Estudio del efecto del aumento de la temperatura asociado a las condiciones del cambio climático sobre el olivo (Olea europaea L.), mediante el uso integrado de técnicas de fisiología y bioquímica y bioquímica clásica, proteómica, metabolómica y transcriptómica

This Doctoral Thesis is focused on olive tree (Olea europaea L. subsp. europaea), the dominant tree crop over large areas of the Western Mediterranean Basin, with a production of 20,872,788 tonnes in 2018. Olive oil is appreciated worldwide because of its chemical composition, nutraceutical value, and organoleptic properties. Like most crops, the olive tree is facing important problems and challenges related to adverse environmental conditions, either biotic or abiotic ones, a situation that can be worsened in a climate change scenario. Hence, understanding the physiological and molecular basis/mechanism of olive tree adaptability and resilience to different environmental stresses, especially to high temperature, is crucial for predicting the consequences of climate warming, for implementing agricultural practices, designing classic or biotechnological breeding programs, and, lately, increasing its production in order to cope with the current demand. In the present work, both the effect and the responses to an increase of 4 °C above ambient temperature have been studied at phenological, physiological and molecular,-omics, levels, in an attempt to integrate all the results in the novel Systems Biology direction. The use of the –omics techniques (transcriptomics, proteomics, and metabolomics), and its integration with physiological and phenological studies constitute the great novelty of this work. It has required an important effort in optimizing different protocols for transcriptome (RNA isolation and qRT-PCR), proteome (shotgun nLC-MS/MS Orbitrap), and metabolome (UHPLC-MS/MS Q-tof) profiling, together with computational tools for the identification and quantification of the different biomolecules, their functional classification, statistical analysis, clustering and networking. Both newly developed tools and knowledge generated will establish the basis for ongoing and future studies to obtain a better understanding of the molecular mechanisms mediating phenotypes of interest (productive, tolerant to environmental cues, nutraceutical value and the selectionof genotypes more resilient to climate warming). The manuscript has been organized and presented in nine chapters as follows: Chapter I summarises the state of the art of the topics covered in this thesis: the experimental system, objectives, hypothesis and methodologies. It starts with the description of the olive tree as a plant species and crop, its economic and cultural importance, problems and challenges related to its cultivation. The chapter continues with the description of, and publications on, environmental stresses with an emphasis on warming temperatures and the denominated “Climate Change”. Responses and mechanisms of tolerance to temperature stress and climate warming will be discussed. This chapter finishes with the description of the methodology employed and the integration of the different approaches. Chapter II presents the general and specific objectives of the Thesis. Chapter III corresponds to two published papers on the warming effect on olive tree phenology: “An approach to global warming effects on flowering and fruit set of olive trees growing under field conditions” and “Global warming effects on yield and fruit maturation of olive trees growing under field conditions” Both were published in Scientia Horticulturae. They present alterations in the flowering and ripening processes, caused by the permanent thermal increase of 4oC above ambient temperatura (AT) applied to adult olive trees (cv. `Picual´) growth underfield conditions. In this chapter, three years were surveyed, analysed and data analysed in relation to climate conditions. In general, earlier and prolonged flowering and ripening were observed under warmer (AT+4oC) conditions. Temperature increase reduced the quantity of perfect (well- developed) flowers and fruits. A decrease in oil quality-related chemical parameters (fat, phenolics, and anthocyanins) was observed under thermal increase conditions. Chapter IV corresponds to two published papers on the effect of different temperature regimes in roots and shoots of olive tree seedlings (cv. Arbequina), a plant growth chamber experiment: “Effect of moderate high temperature on the vegetative growth and potassium allocation in olive plants” and “Effects of olive root warming on potassium transport and plant growth” They were published in Journal of Plant Physiology. The effect of differential thermal 37oC /25oC, exposurebetween root and aerial parton growth, and K uptake and transport were evaluated. At 37oC root, 25oC shoot, potassium uptake increased. 