Introducing the halophyte Salicornia europaea to investigate combined impact of salt and tidal submergence conditions.

Tolerance mechanisms to single abiotic stress events are being investigated in different plant species, but how plants deal with multiple stress factors occurring simultaneously is still poorly understood. Here, we introduce Salicornia europaea as a species with an extraordinary tolerance level to both flooding and high salt concentrations. Plants exposed to 0.5MNaCl (mimicking sea water concentrations) grew larger than plants not exposed to salt. Adding more salt reduced growth, but concentrations up to 2.5MNaCl were not lethal. Regular tidal flooding with salt water (0.5MNaCl) did not affect growth or chlorophyll fluorescence, whereas continuous flooding stopped growth while plants survived. Quantitative polymerase chain reaction (qPCR) analysis of plants exposed to 1% oxygen in air revealed induction of selected hypoxia responsive genes, but these genes were not induced during tidal flooding, suggesting that S. europaea did not experience hypoxic stress. Indeed, plants were able to transport oxygen into waterlogged soil. Interestingly, sequential exposure to salt and hypoxic air changed the expression of several but not all genes as compared to their expression upon hypoxia only, demonstrating the potential to use S . europaea to investigate signalling-crosstalk between tolerance reactions to multiple environmental perturbations.

Fermented biochar has a markedly different effect on fate of pesticides in soil than compost, straw, and a mixed biochar-product.

Current knowledge about how biochars affect the fate of pesticides in soil is based on studies that used pure biochars. After finding that an additional biological post-pyrolysis treatment, such as co-composting or lactic fermentation, is required for biochars for superior performance in temperate arable soils, a knowledge gap formed of how such further processed biochar products would affect the fate of pesticides in soil. This study compared the effects of a novel fermented biochar alone or mixed with biogas residues on the fate of two pesticides, 4-chloro-2-methylphenoxyacetic acid (MCPA) and metalaxyl-M, in a temperate arable soil to the traditional organic amendments wheat straw and compost. The fate of 14C-labeled MCPA was markedly affected in different ways. Fermented biochar effectively reduced the water-extractability and mineralization due to adsorption that was comparable to adsorption strengths reported for pure biochars. However, this effect was weak for the biochar mixed with biogas residues. Straw reduced water-extractable amounts due to increased biodegradation and formation of likely biogenic non-extractable residues of MCPA. In contrast, compost decelerated mineralization and increased the water solubility of the MCPA residues due to released dissolved organic matter. The amendments' effects were minor regarding 14C-metalaxyl-M, except for the fermented biochar which again reduced water-extractability and delayed degradation due to adsorption. Thus, the effects of the organic amendments differed for the two pesticide compounds with only the fermented biochar's effect being similar for both. However, this effect was no longer present in the mixed product containing 20% biochar. Our findings clearly show that biologically treated biochar-containing products can affect the fate of pesticides in soil very differently, also when compared to traditional organic amendments. Such impacts and their desirable and undesirable ecotoxicological implications need to be considered before the large-scale application of biochars to temperate arable soils.

The Arabidopsis thaliana onset of leaf death 12 mutation in the lectin receptor kinase P2K2 results in an autoimmune phenotype.

BACKGROUND: Plant immunity relies on the perception of immunogenic signals by cell-surface and intracellular receptors and subsequent activation of defense responses like programmed cell death. Under certain circumstances, the fine-tuned innate immune system of plants results in the activation of autoimmune responses that cause constitutive defense responses and spontaneous cell death in the absence of pathogens. RESULTS: Here, we characterized the onset of leaf death 12 (old12) mutant that was identified in the Arabidopsis accession Landsberg erecta. The old12 mutant is characterized by a growth defect, spontaneous cell death, plant-defense gene activation, and early senescence. In addition, the old12 phenotype is temperature reversible, thereby exhibiting all characteristics of an autoimmune mutant. Mapping the mutated locus revealed that the old12 phenotype is caused by a mutation in the Lectin Receptor Kinase P2-TYPE PURINERGIC RECEPTOR 2 (P2K2) gene. Interestingly, the P2K2 allele from Landsberg erecta is conserved among Brassicaceae. P2K2 has been implicated in pathogen tolerance and sensing extracellular ATP. The constitutive activation of defense responses in old12 results in improved resistance against Pseudomonas syringae pv. tomato DC3000. CONCLUSION: We demonstrate that old12 is an auto-immune mutant and that allelic variation of P2K2 contributes to diversity in Arabidopsis immune responses.

