Publikationen - Molekulare Signalverarbeitung

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Benzylisoquinoline alkaloids (BIAs) are a group of nitrogen-containing plant secondary metabolites comprised of an estimated 2500 identified structures. In BIA metabolism, (S)-reticuline is a key branch-point intermediate that can be directed into several alkaloid subtypes with different structural skeleton configurations. The morphinan alkaloids are one subclass of BIAs produced in only a few plant species, most notably and abundantly in the opium poppy (Papaver somniferum). Comparative transcriptome analysis of opium poppy and several other Papaver species that do not accumulate morphinan alkaloids showed that known genes encoding BIA biosynthetic enzymes are expressed at higher levels in P. somniferum. Three unknown cDNAs that are co-ordinately expressed with several BIA biosynthetic genes were identified as enzymes in the pathway. One of these enzymes, salutaridine reductase (SalR), which is specific for the production of morphinan alkaloids, was isolated and heterologously overexpressed in its active form not only from P. somniferum, but also from Papaver species that do not produce morphinan alkaloids. SalR is a member of a class of short chain dehydrogenase/reductases (SDRs) that are active as monomers and possess an extended amino acid sequence compared with classical SDRs. Homology modelling and substrate docking revealed the substrate binding site for SalR. The amino acids residues conferring salutaridine binding were compared to several members of the SDR family from different plant species, which non-specifically reduce (−)-menthone to (+)-neomenthol. Previously, it was shown that some of these proteins are involved in plant defence. The recruitment of specific monomeric SDRs from monomeric SDRs involved in plant defence is discussed.

The peroxidation of polyunsaturated fatty acids, common to all eukaryotes, is mostly catalyzed by members of the lipoxygenase enzyme family of non-heme iron containing dioxygenases. Lipoxygenase products can be metabolized further in the oxylipin pathway by several groups of CYP74 enzymes. One prominent oxylipin is jasmonic acid (JA), a product of the 13-allene oxide synthase branch of the pathway and known as signaling substance that plays a role in vegetative and propagative plant development as well as in plant responses to wounding and pathogen attack. In barley roots, JA level increases upon colonization by arbuscular mycorrhizal fungi. Apart from this first result regarding JA, no information is available on the relevance of lipidperoxide metabolism in arbuscular mycorrhizal symbiosis. Thus we analyzed fatty acid and lipidperoxide patterns in roots of Medicago truncatula during mycorrhizal colonization. Levels of fungus-specific fatty acids as well as palmitic acid (16:0) and oleic acid (18:1 n − 9) were increased in mycorrhizal roots. Thus the degree of arbuscular mycorrhizal colonization of roots can be estimated via analysis of fungal specific esterified fatty acids. Otherwise, no significant changes were found in the profiles of esterified and free fatty acids. The 9- and 13-LOX products of linoleic and α-linolenic acid were present in all root samples, but did not show significant differences between mycorrhizal and non-mycorrhizal roots, except JA which showed elevated levels in mycorrhizal roots. In both types of roots levels of 13-LOX products were higher than those of 9-LOX products. In addition, three cDNAs encoding CYP74 enzymes, two 9/13-hydroperoxide lyases and a 13-allene oxide synthase, were isolated and characterized. The transcript accumulation of these three genes, however, was not increased in mycorrhizal roots of M. truncatula.

The biosynthesis of benzophenanthridine alkaloids, phytoalexins of Eschscholzia californica, in cultured cells can be induced by a glycoprotein preparation from yeast, methyljasmonate, artificial acidification with permeant acids, or mild osmotic stress. Each of these stimuli strongly attenuated the subsequent response to the same stimulus (homologous desensitization). Elicitor contact and artificial acidification mutually desensitized the cells for either signal. In contrast, elicitor-treated cells maintained their responsiveness to methyljasmonate or hyperosmolarity (sorbitol). Elicitor concentrations that nearly saturated the alkaloid response did not cause a detectable increase of jasmonate content. Transient acidification of the cytoplasm is a necessary step of signaling by low elicitor concentrations but was not detectable after jasmonate treatment. Seen together, the data indicate the existence of a jasmonate-dependent and jasmonate-independent (ΔpH controlled) signal pathway towards the expression of benzophenanthridine biosynthesis. Selective desensitization allows either stimulus to activate a distinct share of the biosynthetic capacity of the cell and limits the accumulation of toxic defense metabolites.Yeast elicitor and jasmonate trigger alkaloid production via different signal pathways that show selective desensitization. Elicitor treatment (bottom cells) but not jasmonate (top cells) evokes intracellular pH shifts.

Occurrence of an essential enzyme in jasmonate (JA) biosynthesis, the allene oxide cyclase, (AOC) was analyzed in different developmental stages and various organs of Arabidopsis thaliana plants by immuno blot analysis and immunocytological approaches. Levels of AOC and of the two preceding enzymes in JA biosynthesis increased during seedling development accompanied by increased levels of JA and 12-oxophytodienoic acid levels after 4 and 8 weeks. Most tissues including all vascular bundles and that of flower buds contain AOC protein. Flowers shortly before opening, however, contain AOC protein preferentially in ovules, stigma cells and vascular bundles, whereas in anthers and pollen AOC could not be detected. The putative roles of AOC and JA in development are discussed.The allene oxide cyclase (AOC) is an important enzyme in jasmonate biosynthesis. Levels and occurrence of AOC in different organs and tissues are altered during development of Arabidopsis thaliana.