37 °C in both organs inhibited growth, reduced K uptake and transport, and diminished water content. Chapters V, VI and VII are devoted to the different –omics approaches (proteomics, transcriptomics and metabolomics) employed in the molecular study of the effect and responses to warming temperature (+ 4oC) in fruit pulp at three ripening stages: green (0), turning red (called veraison) (3) and purple epicarp (4). A discrepancy between the colouring of the skin, traditionally observed as a reference for phenology studies and harvesting, and the olive pulp’s internal ripeness stage and chemical composition has been found. The veraison stage was the one most affected by the temperature increase. Overall, a reduction in the abundance of major compounds related to olive oil quality (lipids, phenylpropanoids, flavonoids, and terpenes) was observed under warming conditions. Chapter VIII contains an integration of all the data, and a general discussion, with focus on those most relevant parameters from an agronomic point of view, those related to crop productivity, either from a quantitative (fruit production) or qualitative (fruit chemical composition) perspective. Based on the present study, a predictive hypothesis on the olive tree situation and production in a future climate change scenario is presented, proposing alternative management practices to deal with the social demand for this crop. Chapter IX corresponds to the conclusions.Esta Tesis Doctoral se centra en el olivo (Olea europaea L. subsp. europaea), el cultivo arbóreo dominante en grandes áreas de la cuenca del Mediterráneo Occidental, con una producción de 20.872.788 toneladas en 2018. El aceite de oliva es apreciado en todo el mundo por su composición química, su valor nutracéutico y sus propiedades organolépticas. Como la mayoría de los cultivos, el olivo se enfrenta a importantes problemas y desafíos relacionados con las condiciones ambientales adversas, ya sean bióticas o abióticas, situación que puede empeorar en un escenario de cambio climático. Por lo tanto, comprender la base/mecanismo fisiológico y molecular de la adaptabilidad y resiliencia del olivo a las diferentes tensiones ambientales, especialmente a altas temperaturas, es crucial para predecir las consecuencias del calentamiento climático, implementar prácticas agrícolas, diseñar programas de mejoramiento genético clásicos o biotecnológicos y, en los últimos tiempos, aumentar su producción a fin de hacer frente a la demanda actual. En el presente trabajo se han estudiado tanto el efecto como las respuestas a un aumento de 4 °C por encima de la temperatura ambiente a nivel fenológico, fisiológico y molecular, -ómico, en un intento de integrar todos los resultados en la nueva dirección de la Biología de Sistemas. El uso de las técnicas de la -ómica (transcriptómica, proteómica y metabolómica) y su integración con estudios fisiológicos y fenológicos constituyen la gran novedad de este trabajo. Se ha solicitado un importante esfuerzo en la optimización de diferentes protocolos de perfiles de transcriptoma (aislamiento de ARN y qRT-PCR), proteoma (Orbitrap de escopeta nLC-MS/MS) y metaboloma (UHPLC-MS/MS Q-tof), junto con herramientas computacionales para la identificación y cuantificación de las diferentes biomoléculas, su clasificación funcional, análisis estadístico, clustering y networking. Tanto las herramientas de nuevo desarrollo como los conocimientos generados sentarán las bases de los estudios en curso y futuros para obtener una mejor comprensión de los mecanismos moleculares que median los fenotipos de interés (productivos, tolerantes a las señales ambientales, de valor nutracéutico y a la selección de genotipos más resistentes al calentamiento climático). El manuscrito ha sido organizado y presentado en nueve capítulos como a continuación se