Effects of Ca2+ on phytoalexin induction by fungal elicitor in soybean cells

A glucan elicitor from the cell walls of the fungus Phytophthora megasperma f.sp. glycinea caused increases in the activities of the phytoalexin biosynthetic enzymes, phenylalanine ammonia-lyase and chalcone synthase, and induced the production of the phytoalexin, glyceollin, in soybean (Glycine max) cell suspension cultures when tested in culture medium containing 1.2 mmol/liter Ca2+. Removal of extracellular Ca2+ by treatment with ethylene glycol bis(beta-aminoethyl ether)-N, N'-tetraacetic acid followed by washing the cells with Ca2+-free culture medium abolished the elicitor-mediated phytoalexin response. This suppression was largely reversed on readdition of Ca2+. Elicitor-mediated enhancement of biosynthetic enzyme activities and accumulation of glyceollin was strongly inhibited by La3+; effective concentrations for 50% inhibition were (mumol/liter) 40 for phenylalanine ammonia-lyase, 100 for chalcone synthase, and 30 for glyceollin. Verapamil caused similar effects only at concentrations higher than 0.1 mmol/liter, whereas trifluoperazine and 8-(diethylamino)-octyl-3,4,5-trimethoxybenzoate did not affect enzyme induction by the elicitor in the concentration range tested. Uptake of alpha-amino isobutyric acid into soybean cells, which was rapidly inhibited in the presence of the glucan elicitor, was not affected by La3+ nor was uptake inhibition by the elicitor relieved by La3+. The Ca2+ ionophore, A23187, enhanced phytoalexin biosynthetic enzyme activities and glyceollin accumulation in a dose-dependent manner, with 50% stimulation (relative to the elicitor) occurring at about 5 mumol/liter. The results suggest that the glucan elicitor causes changes in metabolite fluxes across the plasma membrane of soybean cells, among which changes in Ca2+ fluxes appear to be important for the stimulation of the phytoalexin response.     

Solubilization of soybean membrane binding sites for fungal beta-glucans that elicit phytoalexin accumulation

Soybean membranes contain high-affinity binding sites for fungal beta-glucans. These sites may play a role in the recognition by soybean tissues of fungal phytoalexin elicitors. We have solubilized beta-glucan-binding activity from microsomal membranes using two C12-alkyl zwitterionic detergents, Zwittergent 3-12 (ZW 3-12) and the lysolecithin analog 1-dodecanoyl propanediol-3-phosphorylcholine [corrected] (ES12H). The solubilized binding sites displayed identical affinity for beta-glucans as that found in membranes (KD = 11-34 nM). Detergent-protein micelles with glucan binding activity eluted with approximate Mr values of 300,000 in ZW 3-12 and 380,000 in ES12H in gel permeation chromatography. Maximal binding activity eluted from a chromatofocusing column in the pH range between 6.2 and 6.6 with both ES12H and ZW 3-12, suggesting an apparent pI close to neutral.     

Identification of a high-affinity binding protein for a hepta-beta-glucoside phytoalexin elicitor in soybean.

A putative receptor protein for a hepta-beta-glucoside phytoalexin elicitor was identified by photoaffinity labeling of detergent-solubilized proteins from soybean root membranes. Incubation of partially purified beta-glucan-binding proteins with a photolabile 125I-labeled 2-(4-azidophenyl)ethyl-amino conjugate of the heptaglucoside elicitor, followed by irradiation with ultraviolet light (366 nm) resulted in specific labeling of a 70-kDa band in SDS/PAGE. Half-maximal inhibition of the 125I-labeling of the protein band by underivatized hepta-beta-glucoside was achieved by 15 nM heptaglucoside. Analysis of the affinity of radiolabel incorporation into the protein by ligand-saturation experiments, gave an apparent Kd value of 3 nM, in full agreement with the results from radioligand-binding studies. Good correlation was also observed between the amount of radiolabel incorporated into the protein and the binding activity of the fractions obtained at different stages in the purification of heptaglucoside-binding activity. Photoaffinity labeling of proteins purified by glucan-affinity chromatography showed the 70-kDa band as the main component along with weak 125I-labeling of a 100-kDa band. The 70-kDa band was also the major protein visualized by silver staining after SDS/PAGE of this fraction, suggesting that it is the predominant form of the heptaglucoside-binding proteins in detergent-solubilized soybean membranes.     

