Person:
Zulet González, Ainhoa

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https://academica-e.unavarra.es/handle/2454/50219

Last Name

Zulet González

First Name

Ainhoa

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Ciencias del Medio Natural

ORCID

0000-0003-3714-3825

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811122

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Now showing 1 - 10 of 11

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 Publikoa
A 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.
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 Publikoa
The 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.
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 Publikoa
En 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.
Open Access
The pattern of shikimate pathway and phenylpropanoids after inhibition by glyphosate or quinate feeding in pea roots
(Elsevier, 2017) Zabalza Aznárez, Ana; Orcaray Echeverría, Luis; Fernández Escalada, Manuel; Zulet González, Ainhoa; Royuela Hernando, Mercedes; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
The shikimate pathway is a metabolic route for the biosynthesis of aromatic amino acids (AAAs) (i.e. phenylalanine, tyrosine, and tryptophan). A key enzyme of shikimate pathway (5-enolpyruvylshikimate-3-phosphate synthase, EPSPS) is the target of the widely used herbicide glyphosate. Quinate is a compound synthesized in plants through a side branch of the shikimate pathway. Glyphosate provokes quinate accumulation and exogenous quinate application to plants shows a potential role of quinate in the toxicity of the herbicide glyphosate. Based on this, we hypothesized that the role of quinate accumulation in the toxicity of the glyphosate would be mediated by a deregulation of the shikimate pathway. In this study the effect of the glyphosate and of the exogenous quinate was evaluated in roots of pea plants by analyzing the time course of a full metabolic map of several metabolites of shikimate and phenylpropanoid pathways. Glyphosate application induced an increase of the 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHPS, first enzyme of the shikimate pathway) protein and accumulation of metabolites upstream of the enzyme EPSPS. No common effects on the metabolites and regulation of shikimate pathway were detected between quinate and glyphosate treatments, supporting that the importance of quinate in the mode of action of glyphosate is not mediated by a common alteration of the regulation of the shikimate pathway. Contrary to glyphosate, the exogenous quinate supplied was probably incorporated into the main trunk from the branch pathway and accumulated in the final products, such as lignin, concomitant with a decrease in the amount of DAHPS protein.
Open Access
Unravelling the phytotoxic effects of glyphosate on sensitive and resistant Amaranthus Palmeri populations by GC-MS and LC-MS metabolic profiling
(MDPI, 2023) Zulet González, Ainhoa; Gorzolka, Karin; Döll, Stefanie; Gil Monreal, Miriam; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Glyphosate, the most successful herbicide in history, specifically inhibits the activity of the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS; EC 2.5.1.19), one of the key enzymes in the shikimate pathway. Amaranthus palmeri is a driver weed in agriculture today that has evolved glyphosate-resistance through increased EPSPS gene copy number and other mechanisms. Non-targeted GC–MS and LC–MS metabolomic profiling was conducted to examine the innate physiology and the glyphosate-induced perturbations in one sensitive and one resistant (by EPSPS amplification) population of A. palmeri. In the absence of glyphosate treatment, the metabolic profile of both populations was very similar. The comparison between the effects of sublethal and lethal doses on sensitive and resistant populations suggests that lethality of the herbicide is associated with an amino acid pool imbalance and accumulation of the metabolites of the shikimate pathway upstream from EPSPS. Ferulic acid and its derivatives were accumulated in treated plants of both populations, while quercetin and its derivative contents were only lower in the resistant plants treated with glyphosate.
Open Access
Physiological performance of glyphosate and imazamox mixtures on Amaranthus palmeri sensitive and resistant to glyphosate
(Nature Research, 2019) Fernández Escalada, Manuel; Zulet González, Ainhoa; Gil Monreal, Miriam; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
