Phytoalexin Induction Research Articles

THE PHYTOALEXIN RESPONSE: ELICITATION, SIGNALLING AND CONTROL OF HOST GENE EXPRESSION

Summary1. Induced resistance of plants to incompatible races of their microbial pathogens is often characterized by rapid cell death (the hypersensitive response) and the accumulation of low‐Mr antimicrobial compounds termed phytoalexins. There is much indirect evidence to support a major role for phytoalexin induction as an event determining host resistance, and genetical analysis of race‐specific pathogen‐host interactions predicts that induced resistance occurs via interaction between a pathogen avirulence gene product and a host resistance gene product. Elicitors (inducing agents) of phytoalexin accumulation are possible candidates for the active products directly or indirectly resulting from the expression of microbial avirulence genes. The present review considers the process of phytoalexin induction, at the molecular level, from the fungal elicitor to the early changes in host gene expression associated with its action.2. A number of microbial polysaccharides, glycoproteins, pectic enzymes, peptides and fatty acids have potent elicitor activity.3. The results of studies on the purification and characterization of microbial elicitors depend on the nature of the bioassay used. Methods for elicitor extraction may lead to artifacts or loss of race specificity.4. Elicitors isolated from plant pathogens may be race‐specific or race‐non‐specific. In cases where only race‐non‐specific elicitors can be shown, race‐specific induction of phytoalexins may result from the action of enhancer or suppressor molecules. Fungal glucans have been proposed as candidates to act in both these roles.5. Molecular genetic approaches to the identification of phytopathogenic bacterial avirulence genes may help to prove or disprove the role of elicitors of the phytoalexin response as agents responsible for the induction of host resistance. Similar analyses, involving genetic transformation, should soon be possible for phytopathogenic fungi.6. Studies on the nature of host receptors for microbial elicitors are still in their infancy. Such receptors are probably localized in the plant plasma membrane, and elicitation results in often striking changes in host membrane properties.7. Cyclic 3′,5′‐adenosine monophosphate and polyamines do not appear to act in plants as intracellular transducers of the phytoalexin response. Interest is now being shown in a possible role for calcium in intracellular signalling.8. Plant cells contain endogenous elicitor molecules whose synthesis or release may play a role in the intercellular transmission of the phytoalexin response. The main candidates for endogenous elicitors are pectic fragments of the host cell wall, although no direct evidence for their involvement in plant‐pathogen interactions is available. Pectic fragments may act as synergists in parallel with, rather than as couplers in series with, microbial elicitors. The role of ethylene as a response coupler for induced resistance phenomena is difficult to assess.9. Some elicitors may themselves move to the host cell nucleus, although whether this is their effective site of action remains unclear.10. Induction of the phytoalexin response is associated with specific changes in host gene expression related to the selective induction of new mRNA species and enzyme activities.11. Progress is now being made in the characterization of enzymes specific for phytoalexin biosynthesis. Enzyme induction and/or infection appears to involve rapid modulation of gene transcription, although some post‐translational events may also be involved in determining induction patterns.12. Work is now commencing on the sequencing of the genes encoding elicitor‐inducible enzymes. Multigene families have been identified for phenylalanine ammonia‐lyase and chalcone synthase.13. Phytoalexin accumulation may be accompanied by the rapid induction of ethylene biosynthesis and the expression of genes encoding activities involved in the synthesis of hydroxyproline‐rich glycoproteins.14. Pectic endogenous elicitors of phytoalexin accumulation are also active as inducers of systemic proteinase inhibitor synthesis in some solanaceous species.15. Results of studies on the mechanisms of signal transmission and differential gene expression in relation to active defence reactions should be of relevance to many other aspects of the plant's response to environmental stimuli at the physiological, biochemical and molecular genetical levels.

Phytoalexin Production by Isolated Soybean Protoplasts

Abstract Protoplasts isolated enzymatically from suspension-cultured cells of soybean (Glycine max L. Merr., cv. Harosoy 63) were used to study the production of the isoflavonoid-derived phytoale­xin, glyceollin. A large enhancement in the in vivo rates of synthesis and catalytic activities of two of the enzymes associated with glyceollin biosynthesis, phenylalanine ammonia-lyase and chalcone synthase, preceded phytoalexin accumulation during early stages of culture of isolated protoplasts while cell wall regeneration occurred. A glucan elicitor from cell walls of the fungus Phytophthora megasperm a f. sp. glycinea, an effective inducer of the phytoalexin response in cultured cells, was not capable of enhancing phytoalexin formation in protoplasts. Lack of respon­siveness of the protoplasts to the glucan elicitor could either be associated with their stressed metabolic state in which the response system is already saturated or with the removal from cultured cells of an essential factor of the glucan elicitor-mediated phytoalexin induction during protoplast isolation. At least two components of the protoplast isolation medium, the osmoticum and the fungal endopolygalacturonase, have the potential to initiate the observed phytoalexin synthesis during protoplast isolation. Our results indicate that under the methods employed isolated soybean protoplasts display a stress response which other types of soybean cells show following microbial attack or treatment with elicitor.