detallan: El capítulo I resume el “estado del arte” de los temas tratados en esta tesis, el sistema experimental, los objetivos, las hipótesis y las metodologías. Comienza con la descripción del olivo, como especie vegetal y cultivo, su importancia económica y cultural, problemas y retos relacionados con su cultivo. El capítulo continúa con la descripción y publicaciones sobre el estrés ambiental con énfasis en el calentamiento de las temperaturas y el denominado "Cambio Climático". Se discutirán las respuestas y los mecanismos de tolerancia al estrés de la temperatura y al calentamiento climático. Este capítulo finaliza con la descripción de la metodología empleada y la integración de los diferentes enfoques. En el capítulo II se presentan los objetivos generales y específicos de la Tesis. El capítulo III corresponde a dos artículos publicados sobre el efecto de calentamiento en la fenología del olivo: "Una aproximación a los efectos del calentamiento global sobre la floración y el cuajado de los olivos que crecen en condiciones de campo". y "Efectos del calentamiento global sobre el rendimiento y la maduración de los frutos del cultivo del olivo en condiciones de campo". Ambos fueron publicados en Scientia Horticulturae. Presenta alteraciones en los procesos de floración y maduración, causadas por un aumento térmico permanente de 4 °C por encima de la temperatura ambiente (AT) aplicada a olivos adultos (cv. `Picual´) en condiciones de campo. En este capítulo se han estudiado, analizado y analizado los datos relativos a las condiciones climáticas durante tres años. En general, se observó una floración y maduración más temprana y prolongada en condiciones más cálidas (AT+4 °C). El aumento de la temperatura redujo la cantidad de flores y frutos perfectos (bien desarrollados). Se observó una disminución de los parámetros químicos relacionados con la calidad del aceite (rendimiento graso, fenólicos y antocianos) en condiciones de aumento térmico. El capítulo IV corresponde a dos artículos publicados sobre el efecto de los diferentes regímenes de temperatura en las raíces y brotes de las plántulas de olivo (cv. `Arbequina´), un experimento de cámara de crecimiento vegetal: "Efecto de las altas temperaturas moderadas sobre el crecimiento vegetativo y la asignación de potasio en las plantas de olivo". y "Efectos del calentamiento de la raíz del olivo en el transporte de potasio y el crecimiento de las plantas" Ambos fueron publicados en Journal of Plant Physiology. Se evaluó el efecto de la diferencia térmica de 25 °C/37 °C, la exposición entre la raíz y la parte aérea sobre el crecimiento, y la absorción y el transporte de K+. En la raíz a 25 °C, brote a 37 °C, la absorción de potasio aumentó. 37 °C en ambos órganos inhibió el crecimiento, redujo la absorción de K+ y el transporte, y disminuyó el contenido de agua. Los capítulos V, VI y VII están dedicados a los diferentes enfoques -ómicos (proteómica, transcriptómica y metabolómica) empleados en el estudio molecular del efecto y las respuestas al calentamiento de la temperatura (+4 °C) en la pulpa del fruto en tres etapas de maduración: verde (0), rojo (llamado envero) (3) y epicarpio púrpura (4). Se ha encontrado una discrepancia entre la coloración de la piel, tradicionalmente observada como referencia para los estudios fenológicos y la recolección, y la fase de maduración interna de la pulpa de la aceituna y la composición química. La fase de envero fue la más afectada por el aumento de temperatura. En general, se observó una reducción en la abundancia de los principales compuestos relacionados con la calidad del aceite de oliva (lípidos, fenilpropanoides, flavonoides y terpenos) en condiciones de calentamiento. El capítulo VIII contiene una integración de todos los datos y una discusión general, centrada en los parámetros más relevantes desde el punto de vista agronómico, relacionados con la productividad de los cultivos, ya sea desde el punto de vista cuantitativo (producción de fruta) o cualitativo (composición química de la aceituna). A partir del presente estudio se presentan hipótesis predictivas sobre la situación del olivo y la producción en un futuro escenario de cambio climático, proponiendo prácticas de gestión alternativas para hacer frente a la demanda social de este cultivo. El capítulo IX corresponde a las conclusiones