Endoplasmic reticulum-bound ANAC013 factor is cleaved by RHOMBOID-LIKE 2 during the initial response to hypoxia in Arabidopsis thaliana.

Aerobic reactions are essential to sustain plant growth and development. Impaired oxygen availability due to excessive water availability, e.g., during waterlogging or flooding, reduces plant productivity and survival. Consequently, plants monitor oxygen availability to adjust growth and metabolism accordingly. Despite the identification of central components in hypoxia adaptation in recent years, molecular pathways involved in the very early activation of low-oxygen responses are insufficiently understood. Here, we characterized three endoplasmic reticulum (ER)-anchored Arabidopsis ANAC transcription factors, namely ANAC013, ANAC016, and ANAC017, which bind to the promoters of a subset of hypoxia core genes (HCGs) and activate their expression. However, only ANAC013 translocates to the nucleus at the onset of hypoxia, i.e., after 1.5 h of stress. Upon hypoxia, nuclear ANAC013 associates with the promoters of multiple HCGs. Mechanistically, we identified residues in the transmembrane domain of ANAC013 to be essential for transcription factor release from the ER, and provide evidence that RHOMBOID-LIKE 2 (RBL2) protease mediates ANAC013 release under hypoxia. Release of ANAC013 by RBL2 also occurs upon mitochondrial dysfunction. Consistently, like ANAC013 knockdown lines, rbl knockout mutants exhibit impaired low-oxygen tolerance. Taken together, we uncovered an ER-localized ANAC013-RBL2 module, which is active during the initial phase of hypoxia to enable fast transcriptional reprogramming.

Mitochondrial alternative NADH dehydrogenases NDA1 and NDA2 promote survival of reoxygenation stress in Arabidopsis by safeguarding photosynthesis and limiting ROS generation.

Plant submergence stress is a growing problem for global agriculture. During desubmergence, rising O2 concentrations meet a highly reduced mitochondrial electron transport chain (mETC) in the cells. This combination favors the generation of reactive oxygen species (ROS) by the mitochondria, which at excess can cause damage. The cellular mechanisms underpinning the management of reoxygenation stress are not fully understood. We investigated the role of alternative NADH dehydrogenases (NDs), as components of the alternative mETC in Arabidopsis, in anoxia-reoxygenation stress management. Simultaneous loss of the matrix-facing NDs, NDA1 and NDA2, decreased seedling survival after reoxygenation, while overexpression increased survival. The absence of NDAs led to reduced maximum potential quantum efficiency of photosystem II linking the alternative mETC to photosynthetic function in the chloroplast. NDA1 and NDA2 were induced upon reoxygenation, and transcriptional activation of NDA1 was controlled by the transcription factors ANAC016 and ANAC017 that bind to the mitochondrial dysfunction motif (MDM) in the NDA1 promoter. The absence of NDA1 and NDA2 did not alter recovery of cytosolic ATP levels and NADH : NAD+ ratio at reoxygenation. Rather, the absence of NDAs led to elevated ROS production, while their overexpression limited ROS. Our observations indicate that the control of ROS formation by the alternative mETC is important for photosynthetic recovery and for seedling survival of anoxia-reoxygenation stress.

Acquisition of hypoxia inducibility by oxygen sensing N-terminal cysteine oxidase in spermatophytes.

