Benzylisoquinoline alkaloids (BIAs) are important secondary plant metabolites and include medicinally relevant drugs, such as morphine or codeine. As the de novo synthesis of BIA backbones is (still) unfeasible, to date the opium poppy plant Papaver somniferum L. represents the main source of BIAs. The formation of BIAs is induced in poppy plants by stress conditions, such as wounding or salt treatment; however, the details about regulatory processes controlling BIA formation in opium poppy are not well studied. Environmental stresses, such as wounding or salinization, are transduced in plants by phospholipid-based signaling pathways, which involve different classes of phospholipases. Here we investigate whether pharmacological inhibition of phospholipase A2 (PLA2, inhibited by aristolochic acid (AA)) or phospholipase D (PLD; inhibited by 5-fluoro-2-indolyl des-chlorohalopemide (FIPI)) in poppy plants influences wound-induced BIA accumulation and the expression of key biosynthetic genes. We show that inhibition of PLA2 results in increased morphinan biosynthesis concomitant with reduced production of BIAs of the papaverine branch, whereas inhibition of PLD results in increased production of BIAs of the noscapine branch. The data suggest that phospholipid-dependent signaling pathways contribute to the activation of morphine biosynthesis at the expense of the production of other BIAs in poppy plants. A better understanding of the effectors and the principles of regulation of alkaloid biosynthesis might be the basis for the future genetic modification of opium poppy to optimize BIA production.

The essential trace element selenium (Se) is controversially discussed concerning its role in health and disease. Its various physiological functions are largely mediated by Se incorporation in the catalytic center of selenoproteins. In order to gain insights into the impact of Se deficiency and of supplementation with different Se compounds (selenite, selenate, selenomethionine) at defined concentrations (recommended, 150 μg/kg diet; excessive, 750 μg/kg diet) in murine colon tissues, a 20‐week feeding experiment was performed followed by analysis of the protein expression pattern of colon tissue specimens by 2D‐DIGE and MALDI‐TOF MS. Using this approach, 24 protein spots were identified to be significantly regulated by the different Se compounds. These included the antioxidant enzyme peroxiredoxin‐5 (PRDX5), proteins with binding capabilities, such as cofilin‐1 (COF1), calmodulin, and annexin A2 (ANXA2), and proteins involved in catalytic processes, such as 6‐phosphogluconate dehydrogenase (6PGD). Furthermore, the Se compounds demonstrated a differential impact on the expression of the identified proteins. Selected target structures were validated by qPCR and Western blot which mainly confirmed the proteomic profiling data. Thus, novel Se‐regulated proteins in colon tissues have been identified, which expand our understanding of the physiologic role of Se in colon tissue.

Drought is one of the most important environmental stressors resulting in increasing losses of crop plant productivity all over the world. Therefore, development of new approaches to increase the stress tolerance of crop plants is strongly desired. This requires precise and adequate modeling of drought stress. As this type of stress manifests itself as a steady decrease in the substrate water potential (ψw), agar plates infused with polyethylene glycol (PEG) are the perfect experimental tool: they are easy in preparation and provide a constantly reduced ψw, which is not possible in soil models. However, currently, this model is applicable only to seedlings and cannot be used for evaluation of stress responses in mature plants, which are obviously the most appropriate objects for drought tolerance research. To overcome this limitation, here we introduce a PEG-based agar infusion model suitable for 6–8-week-old A. thaliana plants, and characterize, to the best of our knowledge for the first time, the early drought stress responses of adult plants grown on PEG-infused agar. We describe essential alterations in the primary metabolome (sugars and related compounds, amino acids and polyamines) accompanied by qualitative and quantitative changes in protein patterns: up to 87 unique stress-related proteins were annotated under drought stress conditions, whereas further 84 proteins showed a change in abundance. The obtained proteome patterns differed slightly from those reported for seedlings and soil-based models.