Plants and Environment

Changing environmental condition and global population demands understanding the plant responses to hostile environment. Significant progress has been made over the past few decades through amalgamation of molecular breeding with non-conventional breeding. Understanding the cellular and molecular mechanisms to stress tolerance has received considerable scientific scrutiny because of the uniqueness of such processes to plant biology, and also its importance in the campaign "Freedom From Hunger". The main intention of this publication is to provide a state-of-the-art and up-to-date knowledge of recent developments in understanding of plant responses to major abiotic stresses, limitations and the current status of crop improvement. A better insight will help in taking a multidisciplinary approach to address the issues affecting plant development and performance under adverse conditions. I trust this book will act as a platform to excel in the field of stress biology

Biochemical and molecular mechanisms of salt stress tolerance in and Olea europaea

Tese de doutoramento em Ciências (área de especialização em Biologia)The current work focused in the research subject of membrane transport and plant - environment interactions and two plant models were the target of the studies: Olea europaea and Populus euphratica. Olive tree is an emblematic species and one of the most important fruit crops in the Mediterranean basin. The halophytic and salt and drought stress tolearant plant P. euphratica, which occurs naturally in semiarid areas, has recently been used as a model to study plant defense mechanisms against salt stress. In both plant species we aimed to contribute to the elucidation of the biochemical mechanisms involved in salt response, in particular those involving transmembrane transport steps of photoassimilates and tonoplast transport of protons and salt. The mechanism on how sodium is accumulated in the vacuole in response to salt in P. euphratica, and how salt stress may affect the generation and maintenance of a transmembrane proton gradient across the tonoplast were investigated. Biochemical data corroborated the involvement of Na+/H+ exchange activity in cell suspensions at the tonoplast level, whose activity increased 6-fold in NaCl-treated cells. Accordingly, confocal and epifluorescence microscopy analyses with the Na+-sensitive probe Sodium Green showed that suspension-cultured cells subjected to a salt pulse accumulated Na+ in the vacuole. In tonoplast vesicles the V-H+-PPase activity decreased with exposure to NaCl, in contrast to the observed sodium-induced increase in the activity of vacuolar H+-ATPase. The increase of both the transmembrane H+ gradient - generated by tonoplast proton pumps - and the Na+/H+ antiport activity in response to salt strongly suggested that Na+ accumulation into the vacuole contributes to salt tolerance in P. euphratica, in line with the confocal microscopy observations. In O. europaea, key biochemical and molecular steps involved in the partitioning of sugars and polyols, and how polyols may enhance salt and drought stress resistance were addressed at the protein activity and gene expression levels. Polyols are the reduced form of aldoses and ketoses, present in several species. In O. europaea leaves, mannitol was found to be the main soluble carbohydrate, followed by the monosaccharide glucose. Fructose was not detected, probably because it acted as precursor for mannitol biosynthesis. Transport experiments with [14C]mannitol showed that a polyol:H+ symport system operates in O. europaea heterotrophic cultured cells (Km = 1.3 mM). Subsequent work led to the cloning of a cDNA sequence of a mannitol carrier which was named OeMaT1 (O. europaea mannitol transporter 1). In parallel experiments, salt strongly repressed mannitol dehydrogenase activity, the first enzyme responsible for intracellular mannitol oxidation, and down-regulated OeMTD1 (O. europaea mannitol dehydrogenase 1) transcripts. This should allow for the intracellular accumulation of mannitol in order to compensate for the decrease of external water activity, thus conferring a response mechanism to salinity in O. europaea. Subsequent studies on the molecular mechanisms of glucose utilization by olive cells led to the cloning and functional characterization of the monosaccharide transporter OeMST2. Heterologous expression of this gene in Saccharomyces cerevisiae deficient in glucose transport restored its capacity to grow and to transport glucose. Transcript levels of OeMST2 increased during fruit maturation, confirming that OeMST2 catalyzes the membrane transport process of hexoses during sugar unloading in the fruits. In addition to this saturable energy dependent transport systems, in a variety of cell types, including plant cells, sugars, polyols and other solutes may be incorporated according to a diffusion-like kinetics, in spite of the real nature of this transport mechanism having been elusive. The measurement of [14C]glucose transport by cells and membrane vesicles in the presence of specific inhibitors, the measurement of activation energies of glucose uptake, among other biochemical approaches, led us to demonstrate that the low-affinity, high-capacity, diffusion-like glucose uptake in olive cells occurs through a channel-like structure whose transport capacity may be regulated by intracellular protonation and phosphorylation/dephosphorylation. The recent publication in Nature reporting the identification and functional characterization of a new class of sugar transporters, named SWEET, which are postulated to be involved