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Systemic acquired resistance (SAR) is a potent innate immunity system in plants that is effective against a broad range of pathogens. SAR in dicotyledonous plants such as tobacco and Arabidopsis has been partially elucidated and is mediated by salicylic acid (SA). However, the SAR mechanism of monocotyledonous rice plants remains to be clarified, although some similarities between SAR mechanisms in both types have been reported. Here we have characterized N-cyanomethyl-2-chloroisonicotinamide (NCI) as an effective SAR inducer in both plant species. Soil drench application of NCI induces a broad range of disease resistance in tobacco and rice and, more specifically, PR gene expression in tobacco. Both SA measurements in wild-type NCI-treated tobacco and pathogenic infection studies using NahG transgenic tobacco plants indicate that NCI-induced resistance enhancement does not require SA. Therefore, it is suggested that NCI induces SAR by triggering signaling at the same level as or downstream of SA accumulation as do both benzo(1,2,3)thiadiazole-7-carbothioic acid S-methyl ester and 2,6-dichloroisonicotinic acid. The fact that all of these chemicals are effective in rice and tobacco suggests that several common components function in disease resistance in both plant species.

The poly[(R)-3-hydroxyalkanoate] (PHA) synthase gene (phaC(Ac)) of Aeromonas caviae FA440 was modified by adding a peroxisome targeting signal encoding the last 10 amino acids at the carboxyl-terminal of spinach glycolate oxidase. The modified gene was introduced into Arabidopsis thaliana plants by Agrobacterium-mediated transformation. The transgenic Arabidopsis plant expressed the introduced gene and its protein, and it accumulated PHA in its tissues. Gas chromatography-mass spectrometry analysis demonstrated the accumulation of a novel type of PHA, poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxyhexanoate). This strongly suggests that short-chain-length (R)-3-hydroxyacyl-CoAs were generated from intermediates of peroxisomal beta-oxidation. It was revealed by using this transgenic plant that Tween-20 can activate peroxisomal beta-oxidation of short-chain-length fatty acids.

Multiple-gene transformation is required to improve or change plant metabolisms effectively; but this many-step procedure is time-consuming and costing. We succeeded in the metabolic engineering of tobacco plants by introducing multiple genes as a bacteria-type operon into a plastid genome. The tobacco plastid was transformed with a polycistron consisting of three bacterial genes for the biosynthesis of a biodegradable polyester, polyhydroxybutyrate (PHB). Accumulation of PHB in the leaves of the transgenic tobacco indicated that the introduced genes were polycistronically expressed. This "phyto-fermentation" system can be used in plant production of various chemical commodities and pharmaceuticals.

Probenazole (PBZ; 3-allyloxy-1,2-benzisothiazole-1,1-dioxide), which is the active ingredient in Oryzemate, has been used widely in Asia to protect rice plants against the rice blast fungus Magnaporthe grisea. To study PBZ's mode of action, we analyzed its ability, as well as that of its active metabolite 1, 2-benzisothiazol-3 (2H)-one 1,1-dioxide (BIT) to induce defense gene expression and resistance in Arabidopsis mutants that are defective in various defense signaling pathways. Wild-type Arabidopsis treated with PBZ or BIT exhibited increased expression of several pathogenesis-related genes, increased levels of total salicylic acid (SA), and enhanced resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC 3000 and the oomycete pathogen Peronospora parasitica Emco5. The role of several defense signaling hormones, such as SA, ethylene and jasmonic acid (JA), in activating resistance following PBZ or BIT treatment was analyzed using NahG transgenic plants and etr1-1 and coi1-1 mutant plants, respectively. In addition, the involvement of NPR1, a key component in the SA signaling pathway leading to defense responses, was assessed. PBZ or BIT treatment did not induce disease resistance or PR-1 expression in NahG transgenic or npr1 mutant plants, but it did activate these phenomena in etr1-1 and coi 1-1 mutant plants. Thus SA and NPR1 appear to be required for PBZ- and BIT-mediated activation of defense responses, while ethylene and JA are not. Furthermore, our data suggest that PBZ and BIT comprise a novel class of defense activators that stimulate the SA/NPR1-mediated defense signaling pathway upstream of SA.

Probenazole (PBZ) induces non-race specific resistance in rice plants against rice blast fungus and PBZ1 was identified as a PBZ-inducible gene from rice. The induction of PBZ1 expression in suspension-cultured rice cells was investigated. Northern blot analysis indicated that PBZ1 was induced by PBZ in a dose-dependent manner. Enzyme-linked immunosorbent assay (ELISA) showed a dose and time-dependent accumulation of PBZ1 protein. Both mRNA and protein analysis showed that PBZ1 was not induced by salicylic acid or an active metabolite, 1,2-benzisothiazole-1,1-dioxide.

The first step of C-P compound biosynthesis is a C-P bond formation reaction catalyzed by phosphoenolpyruvate phosphomutase, but this reaction favors the cleavage of the C-P bond. This C-P bond forming reaction is driven by the following reaction catalyzed by phosphonopyruvate (PnPy) decarboxylase. We have cloned and sequenced the gene (bcpC) encoding PnPy decarboxylase, a key enzyme of C-P compound biosynthesis, from the bialaphos (BA) producing microorganism Streptomyces hygroscopicus by complementation methods using Streptomyces wedmorensis NP-7, which is a mutant of a fosfomycin producing strain deficient in this step. The location of this gene in the BA biosynthetic gene cluster was determined by using the expression system in Streptomyces lividans. DNA sequencing of this gene revealed a 1203-bp open reading frame encoding a polypeptide of 401 amino acids.

The acetoacetyl-CoA reductase gene (phbB) of Ralstonia eutropha and the poly[(R)-(-)-3-hydroxyalkanoate] synthase gene (phaCAC) of Aeromonas caviae were introduced into tobacco plant by Agrobacterium mediated transformation method. The resulting transgenic tobacco expressed both introduced genes and the expression of these genes was confirmed by enzymatic analysis and western blotting. GC-MS analysis of the chloroform extract of tobacco leaves indicated that the transgenic plant produced biodegradable polyester, poly-[(R)-(-)-3-hydroxybutyrate]. GPC analysis indicated that the number-average molecular weights (Mn) and polydispersity (Mw/Mn) were 32,000 and 1.90, respectively.

The biosynthetic step following the phosphoenolpyruvate (PEP) phosphomutase reaction which forms a C-P bond of bialaphos was proven by the identification of phosphonopyruvate (PnPy) and phosphonoacetaldehyde (PnAA) as intermediates in the culture broth of Streptomyces hygroscopicus, a producing organism of bialaphos, and by detection of enzymatic decarboxylation of PnPy to PnAA. Purified PnPy decarboxylase turned out to require thiamine diphosphate and Mg2+ as cofactors. PnPy decarboxylase drives the unfavorable forward reaction to form PnPy catalyzed by PEP phosphomutase and is suggested to be essential to C-P compound biosynthesis.

We consider the primary cause of cubital tunnel syndrome caused by osteoarthrosis of the elbow joint to be degenerative osteophytes originating in the humeroulnar joint underneath the ulnar nerve. In the operative procedure the ulnar nerve in the cubital tunnel is fully exposed by cutting the soft tissues covering the nerve, because some of the soft tissues often seem to compress the nerve. Internal neurolysis is performed with microscopy in cases where the constriction is severe. Osteophytes in the humeroulnar joint underneath the nerve are removed. The osteophytes and loose bodies in the anterior and the posterior parts of the joint are also removed to increase joint motion. In 75 of 77 cases recovery of the ulnar nerve palsy and improvement in the range of joint motion were obtained.