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Publications - Cell and Metabolic Biology

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Displaying results 21 to 30 of 409.

Publications

Paudel, G., Bilova, T., Schmidt, R., Greifenhagen, U., Berger, R., Tarakhovskaya, E., Stöckhardt, S., Balcke, G. U., Humbeck, K., Brandt, W., Sinz, A., Vogt, T., Birkemeyer, C., Wessjohann, L. & Frolov, A Osmotic stress is accompanied by protein glycation in Arabidopsis thaliana J. Exp. Bot. 67, 6283-6295, (2016) DOI: 10.1093/jxb/erw395

Among the environmental alterations accompanying oncoming climate changes, drought is the most important factor influencing crop plant productivity. In plants, water deficit ultimately results in the development of oxidative stress and accumulation of osmolytes (e.g. amino acids and carbohydrates) in all tissues. Up-regulation of sugar biosynthesis in parallel to the increasing overproduction of reactive oxygen species (ROS) might enhance protein glycation, i.e. interaction of carbonyl compounds, reducing sugars and α-dicarbonyls with lysyl and arginyl side-chains yielding early (Amadori and Heyns compounds) and advanced glycation end-products (AGEs). Although the constitutive plant protein glycation patterns were characterized recently, the effects of environmental stress on AGE formation are unknown so far. To fill this gap, we present here a comprehensive in-depth study of the changes in Arabidopsis thaliana advanced glycated proteome related to osmotic stress. A 3 d application of osmotic stress revealed 31 stress-specifically and 12 differentially AGE-modified proteins, representing altogether 56 advanced glycation sites. Based on proteomic and metabolomic results, in combination with biochemical, enzymatic and gene expression analysis, we propose monosaccharide autoxidation as the main stress-related glycation mechanism, and glyoxal as the major glycation agent in plants subjected to drought. 

Publications

Scheler, U., Brandt, W., Porzel, A., Rothe, K., Manzano, D., Božić, D., Papaefthimiou, D., Balcke, G. U., Henning, A., Lohse, S., Marillonnet, S., Kanellis, A. K., Ferrer, A. & Tissier, A. Elucidation of the biosynthesis of carnosic acid and its reconstitution in yeast Nat Commun 7, 12942, (2016) DOI: 10.1038/ncomms12942

Rosemary extracts containing the phenolic diterpenes carnosic acid and its derivative carnosol are approved food additives used in an increasingly wide range of products to enhance shelf-life, thanks to their high anti-oxidant activity. We describe here the elucidation of the complete biosynthetic pathway of carnosic acid and its reconstitution in yeast cells. Cytochrome P450 oxygenases (CYP76AH22-24) from Rosmarinus officinalis and Salvia fruticosa already characterized as ferruginol synthases are also able to produce 11-hydroxyferruginol. Modelling-based mutagenesis of three amino acids in the related ferruginol synthase (CYP76AH1) from S. miltiorrhiza is sufficient to convert it to a 11-hydroxyferruginol synthase (HFS). The three sequential C20 oxidations for the conversion of 11-hydroxyferruginol to carnosic acid are catalysed by the related CYP76AK6-8. The availability of the genes for the biosynthesis of carnosic acid opens opportunities for the metabolic engineering of phenolic diterpenes, a class of compounds with potent anti-oxidant, anti-inflammatory and anti-tumour activities.

Publications

Treutler, H., Tsugawa, H., Porzel, A., Gorzolka, K., Tissier, A., Neumann, S. & Balcke, G. U. Discovering regulated metabolite families in untargeted metabolomics studies. Anal Chem 88, 8082-8090, (2016) DOI: 10.1021/acs.analchem.6b01569

The identification of metabolites by mass spectrometry constitutes a major bottleneck which considerably limits the throughput of metabolomics studies in biomedical or plant research. Here, we present a novel approach to analyze metabolomics data from untargeted, data-independent LC-MS/MS measurements. By integrated analysis of MS1 abundances and MS/MS spectra, the identification of regulated metabolite families is achieved. This approach offers a global view on metabolic regulation in comparative metabolomics. We implemented our approach in the web application “MetFamily”, which is freely available at http://msbi.ipb-halle.de/MetFamily/. MetFamily provides a dynamic link between the patterns based on MS1-signal intensity and the corresponding structural similarity at the MS/MS level. Structurally related metabolites are annotated as metabolite families based on a hierarchical cluster analysis of measured MS/MS spectra. Joint examination with principal component analysis of MS1 patterns, where this annotation 

Publications

Pedranzani, H., Rodríguez-Rivera, M., Gutiérrez, M., Porcel, R., Hause, B. & Ruiz-Lozano, J. M. Arbuscular mycorrhizal symbiosis regulates physiology and performance of Digitaria eriantha plants subjected to abiotic stresses by modulating antioxidant and jasmonate levels. Mycorrhiza 26 , 141-152, (2016) DOI: 10.1007/s00572-015-0653-4