Several biotic and abiotic elicitors act synergistically in the induction of phytoalexin accumulation in soybean

Plants often respond to microbial infection by producing antimicrobial compounds called phytoalexins. Plants also produce phytoalexins in response to in vitro treatment with molecules called elicitors. Specific elicitors, including a hexa-β-glucosyl glucitol derived from fungal cell walls, the pectin-degrading enzyme endopolygalacturonic acid lyase, and oligogalacturonides obtained by either partial acid hydrolysis or enzymatic degradation of plant cell walls or citrus polygalacturonic acid, induce soybean (Glycine max. L.) cytoledons to accumulate phytoalexins. The experiments reported here demonstrate that the elicitor-active hexa-β-glucosyl glucitol acts synergistically with several biotic and abiotic elicitors in the induction of phytoalexins in soybean cotyledons. At concentrations below 50 ng/ml, the hexa-β-glucosyl glucitol does not induce significant phytoalexin accumulation. When assayed in combination with either endopolygalacturonic acid lyase or with a decagalacturonide released from citrus polygalacturonic acid by this lyase, however, the observed elicitor activity of the hexa-β-glucosyl glucitol is as much as 35-fold higher than the sum of the responses of these elicitors assayed separately. A similar synergism was also demonstrated for the combination of the hexa-β-glucosyl glucitol with dilute solutions of sodium acetate, sodium formate, or sodium propionate buffers. These buffers are thought to damage or kill plant cells, which may cause the release of oligogalacturonides from the plant cell wall. The results suggest that oligogalacturonides act as signals of tissue damage and, as such, can enhance the response of plant tissues to other elicitor-active molecules during the initiation of phytoalexin accumulation.

Dependence of the level of phytoalexin and enzyme induction by fungal elicitor on the growth stage of Petroselinum crispum cell cultures

The extent of induction of some metabolic activities in cultured parsley cells (Petroselinum crispum) by an elicitor preparation from Phytophthora megasperma f. sp. glycinea varied with the growth stage of the cell culture. On the basis of cell fresh weight, the induction of phytoalexin accumulation was high until cell mass reached a maximum, and then declined to a low level which was indistinguishable from a level caused by an endogenous mechanism operating at this late growth stage. The induction of phenylalanine ammonia-lyase and 4-coumarate:CoA ligase activities by the elicitor showed a high degree of coordination and a sharp maximum preceding the stage of maximal cell mass. 1,3-β-Glucanase activity was induced to about the same level throughout all growth stages, with a large contribution by an endogenous mechanism at late stages.

Tomato‐Fusarium oxysporum f. sp. lycopersici Interaction: “in vitro” Analysis of Several Possible Pathogenic Factors

AbstractThe behaviour of Tomato cultures from known resistant and susceptible cultivars and lines was examined for callus growth on culture media containing increasing concentrations of Fusarium oxysporum f. sp. lycopersici race 1 culture filtrate and phytoalexin synthesis elicited by Fusarium cell wall components. A strict correlation was found between in vivo resistance to the fungus and in vitro hypersensitive response and phytoalexin induction. On the other hand in vitro tolerance to toxic filtrate does not seem in this case a good indicator of in vivo resistance to the pathogen.

Phytoalexin Induction in French Bean

Treatment of hypocotyl sections or cell suspension cultures of dwarf French bean (Phaseolus vulgaris L.) with an abiotic elicitor (denatured ribonuclease A) resulted in increased extractable activity of the enzyme l-phenylalanine ammonia-lyase. This induction could be transmitted from treated cells through a dialysis membrane to cells which were not in direct contact with the elicitor. In hypocotyl sections, induction of isoflavonoid phytoalexin accumulation was also transmitted across a dialysis membrane, although levels of insoluble, lignin-like phenolic material remained unchanged in elicitor-treated and control sections. In bean cell suspension cultures, the induction of phenylalanine ammonia-lyase in cells separated from ribonuclease-treated cells by a dialysis membrane was also accompanied by increases in the activities of chalcone synthase and chalcone isomerase, two enzymes previously implicated in the phytoalexin defense response. Such intercellular transmission of elicitation did not occur in experiments with cells treated with a biotic elicitor preparation heat-released from the cell walls of the bean pathogen Colletotrichum lindemuthianum. The results confirm and extend previous suggestions that a low molecular weight, diffusible factor of host plant origin is involved (in French bean) in the intercellular transmission of the elicitation response to abiotic elicitors.

