metabolism de la Pared Celular y 7,5% a la Actividad ANTIOXIDANTE, Proteínas chaperonas y factors de Defensa (catalasa, tiorredoxina, Proteínas de choque térmico, P450 citocromo). In Ambás Librerías sí DESTACA La Presencia De genes / Proteínas no descritos en algas. Proporciona Información This El Primer Trabajo molecular Que Estudia la tolerancia a desecación en P. columbina y Sus Resultados Ayudan a explicar los patrones clásicos de Distribución descritos párr algas en la zona intermareal. Palabras clave: Pyropia, desecación porción Estrés, EST, macroalgas, transcriptómica, Proteínas.ABSTRACT. In rocky shores, desiccation is triggered by daily tide changes, and experimental evidence suggests that local distribution of algal species across the intertidal rocky zone is related to their capacity to tolerate desiccation. In this context, the permanence of Pyropia columbina in the high intertidal rocky zone is explained by its exceptional physiological tolerance to desiccation. This study explored the metabolic pathways involved in tolerance to desiccation in the Chilean P. columbina, by characterizing its transcriptome under contrasting conditions of hydration. We obtained 1,410 ESTs from two subtracted cDNA libraries in naturally hydrated and desiccated fronds. Results indicate that transcriptome from both libraries contain transcripts from diverse metabolic pathways related to tolerance. Among the transcripts differentially expressed, 15% appears involved in protein synthesis, processing and degradation, 14.4% are related to photosynthesis and chloroplast, 13.1% to respiration and mitochondrial function (NADH dehydrogenase and cytochrome c oxidase proteins), 10.6% to cell wall metabolism, and 7.5% are involved in antioxidant activity, chaperone and defense factors (catalase, thioredoxin, heat shock proteins, cytochrome P450). Both libraries highlight the presence of genes/proteins never described before in algae. This information provides the first molecular work regarding desiccation tolerance in P. columbina, and helps, to some extent, explaining the classical patterns of ecological distribution described for algae across the intertidal zone.http://ref.scielo.org/jm5rc

Antimitotic and hormone effects on green diplohaploid plant production through anther culture on Mediterranean japonica rice

Rice double haploid (DH) plants are produced mainly through anther culture. In order to improve the anther culture protocol, microspores of two japonica rice genotypes (NRVC980385 and H28) were subjected to three growth regulator combinations and four colchicine treatments on induction medium. In addition, a post anther culture procedure using colchicine or oryzalin was tested to induce double haploid plantlets from haploid plantlets. A cold pre-treatment of microspores for 9 days at 10 °C increased callus induction 50-fold in the NRCV980385 genotype. For both genotypes, 2 mg L−1 2,4-D and 1 mg L−1 kinetin on colchicine-free induction medium gave the best culture responses. The culturability of both genotypes changed on colchicine-supplemented induction media. A high genotype dependency was recorded for callus induction, callus regenerating green plantlets and regeneration of green double haploid plantlets. Colchicine at 300 mg L−1 for 48 h enhanced callus induction 100-fold in H28. Colchicine-supplemented media clearly improved green double haploid plantlet regeneration. We showed that the post-anther culture treatment of haploid plantlets at 500 mg L−1 of colchicine permitted fertile double haploid plantlets to be generated. Finally, an enhanced medium-throughput flow cytometry protocol for rice was tested to analyse all the plantlets from anther and post anther culture

Phytohormone Profiling method for rice: effects of GA20ox mutation on the gibberellin content of japonica rice varieties

Gibberellins (GAs) are a very important group of phytohormones involved in seed germination, vegetative growth, flowering, and fruit development, being only 4 of the 136 known bioactives: GA1, GA3, GA4 and GA7. It has been evidenced that mutations in the OsGA20ox-2 gene produce rice (Oryza sativa) dwarf varieties, which were one of the main pillars of the green revolution. In this work two main objectives were proposed: (i) develop a rapid and broad phytohormone profiling method and (ii) to study the effects on the GA content of the GA20ox-2 mutation in several rice developmental stages using three varieties (tall variety, elite variety, mutated variety). A phytohormone extraction using an SPE step and HPLC-MS/MS detection using a QqQ instrument was determined which resulted in limits of detection (LOD) and limits of quantification (LOQ) for GAs that varied between 0.1-0.7 and 0.3-2.3 pg/g of rice sample respectively, allowing highly sensitive phytohormones detection in samples. Moreover, a good reproducibility was obtained for the GAs as relative standard deviations (RSD) for a 40 ng/mL pattern varied between 0.3-0.9%. Notoriously, GA1 was absent in the coleoptile and GA4 was the GA with higher content in the majority of developmental stages. We also observed a huge large content increment increase of the 4 bioactive GAs in the internode of the flag leaf of the mutated variety that allowinged it to attain reach the same height as the elite variety. Therefore, we provide a rapid and broad phytohormonal profiling method and evidence that the GA20ox-2 mutation is not the only factor generating dwarf varieties. To our knowledge, this is the first study that it has been reported such a high number of analyzed gibberellins at the same time in rice (Oryza sativa L. ssp. japonica)Oryza sativa japonica)

