Metabolic profiling of Lolium perenne shows functional integration of metabolic responses to diverse subtoxic conditions of chemical stress.

Plant communities are confronted with a great variety of environmental chemical stresses. Characterization of chemical stress in higher plants has often been focused on single or closely related stressors under acute exposure, or restricted to a selective number of molecular targets. In order to understand plant functioning under chemical stress conditions close to environmental pollution conditions, the C3 grass Lolium perenne was subjected to a panel of different chemical stressors (pesticide, pesticide degradation compound, polycyclic aromatic hydrocarbon, and heavy metal) under conditions of seed-level or root-level subtoxic exposure. Physiological and metabolic profiling analysis on roots and shoots revealed that all of these subtoxic chemical stresses resulted in discrete physiological perturbations and complex metabolic shifts. These metabolic shifts involved stressor-specific effects, indicating multilevel mechanisms of action, such as the effects of glyphosate and its degradation product aminomethylphosphonic acid on quinate levels. They also involved major generic effects that linked all of the subtoxic chemical stresses with major modifications of nitrogen metabolism, especially affecting asparagine, and of photorespiration, especially affecting alanine and glycerate. Stress-related physiological effects and metabolic adjustments were shown to be integrated through a complex network of metabolic correlations converging on Asn, Leu, Ser, and glucose-6-phosphate, which could potentially be modulated by differential dynamics and interconversion of soluble sugars (sucrose, trehalose, fructose, and glucose). Underlying metabolic, regulatory, and signalling mechanisms linking these subtoxic chemical stresses with a generic impact on nitrogen metabolism and photorespiration are discussed in relation to carbohydrate and low-energy sensing.

Low environmentally relevant levels of bioactive xenobiotics and associated degradation products cause cryptic perturbations of metabolism and molecular stress responses in Arabidopsis thaliana.

Anthropic changes and chemical pollution confront wild plant communities with xenobiotic combinations of bioactive molecules, degradation products, and adjuvants that constitute chemical challenges potentially affecting plant growth and fitness. Such complex challenges involving residual contamination and mixtures of pollutants are difficult to assess. The model plant Arabidopsis thaliana was confronted by combinations consisting of the herbicide glyphosate, the fungicide tebuconazole, the glyphosate degradation product aminomethylphosphonic acid (AMPA), and the atrazine degradation product hydroxyatrazine, which had been detected and quantified in soils of field margins in an agriculturally intensive region. Integrative analysis of physiological, metabolic, and gene expression responses was carried out in dose-response experiments and in comparative experiments of varying pesticide combinations. Field margin contamination levels had significant effects on plant growth and metabolism despite low levels of individual components and the presence of pesticide degradation products. Biochemical and molecular analysis demonstrated that these less toxic degradation products, AMPA and hydroxyatrazine, by themselves elicited significant plant responses, thus indicating underlying mechanisms of perception and transduction into metabolic and gene expression changes. These mechanisms may explain observed interactions, whether positive or negative, between the effects of pesticide products (AMPA and hydroxyatrazine) and the effects of bioactive xenobiotics (glyphosate and tebuconazole). Finally, the metabolic and molecular perturbations induced by low levels of xenobiotics and associated degradation products were shown to affect processes (carbon balance, hormone balance, antioxidant defence, and detoxification) that are likely to determine environmental stress sensitivity.

Physiology and toxicology of hormone-disrupting chemicals in higher plants.

Higher plants are exposed to natural environmental organic chemicals, associated with plant-environment interactions, and xenobiotic environmental organic chemicals, associated with anthropogenic activities. The effects of these chemicals result not only from interaction with metabolic targets, but also from interaction with the complex regulatory networks of hormone signaling. Purpose-designed plant hormone analogues thus show extensive signaling effects on gene regulation and are as such important for understanding plant hormone mechanisms and for manipulating plant growth and development. Some natural environmental chemicals also act on plants through interference with the perception and transduction of endogenous hormone signals. In a number of cases, bioactive xenobiotics, including herbicides that have been designed to affect specific metabolic targets, show extensive gene regulation effects, which are more in accordance with signaling effects than with consequences of metabolic effects. Some of these effects could be due to structural analogies with plant hormones or to interference with hormone metabolism, thus resulting in situations of hormone disruption similar to animal cell endocrine disruption by xenobiotics. These hormone-disrupting effects can be superimposed on parallel metabolic effects, thus indicating that toxicological characterisation of xenobiotics must take into consideration the whole range of signaling and metabolic effects. Hormone-disruptive signaling effects probably predominate when xenobiotic concentrations are low, as occurs in situations of residual low-level pollutions. These hormone-disruptive effects in plants may thus be of importance for understanding cryptic effects of low-dosage xenobiotics, as well as the interactive effects of mixtures of xenobiotic pollutants.