N-terminal cysteine oxidases (NCOs) use molecular oxygen to oxidise the amino-terminal cysteine of specific proteins, thereby initiating the proteolytic N-degron pathway. To expand the characterisation of the plant family of NCOs (plant cysteine oxidases [PCOs]), we performed a phylogenetic analysis across different taxa in terms of sequence similarity and transcriptional regulation. Based on this survey, we propose a distinction of PCOs into two main groups. A-type PCOs are conserved across all plant species and are generally unaffected at the messenger RNA level by oxygen availability. Instead, B-type PCOs appeared in spermatophytes to acquire transcriptional regulation in response to hypoxia. The inactivation of two A-type PCOs in Arabidopsis thaliana, PCO4 and PCO5, is sufficient to activate the anaerobic response in young seedlings, whereas the additional removal of B-type PCOs leads to a stronger induction of anaerobic genes and impairs plant growth and development. Our results show that both PCO types are required to regulate the anaerobic response in angiosperms. Therefore, while it is possible to distinguish two clades within the PCO family, we conclude that they all contribute to restrain the anaerobic transcriptional programme in normoxic conditions and together generate a molecular switch to toggle the hypoxic response.

Remobilization of pollutants during extreme flood events poses severe risks to human and environmental health.

While it is well recognized that the frequency and intensity of flood events are increasing worldwide, the environmental, economic, and societal consequences of remobilization and distribution of pollutants during flood events are not widely recognized. Loss of life, damage to infrastructure, and monetary cleanup costs associated with floods are important direct effects. However, there is a lack of attention towards the indirect effects of pollutants that are remobilized and redistributed during such catastrophic flood events, particularly considering the known toxic effects of substances present in flood-prone areas. The global examination of floods caused by a range of extreme events (e.g., heavy rainfall, tsunamis, extra- and tropical storms) and subsequent distribution of sediment-bound pollutants are needed to improve interdisciplinary investigations. Such examinations will aid in the remediation and management action plans necessary to tackle issues of environmental pollution from flooding. River basin-wide and coastal lowland action plans need to balance the opposing goals of flood retention, catchment conservation, and economical use of water.

Molecular oxygen as a signaling component in plant development.

While traditionally hypoxia has been studied as a detrimental component of flooding stress, the last decade has flourished with studies reporting the involvement of molecular oxygen availability in plant developmental processes. Moreover, proliferating and undifferentiated cells from different plant tissues were found to reside in endogenously generated hypoxic niches. Thus, stress-associated acute hypoxia may be distinguished from constitutively generated chronic hypoxia. The Cys/Arg branch of the N-degron pathway assumes a central role in integrating oxygen levels resulting in proteolysis of transcriptional regulators that control different aspects of plant growth and development. As a target of this pathway, group VII of the Ethylene Response Factor (ERF-VII) family has emerged as a hub for the integration of oxygen dynamics in root development and during seedling establishment. Additionally, vegetative shoot meristem activity and reproductive transition were recently associated with oxygen availability via two novel substrates of the N-degron pathways: VERNALISATION 2 (VRN2) and LITTLE ZIPPER 2 (ZPR2). Together, these observations support roles for molecular oxygen as a signalling molecule in plant development, as well as in essential metabolic reactions. Here, we review recent findings regarding oxygen-regulated development, and discuss outstanding questions that spring from these discoveries.

Comparing straw, compost, and biochar regarding their suitability as agricultural soil amendments to affect soil structure, nutrient leaching, microbial communities, and the fate of pesticides.

The emission of nutrients and pesticides from agricultural soils endangers natural habitats. Here, we review to which extent carbon-rich organic amendments help to retain nutrients and pesticides in agricultural soils and to reduce the contamination of surrounding areas and groundwater. We compare straw, compost, and biochar to see whether biochar outperforms the other two more traditional and cheaper materials. We present a list of criteria to evaluate the suitability of organic materials to be used as soil amendments and discuss differences in elemental compositions of straw, compost, and biochar to understand, how soil microorganisms utilize those materials. We review their effects on physical and chemical soil characteristics, soil microbial communities, as well as effects on the transformation and retention of nutrients and pesticides in detail. It becomes clear that for all three amendments their effects can vary greatly depending on numerous aspects, such as the type of soil, application rate, and production procedure of the organic material. Biochar is most effective in increasing the sorption capacity of soils but does not outperform straw and compost with regards to the other aspects investigated. Nevertheless, the possibility to design biochar properties makes it a very promising material. Finally, we provide critical comments about how to make studies about organic amendments more comparable (comprehensive provision of material properties), how to improve concepts of future work (meta-analysis, long-term field studies, use of deep-insight microbial DNA sequencing), and what needs to be further investigated (the link between structural and functional microbial parameters, the impact of biochar on pesticide efficiency).

