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Proteomics of the effect of heavy metals: induced stress in transgenic tobacco plants, with different expression levels of trehalose-6-phosphate synthase

By 1984- Luis Filipe Pinheiro Domingues


Tese de mestrado. Biologia (Biologia Celular e Biotecnologia). Universidade de Lisboa, Faculdade de Ciências, 2012Presently, heavy metal contamination is an important pollution problem in world agriculture. The specific issues concerning heavy metal, in farming issues, are mainly due to their accumulation in the food chain with direct consequences to the consumer health. Cadmium (Cd), particularly, is one of the most challenging pollutants, due to its high water solubility, mobility, persistence and toxicity, even in low concentrations. Trehalose is a sugar that acts as an osmoprotector, and has the capacity to stabilize membranes and proteins during desiccation, heat and osmotic stress. Trehalose-6- phosphate synthase, one of the enzymes of the biosynthetic pathway has been the major target for genetic engineering, as it is considered to be the decisive factor in the trehalose biosynthesis. The aim of the present work is to study the effects of a Cd exposure of two transgenic Nicotiana tbacum lines (B5H and B1F), which had already shown enhanced tolerance to abiotic stress, in comparison to wild type. We used a proteomics approach, namely 2-DE-DIGE.and identification of differentially expressed proteins by mass spectrometry. In this work we could identify fifteen different proteins, such as carbonic anhydrase, RuBisCO and phosphoglycerate kinase, with differential expression. Results demonstrate that both B1F and B5H present an enhanced tolerance to cadmium induced stress, showing signals which denote that they may not be as affected as Wt plants by the concentration of cadmium to which they were subjected. The physiological significance of the expression profiles obtained is discussed. Genetic engineering with the trehalose-6-phosphate synthase gene seems therefore to be of use in obtaining transgenic plants of commercial use for possible growth in contaminated soils or used in phytoremediation.A contaminação dos solos por metais pesados é um importante problema ambiental. Um dos principais problemas da contaminação de solos agrícolas prende-se com a acumulação destes metais na cadeia alimentar com consequências directas, graves, na saúde do consumidor. O Cádmio (Cd), em particular, é um dos poluentes mais problemáticos, devido à sua elevada solubilidade em água, mobilidade, persistência e toxicidade, mesmo em baixas concentrações. A trealose é um açúcar que actua como osmoprotector e tem a capacidade de estabilizar membranas e proteínas durante stresses provocados por dissecação, temperatura, e stress osmótico. A Trealose-6-fosfato-sintase, uma das enzimas da via biossintética da trehalose tem sido o alvo principal para a engenharia genética, uma vez que e considerado o factor determinante na biossíntese de trealose. Com o presente trabalho pretende-se, utilizando uma abordagem proteómica, baseada em 2-DE-DIGE seguida de identificação de proteinas por espectrometria de massa, estudar os efeitos da uma exposição a Cd em duas linhas de Nicotiana tbacum (B5H e B1F), que já tinham demonstrado tolerância aumentada a stress abiótico induzido por metais pesados, em comparação com plantas Wild type. Neste trabalho foi possivel identificar 15 proteinas diferentes, tais como a anhidrase carbónica, a RuBisCO e a fosfoglicerato kinase, com expressão diferencial. Os resultados demonstram que tanto as plantas B1F como as B5H apresentam uma tolerância aumentada ao stresse induzido por cádmio., mostrando sinais que denotam que eles podem não ser tão afetados quanto plantas Wt pela concentração de cádmio a que foram submetidas. O significado fisiológico dos perfis de expressão obtidos é discutido. A engenharia genética com o gene da sintetase de trealose-6-fosfato parece, portanto, ser util na obtenção de plantas transgénicas de uso comercial para uma possivel plantação em solos contaminados ou utilizados em fitorremediação

Topics: Tabaco, Metais pesados, Transgénicos, Biotecnologia, Teses de mestrado - 2012
Year: 2012
OAI identifier: oai:repositorio.ul.pt:10451/8200

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