Proteomic approaches using two-dimensional gel electrophoresis (2-DE) were adopted to identify proteins from rice leaf that are differentially expressed in response to the rice blast fungus, Magnaporthe grisea. Microscopic observation of inoculated leaf with M. grisea revealed that callose deposition and hypersensitive response was clearly visible in incompatible interactions but excessive invading hypha with branches were evident in compatible interactions. Proteins were extracted from leaves 24, 48, and 72 hours after rice blast fungus inoculation. Eight proteins resolved on the 2-DE gels were induced or increased in the inoculated leaf. Matrix-assisted laser desorption/ionization-time of flight analysis of these differentially displayed proteins showed them to be two receptor-like protein kinases (RLK), two beta-1.3-glucanases (Glu1, Glu2), thaumatin-like protein (TLP), peroxidase (POX 22.3), probenazole-inducible protein (PBZ1), and rice pathogenesis-related 10 (OsPR-10). Of these proteins, RLK, TLP, PBZ, and OsPR-10 proteins were induced more in the incompatible interactions than in compatible ones. A phytohormone, jasmonic acid also induced all eight proteins in leaves. To confirm whether the expression profile is equal to the 2-DE data, seven cDNA clones were used as probes in Northern hybridization experiments using total RNA from leaf tissues inoculated with incompatible and compatible rice blast fungal races. The genes encoding POX22.3, Glu1, Glu2, TLP, OsRLK, PBZ1, and OsPR-10 were activated in inoculated leaves, with TLP, OsRLK, PBZ1, and OsPR-10 being expressed earlier and more in incompatible than in compatible interactions. These results suggest that early and high induction of these genes may provide host plants with leading edges to defend themselves. The localization of two rice PR-10 proteins, PBZ1 and OsPR-10, was further examined by immunohistochemical analysis. PBZ1 accumulated highly in mesophyll cells under the attachment site of the appressorium. In contrast, OsPR-10 expression was mainly localized to vascular tissue.

Obg is a ribosome-associated GTPase essential for bacterial viability and is conserved in most organisms, from bacteria to eukaryotes. Obg is also expressed in plants, which predicts an important role for this molecule in plant viability; however, the functions of the plant Obg homologs have not been reported. Here, we first identified Arabidopsis AtObgC as a plant chloroplast-targeting Obg and elucidated its molecular biological and physiological properties. AtObgC encodes a plant-specific Obg GTPase that contains an N-terminal region for chloroplast targeting and has intrinsic GTP hydrolysis activity. A targeting assay using a few AtObgC N-terminal truncation mutants revealed that AtObgC localizes to chloroplasts and its transit peptide consists of more than 50 amino acid residues. Interestingly, GFP-fused full-length AtObgC exhibited a punctate staining pattern in chloroplasts of Arabidopsis protoplasts, which suggests a dimerization or multimerization of AtObgC. Moreover, its Obg fold was indispensable for the generation of the punctate staining pattern, and thus, was supposed to be important for such oligomerization of AtObgC by mediating the protein-protein interaction. In addition, the T-DNA insertion AtObgC null mutant exhibited an embryonic lethal phenotype that disturbed the early stage of embryogenesis. Altogether, our results provide a significant implication that AtObgC as a chloroplast targeting GTPase plays an important role at the early embryogenesis by exerting its function in chloroplast protein synthesis.

No abstract

The chloroplastic NADP-malic enzyme (NADP-ME) is a key enzyme of the C4 photosynthesis pathway in NADP-ME type C4 plants such as maize. To express the chloroplastic NADP-ME in leaves of a C3 plant, rice, full-length cDNAs encoding the rice C3-specific isoform and the maize C4-specific isoform of the enzyme were expressed under the control of the rice CAB: promoter. Transformants carrying the rice cDNA showed the NADP-ME activities in the leaves less than several-fold that of non-transformants, while those carrying the maize cDNA showed activities up to 30-fold that of non-transformants or about 60% of the NADP-ME activity of maize leaves. These results indicate that expression of the rice C3-specific NADP-ME is suppressed at co- and/or post-transcriptional levels by some regulation mechanisms intrinsic to rice, while that of the foreign C4-specific isoform can escape from such suppression. The accumulation of the maize C4-specific NADP-ME led to bleaching of leaf color and growth hindrance in rice plants under natural light. These deteriorative effects resulted from enhanced photoinhibition of photosynthesis due to an increase in the level of NADPH inside the chloroplast by the action of the maize enzyme.

No abstract