Multiparametric real-time sensing of cytosolic physiology links hypoxia responses to mitochondrial electron transport.

Hypoxia regularly occurs during plant development and can be induced by the environment through, for example, flooding. To understand how plant tissue physiology responds to progressing oxygen restriction, we aimed to monitor subcellular physiology in real time and in vivo. We establish a fluorescent protein sensor-based system for multiparametric monitoring of dynamic changes in subcellular physiology of living Arabidopsis thaliana leaves and exemplify its applicability for hypoxia stress. By monitoring cytosolic dynamics of magnesium adenosine 5'-triphosphate, free calcium ion concentration, pH, NAD redox status, and glutathione redox status in parallel, linked to transcriptional and metabolic responses, we generate an integrated picture of the physiological response to progressing hypoxia. We show that the physiological changes are surprisingly robust, even when plant carbon status is modified, as achieved by sucrose feeding or extended night. Inhibition of the mitochondrial respiratory chain causes dynamics of cytosolic physiology that are remarkably similar to those under oxygen depletion, highlighting mitochondrial electron transport as a key determinant of the cellular consequences of hypoxia beyond the organelle. A broadly applicable system for parallel in vivo sensing of plant stress physiology is established to map out the physiological context under which both mitochondrial retrograde signalling and low oxygen signalling occur, indicating shared upstream stimuli.

HBI1 Mediates the Trade-off between Growth and Immunity through Its Impact on Apoplastic ROS Homeostasis.

Plants continuously need to adapt to their environment and prioritize either growth or defense responses to secure survival and reproduction. Trade-offs between growth and defense are often attributed to the allocation of energy for growth to adaptation responses. Still, the exact mechanisms underlying growth and defense trade-offs are poorly understood. Here, we demonstrate that the growth-related transcription factor HOMOLOG OF BEE2 INTERACTING WITH IBH 1 (HBI1) regulates apoplastic reactive oxygen species (ROS) homeostasis by differentially controlling the expression of NADPH oxidases (NOXs) and peroxidases (POXs). The HBI1 target genes RESPIRATORY BURST OXIDASE HOMOLOG A (RbohA) and RbohC have contrasting effects on the regulation of cell size. In addition, the HBI1-controlled NOXs and POXs oppositely regulate susceptibility toward Pseudomonas syringae. Our findings reveal that the incompatibility between growth and defense programs can be attributed to the way apoplastic ROS homeostasis is modulated during both processes.

The ACBP1-RAP2.12 signalling hub: A new perspective on integrative signalling during hypoxia in plants.

During their lifetime, plants are frequently exposed to a variety of stresses which negatively impact on growth and vitality. In order to respond specifically to a given stress situation, integration of multiple signal inputs is of utmost importance. Recently, we demonstrated that recognition and adaptation to low-oxygen stress requires integration of signals from energy metabolism, lipid metabolism and oxygen availability. Low oxygen which results in an energy crisis causes a shift in lipid intermediate ratios. Binding of C18:1-CoA by ACYL-COA BINDING PROTEIN 1 (ACBP1) at the plasma membrane concomitantly leads to release and nuclear accumulation of the ERFVII transcription factor RELATED TO APETALA 2.12 (RAP2.12) which is central to the activation of anaerobic metabolism during stress. Moreover, RAP2.12 protein stability is oxygen-dependently regulated and its oxidation results in degradation by the N-end rule pathway. Here, we illuminate the concept of multiple-signal integration under hypoxia and discuss signal inputs merging at the ACBP1-ERFVII signaling hub.

An apical hypoxic niche sets the pace of shoot meristem activity.