The herbicides glyphosate and imazamox inhibit the biosynthetic pathway of aromatic amino acids (AAA) and branched-chain amino acids (BCAA), respectively. Both herbicides share several physiological effects in the processes triggered in plants after herbicide application that kills the plant, and mixtures of both herbicides are being used. The aim of this study was to evaluate the physiological effects in the mixture of glyphosate and imazamox in glyphosate-sensitive (GS) and -resistant (GR) populations of the troublesome weed Amaranthus palmeri. The changes detected in the physiological parameters after herbicide mixtures application were similar and even less to the changes detected after individual treatments. This pattern was detected in shikimate, amino acid and carbohydrate content, and it was independent of the EPSPS copy number, as it was detected in both populations. In the case of the transcriptional pattern of the AAA pathway after glyphosate, interesting and contrary interactions with imazamox treatment were detected for both populations; enhancement of the effect in the GS population and alleviation in the GR population. At the transcriptional level, no cross regulation between AAA and BCAA inhibitors was confirmed. This study suggests that mixtures are equally or less toxic than herbicides alone, and would implicate careful considerations when applying the herbicide mixtures.
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 Publikoa
Este 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.
Open Access
Quinate-enhanced glyphosate toxicity is related to the accumulation of quinate derivatives
(Elsevier, 2024) Zulet González, Ainhoa; Gil Monreal, Miriam; Gorzolka, Karin; Royuela Hernando, Mercedes; Zabalza Aznárez, Ana; Institute for Multidisciplinary Research in Applied Biology - IMAB
Glyphosate is the most widely used herbicide and works by inhibiting the enzyme 5-enolpyruvylshikimate 3-phosphate synthase (EPSPS) of the shikimate pathway, preventing the aromatic amino acid biosynthesis. When applied to plants, it provokes the accumulation of quinate, a metabolite synthesized through a side branch of the shikimate pathway. The joint application of glyphosate and quinate enhanced glyphosate efficacy on Amaranthus palmeri, requiring one-quarter of the recommended dose of glyphosate for complete control. Expression of the genes of the shikimate pathway and non-targeted GC-MS metabolic profiling were conducted to compare the physiological response after glyphosate, quinate or the combination of both. A perturbed gene expression of the shikimate pathway was detected after quinate applied alone, while no relevant changes in the metabolome were detected. The sub-lethal glyphosate treatment induced gene expression in the shikimate pathway, accumulation of the metabolites located upstream EPSPS and disturbances in the amino acid content. The exacerbation of the phytotoxicity in the lethal combined treatment was not related to any specific change in the expression level of the shikimate pathway. Metabolic profiling indicated that the accumulation of quinate and quinate derivatives detected after quinate applied alone was severely enhanced after the combined treatment of quinate and glyphosate.
Open Access
Physiological approach to the use of the natural compound quinate in the control of sensitive and resistant Papaver rhoeas
(MDPI, 2020) Zabalza Aznárez, Ana; Zulet González, Ainhoa; Barco Antoñanzas, María; Eceiza, Mikel Vicente; Gil Monreal, Miriam; Royuela Hernando, Mercedes; Institute for Multidisciplinary Research in Applied Biology - IMAB; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Quinate (1,3,4,5-tetrahydroxycyclohexanecarboxylate) is a compound synthesized in plants through a side-branch of the shikimate biosynthesis pathway, which is accumulated after glyphosate and acetolactate synthase inhibiting herbicides (ALS-inhibitors) and has phytotoxic potential. The objective of this study was to evaluate the phytotoxicity of quinate on several weed species. Among the species evaluated, Cynodon dactylon, Bromus diandrus, Lolium rigidum, Sinapis alba, and Papaver rhoeas, P. rhoeas was the most sensitive, and its growth was controlled with quinate concentrations above 100 mM at the phenological stage of 6–8 true leaves. A physiological study, including the shikimate pathway and the physiological markers of ALS-inhibitors (carbohydrates and amino acids), was performed in the sensitive and resistant plants treated with sulfonylureas or quinate. The typical physiological effects of ALS-inhibitors were detected in the sensitive population (free amino acid and carbohydrate accumulation) and not detected in the resistant population. The mode of action of quinate appeared to be related to general perturbations in their carbon/nitrogen metabolism rather than to specific changes in the shikimate pathway. These results suggest the possibility of using quinate in the weed control management of P. rhoeas.
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 Publikoa
Glyphosate 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.