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Publications - Stress and Develop Biology

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Publications

Böttcher, C.; Westphal, L.; Schmotz, C.; Prade, E.; Scheel, D.; Glawischnig, E.; The Multifunctional Enzyme CYP71B15 (PHYTOALEXIN DEFICIENT3) Converts Cysteine-Indole-3-Acetonitrile to Camalexin in the Indole-3-Acetonitrile Metabolic Network of Arabidopsis thaliana Plant Cell 21, 1830-1845, (2009) DOI: 10.1105/tpc.109.066670

Accumulation of camalexin, the characteristic phytoalexin of Arabidopsis thaliana, is induced by a great variety of plant pathogens. It is derived from Trp, which is converted to indole-3-acetonitrile (IAN) by successive action of the cytochrome P450 enzymes CYP79B2/B3 and CYP71A13. Extracts from wild-type plants and camalexin biosynthetic mutants, treated with silver nitrate or inoculated with Phytophthora infestans, were comprehensively analyzed by ultra-performance liquid chromatography electrospray ionization quadrupole time-of-flight mass spectrometry. This metabolomics approach was combined with precursor feeding experiments to characterize the IAN metabolic network and to identify novel biosynthetic intermediates and metabolites of camalexin. Indole-3-carbaldehyde and indole-3-carboxylic acid derivatives were shown to originate from IAN. IAN conjugates with glutathione, γ-glutamylcysteine, and cysteine [Cys(IAN)] accumulated in challenged phytoalexin deficient3 (pad3) mutants. Cys(IAN) rescued the camalexin-deficient phenotype of cyp79b2 cyp79b3 and was itself converted to dihydrocamalexic acid (DHCA), the known substrate of CYP71B15 (PAD3), by microsomes isolated from silver nitrate–treated Arabidopsis leaves. Surprisingly, yeast-expressed CYP71B15 also catalyzed thiazoline ring closure, DHCA formation, and cyanide release with Cys(IAN) as substrate. In conclusion, in the camalexin biosynthetic pathway, IAN is derivatized to the intermediate Cys(IAN), which serves as substrate of the multifunctional cytochrome P450 enzyme CYP71B15.
Publications

Gust, A. A.; Biswas, R.; Lenz, H. D.; Rauhut, T.; Ranf, S.; Kemmerling, B.; Götz, F.; Glawischnig, E.; Lee, J.; Felix, G.; Nürnberger, T.; Bacteria-derived Peptidoglycans Constitute Pathogen-associated Molecular Patterns Triggering Innate Immunity in Arabidopsis J. Biol. Chem. 282, 32338-32348, (2007) DOI: 10.1074/jbc.M704886200

Pathogen-associated molecular pattern (PAMP)-triggered immunity constitutes the primary plant immune response that has evolved to recognize invariant structures of microbial surfaces. Here we show that Gram-positive bacteria-derived peptidoglycan (PGN) constitutes a novel PAMP of immune responses in Arabidopsis thaliana. Treatment with PGN from Staphylococcus aureus results in the activation of plant responses, such as medium alkalinization, elevation of cytoplasmic calcium concentrations, nitric oxide, and camalexin production and the post-translational induction of MAPK activities. Microarray analysis performed with RNA prepared from PGN-treated Arabidopsis leaves revealed enhanced transcript levels for 236 genes, many of which are also altered upon administration of flagellin. Comparison of cellular responses after treatment with bacteria-derived PGN and structurally related fungal chitin indicated that both PAMPs are perceived via different perception systems. PGN-mediated immune stimulation in Arabidopsis is based upon recognition of the PGN sugar backbone, while muramyl dipeptide, which is inactive in this plant, triggers immunity-associated responses in animals. PGN adds to the list of PAMPs that induce innate immune programs in both plants and animals. However, we propose that PGN perception systems arose independently in both lineages and are the result of convergent evolution.
Publications

Qutob, D.; Kemmerling, B.; Brunner, F.; Küfner, I.; Engelhardt, S.; Gust, A. A.; Luberacki, B.; Seitz, H. U.; Stahl, D.; Rauhut, T.; Glawischnig, E.; Schween, G.; Lacombe, B.; Watanabe, N.; Lam, E.; Schlichting, R.; Scheel, D.; Nau, K.; Dodt, G.; Hubert, D.; Gijzen, M.; Nürnberger, T.; Phytotoxicity and Innate Immune Responses Induced by Nep1-Like Proteins Plant Cell 18, 3721-3744, (2006) DOI: 10.1105/tpc.106.044180

We show that oomycete-derived Nep1 (for necrosis and ethylene-inducing peptide1)–like proteins (NLPs) trigger a comprehensive immune response in Arabidopsis thaliana, comprising posttranslational activation of mitogen-activated protein kinase activity, deposition of callose, production of nitric oxide, reactive oxygen intermediates, ethylene, and the phytoalexin camalexin, as well as cell death. Transcript profiling experiments revealed that NLPs trigger extensive reprogramming of the Arabidopsis transcriptome closely resembling that evoked by bacteria-derived flagellin. NLP-induced cell death is an active, light-dependent process requiring HSP90 but not caspase activity, salicylic acid, jasmonic acid, ethylene, or functional SGT1a/SGT1b. Studies on animal, yeast, moss, and plant cells revealed that sensitivity to NLPs is not a general characteristic of phospholipid bilayer systems but appears to be restricted to dicot plants. NLP-induced cell death does not require an intact plant cell wall, and ectopic expression of NLP in dicot plants resulted in cell death only when the protein was delivered to the apoplast. Our findings strongly suggest that NLP-induced necrosis requires interaction with a target site that is unique to the extracytoplasmic side of dicot plant plasma membranes. We propose that NLPs play dual roles in plant pathogen interactions as toxin-like virulence factors and as triggers of plant innate immune responses.
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