Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis

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dc.contributor.advisorCoyne, Vernonen_ZA
dc.contributor.authorReddy, Amelia Francineen_ZA
dc.date.accessioned2014-09-02T17:08:16Z
dc.date.available2014-09-02T17:08:16Z
dc.date.issued2012en_ZA
dc.description.abstractGracilaria gracilis occurs naturally in Saldanha Bay, and was important to the South African export industry as it is a source of two important types of agar, namely food grade and sugar reactive agar. However, a number of abiotic and biotic factors, such as nutrient limitation, has virtually destroyed the G. gracilis resource. An understanding of the physiological response of the alga to nutrient limitation will aid in re-establishing and sustaining G. gracilis populations. However, modelling algal physiology and growth in response to abiotic stresses such as nutrient limitation requires an understanding of the underlying metabolic processes. The present study aimed to address this by investigating nitrogen metabolism and the mechanisms regulating nitrogen metabolism in G. gracilis. This was achieved by profiling changes in gene and protein expression, and activity of two major nitrogen metabolic enzymes, nitrate reductase and glutamine synthetase. Long term culture of G. gracilis in nitrogen replete and lacking conditions indicated that nutrient limitation causes a reduction in intracellular nitrogen and nitrogen protein stores such as phycoerythrin. When various sources were introduced to the culture medium to replenish nitrogen starved G. gracilis, changes in nitrate reductase and glutamine synthetase mRNA, protein and activity seemed to be dependent on the nutrient history of the cells, intracellular and extracellular nitrogen concentrations, metabolites of nitrogen assimilation and other metabolic processes such as carbon metabolism and photosynthesis. