Expressão de genes de resposta a estresses ambientais em mutantes defectivos de milho e Arabidopsis

AUTOR(ES)
DATA DE PUBLICAÇÃO

2008

RESUMO

Reverse genetics and gene expression analysis has been the main approach adopted in order to carry out the functional characterization of genes. Results from such studies have typically demonstrated that same gene can be involved in different cell functions. In this context, analysis of trehalose metabolism in plants has revealed that this pathway plays an important regulatory role during development as well as in the response to diverse environmental stresses, mainly salinity. Although several works have attempted to clarify these mechanisms, the exact step in the pathway that actually has effect on the regulation of gene expression still remains unclear. Evidences strongly suggest that this role is associated with trehalose-6-phosphate synthase enzyme or trehalose-6-phosphate and trehalose pathways by-products. Accordingly, the first chapter of the presented thesis aimed to characterize the trehalose metabolic pathway in Arabidopsis thaliana by inhibiting trehalose degradation. Defective mutants for trehalase gene were found to have a 130-fold decrease in the transcription rate of this gene. This mutation led to a sensibility to trehalose and a deficient development of the root system. Nevertheless, the phenotype could be reverted by exogenous addition of sucrose to the medium. Otherwise, in substrate, no alteration in the plant development took place. Salinity tolerance trials revealed that trehalase gene inactivation did not protect Arabidopsis plants against damages caused by salt. Interestingly, mutation affected plant germination in osmotic and saline stresses conditions. Addiotionally, studies about gene expression in wide type plants, treated with trehalose and validamicin A, through microarray technology, allowed to verify that trehalose affects transcription factors expression, especially AP2/EREBP family. Through electronic Northern, it was verified that 14 genes encoding transcription factors are expressed, consistently, in roots and 35 are expressed in seeds. Among them, 10 genes are strongly induced by salinity. Such results suggest that trehalose metabolism may participate in both regulation of plant development and plant response to stress by modulating transcription factors expression. In the second chapter, a maize dek mutant was analyzed. Such mutants have been utilized, mainly, to identify genes involved in seeds development. Nevertheless, responses adopted by the plant in function of disturbances caused by mutations are little studied. Accordingly, this chapter aimed to characterize induced proteins in response to dek 827K pro1 mutation. For this end, protein profile of both wide type and defective seeds of dek 827k pro1 mutants was compared. In this analysis, a protein of about 35 kDa induced in defective seeds, was identified. Comparison of the NH2-terminal sequence of this protein in databanks revealed that the protein is similar to β -1,3-glucanases. The study of the expression of this gene has demonstrated that ZmGlucA is a seed-specific protein, being differentially expressed during the grain development. Nevertheless, in defective seeds, the expression of this gene is induced, taking place a high accumulation of this protein in dried seeds. The molecular grouping and the electronic Northern revealed that ZmGlucA is similar to rice and Arabidopsis β -1,3-glucanases regulated by environmental factors and phytormones. Several studies demonstrate that this protein family is involved in various biological processes. However, there are few works demonstrating that such enzyme is involved in seeds development. Similarity between ZmGlucA and responsive genes to different stimulus suggests that, in defective seeds, cells perceive disturbances caused by mutation as a stress and respond producing defense proteins. These data suggest that ZmGlucA participates in both defense and development of maize seeds.

ASSUNTO(S)

estresse ambiental -1,3-glucanase trehalose environmental stress β -1,3-glucanases trehalase trealose β salinidade trealase ciencias biologicas salinity

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