Drought, salinity and alkalinity are distinct forms of osmotic stress with serious impacts on rice productivity. We investigated, for a salt-sensitive rice cultivar, the response to osmotically equivalent doses of these stresses. Drought, experimentally mimicked by mannitol (single factor: osmotic stress), salinity (two factors: osmotic stress and ion toxicity), and alkalinity (three factors: osmotic stress, ion toxicity, and depletion of nutrients and protons) produced different profiles of adaptive and damage responses, both locally (in the root) as well as systemically (in the shoot). The combination of several stress factors was not necessarily additive, and we even observed cases of mitigation, when two (salinity), or three stressors (alkalinity) were compared to the single stressor (drought). The response to combinations of individual stress factors is therefore not a mere addition of the partial stress responses, but rather represents a new quality of response. We interpret this finding in a model, where the output to signaling molecules is not determined by their abundance per se, but qualitatively depends on their adequate integration into an adaptive signaling network. This output generates a systemic signal that will determine the quality of the shoot response to local concentrations of ions.

PTMs are defined as covalent additions to functional groups of amino acid residues in proteins like phosphorylation, glycosylation, S‐nitrosylation, acetylation, methylation, lipidation, SUMOylation as well as oxidation. Oxidation of proteins has been characterized as a double‐edged sword. While oxidative modifications, in particular of cysteine residues, are widely involved in the regulation of cellular homeostasis, oxidative stress resulting in the oxidation of biomolecules along with the disruption of their biological functions can be associated with the development of diseases, such as cancer, diabetes, and neurodegenerative diseases, respectively. This is also the case for advanced glycation end products, which result from chemical reactions of keto compounds such as oxidized sugars with proteins. The role of oxidative modifications under physiological and pathophysiological conditions remains largely unknown. Recently, novel technologies have been established that allow the enrichment, identification, and characterization of specific oxidative PTMs (oxPTMs). This is essential to develop strategies to prevent and treat diseases that are associated with oxidative stress. Therefore this review will focus on (i) the methods and technologies, which are currently applied for the detection, identification, and quantification of oxPTMs including the design of high throughput approaches and (ii) the analyses of oxPTMs related to physiological and pathological conditions.

We applied an extended charge‐based fractional diagonal chromatography (ChaFRADIC) workflow to analyze the N‐terminal proteome of Arabidopsis thaliana seedlings. Using iTRAQ protein labeling and a multi‐enzyme digestion approach including trypsin, GluC, and subtilisin, a total of 200 μg per enzyme, and measuring only one third of each ChaFRADIC‐enriched fraction by LC‐MS, we quantified a total of 2791 unique N‐terminal peptides corresponding to 2249 different unique N‐termini from 1270 Arabidopsis proteins. Our data indicate the power, reproducibility, and sensitivity of the applied strategy that might be applicable to quantify proteolytic events from as little as 20 μg of protein per condition across up to eight different samples. Furthermore, our data clearly reflect the methionine excision dogma as well as the N‐end rule degradation pathway (NERP) discriminating into a stabilizing or destabilizing function of N‐terminal amino acid residues. We found bona fide NERP destabilizing residues underrepresented, and the list of neo N‐termini from wild type samples may represent a helpful resource during the evaluation of NERP substrate candidates. All MS data have been deposited in the ProteomeXchange with identifier PXD001855 (http://proteomecentral.proteomexchange.org/dataset/PXD001855).