Rhizosphere salinity and phytoalexin accumulation in soybean

Summary Rhizosphere salinity decreased the capacity of soybean to accumulate a pterocarpanoid phytoalexin (glyceollin) in the stem in response toPhytophthora megasperma var.sojae. Rapid (48h) accumulation was depressed by NaCl, Na₂SO₄, CaCl₂ and MgSO₄ applications. Time-course accumulations was slowed by applications. Time-course accumulation was slowed by application of 0.131M NaCl. Glyceollin accumulation was also reduced in plants subjected to a period of high salinity stress (0.177M NaCl, 72 h) after a period of nonsalinized growth. Calcium chloride completely suppressed glyceollin accumulation in normally-resistant plants but no susceptibility to the fungus was observed.     

Phytoalexin synthesis in soybean cells: elicitor induction of phenylalanine ammonia-lyase and chalcone synthase mRNAs and correlation with phytoalexin accumulation

A glucan elicitor from cell walls of the fungus Phytophthora megasperma f. sp. glycinea, a pathogen of soybean (Glycine max), induced large and rapid increases in the activities of enzymes of general phenylpropanoid metabolism, phenylalanine ammonia-lyase, and of the flavonoid pathway, acetyl-CoA carboxylase and chalcone synthase, in suspension-cultured soybean cells. The changes in phenylalanine ammonia-lyase and chalcone synthase activities were correlated with corresponding changes in the mRNA activities encoding these enzymes, as determined by enzyme synthesis in vitro in a mRNA-dependent reticulocyte lysate. The time courses of the elicitor-induced changes in mRNA activities for both enzymes were very similar with respect to each other. Following the onset of induction, the two mRNA activities increased significantly at 3 h, reached highest levels at 5 to 7 h, and subsequently returned to low values at 10 h. Similar degrees of induction of mRNA activities and of the catalytic activities of phenylalanine ammonia-lyase and chalcone synthase were observed in response to three diverse microbial compounds, the glucan elicitor from P. megasperma, xanthan, an extracellular polysaccharide from Xanthomonas campestris, and endopolygalacturonase from Aspergillus niger. However, whereas the glucan elicitor induced the accumulation of large amounts of the phytoalexin, glyceollin, in soybean cells, endopolygalacturonase induced only low, albeit significant, amounts; xanthan did not enhance glyceollin accumulation under the conditions of this study. This result might imply that enzymes other than phenylalanine ammonia-lyase or chalcone synthase exert an important regulatory function in phytoalexin synthesis in soybean cells.     

Nitric oxide synthase-mediated phytoalexin accumulation in soybean cotyledons in response to the Diaporthe phaseolorum f. sp. meridionalis elicitor

Phytoalexin biosynthesis is part of the defense mechanism of soybean (Glycine max) plants against attack by the fungus Diaporthe phaseolorum f. sp. meridionalis (Dpm), the causal agent of stem canker disease. The treatment of soybean cotyledons with Dpm elicitor or with sodium nitroprusside (SNP), a nitric oxide (NO) donor, resulted in a high accumulation of phytoalexins. This response did not occur when SNP was replaced by ferricyanide, a structural analog of SNP devoid of the NO moiety. Phytoalexin accumulation induced by the fungal elicitor, but not by SNP, was prevented when cotyledons were pretreated with NO synthase (NOS) inhibitors. The Dpm elicitor also induced NOS activity in soybean tissues proximal to the site of inoculation. The induced NOS activity was Ca(2+)- and NADPH-dependent and was sensitive to the NOS inhibitors N(G)-nitro-L-arginine methyl ester, aminoguanidine, and L-N(6)-(iminoethyl) lysine. NOS activity was not observed in SNP-elicited tissues. An antibody to brain NOS labeled a 166-kD protein in elicited and nonelicited cotyledons. Isoflavones (daidzein and genistein), pterocarpans (glyceollins), and flavones (apigenin and luteolin) were identified after exposure to the elicitor or SNP, although the accumulation of glyceollins and apigenin was limited in SNP-elicited compared with fungal-elicited cotyledons. NOS activity preceded the accumulation of these flavonoids in tissues treated with the Dpm elicitor. The accumulation of these metabolites was faster in SNP-elicited than in fungal-elicited cotyledons. We conclude that the response of soybean cotyledons to Dpm elicitor involves NO formation via a constitutive NOS-like enzyme that triggers the biosynthesis of antimicrobial flavonoids.     

