Elicitation of Casbene Synthetase Activity in Castor Bean

Abstract

castor bean plant as a phytoalexin (18). A number of the biochemical characteristics of casbene biosynthesis have been elucidated (26).Substances which trigger the production of phytoalexins in higher plants are termed elicitors. A wide variety of fungal molecules, including polysaccharide wall components, polypeptides, glycoproteins and lipids, have been implicated as elicitors (25). Two lines of investigation have shown that fungal enzymes can also act as elicitors. Swinburne (22,23) has implicated a fungal proteinase as an elicitor of benzoic acid production in infected apple fruit. In addition, recent work in our laboratory (11,12) has shown that homogeneous a-1,4-endopolygalacturonase from culture filtrates of the fungus Rhizopus stolonifer elicits casbene synthetase activity in castor bean (R communis L.) seedlings. In both cases, heat-treatment of the enzyme preparations leads to equivalent losses of both enzymic activity and elicitor ability. This suggests that the elicitor abilities of these enzymes may be dependent on their catalytic activities.The work described in this paper demonstrates that pectic cell wall fragments released from particulate fractions of castor bean seedling homogenates through the action of R stolonifer endopolygalacturonase possess casbene synthetase elicitor activity and supports a model in which these elicitor fragments serve as obligate intermediates in the process of elicitation by the enzyme in vivo.Many species of higher plants have been shown to produce antifungal compounds known as phytoalexins (4,6,10). The presence of low or undetectable levels of phytoalexins in healthy plant tissues and the greatly increased accumulation of these substances in the region of fungal infection are distinctive features of phytoalexin production. Because of these characteristics and their general toxicities for a broad range of fungi, phytoalexins are believed to play a defensive role against invasion of the plant by fungal predators.Casbene is one of the five cyclic diterpene hydrocarbons produced from GGPP2 in cell-free extracts of young seedlings of the castor bean (Ricinus communis L.) (14,15 Figure 1. At the lower concentrations of elicitor, the amount of casbene synthetase activity follows a nearly linear relationship with the amount of elicitor tested. Higher levels of elicitor, however, saturate the response. The amount of EPGase tested in elicitor bioassays was usually between 0.2 and 0.3 units in order to produce responses which were in the approximately linear region of the dose response curve.Synthesis of Radiolabeled Geranylgeranyl Pyropbosphate. 4,4'-[3H]Geranylgeraniol (10.0 mCi/mmol) was synthesized from farnesylacetone in a manner similar to that described by Upper andWest (24) as later modified by Simcox (17). The tritium label was introduced into farnesylacetone by refluxing a solution of famesylacetone and tritiated water in dioxane for 40 h. A small amount of NaOH was added to the mixture prior to reflux to catalyze the exchange of hydrogens bound to carbons alpha to...

Host-Pathogen Interactions

“…This is similar to the elicitor activity of an endopolygalacturonase from Rhizopus stolonifer, which has been shown to be near maximally active as an elicitor of casbene synthetase in castor bean (Ricinus communis) at 0.05 units/ml enzyme activity (7). This suggested that the two enzymes may elicit by similar mechanisms.…”

“…It has also been shown that this R. stolonifer pectindegrading enzyme can release heat-stable elicitors from citrus pectin, polygalacturonic acid, and a particulate fraction obtained from castor bean seedlings (7). The results with the R. stolonifer and E. carotovora enzymes, considered in conjunction with the observation that pectic oligosaccharide elicitors can be obtained from several different types of plant cell walls by partial acid hydrolysis (14), suggests a general concept of elicitation by microbial pectin-degrading enzymes (7,14,34). Briefly, this concept proposes that plants have evolved a defense mechanism that responds to pectin-degrading enzymes in a manner that does not depend on recognizing the enzyme molecules themselves.…”

Host-Pathogen Interactions

“…Furthermore, these pectin-degrading en-zymes released elicitor-active oligogalacturonides from plant cell walls, citrus pectin, and polygalacturonic acid (4,(7)(8)(9)18 …”

“…(4,18,20,21) with an endopolygalacturonase produced by the fungus Rhizopus stolonifer. The R. stolonifer endopolygalacturonase induced accumulation of casbene synthetase (an enzyme involved in the synthesis of the phytoalexin, casbene) in castor bean seedlings (20) and released heat-stable, GalUA-rich elicitors of casbene synthetase from particulate fractions of castor bean seedling homogenates, citrus pectin, and polygalacturonic acid (4,18).…”

Exogenous application of liquid diet, previously fed upon by pea aphidsAcyrthosiphon pisum(Harris), to broad bean leaves induces volatiles attractive to the specialist parasitic waspAphidius ervi(Haliday)

“…This suggests that molecules other than enzymes may be involved in the elicitors. Oligosaccharides induce production of defensive compounds under the infection of phytopathogens in the Castor bean, Ricinus commais (Bruce and West 1982). A peptide, inceptin, is one of the elicitors which induce volatile production in Cowpea plants (Schmelz et al 2006).…”