Browsing by Author "Royuela Hernando, Mercedes"
Now showing 1 - 20 of 37
Results Per Page
Sort Options
Publication Open Access An aerated axenic hydroponic system for the application of root treatments: exogenous pyruvate as a practical case(BioMed Central, 2018) Gil Monreal, Miriam; Fernández Escalada, Manuel; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Ciencias; Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaBackground: Hydroponic systems are a convenient platform for plant cultivation when treatments are applied to the roots because they provide precise control of the composition of the growth medium, ensuring the availability of different compounds. A problem arises when axenic conditions are needed but the treatment of choice (exogenous organic acids or sugars) promote the growth of unwanted microorganisms. Moreover, axenic conditions are usually applied in liquid and semi-liquid growing systems, where oxygen availability can be compromised, if no aeration is provided. Results: The driver for the development of this hydroponic system was the application of the organic acid pyruvate to the roots of plants grown under aerated axenic conditions. No contamination was detected in the nutrient solution, even after the addition of pyruvate. The system was validated in pea plants treated with either pyruvate or herbicides inhibiting amino acid biosynthesis. The effects on ethanol fermentation were compared by analysing the enzymatic activity, protein content and transcriptional levels in plants treated with either pyruvate or herbicides. Conclusions: The developed system enables the study of the exogenous application of organic acids in the nutrient solution under axenic conditions and without oxygen limitation. This system allows the study of the effect of any type of treatments applied to roots under aerated axenic hydroponic systems at physiological and molecular levels. The role of pyruvate in the induction of fermentation by herbicides cannot be simply explained by an increase in substrate availability.Publication Embargo Alterations of oxidative status induced by amino acid synthesis-inhibiting herbicides in sensitive and target-site resistantpopulations of Amaranthus palmeri(2023) Eceiza, Mikel Vicente; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Ciencias; Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, UPNA20-6138Entre los métodos de control de malas hierbas, los herbicidas aparecen como métodos rápidos y efectivos, y conforman el método más utilizado para el control de malas hierbas. Dos de los grupos de herbicidas más importantes son los inhibidores de la 5-enolpiruvilsiquimato-3-fosfatosintasa (EPSPS) y la acetolactato sintasa (ALS). El único inhibidor de la EPSPS que se conoce es el glifosato, el herbicida más utilizado a nivel mundial; mientras, los inhibidores de la ALS incluyen numerosas sustancias activas. La EPSPS y la ALS son enzimas importantes que forman parte de la biosíntesis de aminoácidos aromáticos y aminoácidos ramificados, respectivamente. Cómo el glifosato o los inhibidores de la ALS bloquean estas rutas de biosíntesis de aminoácidos está estudiado a fondo, pero la secuencia de eventos fisiológicos que ocurre entre la aplicación del herbicida y la muerte de la planta no se conoce del todo. Los efectos fisiológicos establecidos hoy en día para ambos grupos de herbicidas son bastante parecidos (a pesar de que las enzimas diana sean diferentes). Algunos estudios muestran que entre estos efectos fisiológicos provocados por el glifosato y los inhibidores de la ALS está el estrés oxidativo, pero su origen (si está ligado a la inhibición de la EPSPS o ALS o es un efecto secundario aparte) y la importancia del daño oxidativo y las alteraciones en los sistemas antioxidantes (como el glutatión) inducidos en la cascada fisiológica que termina con la muerte de la planta no se conocen. En este contexto, el objetivo principal de esta tesis fue evaluar el papel del estrés oxidativo y el metabolismo del glutatión en el modo de acción del glifosato y los inhibidores de la ALS, tratando de elucidar si está relacionado con el mecanismo de acción de ambos grupos de herbicidas. Se crecieron cuatro poblaciones de la mala hierba Amaranthus palmeri S. Wats.: sensible a glifosato (GFS), resistente a glifosato (GFR), sensible a inhibidores de la ALS (AIS) y resistente a inhibidores de la ALS (AIR). En las dos poblaciones resistentes, la resistencia venía dada por mecanismos target-site (TSR). Las poblaciones GFS y GFR fueron tratadas con diferentes dosis de glifosato, mientras que las poblaciones AIS y AIR fueron tratadas con dosis diferentes de nicosulfuron (una sulfonilurea, inhibidor de la ALS). Se realizaron varios análisis fisiológicos y se compararon, en las hojas de estas poblaciones: acumulación de especies reactivas de oxígeno (ROS), daño oxidativo, sistemas antioxidantes (con especial énfasis en