Xenobiotic sensing and signalling in higher plants.

Anthropogenic changes and chemical pollution confront plant communities with various xenobiotic compounds or combinations of xenobiotics, involving chemical structures that are at least partially novel for plant species. Plant responses to chemical challenges and stimuli are usually characterized by the approaches of toxicology, ecotoxicology, and stress physiology. Development of transcriptomics and proteomics analysis has demonstrated the importance of modifications to gene expression in plant responses to xenobiotics. It has emerged that xenobiotic effects could involve not only biochemical and physiological disruption, but also the disruption of signalling pathways. Moreover, mutations affecting sensing and signalling pathways result in modifications of responses to xenobiotics, thus confirming interference or crosstalk between xenobiotic effects and signalling pathways. Some of these changes at gene expression, regulation and signalling levels suggest various mechanisms of xenobiotic sensing in higher plants, in accordance with xenobiotic-sensing mechanisms that have been characterized in other phyla (yeast, invertebrates, vertebrates). In higher plants, such sensing systems are difficult to identify, even though different lines of evidence, involving mutant studies, transcription factor analysis, or comparative studies, point to their existence. It remains difficult to distinguish between the hypothesis of direct xenobiotic sensing and indirect sensing of xenobiotic-related modifications. However, future characterization of xenobiotic sensing and signalling in higher plants is likely to be a key element for determining the tolerance and remediation capacities of plant species. This characterization will also be of interest for understanding evolutionary dynamics of stress adaptation and mechanisms of adaptation to novel stressors.

Local-scale dynamics of plant-pesticide interactions in a northern Brittany agricultural landscape.

Soil pollution by anthropogenic chemicals is a major concern for sustainability of crop production and of ecosystem functions mediated by natural plant biodiversity. Understanding the complex effects of soil pollution requires multi-level and multi-scale approaches. Non-target and agri-environmental plant communities of field margins and vegetative filter strips are confronted with agricultural xenobiotics through soil contamination, drift, run-off and leaching events that result from chemical applications. Plant-pesticide dynamics in vegetative filter strips was studied at field scale in the agricultural landscape of a long-term ecological research network in northern Brittany (France). Vegetative filter strips effected significant pesticide abatement between the field and riparian compartments. However, comparison of pesticide usage modalities and soil chemical analysis revealed the extent and complexity of pesticide persistence in fields and vegetative filter strips, and suggested the contribution of multiple sources (yearly carry-over, interannual persistence, landscape-scale contamination). In order to determine the impact of such persistence, plant dynamics was followed in experimentally-designed vegetative filter strips of identical initial composition (Agrostis stolonifera, Anthemis tinctoria/Cota tinctoria, Centaurea cyanus, Fagopyrum esculentum, Festuca rubra, Lolium perenne, Lotus corniculatus, Phleum pratense, Trifolium pratense). After homogeneous vegetation establishment, experimental vegetative filter strips underwent rapid changes within the following two years, with Agrostis stolonifera, Festuca rubra, Lolium perenne and Phleum pratense becoming dominant and with the establishment of spontaneous vegetation. Co-inertia analysis showed that plant dynamics and soil residual pesticides could be significantly correlated, with the triazole fungicide epoxiconazole, the imidazole fungicide prochloraz and the neonicotinoid insecticide thiamethoxam as strong drivers of the correlation. However, the correlation was vegetative-filter-strip-specific, thus showing that correlation between plant dynamics and soil pesticides likely involved additional factors, such as threshold levels of residual pesticides. This situation of complex interactions between plants and soil contamination is further discussed in terms of agronomical, environmental and health issues.

Species- and organ-specific responses of agri-environmental plants to residual agricultural pollutants.