Structure-activity relationships of oligo-beta-glucoside elicitors of phytoalexin accumulation in soybean.

The abilities of a family of chemically synthesized oligo-beta-glucosides, ranging in size from hexamer to decamer, to induce phytoalexin accumulation in soybean cotyledons were investigated to determine which structural elements of the oligoglucosides are important for their biological activity. The results of the biological assays established that the following structural motif is necessary for the oligo-beta-glucosides to have high elicitor activity: [formula; see text] The branched trisaccharide at the nonreducing end of the oligoglucosides was found to be essential for maximum elicitor activity. Substitution of either the nonreducing terminal backbone glucosyl residue or the side-chain glucosyl residue closest to the nonreducing end with glucosaminyl or N-acetylglucosaminyl residues reduced the elicitor activity of the oligoglucosides between 10-fold and 10,000-fold. Elicitor activity was also reduced 1000-fold if the two side-chain glucosyl residues were attached to adjacent backbone glucosyl residues rather than to glucosyl residues separated by an unbranched residue. In contrast, modifications of the reducing terminal glucosyl residue of an elicitor-active hepta-beta-glucoside by conjugation with tyramine and subsequent iodination had no significant effect on the elicitor activity of the hepta-beta-glucoside. These results demonstrate that oligo-beta-glucosides must have a specific structure to trigger the signal transduction pathway, which ultimately leads to the de novo synthesis of phytoalexins in soybean.     

Effects of fungal elicitor on lignin biosynthesis in cell suspension cultures of soybean

Soybean (Glycine max L.) cells cultured in B5 medium produce extremely low amounts of lignin. However, modification in the growth medium, by lowering the concentration of NO⁻₃ and PO²⁻₄, results in the lignification of these cells without affecting levels of cell wall-esterified 4-coumaric and ferulic acid. The production of an extracellular, macromolecular complex by the cultured soybean cells (Moore TS Jr 1973 Plant Physiol 51: 529-536) allows a rapid, nondestructive solubilization of the lignin which can be estimated by reaction with phloroglucinol in free solution. This system has been used to study the effects of fungal elicitor on the synthesis of lignin in soybean cells. The inclusion of very low levels of an elicitor fraction from the cell walls of Phytophthora megasperma in the medium in which lignification of the soybean cells occurs suppressed both the accumulation of extracellular lignin and phloroglucinol staining of the cell walls without affecting the levels of bound hydroxycinnamic acids. The activity profiles of phenylalanine ammonia-lyase (EC 4.3.1.5) and isoenzymes of 4-coumarate:CoA ligase (EC 6.2.1.12) were compared in lignifying and elicitor-treated cell cultures as was the activity of chalcone synthase, an enzyme of flavonoid biosynthesis. The measured activities of these enzymes in cell cultures treated with elicitor were considerably lower than in untreated cells.     

A specific, high-affinity binding site for the hepta-beta-glucoside elicitor exists in soybean membranes.

The presence of a specific binding site for a hepta-beta-glucoside elicitor of phytoalexin accumulation has been demonstrated in soybean microsomal membranes. A tyramine conjugate of the elicitor-active hepta-beta-glucoside was prepared and radiolabeled with 125I. The labeled hepta-beta-glucoside-tyramine conjugate was used as a ligand in binding assays with a total membrane fraction prepared from soybean roots. Binding of the radiolabeled hepta-beta-glucoside elicitor was saturable, reversible, and with an affinity (apparent Kd = 7.5 x 10(-10) M) comparable with the concentration of hepta-beta-glucoside required for biological activity. A single class of hepta-beta-glucoside binding sites was found. The binding site was inactivated by proteolysis and by heat treatment, suggesting that the binding site is a protein or glycoprotein. Competitive inhibition of binding of the radiolabeled hepta-beta-glucoside elicitor by a number of structurally related oligoglucosides demonstrated a direct correlation between the binding affinities and the elicitor activities of these oligoglucosides. Thus, the hepta-beta-glucoside-binding protein fulfills criteria expected of a bona fide receptor for the elicitor-active oligosaccharin.     