el metabolismo del glutatión) y señalización hormonal.Casi no hubo diferencias entre las plantas sensibles y resistentes no tratadas en los parámetros analizados, mostrando que la TSR no viene acompañada por un estado oxidativo basal diferente. Las plantas GFS y AIS tratadas murieron con todas las dosis de glifosato o nicosulfuron; en cambio, todas las plantas resistentes sobrevivieron. Las plantas GFS y AIS tratadas mostraron una producción y acumulación de ROS mayor que las no tratadas, un efecto que no ocurrió en las resistentes. Esta sobreproducción de ROS en las plantas sensibles tratadas provocó una peroxidación lipídica proporcional a la dosis de herbicida, un indicador inequívoco de estrés oxidativo, y cambios en los sistemas antioxidantes. Solo las plantas sensibles de las dos poblaciones mostraron un incremento de la actividad y expresión génica de la glutatión Stransferasa (GST), indicando la implicación del glutatión como antioxidante. La presencia de estrés oxidativo solo en las plantas sensibles tratadas con glifosato o nicosulfuron y su ausencia en las plantas TSR indica que el estrés oxidativo está relacionado con la inhibición de la EPSPS o la ALS, aunque los mecanismos exactos permanecen sin esclarecer. El daño oxidativo se probó como otro efecto común de ambos herbicidas, pero fue demasiado leve para provocar por sí solo la muerte de la planta. Por otra parte, los dos herbicidas alteraron el perfil hormonal, pero indujeron hormonas diferentes. Así, los cambios observados en el perfil hormonal muestran un efecto diferente de la acción herbicida. Los mecanismos que provocan estos cambios hormonales siguen siendo desconocidos, pero la inducción de ciertas hormonas puede estar relacionada con el estrés oxidativo.Publication Open Access Both foliar and residual applications of herbicides that inhibit amino acid biosynthesis induce alternative respiration and aerobic fermentation in pea roots(Wiley, 2016) Armendáriz García, Óscar; Gil Monreal, Miriam; Zulet González, Amaia; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaThe objective of this work was to ascertain whether there is a general pattern of carbon allocation and utilisation in plants following herbicide supply, independent of the site of application: sprayed on leaves or supplied to nutrient solution. The herbicides studied were the amino acid biosynthesis-inhibiting herbicides (ABIH): glyphosate, an inhibitor of aromatic amino acid biosynthesis, and imazamox, an inhibitor of branched-chain amino acid biosynthesis. All treated plants showed impaired carbon metabolism; carbohydrate accumulation was detected in both leaves and roots of the treated plants. The accumulation in roots was due to lack of use of available sugars as growth was arrested, which elicited soluble carbohydrate accumulation in the leaves due to a decrease in sink strength. Under aerobic conditions, ethanol fermentative metabolism was enhanced in roots of the treated plants. This fermentative response was not related to a change in total respiration rates or cytochrome respiratory capacity, but an increase in alternative oxidase capacity was detected. Pyruvate accumulation was detected after most of the herbicide treatments. These results demonstrate that both ABIH induce the less-efficient, ATP-producing pathways, namely fermentation and alternative respiration, by increasing the key metabolite, pyruvate. The plant response was similar not only for the two ABIH but also after foliar or residual application.Publication Open Access Branched-chain amino acid biosynthesis inhibitors: herbicide efficacy is associated with an induced carbon–nitrogen imbalance(Elsevier, 2013) Zabalza Aznárez, Ana; Zulet González, Amaia; Igal Díaz de Cerio, María; Gil Monreal, Miriam; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen ZientziakAcetolactate synthase (ALS; EC 4.1.3.18) and ketol-acid reductoisomerase (KARI; EC 1.1.1.86) are two consecutive enzymes in the biosynthesis of branched-chain amino acids. Several commercial herbicides inhibit ALS as their primary site of action. KARI has also attracted attention as a potential target for herbicides. Although potent and selective inhibitors of KARI have been discovered, these inhibitors display less herbicidal activity than ALS-inhibiting herbicides. To obtain a better understanding of these findings, we have compared the physiological effects induced in pea plants after KARI or ALS inhibition. Although, both types of inhibitors induce growth arrest and photosynthesis inhibition, plant death occurs more rapidly under ALS inhibition than KARI inhibition. Carbohydrates accumulated in the leaves and roots following treatments with both inhibitors. The carbohydrate accumulation in the leaves occurred as a consequence of a decrease in sink strength. In contrast, the free amino acid content was only affected through ALS inhibition. These results indicate that