in phloem transport and plant nectar production, further strengthened the involvement of low-affinity sugar facilitators in plants.O presente trabalho focou-se no tema do transporte transmembranar de solutos em plantas, em particular no estudo dos mecanismos de transporte envolvidos na interação das plantas com o ambiente. As espécies modelo Olea europaea e Populus euphratica foram os alvos destes estudos ao nível bioquímico e molecular. A oliveira é uma espécie emblemática desde tempos ancestrais e uma das árvores de fruto mais importantes na bacia Mediterrânica. P. euphratica é uma espécie arbórea resistente ao sal e à secura, presente naturalmente em zonas semiáridas, e tem sido usada recentemente como uma planta-modelo para o estudo dos mecanismos de resistência de plantas ao stress salino. No presente trabalho tentámos contribuir para a elucidação dos mecanismos bioquímicos envolvidos na resposta de ambas as plantas à elevada salinidade, mais concretamente aqueles que envolvem o transporte transmembranar de fotoassimilados e a compartimentação no vacúolo de protões e NaCl. Em particular, foram investigados os mecanismos de compartimentação de sódio no vacúolo de P. euphratica em resposta à elevada salinidade, bem como o efeito do stresse salino na geração e manutenção de um gradiente transmembranar de protões através do tonoplasto. Os resultados mostraram o envolvimento de um sistema de antiporte do tipo Na+/H+ ao nível do tonoplasto de culturas celulares de P. euphratica, cuja actividade aumentou significativamente em células tratadas com NaCl. Estudos de microscopia confocal e de fluorescência com a sonda fluorescente Sodium Green mostraram que o NaCl se acumula no vacúolo quando as células são expostas a um pulso de sal. Em vesículas de tonoplasto purificadas de células expostas ao sal observou-se uma diminuição da actividade da bomba de protões vacuolar V-H+-PPase, contrastando com o aumento da actividade da V-H+-ATPase. No seu conjunto, os resultados sugerem que o aumento do gradiente transmembranar de H+ gerado pelas bombas de protões do tonoplasto, bem como o aumento da atividade do sistema de antiporte Na+/H+, contribuem para a acumulação de sódio no vacúolo em P. euphratica em resposta ao sal. Estas observações corroboram os resultados de microscopia confocal que mostraram a compartimentação de sódio no vacúolo. Em O. europaea foram estudados ao nível bioquímico e molecular transportadores membranares e enzimas envolvidos no metabolismo de açúcares e de polióis, no sentido de clarificar o papel dos fotoassimilados nos mecanismos de resposta à salinidade e à secura. Os polióis são formas reduzidas de aldoses e cetoses, presentes em mais de 100 espécies de plantas. Os resultados mostraram que o poliol manitol consiste no principal hidrato de carbono solúvel em folhas de O. europaea, seguido do monossacarídeo glucose. A frutose não foi detectada, provavelmente por ser utilizada como precursor biossintético de manitol. Experiências com [14C]manitol demonstraram o envolvimento de um sistema de transporte do tipo simporte poliol:H+ em culturas celulares heterotróficas de O. europaea (Km = 1.3 mM). Em paralelo, foi clonada a sequência de cDNA de um transportador de manitol, denominado OeMaT1 (O. europaea mannitol transporter 1). A adição de um pulso de sal a culturas celulares reprimiu a actividade da manitol desidrogenase, enzima responsável pelo primeiro passo de oxidação intracelular do manitol, e inibiu a transcrição do gene OeMTD1 (O. europaea mannitol dehydrogenase 1). Este mecanismo de regulação do transporte e metabolismo intracelular deve contribuir para a acumulação intracelular de manitol de modo a compensar a diminuição da actividade da água no espaço extracelular, constituindo um mecanismo de resposta à salinidade em O. europaea. Estudos subsequentes sobre os mecanismos moleculares de utilização de glucose em culturas celulares de oliveira permitiram a clonagem e caracterização funcional do transportador de monossacarídeos OeMST2. A expressão heteróloga deste gene numa estirpe mutante de Saccharomyces cerevisiae deficiente no transporte de glucose restaurou a sua capacidade de crescer em e de transportar glucose. Ao nível da planta foi observado que os níveis de transcritos do gene OeMST2 aumentam durante a maturação da azeitona, sugerindo que o OeMST2 está envolvido no descarregamento de açúcares do floema para o fruto. Diversos estudos desenvolvidos numa ampla variedade de modelos celulares têm mostrado que os açúcares, polióis e outros solutos podem ser incorporados de acordo com uma cinética de primeira ordem (do tipo difusional), para além dos mecanismos saturáveis, dependentes de energia, como os descritos neste trabalho para os transportadores de polóis e monossacarídeos. Contudo, a natureza bioquímica e molecular destes mecanismos não saturáveis permanece ainda pouco esclarecida. No presente trabalho desenvolvemos experiências de transporte com substratos radioativos em células e vesículas de membrana plasmática na presença de inibidores específicos, no sentido de procurar compreender a natureza das cinéticas de primeira ordem observadas em culturas celulares de oliveira. No seu conjunto, os resultados sugeriram o envolvimento de proteínas do tipo canal cuja capacidade de transporte pode ser regulada por protonação intracelular e fosforilação/desfosforilação. A publicação na prestigiada revista Nature sobre a recente identificação e caracterização funcional de uma nova classe de transportadores de açúcares denominada SWEET mostrou que permeases de baixa afinidade podem estar envolvidas no carregamento e descarregamento do floema