Phytoalexin induction in Beta and Spinacia

Abstract The production of the phytoalexins betagarin and betavulgarin in the three sections and various ploidy levels of the genus Beta have been studied. Variation occurred within some species. Spinacia produced no detectable phytoalexins.

Phytoalexin induction and its chemosystematic significance in the genus Trigonella

Using the drop-diffusate technique, a number of isoflavonoid phytoalexins have been obtained from the excised, fungus-inoculated leaflets of 41 species belonging to the legume genus Trigonella. Leaf diffusates variously contained pterocarpan (medicarpin and maackiain) and isoflavan (vestitol and sativan) derivatives previously associated with genera closely allied to Trigonella. In diffusates from T. calliceras, medicarpin was accompanied by a phytoalexin (designated TC-1) provisionally identified as a new hydroxylated pterocarpan. Most of the Trigonella species were also examined for their ability to release coumarin upon tissue maceration. The combined phytoalexin/coumarin data suggest that three major intrageneric chemical divisions occur in Trigonella; two of these apparently link the genus to Medicado/Factorovskya and Melilotus respectively, whilst the third provides some evidence for a connection with Trifolium. The taxonomic aspects of these findings are discussed in the light of earlier morphological studies which provided evidence for a distinct floral dichotomy amongst Trigonella species.

Chitosan as a Component of Pea-Fusarium solani Interactions

Chitosan, a polymer of beta-1,4-linked glucosamine residues with a strong affinity for DNA, was implicated in the pea pod-Fusarium solani interaction as an elicitor of phytoalexin production, an inhibitor of fungal growth and a chemical which can protect pea tissue from infection by F. solani f. sp. pisi. Purified Fusarium fungal cell walls can elicit phytoalexin production in pea pod tissue. Enzymes from acetone powders of pea tissue release eliciting components from the F. solani f. sp. phaseoli cell walls. Hydrochloric acid-hydrolyzed F. solani cell walls are about 20% glucosamine. The actual chitosan content of F. solani cell walls is about 1%. However, chitosan assays and histochemical observations indicate that chitosan content of F. solani spores and adjacent pea cells increases following inoculation. Dormant F. solani spores also accumulate chitosan. Concentrations of nitrous acid-cleaved chitosan as low as 0.9 microgram per milliliter and 3 micrograms per milliliter elicit phytoalexin induction and inhibit germination of F. solani macroconidia, respectively. When chitosan is applied to pea pod tissue with or prior to F. solani f. sp. pisi, the tissue is protected from infection.

Metabolites of the pathogenic fungusVerticillium dahliae. X. Induction of phytoalexins in the cotton plant by metabolites in the pathogen

The capacity of ten metabolites isolated from the culture liquid and mycelium of the phytopathogenic fungusVerticillium dahliae Kleb. and identified by chemical and spectral methods or of known chemical nature for inducing the synthesis and accumulation of phytoalexins in the cotton plant has been investigated for the first time. It has been established that the maximum inducing capacity among the metabolites investigated in the concentrations tried is possessed by elaidic acid and di-2-ethylhexyl phthalate. A considerable amount of phytoalexins is also induced under the action on the cotton plant of PCL-1 and a substance of carbohydrate nature. In view of the inducing capacity of the lipid component the protein-lipoid-polysaccharide complex we have studied its total lipid and fatty acid compositions. It has been shown that the compositions of the classes of neutral lipids of the protein-lipoid-polysaccharide complex of the mycelium and the culture liquid do not differ fundamentally, while there are certain differences in the qualitative composition and quantitative amounts of fatty acids of the protein-lipoid-poly-saccharide complex and of the culture liquid, with the exception of the C16:0 and C18:0 acids, the amounts of which are approximately the same.

A chemical dichotomy in phytoalexin induction within the tribe vicieae of the leguminosae

Using either leaf or cotyledon tissues, the phytoalexin response of over 60 representative members of the genera Lathyrus, Lens, Pisum and Vicia was determined. Twenty-nine Vicia species produced furanoacetylenes, as did the two Lens species examined. By contrast, Pisum and Lathyrus failed to produce any of these phytoalexins, but formed pisatin instead. Thus, pisatin was formed as the major phytoalexin in 29 of 31 Lathyrus species tested, representing 10 sections within the genus. This dichotomy in phytoalexin response is discussed in relation to the taxonomy of the tribe Vicieae.

Isoflavonoid phytoalexins of the genus Medicago

Detached leaves of 25 annual and perennial Medicago species have been inoculated with the fungus, Helminthosporium carbonum, and examined for subsequent production of isoflavonoid phytoalexins. Hydroxylated pterocarpan (medicarpin) and isoflavan (vestitol, sativan and isosativan) phytoalexins were obtained from most of the species investigated. Although large quantities of two flavonoid derivatives, isoliquiritigenin and liquiritigenin, were isolated from leaves of M. lupulina, this species was apparently unable to produce isoflavonoid compounds. Traces of 7-hydroxy-2′,4′-dimethoxyisoflavanone (a substance not previously recorded in the tribe Trifolieae) were obtained (together with medicarpin, vestitol and sativan) from M. sativa cv Du Puits. Some taxonomic aspects of phytoalexin induction are discussed with reference to the generic location of certain controversial Medicago species including M. (Trigonella) radiata.