although KARI and ALS inhibition block the same biosynthetic pathway and exert common effects on carbon metabolism, nitrogen metabolism is more affected via ALS than KARI inhibition. Thus, metabolic alterations in nitrogen metabolism induced through ALS inhibitors might contribute to the increased efficacy of these chemicals as herbicides.Publication Open Access Characterization of the Amaranthus palmeri physiological response to glyphosate in susceptible and resistant populations(American Chemical Society, 2016) Fernández Escalada, Manuel; Gil Monreal, Miriam; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaThe herbicide glyphosate inhibits the plant enzyme 5-enolpyruvylshikimate3-phosphate synthase (EPSPS) in the aromatic amino acid (AAA) biosynthetic pathway. The physiologies of an Amaranthus palmeri population exhibiting resistance to glyphosate by EPSPS gene amplification (NC-R) and a susceptible population (NC-S) were compared. The EPSPS copy number of NC-R plants was 47.5-fold the copy number of NC-S plants. Although the amounts of EPSPS protein and activity were higher in NC-R plants than in NC-S plants, the AAA concentrations were similar. The increases in total free amino acid and in AAA contents induced by glyphosate were more evident in NC-S plants. In both populations, the EPSPS protein increased after glyphosate exposure, suggesting regulation of gene expression. EPSPS activity seems tightly controlled in vivo. Carbohydrate accumulation and a slight induction of ethanol fermentation were detected in both populations.Publication Open Access Cysteine proteases are activated in sensitive Amaranthus palmeri populations upon treatment with herbicides inhibiting amino acid biosynthesis(Wiley, 2023) Barco Antoñanzas, María; Font Farre, María; Eceiza, Mikel Vicente; Gil Monreal, Miriam; Van der Hoorn, Reiner; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaThe herbicides glyphosate and pyrithiobac inhibit the enzyme 5-enolpyruvylshikimate3-phosphate synthase (EPSPS) in the aromatic amino acid biosynthetic pathway and acetolactate synthase (ALS) in the branched-chain amino acid biosynthetic pathway, respectively. Here we characterise the protease activity profiles of a sensitive (S), a glyphosate-resistant (GR) and a multiple-resistant (MR) population of Amaranthus palmeri in response to glyphosate and pyrithiobac. Amino acid accumulation and cysteine protease activities were induced with both herbicides in the S population and with pyrithiobac in the GR population, suggesting that the increase in cysteine proteases is responsible for the increased degradation of the available proteins and the observed increase in free amino acids. Herbicides did not induce any changes in the proteolytic activities in the populations with target-site resistance, indicating that this effect was only induced in sensitive plants.Publication Open Access Efecto del glifosato en la expresión génica de la ruta del siquimato en Amaranthus palmeri(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2017) Fernández Escalada, Manuel; Zulet González, Ainhoa; Gil Monreal, Miriam; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaEste estudio ha abordado el efecto del herbicida glifosato en la regulación de la ruta del siquimato en la especie Amaranthus palmeri, comparando una población sensible a glifosato con una población resistente. Se han determinado, mediante PCR cuantitativa (qPCR), el efecto de dos dosis de glifosato sobre la expresión génica de los enzimas de la ruta del siquimato. También se ha determinado la actividad enzimática de los enzimas post-corismato antranilato sintasa (AS) y corismato mutasa (CM). Se ha podido observar que el gran aumento en el número de copias génicas del enzima EPSPS en la población resistente no tiene un efecto pleiotrópico basal en la expresión de los demás genes de esta ruta de síntesis de los aminoácidos aromáticos. Tras el tratamiento con glifosato, se observó, en ambas poblaciones, una inducción general de la expresión de los genes de la ruta del siquimato, dependiente de la dosis de glifosato. Es destacable que la aplicación del glifosato provoca el aumento de la expresión y actividad AS, y la tendencia opuesta en expresión CM, lo que conlleva un flujo preferente hacia la formación de triptófano en lugar de tirosina y fenilalanina.Publication Open Access Efecto del glifosato y de diferentes intermediarios metabólicos en la regulación de la ruta del siquimato(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2017) Zulet González, Ainhoa; Fernández Escalada, Manuel; Gil Monreal, Miriam; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaEn este estudio se han abordado los mecanismos de regulación de la ruta del siquimato en la biosíntesis de los aminoácidos aromáticos. Para ello se han comparado el contenido de siquimato y de los enzimas EPSPS (5-enolpiruvilsiquimato- 3-fosfato sintasa) y DAHPS (3-Deoxi-D-arabino-heptulosonato-7-fosfato sintasa, enzima de entrada a la ruta) en hojas de dos poblaciones de Amaranthus palmeri (una sensible y otra resistente al glifosato) incubadas con glifosato y/o con diferentes intermediarios de la ruta (siquimato, quinato, fosfoenolpiruvato, y aminoácidos aromáticos). En ambas poblaciones se dieron similares efectos regulatorios de los intermediarios, evidenciando que la sobreexpresión de EPSPS no modifica de manera significativa la regulación general de la ruta. El enzima DAHPS se confirma como enzima clave en la regulación y en la respuesta al glifosato; su síntesis es inhibida por los productos finales de la ruta y por el quinato, y su regulación puede explicar la acumulación de siquimato tras la aplicación de glifosato.Publication Open Access Effects of EPSPS Copy Number Variation (CNV) and glyphosate application on the aromatic and branched chain amino acid synthesis pathways in Amaranthus palmeri(Frontiers Media, 2017) Fernández Escalada, Manuel; Zulet González, Ainhoa; Gil Monreal, Miriam; Zabalza Aznárez, Ana; Ravet, Karl; Gaines, Todd; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaA key enzyme of the shikimate pathway, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS; EC 2.5.1.19), is the known target of the widely used herbicide glyphosate. Glyphosate resistance in Amaranthus palmeri, one of the most troublesome weeds in agriculture, has evolved through increased EPSPS gene copy number. The aim of this work was to study the pleiotropic effects of (i) EPSPS increased transcript abundance due to gene copy number variation (CNV) and of (ii) glyphosate application on the aromatic amino acid (AAA) and branched chain amino acid (BCAA) synthesis pathways. Hydroponically grown glyphosate sensitive (GS) and glyphosate resistant (GR) plants were treated with glyphosate 3 days after treatment. In absence of glyphosate treatment, high EPSPS gene copy number had only a subtle effect on transcriptional regulation of AAA and BCAA pathway genes. In contrast, glyphosate treatment provoked a general accumulation of the transcripts corresponding to genes of the AAA pathway leading to synthesis of chorismate in both GS and GR. After chorismate, anthranilate synthase transcript abundance was higher while chorismate mutase transcription showed a small decrease in GR and remained stable in GS, suggesting a regulatory branch point in the pathway that favors synthesis toward tryptophan over phenylalanine and tyrosine after glyphosate treatment. This was confirmed by studying enzyme activities in vitro and amino acid analysis. Importantly, this upregulation was glyphosate dose dependent and was observed similarly in both GS and GR populations. Glyphosate treatment also had a slight effect on the expression of BCAA genes but no general effect on the pathway could be observed. Taken together, our observations suggest that the high CNV of EPSPS in A. palmeri GR populations has no major pleiotropic effect on the expression of AAA biosynthetic genes, even in response to glyphosate treatment. This finding supports the idea that the fitness cost associated with EPSPS CNV in A. palmeri may be limited.Publication Open Access Enhancement of glyphosate efficacy on Amaranthus palmeri by exogenous quinate application(Elsevier, 2019) Zulet González, Ainhoa; Fernández Escalada, Manuel; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaGlyphosate is a widely used herbicide targeting the enzyme 5-enolpyruvylshikimate 3-phosphate synthase (EPSPS) in the aromatic amino acid biosynthesis pathway (shikimate pathway) and provoking accumulation of quinate, a secondary metabolite synthesized through a side branch of this pathway. The objective of this work was to evaluate whether the efficacy of glyphosate activity in Amaranthus palmeri is enhanced by quinate application one day after herbicide treatment. To this end, one glyphosate-sensitive and one glyphosate-resistant (due to EPSPS gene amplification) population of A. palmeri were used. The 3- day time course study of the quinate treatment alone showed quinate, Tyr and Phe accumulation in both populations. When the herbicide was applied alone at 0.25× the recommended dose, no phytotoxicity or glyphosate effects were detected in the sensitive population 3 days after treatment, but the combined treatment with quinate was lethal, and markers of herbicide activity at the amino acid level could be detected. In the resistant population, an important metabolic perturbation in the flux of the shikimate pathway was detected in the combined treatment. These results raise the possibility of the joint application of quinate and glyphosate to enhance glyphosate efficacy while lowering doses in the sensitive population.Publication Open Access ERF-VII transcription factors induce ethanol fermentation in response to amino acid biosynthesis-inhibiting herbicides(Oxford University