Molecular control to salt tolerance mechanisms of woody plants: Recent achievements and perspectives

Key message: Woody plants have salt-tolerant mechanisms similar to those developed by non-woody plants. Among others, compartmentalization of ions, production of compatible solutes, synthesis of specific proteins and metabolites, and induction of transcriptional factors are the most relevant. Woody plant-associated microbial interactions as well as naturally stress-adapted trees are resources that deserve to be deepened to fully understand the tolerance mechanisms. Context: The high variability of salinity responses found in woody plants implies a high potentiality for germplasm selection and breeding. Salt tolerance mechanisms of plants are regulated by numerous genes, which control ion homeostasis, production of compatible solutes and specific proteins, and activation or repression of specific transcription factors. Despite the fact that numerous studies have been done on herbaceous model plants, knowledge about salt tolerance mechanisms in woody plants is still scarce. Aims: The present review critically evaluates molecular control of salt tolerance mechanisms of woody plants, focusing on the regulation and compartmentalization of ions, production of compatible solutes, activation of transcription factors, and differential expression of stress response-related proteins, including omics-based approaches and the role of plant-microbial interactions. The potential identification of genes from naturally stress-adapted woody plants and the integration of the massive omics data are also discussed. Conclusion: In woody plants, salt tolerance mechanisms seem not to diverge to those identified in non-woody plants. More comparative studies between woody and non-woody salt tolerance plants will be relevant to identify potential molecular mechanisms specifically developed for wood plants. In this sense, the activation of metabolic pathways and molecular networks by novel genetic engineering techniques is key to establish strategies to improve the salt tolerance in woody plant species and to contribute to more sustainable agricultural and forestry systems.Fil: Llanes, Analia Susana. Universidad Nacional de Rio Cuarto. Facultad de Cs.exactas Fisicoquimicas y Naturales. Instituto de Investigaciones Agrobiotecnologicas. - Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Cordoba. Instituto de Investigaciones Agrobiotecnologicas.; Argentina. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Ciencias Naturales. Laboratorio de Fisiología Vegetal y de la Interacción Planta-microorganismo; ArgentinaFil: Palchetti, Maria Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Vilo, Claudia. Universidad de La Serena; Chile. Universidad de Antofagasta; ChileFil: Ibáñez, Cristian. Universidad de La Serena; Chil