真菌激发子对桑黄胞内代谢产物积累的影响

利用前期通过大量预试验筛选获得的3种真菌多糖激发子,在桑黄Sanghuangporus sanghuang发酵液中进行添加,研究其对桑黄胞内黄酮、三萜及多糖等代谢产物积累的影响,并对激发效果好的激发子继续进行添加量和激发时间的优化。结果显示:参试的3种真菌多糖激发子在一定程度上均能促进桑黄胞内黄酮和胞内三萜类成分的积累,但对胞内多糖积累的影响不明显;来源菌株为薄多年卧孔菌Perenniporia tenuis的激发子表现最突出,激发效果显著且稳定性良好。激发条件优化的结果表明:在接种的同时按照50mg/L的终浓度进行激发子的添加,激发6d,桑黄胞内黄酮含量达最高值0.7%,比对照提高79.49%;在接种1d后,按照50mg/L的终浓度进行激发子的添加,激发5d,对桑黄胞内三萜类成分代谢的激发效果最显著,胞内三萜含量最高可达5.62%,是对照组的3.43倍。     

Comparison of proteinase inhibitor-inducing activities and phytoalexin elicitor activities of a pure fungal endopolygalacturonase, pectic fragments, and chitosans

Rhizopus stolonifer endopolygalacturonase, an elicitor of casbene synthetase activity in castor bean seedlings, was found to be a potent elicitor of the phytoalexin pisatin in pea pods and of proteinase Inhibitor I in tomato leaves. The enzyme was an active elicitor or inducer only in its active native state; heat-denatured enzyme was inactive in all three systems. The activities of (a) the tomato pectic polysaccharide proteinase inhibitor-inducing factor, (b) a partially acid hydrolyzed proteinase inhibitor-inducing factor, (c) citrus pectic fragments, and (d) chitosan, were also compared in the three bioassay systems. The four oligosaccharide preparations were active in all three systems, but with different degrees of potency. In tomato leaves and pea pods, chitosans were most active, whereas in castor beans, the citrus pectic fragments were the best elicitors. The data presented support the hypothesis that plant and fungal cell wall fragments are important signals in mobilizing a wide variety of biochemically different types of plant defense responses, and that endopolygalacturonases play a key role in releasing the plant cell wall fragments during pest attacks.     

Tropospheric ozone as a fungal elicitor

Tropospheric ozone has been proven to trigger biochemical plant responses that are similar to the ones induced by an attack of fungal pathogens, i.e. it resembles fungal elicitors. This suggests that ozone can represent a valid tool for the study of stress responses and induction of resistance to pathogens. This review provides an overview of the implications of such a phenomenon for basic and applied research. After an introduction about the environmental implications of tropospheric ozone and plant responses to biotic stresses, the biochemistry of ozone stress is analysed, pointing out its similarities with plant responses to pathogens and its possible applications.     

A highly convergent and effective synthesis of the phytoalexin elicitor hexasaccharide.

The peracetylated hexasaccharide 1,2,4-tri-O-acetyl-3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-6- O- (2,3,4-tri-O-acetyl-6-O-(2,4-di-O-acetyl-3,6-di-O-(2,3,4,6-tetra-O-acety l- beta-D-glucopyranosyl)-beta-D-glucopyranosyl)-beta-D-glucopyranosyl)-alp ha, beta-D-glucopyranose 21 was synthesized in a blockwise manner, employing trisaccharide trichloroacetimidate 2,4-di-O-acetyl-3,6-di-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)- alpha-D-glucopyranosyl trichloroacetimidate 17 as the glycosyl donor, and trisaccharide 4-O-acetyl-3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-6-O-(2,3,4 -tri -O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S)ethylidene-alpha-D-glucopyra nose 18 as the acceptor. The donor 17 and acceptor 18 were readily prepared from trisaccharides 3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-6-O-(2,3,4-tri-O-acet yl- 6-O-chloroacetyl-beta-D-glucopyranosyl)-1,2-O-(R,S)ethylidene-alpha-D- glucopyranose 10 and 3,6-di-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S) ethylidene-alpha-D-glucopyranose 11, respectively, which were obtained from rearrangement of orthoesters 3,4-di-O-acetyl-6-O-chloroacetyl-alpha-D-glucopyranose 1,2-(3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S) ethylidene-alpha-D-glucopyranosid-6-yl orthoacetate) 8 and 3,4,6-tri-O-acetyl-alpha-D-glucopyranose 1,2-(3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S) ethylidene-alpha-D-glucopyranosid-6-yl orthoacetate) 9, respectively. The orthoesters were prepared from selective coupling of the disaccharide 3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S) ethylidene-alpha-D-glucopyranose 4 with 'acetobromoglucose' (tetra-O-acetyl-alpha-D-glucopyranosyl bromide) and 6-O-chloroacetylated 'acetobromoglucose', respectively. To confirm the selectivity of the orthoester formation and rearrangement, the disaccharide 4-O-acetyl-3-O-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl)-1,2-O-(R,S ) ethylidene-alpha-D-glucopyranose 7 was prepared from 4 by selective tritylation, acetylation and detritylation. The title compound, an elicitor-active D-glucohexaose 3-O-(beta-D-glucopyranosyl)-6-O-(6-O-(3,6-di-O-(beta-D-glucopyranosyl)-b eta -D-glucopyranosyl)-beta-D-glucopyranosyl)-alpha,beta-D-glucopyranose 1, was finally obtained by Zemplén deacetylation of 21 in quantitative yield.     