Press, 2019) Gil Monreal, Miriam; Giuntoli, Beatrice; Zabalza Aznárez, Ana; Licausi, Francesco; Royuela Hernando, Mercedes; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaHerbicides inhibiting either aromatic or branched-chain amino acid biosynthesis trigger similar physiological responses in plants, despite their different mechanism of action. Both types of herbicides are known to activate ethanol fermentation by inducing the expression of fermentative genes; however, the mechanism of such transcriptional regulation has not been investigated so far. In plants exposed to low-oxygen conditions, ethanol fermentation is transcriptionally controlled by the ethylene response factors-VII (ERF-VIIs), whose stability is controlled in an oxygen-dependent manner by the Cys-Arg branch of the N-degron pathway. In this study, we investigated the role of ERF-VIIs in the regulation of the ethanol fermentation pathway in herbicide-treated Arabidopsis plants grown under aerobic conditions. Our results demonstrate that these transcriptional regulators are stabilized in response to herbicide treatment and are required for ethanol fermentation in these conditions. We also observed that mutants with reduced fermentative potential exhibit higher sensitivity to herbicide treatments, thus revealing the existence of a mechanism that mimics oxygen deprivation to activate metabolic pathways that enhance herbicide tolerance. We speculate that this signaling pathway may represent a potential target in agriculture to affect tolerance to herbicides that inhibit amino acid biosynthesis.Publication Restricted Estudio de posible resistencia de vallico (Lolium multiflorum L) a diferentes materias activas(2006) Álvarez Chivite, Raquel; Royuela Hernando, Mercedes; Escuela Técnica Superior de Ingenieros Agrónomos; Nekazaritza Ingeniarien Goi Mailako Eskola Teknikoa; Ciencias del Medio Natural; Natura Ingurunearen ZientziakPublication Open Access Fermentation and alternative oxidase contribute to the action of amino acid biosynthesis-inhibiting herbicides(Elsevier, 2015) Zulet González, Amaia; Gil Monreal, Miriam; Zabalza Aznárez, Ana; Dongen, Joost T. van; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaAcetolactate synthase inhibitors (ALS-inhibitors) and glyphosate (GLP) are two classes of herbicide that act by the specific inhibition of an enzyme in the biosynthetic pathway of branched-chain or aromatic amino acids, respectively. The physiological effects that are detected after application of these two classes of herbicides are not fully understood in relation to the primary biochemical target inhibition, although they have been well documented. Interestingly, the two herbicides’ toxicity includes some common physiological effects suggesting that they kill the treated plants by a similar pattern despite targeting different enzymes. The induction of aerobic ethanol fermentation and alternative oxidase (AOX) are two examples of these common effects. The objective of this work was to gain further insight into the role of fermentation and AOX induction in the toxic consequences of ALS-inhibitors and GLP. For this, Arabidopsis T-DNA knockout mutants of alcohol dehydrogenase (ADH) 1 and AOX1a were used. The results found in wild-type indicate that both GLP and ALS-inhibitors reduce ATP production by inducing fermentation and alternative respiration. The main physiological effects in the process of herbicide activity upon treated plants were accumulation of carbohydrates and total free amino acids. The effects of the herbicides on these parameters were less pronounced in mutants compared to wild-type plants. The role of fermentation and AOX regarding pyruvate availability is also discussed.Publication Open Access Fisiología de la resistencia múltiple a los herbicidas glifosato e inhibidores de acetolactato sintasa en amaranthus palmeri(2022) Barco Antoñanzas, María; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias; Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaEste trabajo se plantea en el marco de la necesidad de profundizar en el conocimiento de la fisiología de las plantas resistentes a herbicidas, lo que requiere, no solo identificar y dilucidar las bases moleculares de las resistencias, sino también abordar el plano fisiológico, analizando la respuesta a nivel de metabolismo a la aplicación de estos herbicidas. La utilización de plantas que ya han desarrollado resistencia múltiple a estos herbicidas es una buena herramienta para desarrollar métodos para su detección, aportar claves para su manejo y prevenir su aparición. Además, el establecimiento de la acción herbicida en plantas resistentes permite comprender cómo y por qué son letales estos herbicidas (conocimiento del modo de acción herbicida), ayudando a que su utilización sea más racional y sostenible. Se ha profundizado en la fisiología de una población de A. palmeri con resistencia