As a result of the phenylpropanoid pathway, many Brassicaceae produce considerable amounts of soluble hydroxycinnamate conjugates, mainly sinapate esters. From oilseed rape (Brassica napus), we cloned two orthologs of the Arabidopsis (Arabidopsis thaliana) gene REDUCED EPIDERMAL FLUORESCENCE1 (REF1) encoding a coniferaldehyde/sinapaldehyde dehydrogenase. The enzyme is involved in the formation of ferulate and sinapate from the corresponding aldehydes, thereby linking lignin and hydroxycinnamate biosynthesis as a potential branch-point enzyme. We used RNA interference to silence REF1 genes in seeds of oilseed rape. Nontargeted metabolite profiling showed that BnREF1-suppressing seeds produced a novel chemotype characterized by reduced levels of sinapate esters, the appearance of conjugated monolignols, dilignols, and trilignols, altered accumulation patterns of kaempferol glycosides, and changes in minor conjugates of caffeate, ferulate, and 5-hydroxyferulate. BnREF1 suppression affected the level of minor sinapate conjugates more severely than that of the major component sinapine. Mapping of the changed metabolites onto the phenylpropanoid metabolic network revealed partial redirection of metabolic sequences as a major impact of BnREF1 suppression.

In phytopathology quantitative measurements are rarely used to assess crop plant disease symptoms. Instead, a qualitative valuation by eye is often the method of choice. In order to close the gap between subjective human inspection and objective quantitative results, the development of an automated analysis system that is capable of recognizing and characterizing the growth patterns of fungal hyphae in micrograph images was developed. This system should enable the efficient screening of different host–pathogen combinations (e.g., barley—Blumeria graminis, barley—Rhynchosporium secalis) using different microscopy technologies (e.g., bright field, fluorescence). An image segmentation algorithm was developed for gray-scale image data that achieved good results with several microscope imaging protocols. Furthermore, adaptability towards different host–pathogen systems was obtained by using a classification that is based on a genetic algorithm. The developed software system was named HyphArea, since the quantification of the area covered by a hyphal colony is the basic task and prerequisite for all further morphological and statistical analyses in this context. By means of a typical use case the utilization and basic properties of HyphArea could be demonstrated. It was possible to detect statistically significant differences between the growth of an R. secalis wild-type strain and a virulence mutant.

Fungi cause severe diseases on a broad range of crop and ornamental plants, leading to significant economical losses. Plant pathogenic fungi exhibit a huge variability in their mode of infection, differentiation and function of infection structures and nutritional strategy. In this review, advances in understanding mechanisms of biotrophy, necrotrophy and hemibiotrophic lifestyles are described. Special emphasis is given to the biotrophy-necrotrophy switch of hemibiotrophic pathogens, and to biosynthesis, chemical diversity and mode of action of various fungal toxins produced during the infection process.

The effect of temporary dark exposure on adventitious root formation (ARF) in Petunia×hybrida ‘Mitchell’ cuttings was investigated. Histological and metabolic changes in the cuttings during the dark treatment and subsequent rooting in the light were recorded. Excised cuttings were exposed to the dark for seven days at 10°C followed by a nine-day rooting period in perlite or were rooted immediately for 16 days in a climate chamber at 22/20°C (day/night) and a photosynthetic photon flux density (PPFD) of 100 μmol m−2 s−1. Dark exposure prior to rooting increased, accelerated and synchronized ARF. The rooting period was reduced from 16 days (non-treated cuttings) to 9 days (treated cuttings). Under optimum conditions, despite the reduced rooting period, dark-exposed cuttings produced a higher number and length of roots than non-treated cuttings. An increase in temperature to 20 °C during the dark treatment or extending the cold dark exposure to 14 days caused a similar enhancement of root development compared to non-treated cuttings. Root meristem formation had already started during the dark treatment and was enhanced during the subsequent rooting period. Levels of soluble sugars (glucose, fructose and sucrose) and starch in leaf and basal stem tissues significantly decreased during the seven days of dark exposure. This depletion was, however, compensated during rooting after 6 and 24h for soluble sugars in leaves and the basal stem, respectively, whereas the sucrose level in the basal stem was already increased at 6 h. The association of higher carbohydrate levels with improved rooting in previously dark-exposed versus non-treated cuttings indicates that increased post-darkness carbohydrate availability and allocation towards the stem base contribute to ARF under the influence of dark treatment and provide energy for cell growth subject to a rising sink intensity in the base of the cutting.