Development of a radioimmunoassay for the soybean phytoalexin glyceollin I

A radioimmunoassay for glyceollin I, the major phytoalexin produced by soybean (Glycine max [L.] Merr.), has been developed. Antibodies were raised in rabbits against a glyceollin I-bovine serum albumin conjugate. The antisera were used to establish a radioimmunoassay for glyceollin I using [¹²⁵I]glyceollin I as the tracer. A logit plot of a standard concentration series yielded a straight line in the range of 1 to 100 picomoles (0.34-34 nanograms) of glyceollin I. The structurally related pterocarpan phytoalexins, glyceollins II and III, glyceollidin II and glycinol, which also accumulate in infected soybean tissue, show a low cross-reactivity in the radioimmunoassay (0.5-5% at 50% displacement of the tracer). Two related isoflavones present constitutively in soybean tissue, daidzein and genistein, have cross-reactivities of less than 0.84% and 1.1%, respectively. The radioimmunoassay permitted the quantitative determination of glyceollin I in 15-micrometer microtome sections of soybean hypocotyl tissue infected with zoospores of Phytophthora megasperma f. sp. glycinea.     

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--Glucanase activity was induced to about the same level throughout all growth stages, with a large contribution by an endogenous mechanism at late stages.Abbreviations PAL Phenylalanine ammonia-lyase (EC 4.3.1.5) - 4CL 4-Coumarate:CoA ligase (EC 6.2.1.12)     

Cholic acid, a bile acid elicitor of hypersensitive cell death, pathogenesis-related protein synthesis, and phytoalexin accumulation in rice

When plants interact with certain pathogens, they protect themselves by generating various defense responses. These defense responses are induced by molecules called elicitors. Since long ago, composts fermented by animal feces have been used as a fertilizer in plant cultivation, and recently, have been known to provide suppression of plant disease. Therefore, we hypothesized that the compounds from animal feces may function as elicitors of plant defense responses. As a result of examination of our hypothesis, an elicitor of rice defense responses was isolated from human feces, and its structure was identified as cholic acid (CA), a primary bile acid in animals. Treatment of rice (Oryza sativa) leaves with CA induced the accumulation of antimicrobial compounds (phytoalexins), hypersensitive cell death, pathogenesis-related (PR) protein synthesis, and increased resistance to subsequent infection by virulent pathogens. CA induced these defense responses more rapidly than did fungal cerebroside, a sphingolipid elicitor isolated from the rice pathogenic fungus Magnaporthe grisea. Furthermore, fungal cerebroside induced both types of rice phytoalexins, phytocassanes and momilactones, whereas CA mainly induced phytocassanes, but not momilactones. In the structure-activity relationship analysis, the hydroxyl groups at C-7 and C-12, and the carboxyl group at C-24 of CA contributed to the elicitor activity. These results indicate that CA is specifically recognized by rice and is a different type of elicitor from fungal cerebroside. This report demonstrated that bile acid induced defense responses in plants.     