Exploring the potential of Meyerozyma guilliermondii on physiological performances and defense response against Fusarium Crown Rot on Durum Wheat

Coating seeds with bio-control agents is a potentially effective approach to reduce the usage of pesticides and fertilizers applied and protect the natural environment. This study evaluated the effect of seed coating with Meyerozyma guilliermondii, strain INAT (MT731365), on seed germination, plant growth and photosynthesis, and plant resistance against Fusarium culmorum, in durum wheat under controlled conditions. Compared to control plants, seed coating with M. guilliermondii promoted the wheat growth (shoot and roots length and biomass), and photosynthesis and transpiration traits (chlorophyll, ɸPSII, rates of photosynthesis and transpiration, etc.) together with higher nitrogen balance index (NBI) and lower flavonols and anthocyanins. At 21 days post infection with Fusarium, M. guilliermondii was found to reduce the disease incidence and the severity, with reduction rates reaching up to 31.2% and 30.4%, respectively, as well as to alleviate the disease damaging impact on photosynthesis and plant growth. This was associated with lower ABA, flavonols and anthocyanins, compared to infected control. A pivotal function of M. guilliermondii as an antagonist of F. culmorum and a growth promoter is discussed

A novel aspect of essential oils: Coating seeds with thyme essential oil induces drought resistance in wheat

Coating seeds with biostimulants is among the promising approaches in crop production to increase crop tolerance to drought stress. In this study, we evaluated the potential of coating durum wheat seeds of the cultivar 'Karim' with thyme essential oil on enhancing seed germination and seedling growth, and on plant growth promotion and induction of drought resistance. Coated seeds were pre-germinated, grown in hydroponics, and grown in pots under controlled well-watered and progressive water/nutrient stress conditions. Seed coating with thyme oil increased germination rate and enhanced seedling growth development in hydroponics. In the pot experiment, thyme oil increased, when well watered, root and shoot development, chlorophyll, nitrogen balance index (NBI), abscisic acid (ABA), anthocyanins and flavonoids in leaves, decreased nitrogen isotope composition (δ15N) and increased carbon isotope composition (δ13C) of shoots. Increasing water/nutrient stress in control plants induced higher accumulation of ABA and anthocyanins coupled with a transient decrease in chlorophyll and NBI, a decrease in shoot and root development, the Normalized Difference Vegetation Index (NDVI), shoot C content, δ15N, and an increase in δ13C, revealing the avoidance strategy adopted by the cultivar. Thyme oil had the potential to enhance the avoidance strategy by inducing roots elongation, reducing the loss of shoot and roots dry matter and chlorophyll, maintaining balanced NBI, an decreasing anthocyanins, flavonoids, and δ13C via maintaining lower ABA-mediated-stomatal closure. Thyme oil increased shoot N content and δ15N indicating preferential uptake of the 15N enriched NH4+. Coating seeds with thyme oil is suggested as a promising alternative approach to improve plant's water and nutrient status and to enhance drought resistance

Variety improvement in rice (Oryza sativa L.): proteomic, hormonal and in vitro studies