Complex multicellular organisms evolved on Earth in an oxygen-rich atmosphere1; their tissues, including stem-cell niches, require continuous oxygen provision for efficient energy metabolism2. Notably, the maintenance of the pluripotent state of animal stem cells requires hypoxic conditions, whereas higher oxygen tension promotes cell differentiation3. Here we demonstrate, using a combination of genetic reporters and in vivo oxygen measurements, that plant shoot meristems develop embedded in a low-oxygen niche, and that hypoxic conditions are required to regulate the production of new leaves. We show that hypoxia localized to the shoot meristem inhibits the proteolysis of an N-degron-pathway4,5 substrate known as LITTLE ZIPPER 2 (ZPR2)-which evolved to control the activity of the class-III homeodomain-leucine zipper transcription factors6-8-and thereby regulates the activity of shoot meristems. Our results reveal oxygen as a diffusible signal that is involved in the control of stem-cell activity in plants grown under aerobic conditions, which suggests that the spatially distinct distribution of oxygen affects plant development. In molecular terms, this signal is translated into transcriptional regulation by the N-degron pathway, thereby linking the control of metabolic activity to the regulation of development in plants.

Hypoxic Conditions in Crown Galls Induce Plant Anaerobic Responses That Support Tumor Proliferation.

Agrobacterium tumefaciens infection of wounded plant tissues causes the formation of crown gall tumors. Upon infection, genes encoded on the A. tumefaciens tumor inducing plasmid are integrated in the plant genome to induce the biosynthesis of auxin and cytokinin, leading to uncontrolled cell division. Additional sequences present on the bacterial T-DNA encode for opine biosynthesis genes, which induce the production of opines that act as a unique carbon and nitrogen source for Agrobacterium. Crown galls therefore become a very strong sink for photosynthate. Here we found that the increased metabolic demand in crown galls causes an increase in oxygen consumption rate, which leads to a steep drop in the internal oxygen concentration. Consistent with this, plant hypoxia-responsive genes were found to be significantly upregulated in crown galls compared to uninfected stem tissue. Following this observation, we aimed at understanding whether the low-oxygen response pathway, mediated by group VII ethylene response factor (ERF-VII) transcription factors, plays a role in the development of crown galls. We found that quintuple knock-out mutants of all ERF-VII members, which are incapable of inducing the hypoxic response, show reduced crown gall symptoms. Conversely, mutant genotypes characterized by constitutively high levels of hypoxia-associated transcripts, displayed more severe crown gall symptoms. Based on these results, we concluded that uncontrolled cell proliferation of crown galls established hypoxic conditions, thereby requiring adequate anaerobic responses of the plant tissue to support tumor growth.

Low-oxygen response is triggered by an ATP-dependent shift in oleoyl-CoA in Arabidopsis.

Plant response to environmental stimuli involves integration of multiple signals. Upon low-oxygen stress, plants initiate a set of adaptive responses to circumvent an energy crisis. Here, we reveal how these stress responses are induced by combining (i) energy-dependent changes in the composition of the acyl-CoA pool and (ii) the cellular oxygen concentration. A hypoxia-induced decline of cellular ATP levels reduces LONG-CHAIN ACYL-COA SYNTHETASE activity, which leads to a shift in the composition of the acyl-CoA pool. Subsequently, we show that different acyl-CoAs induce unique molecular responses. Altogether, our data disclose a role for acyl-CoAs acting in a cellular signaling pathway in plants. Upon hypoxia, high oleoyl-CoA levels provide the initial trigger to release the transcription factor RAP2.12 from its interaction partner ACYL-COA BINDING PROTEIN at the plasma membrane. Subsequently, according to the N-end rule for proteasomal degradation, oxygen concentration-dependent stabilization of the subgroup VII ETHYLENE-RESPONSE FACTOR transcription factor RAP2.12 determines the level of hypoxia-specific gene expression. This research unveils a specific mechanism activating low-oxygen stress responses only when a decrease in the oxygen concentration coincides with a drop in energy.

Oxygen Sensing via the Ethylene Response Transcription Factor RAP2.12 Affects Plant Metabolism and Performance under Both Normoxia and Hypoxia.