The receptor for the fungal elicitor ethylene-inducing xylanase is a member of a resistance-like gene family in tomato

An ethylene-inducing xylanase (EIX) is a potent elicitor of plant defense responses in specific cultivars of tobacco (Nicotiana tabacum) and tomato (Lycopersicon esculentum). The LeEix locus in tomatoes was characterized by map-based cloning, which led to the identification of a novel gene cluster from which two members (LeEix1 and LeEix2) were isolated. Similar to the tomato Ve resistance genes in tomato plants, the deduced amino acid sequences encoded by LeEix1 and LeEix2 contain a Leu zipper, an extracellular Leu-rich repeat domain with glycosylation signals, a transmembrane domain, and a C-terminal domain with a mammalian endocytosis signal. Silencing expression of the LeEix genes prevented the binding of EIX to cells of an EIX-responsive plant and thus inhibited the hypersensitive response. Overexpression of either LeEix1 or LeEix2 genes in EIX-nonresponsive tobacco plants enabled the binding of EIX, although only LeEix2 could transmit the signal that induced the hypersensitive response. Overexpressing LeEix2 in mammalian COS-7 cells enables binding of EIX, indicating physical interaction between the EIX elicitor and LeEix2 gene product. Structural analysis of the LeEix proteins suggests that they belong to a class of cell-surface glycoproteins with a signal for receptor-mediated endocytosis. Mutating the endocytosis signal in LeEix2 (Tyr 993 to Ala) abolished its ability to induce the hypersensitive response, suggesting that endocytosis plays a key role in the signal transduction pathway.     

Common responses of cultured soybean cells to 2,4-D starvation and fungal elicitor treatment

The patterns of in vivo protein synthesis in soybean cell suspensions were compared by polyacrylamide gel electrophoresis after the cells had been submitted to different stress conditions : treatment with Phytophthora megasperma (Pmg) cell wall elicitors, 2,4-D starvation and heat shock (HS) temperatures. Changes in protein synthesis patterns induced after elicitation of cell suspensions or after infection of soybean hypocotyls by Pmg were found to be similar to changes brought about by auxin starvation of the cells. Changes common to both stress situations involve a prominent 17 kDa peptide family and 27, 29, 35 and about 45 kDa peptides. Moreover, defense reactions, i.e. glyceollin accumulation and synthesis of chalcone synthase (CHS) were also strongly stimulated in auxin-starved cells. On the contrary, although characteristic sets of low molecular weight heat shock (HS) proteins were synthesized by cells grown at 37°C, no clear similarity was observed with peptides characteristic of auxin-starved cells.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - Pmg Phytophthora megasperma Drechs f.sp.glycinea - HS heat shock - PR pathogenesis-related - SDS-PAGE sodium dodecylsulfate-polyacrylamide gel electrophoresis - IEF isoelectrofocusing - iP isoelectric point - kDa kilodalton - P17 17 kDa peptide group of soybean cells cultured in vitro - CHS chalcone synthase     

Glyceollin, a soybean phytoalexin with medicinal properties

This review covers the biosynthesis of glyceollin and its biological activities including antiproliferative/antitumor action (toward B16 melanoma cells, LNCaP prostate cancer cells, and BG-1 ovarian cancer cells), anti-estrogenic action (through estrogen receptors ??- and ??-), antibacterial action (toward Erwinia carotovora, Escherichia coli, Bradyrhizobium japonicum, Sinorhizobium fredii ), antinematode activity, and antifungal activity (toward Fusarium solani, Phakospora pachyrhizi, Diaporthe phaseolorum, Macrophomina phaseolina, Sclerotina sclerotiorum, Phytophthora sojae, Cercospora sojina, Phialophora gregata, and Rhizoctonia solani). Other activities include insulinotropic action and attenuation of vascular contractions in rat aorta.     

Investigation of the mechanism of phytoalexin accumulation in soybean induced by glucan or mercuric chloride

Soybean cotyledons which had been treated with glucan from Phytophthora megasperma f.sp. glycinea or with mercuric chloride were pulse-labeled with ¹⁴CO₂ and then the ¹⁴C-incorporation into the phytoalexins was determined. The kinetics of ¹⁴C-incorporation into phytoalexins (glyceollin isomers and 3,6α,9-trihydroxypterocarpan) was very similar with the two types of elicitors. Metabolic rates of phytoalexins were determined by pulse-chase experiments. The apparent half-life of metabolism was about 100 h for glyceollin with either glucan or HgCl₂. The half-lives for trihydroxypterocarpan were 39 h with glucan and 14 h with HgCl₂. According to our results levels of glyceollins in soybean cotyledons are mainly controlled by their rates of synthesis. Biotic (glucan) and abiotic (HgCl₂) elicitors have similar induction effects. Both types of elicitors could act by effecting the release of endogenous elicitors.