Identification of medicarpin as a phytoalexin in the broad bean plant (Vicia faba L.)

THE analysis of phytoalexin induction has been proposed as a dynamic approach to plant systematics1. It has also been pointed out that production of the furanoacetylene wyerone acid2 by Vicia faba is an intriguing anomaly to the more general accumulation of isoflavanoid phytoalexins by members of the Leguminosae. In our studies on phytoalexin production by V. faba, three related furanoacetylenes (Fig. 1), wyerone acid (a), wyerone (b)3 and wyerone expoxide (c)4, have been identified as the major components of the multiple phytoalexin response of the broad bean plant to fungal infection5. In addition to these phytoalexins other minor inhibitors were detected in bioassays of thin layer chromatograms of methanol extracts of infected tissues5. Only one of these inhibitors reacted with p-nitroaniline6 to give an orange coloration indicating phenolic character. No anti-fungal compounds were detected in extracts of healthy tissues. Here we report the identification of the phenolic inhibitor as medicarpin (3-hydroxy-9-methoxy pterocarpan) (Fig. 1d), a phytoalexin present in a wide range of leguminous species1.

Phytoalexin induction as a new dynamic approach to the study of systematic relationships among higher plants

IT is well established that many higher plants respond to microbial invasion by the de novo production of organic substances called phytoalexins1,2. These compounds are absent from healthy plants and are induced by the attacking microorganisms. Although the role of phytoalexins in disease resistance is not yet entirely clear, considerable evidence suggests that they are of importance in the protection of higher plants from fungal colonisation. Although few surveys have been attempted, there is clearly a taxonomic element in phytoalexin biosynthesis, in that different plant families accumulate chemically different types of compounds3. Thus, the Leguminosae in general produce isoflavonoids, the Solanaceae diterpenes, the Compositae polyacetylenes and so on2; anomalies are rare, for example, the furanoacetylene, wyerone acid, from Vicia faba (Leguminosae)4. As lesser variations also occur within these families, there is the clear possibility of using phytoalexin induction as a tool in taxonomic studies. We report here the first successful application of this technique to the problems of classification at the generic and species level in the Leguminosae.

フィトアレキシン誘導のメカニズム

フィトアレキシン誘導のメカニズム

Phytoalexin Induction in Sweet Potato Roots by Amino Acids

Phytoalexin Induction in Sweet Potato Roots by Amino Acids

Phytoalexin Induction in Sweet Potato Roots by Amino Acids

(1974). Phytoalexin Induction in Sweet Potato Roots by Amino Acids. Agricultural and Biological Chemistry: Vol. 38, No. 12, pp. 2567-2568.

Mode of Pisatin Induction

Increases in phenylalanine ammonia lyase activity and pisatin synthesis were induced in excised pea pods (a) by basic polypeptides such as protamine, histone, lysozyme, cytochrome c, and ribonuclease; (b) by the polyamines spermine, spermidine, cadaverine, and putrescine, and (c) by the synthetic oligopeptides poly-l-lysine, poly-dl-ornithine, and poly-l-arginine.Poly-l-lysine (1 milligram per milliliter, molecular weight 7,200) was utilized as a model inducer of pisatin and phenylalanine ammonia lyase. The poly-l-lysine-induced responses could be inhibited by adding the RNA synthesis inhibitors cordycepin or alpha-amanitin to the pods prior to or at the time of inducer application. Cordycepin added 1.5 hours after inducer no longer completely inhibited induction. The application of poly-l-lysine was shown to characteristically change the rate of RNA synthesis within 30 minutes. Ultrastructural changes in pea nuclei were detected within 3 hours, and gross changes in nuclear morphology were apparent at 14 hours after inducer application. The physical appearance of uranyl acetate-stained chromatin isolated from poly-l-lysine 2 hours after inducer application differed from that of water-treated tissues. The template properties of chromatin extracted from pods 3 hours after inducer application were consistently superior to control chromatin when assayed with Escherichia coli RNA polymerase (without sigma factor). Chromatin from poly-l-lysine-induced tissue also bound 49% more actinomycin D-(3)H.The DNA-complexing properties of inducer compounds and the induced changes in the template and dye-binding properties of pea chromatin formed the basis for a proposed mode of action for phytoalexin induction.

Phytoalexin Induction in Beans Resistant or Susceptible to Fusarium and Thielaviopsis

Phytoalexin Induction in Beans Resistant or Susceptible to Fusarium and Thielaviopsis