Soil pollution by anthropogenic chemicals is a major concern for sustainability of crop production and of ecosystem functions mediated by natural plant biodiversity. The complex effects on plants are however difficult to apprehend. Plant communities of field margins, vegetative filter strips or rotational fallows are confronted with agricultural pollutants through residual soil contamination and/or through drift, run-off and leaching events that result from chemical applications. Exposure to xenobiotics and heavy metals causes biochemical, physiological and developmental effects. However, the range of doses, modalities of exposure, metabolization of contaminants into derived xenobiotics, and combinations of contaminants result in variable and multi-level effects. Understanding these complex plant-pollutant interactions cannot directly rely on toxicological or agronomical approaches that focus on the effects of field-rate pesticide applications. It must take into account exposure at root level, sublethal concentrations of bioactive compounds and functional biodiversity of the plant species that are affected. The present study deals with agri-environmental plant species of field margins, vegetative filter strips or rotational fallows in European agricultural landscapes. Root and shoot physiological and growth responses were compared under controlled conditions that were optimally adjusted for each plant species. Contrasted responses of growth inhibition, no adverse effect or growth enhancement depended on species, organ and nature of contaminant. However, all of the agricultural contaminants under study (pesticides, pesticide metabolites, heavy metals, polycyclic aromatic hydrocarbons) had significant effects under conditions of sublethal exposure on at least some of the plant species. The fungicide tebuconazole and polycyclic aromatic hydrocarbon fluoranthene, which gave highest levels of responses, induced both activation or inhibition effects, in different plant species or in different organs of the same plant species. These complex effects are discussed in terms of dynamics of agri-environmental plants and of ecological consequences of differential root-shoot growth under conditions of soil contamination.

Herbicide-related signaling in plants reveals novel insights for herbicide use strategies, environmental risk assessment and global change assessment challenges.

Herbicide impact is usually assessed as the result of a unilinear mode of action on a specific biochemical target with a typical dose-response dynamics. Recent developments in plant molecular signaling and crosstalk between nutritional, hormonal and environmental stress cues are however revealing a more complex picture of inclusive toxicity. Herbicides induce large-scale metabolic and gene-expression effects that go far beyond the expected consequences of unilinear herbicide-target-damage mechanisms. Moreover, groundbreaking studies have revealed that herbicide action and responses strongly interact with hormone signaling pathways, with numerous regulatory protein-kinases and -phosphatases, with metabolic and circadian clock regulators and with oxidative stress signaling pathways. These interactions are likely to result in mechanisms of adjustment that can determine the level of sensitivity or tolerance to a given herbicide or to a mixture of herbicides depending on the environmental and developmental status of the plant. Such regulations can be described as rheostatic and their importance is discussed in relation with herbicide use strategies, environmental risk assessment and global change assessment challenges.

Genome-Wide Transcriptional Profiling and Metabolic Analysis Uncover Multiple Molecular Responses of the Grass Species Lolium perenne Under Low-Intensity Xenobiotic Stress.

Lolium perenne, which is a major component of pastures, lawns, and grass strips, can be exposed to xenobiotic stresses due to diffuse and residual contaminations of soil. L. perenne was recently shown to undergo metabolic adjustments in response to sub-toxic levels of xenobiotics. To gain insight in such chemical stress responses, a de novo transcriptome analysis was carried out on leaves from plants subjected at the root level to low levels of xenobiotics, glyphosate, tebuconazole, and a combination of the two, leading to no adverse physiological effect. Chemical treatments influenced significantly the relative proportions of functional categories and of transcripts related to carbohydrate processes, to signaling, to protein-kinase cascades, such as Serine/Threonine-protein kinases, to transcriptional regulations, to responses to abiotic or biotic stimuli and to responses to phytohormones. Transcriptomics-based expressions of genes encoding different types of SNF1 (sucrose non-fermenting 1)-related kinases involved in sugar and stress signaling or encoding key metabolic enzymes were in line with specific qRT-PCR analysis or with the important metabolic and regulatory changes revealed by metabolomic analysis. The effects of pesticide treatments on metabolites and gene expression strongly suggest that pesticides at low levels, as single molecule or as mixture, affect cell signaling and functioning even in the absence of major physiological impact. This global analysis of L. perenne therefore highlighted the interactions between molecular regulation of responses to xenobiotics, and also carbohydrate dynamics, energy dysfunction, phytohormones and calcium signaling.