This study evaluates antioxidant responses and jasmonate regulation in Digitaria eriantha cv. Sudafricana plants inoculated (AM) and non-inoculated (non-AM) with Rhizophagus irregularis and subjected to drought, cold, or salinity. Stomatal conductance, photosynthetic efficiency, biomass production, hydrogen peroxide accumulation, lipid peroxidation, antioxidants enzymes activities, and jasmonate levels were determined. Stomatal conductance and photosynthetic efficiency decreased in AM and non-AM plants under all stress conditions. However, AM plants subjected to drought, salinity, or non-stress conditions showed significantly higher stomatal conductance values. AM plants subjected to drought or non-stress conditions increased their shoot/root biomass ratios, whereas salinity and cold caused a decrease in these ratios. Hydrogen peroxide accumulation, which was high in non-AM plant roots under all treatments, increased significantly in non-AM plant shoots under cold stress and in AM plants under non-stress and drought conditions. Lipid peroxidation increased in the roots of all plants under drought conditions. In shoots, although lipid peroxidation decreased in AM plants under non-stress and cold conditions, it increased under drought and salinity. AM plants consistently showed high catalase (CAT) and ascorbate peroxidase (APX) activity under all treatments. By contrast, the glutathione reductase (GR) and superoxide dismutase (SOD) activity of AM roots was lower than that of non-AM plants and increased in shoots. The endogenous levels of cis-12-oxophytodienoc acid (OPDA), jasmonic acid (JA), and 12-OH-JA showed a significant increase in AM plants as compared to non-AM plants. 11-OH-JA content only increased in AM plants subjected to drought. Results show that D. eriantha is sensitive to drought, salinity, and cold stresses and that inoculation with AM fungi regulates its physiology and performance under such conditions, with antioxidants and jasmonates being involved in this process

Publications

Taylor, I., Wang, Y., Seitz, K., Baer, J., Bennewitz, S., Mooney, B. P. & Walker, J. C. Analysis of Phosphorylation of the Receptor-Like Protein Kinase HAESA during Arabidopsis Floral Abscission Plos One 11, e0147203, (2016) DOI: org/10.1371/journal.pone.0147203

Receptor-like protein kinases (RLKs) are the largest family of plant transmembrane signaling proteins. Here we present functional analysis of HAESA, an RLK that regulates floral organ abscission in Arabidopsis. Through in vitro and in vivo analysis of HAE phosphorylation, we provide evidence that a conserved phosphorylation site on a region of the HAE protein kinase domain known as the activation segment positively regulates HAE activity. Additional analysis has identified another putative activation segment phosphorylation site common to multiple RLKs that potentially modulates HAE activity. Comparative analysis suggests that phosphorylation of this second activation segment residue is an RLK specific adaptation that may regulate protein kinase activity and substrate specificity. A growing number of RLKs have been shown to exhibit biologically relevant dual specificity toward serine/threonine and tyrosine residues, but the mechanisms underlying dual specificity of RLKs are not well understood. We show that a phospho-mimetic mutant of both HAE activation segment residues exhibits enhanced tyrosine auto-phosphorylation in vitro, indicating phosphorylation of this residue may contribute to dual specificity of HAE. These results add to an emerging framework for understanding the mechanisms and evolution of regulation of RLK activity and substrate specificity.

Books and chapters

Bilova, T., Greifenhagen, U., Paudel, G., Lukasheva, E., Brauch, D., Osmolovskaya, N., Tarakhovskaya, E., Balcke, G. U., Tissier, A., Vogt, T., Milkowski, C., Birkemeyer, C., Wessjohann, L. & Frolov, A. Glycation of Plant Proteins under Environmental Stress — Methodological Approaches, Potential Mechanisms and Biological Role. In: Abiotic and Biotic Stress in Plants - Recent Advances and Future Perspectives (Shanker, A. K.; Shanker, C.). (2016) ISBN: 978-953-51-2250-0 DOI: 10.5772/61860

 Environmental stress is one of the major factors reducing crop productivity. Due to the oncoming climate changes, the effects of drought and high light on plants play an increasing role in modern agriculture. These changes are accompanied with a progressing contamination of soils with heavy metals. Independent of their nature, environmental alterations result in development of oxidative stress, i.e. increase of reactive oxygen species (ROS) contents, and metabolic adjustment, i.e. accumulation of soluble primary metabolites (amino acids and sugars). However, a simultaneous increase of ROS and sugar concentrations ultimately results in protein glycation, i.e. non-enzymatic interaction of reducing sugars or their degradation products (α-dicarbonyls) with proteins. The eventually resulting advanced glycation end-products (AGEs) are known to be toxic and pro-inflammatory in mammals. Recently, their presence was unambiguously demonstrated in vivo in stressed Arabidopsis thaliana plants. Currently, information on protein targets, modification sites therein, mediators and mechanisms of plant glycation are being intensively studied. In this chapter, we comprehensively review the methodological approaches for plant glycation research and discuss potential mechanisms of AGE formation under stress conditions. On the basis of these patterns and additional in vitro experiments, the pathways and mechanisms of plant glycation can be proposed.

