RESUMO
KEY MESSAGE: Expression of the TaMDC1 in transgenic tomato plants confer resistance to bacterial and fungal pathogens, as well as an insect pest and thus prove in planta function of the wheat cystatin. Cystatins are the polypeptides with cysteine proteinase inhibitory activities. Plant cystatins or phytocystatins are known to contribute to plant resistance against insect pests. Recently, increasing data proved that some of the phytocystatins also have antifungal activities in vitro. Here, we functionally characterized a wheat multidomain cystatin, TaMDC1, using in planta assays. Expression of TaMDC1 in wheat seedlings is up-regulated in response to methyl jasmonate and salicylic acid, indicating that TaMDC1 is involved in biotic stress responses mediated by these plant hormones. The TaMDC1 cDNA was integrated in tomato genome and expressed under cauliflower mosaic virus 35S promoter. Four transgenic plants that show high level of the transgene expression were selected by RNA gel blot and immunoblot analysis and utilized to assess biotic stress resistance against the bacterial pathogen Pseudomonas syringae, the fungal pathogens Botrytis cinerea and Alternaria alternata, and the insect pest Colorado potato beetle (CPB, Leptinotarsa decemlineata). Detached leaf inoculation assays revealed that the tomato plants expressing TaMDC1 showed high levels of resistance against P. syringae and A. alternata, and elevated tolerance against B. cinerea. Sustenance of L. decemlineata larvae to the transgenic plants demonstrated inhibition of CPB larvae growth. Inhibitory activity of TaMDC1 against selected pathogens was also demonstrated by in vitro assays with total protein extracted from transgenic tomato plants. Taken together, the presented data suggest that TaMDC1 is involved in a broad spectrum biotic stress resistance in planta.
Assuntos
Cistatinas/metabolismo , Resistência à Doença/genética , Doenças das Plantas/imunologia , Reguladores de Crescimento de Plantas/metabolismo , Triticum/genética , Acetatos/metabolismo , Animais , Antibacterianos/metabolismo , Antifúngicos/metabolismo , Botrytis/fisiologia , Besouros/fisiologia , Ciclopentanos/metabolismo , Cistatinas/genética , Expressão Gênica , Larva , Solanum lycopersicum/genética , Solanum lycopersicum/microbiologia , Solanum lycopersicum/parasitologia , Oxilipinas/metabolismo , Doenças das Plantas/microbiologia , Doenças das Plantas/parasitologia , Folhas de Planta/genética , Folhas de Planta/imunologia , Folhas de Planta/microbiologia , Folhas de Planta/parasitologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Pseudomonas syringae/fisiologia , Ácido Salicílico/metabolismo , Triticum/imunologia , Triticum/microbiologia , Triticum/parasitologiaRESUMO
Overwintering crops such as winter wheat display significant increase in freezing tolerance during a period of cold acclimation (CA). To gain better understanding of molecular mechanisms of CA, it is important to unravel functions and regulations of CA-associated genes. Differential screening of a cDNA library constructed from cold acclimated crown tissue of winter wheat identified three novel CA-associated cDNA clones. Nucleotide sequence analysis showed that the clones encode a high mobility globular protein (HMGB1), a glycine-rich RNA-binding protein (TaGRP2), and a LEA D-11 dehydrin (DHN14). Accumulation of the three mRNAs during 14 days of CA was differentially regulated. In response to drought, and ABA, DHN14 mRNA rapidly accumulated while HMGB1 and TaGRP2 mRNA levels remained unchanged. The possible functions of each of these genes in cold acclimation are discussed.
Assuntos
Aclimatação/genética , Temperatura Baixa , Regulação da Expressão Gênica de Plantas , Triticum/genética , Sequência de Bases , Northern Blotting , Clonagem Molecular , DNA Complementar/genética , DNA de Plantas/genética , Genes de Plantas , Dados de Sequência Molecular , Proteínas de Plantas/genética , Alinhamento de SequênciaRESUMO
The DNA sequence of a truncated cry1C gene encoding the active fragment of Bacillus thuringiensis (Bt) delta-endotoxin was fully reconstructed by introduction of silent mutations. Each of the truncated wild type and the synthetic genes encoding the active fragment of the protoxin was introduced into haploid tobacco plants under the control of the rbcS promoter. To facilitate selection of transgenic tobacco plants with high insecticidal activity, a fusion gene encoding both rat CYP1A1 cytochrome P450 and yeast NADPH-P450 oxidoreductase was cotransformed with the wild type cry1C gene. The synthetic gene elevated the levels of Cry1C protein and the mRNA in transgenic tobacco plants as well as mortality in Spodoptera litura larvae. The Cry1C protein was accumulated mainly in the leaf tissues of the transgenic tobacco plants. The results reported here imply that the green-tissue-specific expression of the synthetic cry1C gene is useful for the control of S. litura which was rather resistant to the other types of Bt toxins.
