Genetics - Theses

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Author: Fulton, Lynette M. × End date: 2003 × Start date: 2003 × Type: PhD thesis ×

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    Characterisation of genes involved in L-Arabinose metabolism in Arabidopsis
    Fulton, Lynette M. (University of Melbourne, 2003)
    Glycosyl hydrolases are important mediators of plant cell wall modification during plant development. These enzymes catalyse the hydrolytic release of specific sugars, such as L-arabinose, from the polysaccharide-rich cell wall matrix. Here, the cloning and expression analysis of two genes, AtASD1 and AtASD1, encoding putative ?-L-arabinofuranosidases, is reported in Arabidopsis thaliana. AtASD1 and AtASD2 identities were assigned on the basis of homology to plant and microbial family 51 glycoside hydrolases. Despite numerous attempts, no enzyme activity was demonstrated for either protein. Over-expression of AtASD1 in the cell wall mutant, mur1, resulted in a genetic interaction that caused extreme dwarfism in plants. An AtASD2 T-DNA knockout mutant presented no visible phenotypes. Using the experimental approaches of RT-PCR, RNA gel blot analysis and GUS reporter gene expression analysis, AtASD1 and AtASD2 were shown to have different developmental expression profiles. High levels of AtASD1 promoter activity are present in multiple tissues during vegetative and reproductive growth. In comparison, AtASD2 expression is limited to the vasculature of older root tissue and to some floral organs and floral abscission zones. These findings implicate multiple and complex developmental roles for L-arabinosidases in addition to a simple role in carbon homeostasis. A previously described Arabidopsis mutant, ara1, exhibits a visible sensitivity to exogenous L-arabinose and encodes a defective L-arabinose kinase gene allele. The molecular characteristics of an ARA1 homologue, denoted ARA2, are reported. Expression studies revealed ARA1 is expressed during early vegetative growth, in floral organs and in conjunction with ARA2, developing pollen. Attempts to identify an ARA2 T-DNA mutant were made. Thus, a total of four genes that are involved in L-arabinose metabolism in Arabidopsis, are described. In summary, these findings have contributed significant insights into an intricate and highly regulated mechanism, whereby the concerted action of many enzymes mediate development-related cell wall remodelling that ultimately influences plant form and function.