Suppression of mRNAs for lipoxygenase (LOX), allene oxide synthase (AOS), allene oxide cyclase (AOC) and 12-oxo-phytodienoic acid reductase (OPR) in pea reduces sensitivity to the phytotoxin coronatine and disease development by Mycosphaerella pinodes

Using a recently developed model pathosystem involving Medicago truncatula and Mycosphaerella pinodes, causal agent of Mycosphaerella blight on pea to understand host molecular response to a fungal suppressor, we applied the suppressor to leaves of M. truncatula and identified 151 nonredundant cDNA fragments as newly expressed genes. These included genes encoding lipoxygenase (LOX) and enoyl-CoA hydratase, which are presumably involved in jasmonic acid (JA) synthesis. Potential genes encoding plastidic enzymes, including allene oxide synthase (AOS) and allene oxide cyclase (AOC), and other peroxisomal enzymes involved in β-oxidation were predicted from the Medicago Gene Index EST database and tested for altered expression by semiquantitative RT-PCR. The coordinated expression of genes encoding both plastidic and peroxisomal enzymes showed that the suppressor likely conditions certain cellular process(es) through the JA synthesis in M. truncatula. To explore the role of JA or JA-regulated cellular process(es) in conditioning susceptibility, we used an Apple latent spherical virus (ALSV)-based virus-induced gene silencing (VIGS) technology to silence pea genes including LOX, AOS, AOC and 12-oxo-phytodienoic acid reductase (OPR). In LOX-, AOS-, AOC- or OPR-silenced pea plants, disease development induced by M. pinodes was remarkably reduced. Similarly, silencing of mRNA for LOX, AOS, AOC or OPR reduced the sensitivity to a phytotoxin, coronatine, which is believed to act through a JA-dependent process. On the basis of these results, it is conceivable that M. pinodes has evolved a strategy to condition susceptibility by manipulating the physiology of host cells, in particular JA-regulated cellular process(es), to promote disease development in pea.     

Plant cell walls as suppliers of potassium and sodium ions for induced resistance in pea (Pisum sativum L.) and cowpea (Vigna unguiculata L.)

When an elicitor is applied to plants to induce resistance, one of the first detectable events is the efflux of ions from the treated tissue. Here we are the first to demonstrate that an elicitor from Mycosphaerella pinodes evokes leakage of Na⁺ and K⁺ ions from isolated cell walls of pea and cowpea in vitro, as observed for epicotyl tissues. Pharmacological experiments showed that this elicitor-stimulated leakage was sensitive to vanadate and N-(3-methylphenyl)biphenyl-4-sulfonamide (NGXT-191), that inhibit a cell wall-associated ATPase (apyrase). Vanadate or NGXT-191 suppressed elicitor-induced superoxide generation and expression of defense genes in vivo. On the basis of these results, we assume that the leakage of these ions, probably associated with an ATP-dependent process(es) in the cell wall, is likely associated with induced defenses of pea and cowpea.     

Glycosylation of flagellin from Pseudomonas syringae pv. tabaci 6605 contributes to evasion of host tobacco plant surveillance system

Pseudomonas syringae pv. tabaci (Pta) possesses a genetic region composed of two open reading frames (ORFs), fgt1 and fgt2, that are involved in glycosylation of flagellin. The deletion mutant Δfgt1 produced non-glycosylated flagellin, and exhibited reduced ability to cause disease in the host tobacco plant. Flagellin is known to induce plant defense responses, and the recognition of flagellin by Arabidopsis thaliana is mediated by a conserved N-terminal region, flg22, in flagellin and a leucine-rich repeat domain in the FLS2 receptor. Because flg22 localizes inside the flagellum, polymerized flagellum needs to be dissociated to be recognized. Therefore, the effect of glycosylation on flagella stability was investigated. The polymerized flagella from glycosylated flagellins were more resistant to heat treatment than those from non-glycosylated flagellins, suggesting that the glycosylation of flagellin contributes to the structural stability of flagella and prevents exposure of the flg22 region. Polymerized flagella from Pta Δfgt1 flagellin and depolymerized and glycosylated flagellin from Pta wild type induced cell death and callose deposition, and inhibited seedling growth in tobacco more effectively, whereas polymerized flagella from Pta wild-type flagellin caused a low level of these responses. These results suggest Pta might have evolved the flagellin glycosylation system to evade detection and defense response of a host by increasing flagella stability and suppressing their dissociation.