Publications

Hazmana, M., Hause, B., Eiche, E., Riemann, M. & Nick, P. Different forms of osmotic stress evoke qualitatively different responses in rice J PLANT PHYSIOL 202, 45–56, (2016) DOI: 10.1016/j.jplph.2016.05.027

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.

Publications

Bilova, T., Lukasheva, E., Brauch, D., Greifenhagen, U., Paudel, G., Tarakhovskaya, E., Frolova, N., Mittasch, J., Balcke, G. U., Tissier, A., Osmolovskaya, N., Vogt, T., Wessjohann, L. A., Birkemeyer, C., Milkowski, C. & Frolov A Snapshot of the Plant Glycated Proteome: structural, functional, and mechanistic aspects J. Biol. Chem. 291, 7621-7636, (2016) DOI: doi:10.1074/jbc.M115.678581

Glycation is the reaction of carbonyl compounds (reducing sugars and α-dicarbonyls) with amino acids, lipids, and proteins, yielding early and advanced glycation end products (AGEs). The AGEs can be formed via degradation of early glycation intermediates (glycoxidation) and by interaction with the products of monosaccharide autoxidation (autoxidative glycosylation). Although formation of these potentially deleterious compounds is well characterized in animal systems and thermally treated foods, only a little information about advanced glycation in plants is available. Thus, the knowledge of the plant AGE patterns and the underlying pathways of their formation are completely missing. To fill this gap, we describe the AGE-modified proteome of Brassica napus and characterize individual sites of advanced glycation by the methods of liquid chromatography-based bottom-up proteomics. The modification patterns were complex but reproducible: 789 AGE-modified peptides in 772 proteins were detected in two independent experiments. In contrast, only 168 polypeptides contained early glycated lysines, which did not resemble the sites of advanced glycation. Similar observations were made with Arabidopsis thaliana. The absence of the early glycated precursors of the AGE-modified protein residues indicated autoxidative glycosylation, but not glycoxidation, as the major pathway of AGE formation. To prove this assumption and to identify the potential modifying agents, we estimated the reactivity and glycative potential of plant-derived sugars using a model peptide approach and liquid chromatography-mass spectrometry-based techniques. Evaluation of these data sets together with the assessed tissue carbohydrate contents revealed dihydroxyacetone phosphate, glyceraldehyde 3-phosphate, ribulose, erythrose, and sucrose as potential precursors of plant AGEs.

Publications

Hettwer, K., Böttcher, C., Frolov, A., Mittasch, J., Albert, A., von Roepenack-Lahayeb, E., Strack, D. & Milkowski, C. Dynamic metabolic changes in seeds and seedlings of Brassica napus (oilseed rape) suppressing UGT84A9 reveal plasticity and molecular regulation of the phenylpropanoid pathway 124, 46–57, (2016) DOI: 10.1016/j.phytochem.2016.01.014

In Brassica napus, suppression of the key biosynthetic enzyme UDP-glucose:sinapic acid glucosyltransferase (UGT84A9) inhibits the biosynthesis of sinapine (sinapoylcholine), the major phenolic component of seeds. Based on the accumulation kinetics of a total of 158 compounds (110 secondary and 48 primary metabolites), we investigated how suppression of the major sink pathway of sinapic acid impacts the metabolome of developing seeds and seedlings. In UGT84A9-suppressing (UGT84A9i) lines massive alterations became evident in late stages of seed development affecting the accumulation levels of 58 secondary and 7 primary metabolites. UGT84A9i seeds were characterized by decreased amounts of various hydroxycinnamic acid (HCA) esters, and increased formation of sinapic and syringic acid glycosides. This indicates glycosylation and β-oxidation as metabolic detoxification strategies to bypass intracellular accumulation of sinapic acid. In addition, a net loss of sinapic acid upon UGT84A9 suppression may point to a feedback regulation of HCA biosynthesis. Surprisingly, suppression of UGT84A9 under control of the seed-specific NAPINC promoter was maintained in cotyledons during the first two weeks of seedling development and associated with a reduced and delayed transformation of sinapine into sinapoylmalate. The lack of sinapoylmalate did not interfere with plant fitness under UV-B stress. Increased UV-B radiation triggered the accumulation of quercetin conjugates whereas the sinapoylmalate level was not affected.

Books and chapters

Wasternack, C. Jasmonates: synthesis, metabolism, signal transduction and action. In: eLS. Chichester: Wiley (2016) ISBN: ISBN 978-0-4700-1590-2 DOI: 10.1002/9780470015902.a0020138.pub2

Jasmonic acid and other fatty-acid-derived compounds called oxylipins are signals in stress responses and development of plants. The receptor complex, signal transduction components as well as repressors and activators in jasmonate-induced gene expression have been elucidated. Different regulatory levels and cross-talk with other hormones are responsible for the multiplicity of plant responses to environmental and developmental cues.

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