[eng] Rice (Oryza sativa L.) is without doubt one of the major crops worldwide, as its consumption is continuously increasing, especially in low- and lower-middle-income countries where it is the most important staple. This cereal has been domesticated for a long time, result of which several species and varieties are now available. It was key during the green revolution when its production increased in more than 2-fold times, and in addition has a deep cultural background in all the regions where it is grown. Hence, it is of uttermost importance for researchers and breeders to broaden and expand the knowledge on this cereal on all the study and research areas, especially now that we are living in the 21st century, which is marked by climate change. This phenomenon is one of the most menacing as it will reduce the quantity and quality of arable land due to salinization of soil as well as water scarcity which is the single and most important factor that determines global crop yields. In this sense, this thesis addresses to three important topics on rice that will help researchers and breeders for the improvement of rice varieties in the forthcoming future: (i) Analysis of salinity tolerant rice plants subjected to high salt concentration through a combined approach of shotgun proteomics and physiological characterization for the identification of new key proteins involved in the tolerance to this stress. In this study we provide a huge database for future studies of the salinity-stress response in rice varieties harboring the Saltol region, as we detected more than 2000 proteins involved during the early stages of salinity stress in the shoots and roots of the FL478 rice line. Moreover, this study highlights the importance of examining both the shoots and roots because their salinity tolerance traits are different and respond to different requirements of the rice plant. Finally, the information presented here expands our knowledge on adaptive processes under high salinity in rice plants and, notably, in rice roots. (ii) Phytohormones analysis in dwarf varieties through a broad phytohormone profiling method developed during this thesis for the characterization of phytohormone levels during the development of plants with contrasting heights that will allow to have more information for developing new dwarf mutant varieties. In this work, we reported that GA19 seems to have a crucial role in gibberellin availability in rice as its levels were much higher than all the other gibberellins in all tissues. In addition, it has been demonstrated that the GA20ox-2 mutation is not the only factor affecting height in rice, as a mutated variety had an increased growth during the half of its development period. Finally, we established for the first time a simple and broad phytohormone extraction and detection protocol that allows to identify 13 gibberellins and ABA, JA and IAA in several tissues at different phenological stages. (iii) Improvement and enhancement of anther culture protocols in rice for obtaining higher rates of stabilized green double haploid plants using different cold-pretreatments and additives in the growing media such as hormones and antimitotics in the growing media for their subsequent commercialization. It was evidenced that the best cold-pretreatment was 10ºC for 9 days, and that colchicine addition greatly enhances the production of green double haploid plantlets. Moreover, we have introduced for the first time a post-anther culture treatment with good prospects. Finally, several of the rice lines produced were assayed in rice paddy fields where they displayed good agronomical behavior.[spa] El arroz (Oryza sativa L.) es sin duda uno de los principales cultivos del mundo, especialmente en países en vías de desarrollo donde es el alimento básico, teniendo además un importante componente cultural. Fue clave en la revolución verde cuando su producción aumentó más del doble. En el siglo 21 marcado por el cambio climático, se verá reducida la cantidad y calidad de las tierras cultivables debido a la salinización del suelo y la escasez de agua. Por tanto, es de gran importancia para los investigadores y mejoradores ampliar el conocimiento que se tiene sobre este cereal en todas las áreas de estudio. En este sentido, esta tesis abordó tres temas importantes referentes a este cereal: (i) Un estudio del proteoma, de la parte aérea y radicular, de plantas de arroz tolerantes a la salinidad sometidas a una alta concentración de sal con el objetivo de identificar de nuevas proteínas clave involucradas en la tolerancia a este estrés. Proporcionamos una gran base de datos, más de 200 proteínas involucradas en este estrés y destacamos la importancia de las raíces para la tolerancia a este estrés. (ii) Análisis de fitohormonas en variedades enanas a través de un amplio y rápido método desarrollado durante esta tesis que permite, por primera vez en arroz, la extracción y detección de fitohormonas que posibilida la identificación de 13 giberelinas y ABA, JA e IAA en varios tejidos en diferentes etapas fenológicas. Se observó que la GA19 parece tener un papel crucial en la disponibilidad de giberelinas en el arroz, ya que sus niveles son los más elevados en todos los tejidos. La mutación GA20ox-2 no es el único factor que afecta la altura ya que la variedad mutada alcanza la altura de variedades wild-type. (iii) Por último, se reportan dos métodos de aumento de la eficiencia del protocolo de cultivo de anteras, técnica muy útil para la obtención de plantas de arroz dihaploides estables. El aumento del rendimiento se debió a nuevos pretratamientos de frío, así como la modificación de los medios de cultivo mediante diferentes concentraciones de hormonas y colchicina. Se introdujo, por primera vez, un tratamiento de diploidización para plantas haploides

Efficient knockout of Phytoene desaturase gene using CRISPR/Cas9 in melon

CRISPR/Cas9 system has been widely applied in many plant species to induce mutations in the genome for studying gene function and improving crops. However, to our knowledge, there is no report of CRISPR/Cas9-mediated genome editing in melon (Cucumis melo). In our study, phytoene desaturase gene of melon (CmPDS) was selected as target for the CRISPR/Cas9 system with two designed gRNAs, targeting exons 1 and 2. A construct (pHSE-CmPDS) carrying both gRNAs and the Cas9 protein was delivered by PEG-mediated transformation in protoplasts. Mutations were detected in protoplasts for both gRNAs. Subsequently, Agrobacterium-mediated transformation of cotyledonary explants was carried out, and fully albino and chimeric albino plants were successfully regenerated. A regeneration efficiency of 71% of transformed plants was achieved from cotyledonary explants, a 39% of genetic transformed plants were successful gene edited, and finally, a 42-45% of mutation rate was detected by Sanger analysis. In melon protoplasts and plants most mutations were substitutions (91%), followed by insertions (7%) and deletions (2%). We set up a CRISPR/Cas9-mediated genome editing protocol which is efficient and feasible in melon, generating multi-allelic mutations in both genomic target sites of the CmPDS gene showing an albino phenotype easily detectable after only few weeks after Agrobacteriummediated transformation