BACKGROUND: Acidic phytohormones are small molecules controlling many physiological functions in plants. A comprehensive picture of their profiles including the active forms, precursors and metabolites provides an important insight into ongoing physiological processes and is essential for many biological studies performed on plants. RESULTS: A high-throughput sample preparation method for liquid chromatography-tandem mass spectrometry determination of 25 acidic phytohormones classed as auxins, jasmonates, abscisates and salicylic acid was optimised. The method uses a small amount of plant tissue (less than 10 mg fresh weight) and acidic extraction in 1 mol/L formic acid in 10% aqueous methanol followed by miniaturised purification on reverse phase sorbent accommodated in pipette tips organised in a 3D printed 96-place interface, capable of processing 192 samples in one run. The method was evaluated in terms of process efficiency, recovery and matrix effects as well as establishing validation parameters such as accuracy and precision. The applicability of the method in relation to the amounts of sample collected from distantly related plant species was evaluated and the results for phytohormone profiles are discussed in the context of literature reports. CONCLUSION: The method developed enables high-throughput profiling of acidic phytohormones with minute amounts of plant material, and it is suitable for large scale interspecies studies.

• Klíčová slova
• 3D printing, Evolutionarily distant plant species, High-throughput, In-tip microSPE, Liquid chromatography, Mass spectrometry, Miniaturisation, Plant hormones,
• Publikační typ
• časopisecké články MeSH

Plant diseases pose a substantial threat to food availability, accessibility, and security as they account for economic losses of nearly $300 billion on a global scale. Although various strategies exist to reduce the impact of diseases, they can introduce harmful chemicals to the food chain and have an impact on the environment. Therefore, it is necessary to understand and exploit the plants' immune systems to control the spread of pathogens and enable sustainable agriculture. Recently, growing pieces of evidence suggest a functional myriad of lipids to be involved in providing structural integrity, intracellular and extracellular signal transduction mediators to substantial cross-kingdom cell signaling at the host-pathogen interface. Furthermore, some pathogens recognize or exchange plant lipid-derived signals to identify an appropriate host or development, whereas others activate defense-related gene expression. Typically, the membrane serves as a reservoir of lipids. The set of lipids involved in plant-pathogen interaction includes fatty acids, oxylipins, phospholipids, glycolipids, glycerolipids, sphingolipids, and sterols. Overall, lipid signals influence plant-pathogen interactions at various levels ranging from the communication of virulence factors to the activation and implementation of host plant immune defenses. The current review aims to summarize the progress made in recent years regarding the involvement of lipids in plant-pathogen interaction and their crucial role in signal transduction.

• Klíčová slova
• lipids, microbes, oxylipins, pathogens, phosphatidic acid, plants,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Jasmonates (JAs) are lipid-derived signals in plant stress responses and development. A crucial step in JA biosynthesis is catalyzed by allene oxide cyclase (AOC). Four genes encoding functional AOCs (AOC1, AOC2, AOC3 and AOC4) have been characterized for Arabidopsis thaliana in terms of organ- and tissue-specific expression, mutant phenotypes, promoter activities and initial in vivo protein interaction studies suggesting functional redundancy and diversification, including first hints at enzyme activity control by protein-protein interaction. Here, these analyses were extended by detailed analysis of recombinant proteins produced in Escherichia coli. Treatment of purified AOC2 with SDS at different temperatures, chemical cross-linking experiments and protein structure analysis by molecular modelling approaches were performed. Several salt bridges between monomers and a hydrophobic core within the AOC2 trimer were identified and functionally proven by site-directed mutagenesis. The data obtained showed that AOC2 acts as a trimer. Finally, AOC activity was determined in heteromers formed by pairwise combinations of the four AOC isoforms. The highest activities were found for heteromers containing AOC4 + AOC1 and AOC4 + AOC2, respectively. All data are in line with an enzyme activity control of all four AOCs by heteromerization, thereby supporting a putative fine-tuning in JA formation by various regulatory principles.

• Klíčová slova
• Arabidopsis allene oxide cyclase isoforms, activity regulation, heteromerization, protein structure analysis, site-directed mutagenesis,
• Publikační typ
• časopisecké články MeSH