Genetic and genomic approaches to characterize crop varieties

La exposición a contaminantes es uno de los mayores retos a los que debe hacer frente el mundo actual debido a los riesgos que implica, tanto para los seres humanos como para la salud de los ecosistemas en general. Por tanto, se necesitan potentes herramientas analíticas, fundamentalmente ligadas a las respuestas biológicas que desencadena la exposición a contaminantes, para evaluar la calidad ambiental. Las metodologías ómicas, junto con el uso de biomarcadores convencionales, han demostrado tener una gran utilidad en este sentido, en estudios que usan al ratón moruno Musspretus como organismo bioindicador de contaminación ambiental en ecosistemas terrestres del Parque Nacional de Doñana. La complejidad de los ecosistemas y los procesos de sinergia/antagonismo existentes entre los contaminantes dificultan el establecimiento de un vínculo claro e inequívoco entre un contaminante concreto y las respuestas biológicas que desencadena. A pesar de estos inconvenientes, este tipo de estudios es muy necesario ya que integra todas las variables presentes en un determinado ecosistema y su interacción con los organismos estudiados. Para establecer, de una forma clara, las relaciones causa-efecto es necesario llevar a cabo experimentos de exposición controlada en laboratorio a contaminantes individuales, de forma que se controlen todas las variables que puedan afectar a las respuestas biológicas desencadenadas, incluyendo las posibles interacciones de los tóxicos con la dieta. De esta manera, es posible establecer los mecanismos de acción mediante los cuales los contaminantes ejercen su toxicidad sobre los organismos. En este trabajo se ha intentado dilucidar la base molecular subyacente a las respuestas biológicas inducidas en el ratón M. spretus por la exposición al compuesto tóxico p,p’-DDE (diclorodifeniltricloroetileno), un derivado metabólico del DDT (diclorodifeniltricloroetano), pesticida organoclorado prohibido en la década de los 70 por su elevada toxicidad y persistencia en el medioambiente. Para ello, se han utilizado diversas metodologías ómicas, semi-ómicas y biomarcadores convencionales a distintos niveles moleculares (transcritos, proteínas y metabolitos) usando como modelo ratones M. spretus criados en cautividad. Estos estudios se han completado con ensayos de actividad enzimática y estudios histopatológicos de manera que se permita una visión más completa de estos procesos.Exposure to contaminants is one of the main challenges in the world, due to their risks for humans and ecosystems. Powerful analytical tools, mainly linked to the detection of biological responses elicited by the exposure to such contaminants are needed to assess the environmental quality within ecosystems. In this context, omic methodologies, in conjunction with conventional biomarkers, have proven to be very useful as shown in previous studies using Mus spretus mice as environmental pollution bioindicators in terrestrial ecosystems of Doñana National Park, Spain. The complexity of the ecosystems and the synergic/antagonist processes among contaminants make it difficult to set a clear, unambiguous link between a given contaminant and the biological responses elicited. Despite this issue, this type of studies is essential to integrate all variables present in the ecosystem and their interactions with the organisms studied. To clearly establish the cause-effect relationship, it is necessary to carry out experiments of controlled exposure to contaminants under laboratory conditions, where all the variables can be monitored, including the posible interactions of the toxicants with the diet. This way, it is possible to identify the action mechanisms by which the contaminants exert their toxicity in the organisms. This work aims to elucidate the molecular basis underlying the biological responses induced in M. spretus mice by exposure to p,p’-DDE (dichlorodipheniltrichloroethilene). This is a metabolic byproduct of the organochlorine pesticide DDT (dichlorodipheniltrichloroethane), banned in the 70s due to its toxicity and persistence in the environment. For this purpose, omic, semi-omic and conventional biomarker approaches have been used at different molecular levels (transcripts, proteins and metabolites) using raisedin- captivity M. spretus as model. To convey a broader view of the processes, these studies have been complemented with enzymatic activity assays and histopathological analysis

Modulators of Oxidative Stress

The book starts with the editors’ preface summarizing the contributions of each of the following thirteen articles dealing with chemical and pharmacological aspects of the molecular modulators of the oxidative stress in regard to various therapeutic approaches in cardiovascular and neurodegenerative diseases, cancer, and diabetes. The seven articles present data from original research studies enlightening the roles and mechanisms of action of small molecular weight compounds (natural and synthetic; ascorbic acid/vitamin C, deferoxamine, N-acetylcysteine, MitoVitE, α-tocopherol, trolox, and ezetimibe) or proteins (SIRT3) in modulation of oxidative stress. In the six review papers, the authors present and discuss the possible therapeutic potential of novel approaches and compounds that are promising and deserve further investigation, in modulation of oxidative stress

Olive Oil

Olive Oil - New Perspectives and Applications is a collection of reviewed and relevant research chapters, offering a comprehensive overview of recent developments in the field of agricultural, medical, and biological sciences. The book comprises single chapters authored by various researchers and edited by an expert active in the olive oil research area. All chapters are complete in themselves but united under a common research study topic. This publication aims at providing a thorough overview of the latest research efforts by international authors on olive oil and opening new possible research paths for further novel developments