múltiple (RM) a los herbicidas glifosato y piritiobac (inhibidor de ALS) y en los efectos fisiológicos de estos herbicidas. Para ello, se han caracterizado a nivel molecular los mecanismos de resistencia, y se ha abordado la acción herbicida estudiando la respuesta fisiológica a diferentes niveles tras el tratamiento con glifosato, piritiobac o la mezcla de ambos. Este trabajo proporciona nuevos detalles acerca de la caracterización y de la fisiología de la población RM y aporta información novedosa en cuanto a los efectos fisiológicos provocados en las plantas sensibles y resistentes por los herbicidas IBAR e IBAA. Estos resultados son, potencialmente, de interés práctico para el sector agrícola, por la importancia global de las malas hierbas resistentes a glifosato, herbicida más usado a nivel mundial y a inhibidores de ALS, grupo de herbicidas que incluye un gran número de materias activas. Todo ello, unido al incesante incremento de resistencias múltiples a ambos tipos de herbicidas, hace necesarios los estudios que profundicen en el conocimiento fisiológico de este tipo de resistencias para avanzar hacia un manejo más sostenible y duradero.Publication Open Access Flora arvense de Navarra(Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa, 2018) Peralta de Andrés, Francisco Javier; Royuela Hernando, Mercedes; Ciencias; ZientziakEl origen de este libro se encuentra en el sitio web de Flora Arvense de Navarra, que fue desarrollado con el objetivo de facilitar el conocimiento de las malas hierbas de Navarra al alumnado de la Escuela Técnica Superior de Ingenieros Agrónomos de la Universidad Pública de Navarra, además de el de ser útil para personal investigador, técnico y agricultores de Navarra y territorios próximos. La edición de este libro se planteó para permitir la disponibilidad del contenido del sitio web en un soporte independiente de Internet.Publication Open Access Hypoxic treatment decreases the physiological action of the herbicide imazamox on pisum sativum roots(MDPI, 2020) Gil Monreal, Miriam; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMABThe inhibition of acetolactate synthase (ALS; EC 2.2.1.6), an enzyme located in the biosynthetic pathway of branched-chain amino acids, is the target site of the herbicide imazamox. One of the physiological effects triggered after ALS inhibition is the induction of aerobic ethanol fermentation. The objective of this study was to unravel if fermentation induction is related to the toxicity of the herbicide or if it is a plant defense mechanism. Pea plants were exposed to two different times of hypoxia before herbicide application in order to induce the ethanol fermentation pathway, and the physiological response after herbicide application was evaluated at the level of carbohydrates and amino acid profile. The effects of the herbicide on total soluble sugars and starch accumulation, and changes in specific amino acids (branched-chain, amide, and acidic) were attenuated if plants were subjected to hypoxia before herbicide application. These results suggest that fermentation is a plant defense mechanism that decreases the herbicidal effect.Publication Open Access Impairment of carbon metabolism induced by the herbicide glyphosate(Elsevier, 2012) Orcaray Echeverría, Luis; Zulet González, Amaia; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen ZientziakThe herbicide glyphosate reduces plant growth and causes plant death by inhibiting the biosynthesis of aromatic amino acids. The objective of this work was to determine whether glyphosate-treated plants show a carbon metabolism pattern comparable to that of plants treated with herbicides that inhibit branched-chain amino acid biosynthesis. Glyphosate-treated plants showed impaired carbon metabolism with an accumulation of carbohydrates in the leaves and roots. The growth inhibition detected after glyphosate treatment suggested impaired metabolism that impedes the utilization of available carbohydrates or energy at the expected rate. These effects were common to both types of amino acid biosynthesis inhibitors. Under aerobic conditions, ethanolic fermentative metabolism was enhanced in the roots of glyphosate-treated plants. This fermentative response was not related to changes in the respiratory rate or to a limitation of the energy charge. This response, which was similar for both types of herbicides, might be considered a general response to stress conditions.Publication Open Access Increased glyphosate-induced gene expression in the shikimate pathway is abolished in the presence of aromatic amino acids and mimicked by shikimate(Frontiers Media, 2020) Zulet González, Ainhoa; Barco Antoñanzas, María; Gil Monreal, Miriam; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaThe herbicide glyphosate inhibits the plant enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) in the aromatic amino acid (AAA) biosynthetic