Subgroup-VII-ethylene-response-factor (ERF-VII) transcription factors are involved in the regulation of hypoxic gene expression and regulated by proteasome-mediated proteolysis via the oxygen-dependent branch of the N-end-rule pathway. While research into ERF-VII mainly focused on their role to regulate anoxic gene expression, little is known on the impact of this oxygen-sensing system in regulating plant metabolism and growth. By comparing Arabidopsis (Arabidopsis thaliana) plants overexpressing N-end-rule-sensitive and insensitive forms of the ERF-VII-factor RAP2.12, we provide evidence that oxygen-dependent RAP2.12 stability regulates central metabolic processes to sustain growth, development, and anoxic resistance of plants. (1) Under normoxia, overexpression of N-end-rule-insensitive Δ13RAP2.12 led to increased activities of fermentative enzymes and increased accumulation of fermentation products, which were accompanied by decreased adenylate energy states and starch levels, and impaired plant growth and development, indicating a role of oxygen-regulated RAP2.12 degradation to prevent aerobic fermentation. (2) In Δ13RAP2.12-overexpressing plants, decreased carbohydrate reserves also led to a decrease in anoxic resistance, which was prevented by external Suc supply. (3) Overexpression of Δ13RAP2.12 led to decreased respiration rates, changes in the levels of tricarboxylic acid cycle intermediates, and accumulation of a large number of amino acids, including Ala and γ-amino butyric acid, indicating a role of oxygen-regulated RAP2.12 abundance in controlling the flux-modus of the tricarboxylic acid cycle. (4) The increase in amino acids was accompanied by increased levels of immune-regulatory metabolites. These results show that oxygen-sensing, mediating RAP2.12 degradation is indispensable to optimize metabolic performance, plant growth, and development under both normoxic and hypoxic conditions.

Redox regulation in shoot growth, SAM maintenance and flowering.

Reactive oxygen species (ROS) and associated reduction/oxidation (redox) controls involving glutathione, glutaredoxins and thioredoxins play key roles in the regulation of plant growth and development. While many questions remain concerning redox functions in the shoot apical meristem (SAM), accumulating evidence suggests that redox master switches integrate major hormone signals and transcriptional networks in the SAM, and so regulate organ growth, polarity and floral development. Auxin-induced activation of plasma-membrane located NADPH-oxidases and mitochondrial respiratory bioenergetics are likely regulators of the ROS bursts that drive the cell cycle in proliferating regions, with other hormones such as jasmonic acid playing propagating or antagonistic roles in gene regulation. Moreover, the activation of oxygen production by photosynthesis and oxygen-dependent N-end rule controls are linked to the transition from cell proliferation to cell expansion and differentiation. While much remains to be understood, the nexus of available redox controls provides a key underpinning mechanism linking hormonal controls, energy metabolism and bioenergetics to plant growth and development.

Mass spectrometry-based plant metabolomics: Metabolite responses to abiotic stress.

Metabolomics is one omics approach that can be used to acquire comprehensive information on the composition of a metabolite pool to provide a functional screen of the cellular state. Studies of the plant metabolome include analysis of a wide range of chemical species with diverse physical properties, from ionic inorganic compounds to biochemically derived hydrophilic carbohydrates, organic and amino acids, and a range of hydrophobic lipid-related compounds. This complexitiy brings huge challenges to the analytical technologies employed in current plant metabolomics programs, and powerful analytical tools are required for the separation and characterization of this extremely high compound diversity present in biological sample matrices. The use of mass spectrometry (MS)-based analytical platforms to profile stress-responsive metabolites that allow some plants to adapt to adverse environmental conditions is fundamental in current plant biotechnology research programs for the understanding and development of stress-tolerant plants. In this review, we describe recent applications of metabolomics and emphasize its increasing application to study plant responses to environmental (stress-) factors, including drought, salt, low oxygen caused by waterlogging or flooding of the soil, temperature, light and oxidative stress (or a combination of them). Advances in understanding the global changes occurring in plant metabolism under specific abiotic stress conditions are fundamental to enhance plant fitness and increase stress tolerance. © 2015 Wiley Periodicals, Inc. Mass Spec Rev 35:620-649, 2016.