Ethylene and Nitric Oxide Involvement in the Regulation of Fe and P Deficiency Responses in Dicotyledonous Plants

Iron (Fe) and phosphorus (P) are two essential elements for plant growth. Both elements are abundant in soils but with poor availability for plants, which favor their acquisition by developing morphological and physiological responses in their roots. Although the regulation of the genes related to these responses is not totally known, ethylene (ET) and nitric oxide (NO) have been involved in the activation of both Fe-related and P-related genes. The common involvement of ET and NO suggests that they must act in conjunction with other specific signals, more closely related to each deficiency. Among the specific signals involved in the regulation of Fe- or P-related genes have been proposed Fe-peptides (or Fe ion itself) and microRNAs, like miR399 (P), moving through the phloem. These Fe- or P-related phloem signals could interact with ET/NO and confer specificity to the responses to each deficiency, avoiding the induction of the specific responses when ET/NO increase due to other nutrient deficiencies or stresses. Besides the specificity conferred by these signals, ET itself could confer specificity to the responses to Fe- or P-deficiency by acting through different signaling pathways in each case. Given the above considerations, there are preliminary results suggesting that ET could regulate different nutrient responses by acting both in conjunction with other signals and through different signaling pathways. Because of the close relationship among these two elements, a better knowledge of the physiological and molecular basis of their interaction is necessary to improve their nutrition and to avoid the problems associated with their misuse. As examples of this interaction, it is known that Fe chlorosis can be induced, under certain circumstances, by a P over- fertilization. On the other hand, Fe oxides can have a role in the immobilization of P in soils. Qualitative and quantitative assessment of the dynamic of known Fe- and P-related genes expression, selected ad hoc and involved in each of these deficiencies, would allow us to get a profound knowledge of the processes that regulate the responses to both deficiencies. The better knowledge of the regulation by ET of the responses to these deficiencies is necessary to properly understand the interactions between Fe and P. This will allow the obtention of more efficient varieties in the absorption of P and Fe, and the use of more rational management techniques for P and Fe fertilization. This will contribute to minimize the environmental impacts caused by the use of P and Fe fertilizers (Fe chelates) in agriculture and to adjust the costs for farmers, due to the high prices and/or scarcity of Fe and P fertilizers. This review aims to summarize the latest advances in the knowledge about Fe and P deficiency responses, analyzing the similarities and differences among them and considering the interactions among their main regulators, including some hormones (ethylene) and signaling substances (NO and GSNO) as well as other P- and Fe-related signals

Foodomics-Based Approaches Shed Light on the Potential Protective Effects of Polyphenols in Inflammatory Bowel Disease

nflammatory bowel disease (IBD) is a chronic and progressive inflammatory disorder affecting the gastrointestinal tract (GT) caused by a wide range of genetic, microbial, and environ- mental factors. IBD is characterized by chronic inflammation and decreased gut microbial diversity, dysbiosis, with a lower number of beneficial bacteria and a concomitant increase in pathogenic species. It is well known that dysbiosis is closely related to the induction of inflammation and oxidative stress, the latter caused by an imbalance between reactive oxygen species (ROS) production and cellular antioxidant capacity, leading to cellular ROS accumulation. ROS are responsible for intestinal epithelium oxidative damage and the increased intestinal permeability found in IBD patients, and their reduction could represent a potential therapeutic strategy to limit IBD progression and alleviate its symptoms. Recent evidence has highlighted that dietary polyphenols, the natural antioxidants, can maintain redox equilibrium in the GT, preventing gut dysbiosis, intestinal epithelium damage, and radical inflammatory responses. Here, we suggest that the relatively new foodomics approaches, together with new technologies for promoting the antioxidative properties of dietary polyphenols, including novel delivery systems, chemical modifications, and combination strategies, may provide critical insights to determine the clinical value of polyphenols for IBD therapy and a comprehensive perspective for implementing natural antioxidants as potential IBD candidate treatment