pathway, also known as the shikimate pathway. Amaranthus palmeri is a fast-growing weed, and several populations have evolved resistance to glyphosate through increased EPSPS gene copy number. The main objective of this study was to elucidate the regulation of the shikimate pathway and determine whether the regulatory mechanisms of glyphosate-sensitive and glyphosate-resistant plants were different. Leaf disks of sensitive and resistant (due to EPSPS gene amplification) A. palmeri plants were incubated for 24 h with glyphosate, AAA, glyphosate + AAA, or several intermediates of the pathway: shikimate, quinate, chorismate and anthranilate. In the sensitive population, glyphosate induced shikimate accumulation and induced the gene expression of the shikimate pathway. While AAA alone did not elicit any change, AAA applied with glyphosate abolished the effects of the herbicide on gene expression. It was not possible to fully mimic the effect of glyphosate by incubation with any of the intermediates, but shikimate was the intermediate that induced the highest increase (three-fold) in the expression level of the genes of the shikimate pathway of the sensitive population. These results suggest that, in this population, the lack of end products (AAA) of the shikimate pathway and shikimate accumulation would be the signals inducing gene expression in the AAA pathway after glyphosate application. In general, the effects on gene expression detected after the application of the intermediates were more severe in the sensitive population than in the resistant population. These results suggest that when EPSPS is overexpressed, as in the resistant population, the regulatory mechanisms of the AAA pathway are disrupted or buffered. The mechanisms underlying this behavior remain to be elucidated.Publication Open Access Insights into the regulation of nitrogen fixation in pea nodules: lessons from drought, abscisic acid and increased photoassimilate availability(EDP Sciences, 2001) González García, Esther; Gálvez, Loli; Royuela Hernando, Mercedes; Aparicio Tejo, Pedro María; Arrese-Igor Sánchez, César; Ciencias del Medio Natural; Natura Ingurunearen ZientziakNitrogen fixation in legume nodules has been shown to be very sensitive to drought and other environmental constraints. It has been widely assumed that this decline in nitrogen fixation was a consequence of an increase in the so-called oxygen diffusion barrier and a subsequent impairment to bacteroid respiration. However, it has been recently shown that nitrogen fixation is highly correlated with nodule sucrose synthase (SS) activity under drought and other environmental stresses. Whether this correlation reflects a causative relationship or not has not been proven yet. The evidence presented here suggests that SS controls nitrogen fixation under mild drought conditions. However, nitrogen fixation cannot be enhanced only by increasing glycolytic flux, as under these conditions nodules become oxygen limited. Abscisic acid also induces a decline in nitrogen fixation that is independent of SS. The overall results suggest the occurrence of a complex regulation of nodule nitrogen fixation involving, at least, both carbohydrate and oxygen fluxes within the nodule.Publication Open Access The moderate oxidative stress induced by glyphosate is not detected in Amaranthus palmeri plants overexpressing EPSPS(Elsevier, 2022) Gil Monreal, Miriam; Barco Antoñanzas, María; Zabalza Aznárez, Ana; Royuela Hernando, Mercedes; Eceiza, Mikel Vicente; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate PublikoaThe present study aimed to determine whether glyphosate-induced oxidative stress is directly related to the action mechanism of this herbicide (5-enolpyruvylshikimate-3-phosphate synthase or EPSPS inhibition) and analyse the role of oxidative stress in glyphosate toxicity of the weed Amaranthus palmeri S. Wats. Two kinds of populations were studied using EPSPS amplification: glyphosate-sensitive and glyphosate-resistant (by gene amplification). Plants were grown hydroponically and treated with different glyphosate doses, after which several oxidative stress markers were measured in the leaves. Untreated, sensitive and resistant plants showed similar values for the analysed parameters. Treated glyphosate-sensitive plants showed an increase in shikimate, superoxide and H2O2 contents and dose-dependent lipid peroxidation and antioxidant responses; however, none of these effects were observed in resistant plants, indicating that glyphosate-induced oxidative stress is related to EPSPS inhibition. Oxidative stress is associated with an increase in the activity of peroxidases due to EPSPS inhibition, although the link between both processes remains elusive. The fact that some glyphosate doses were lethal but did not induce major oxidative damage provides evidence that glyphosate toxicity is independent of oxidative stress.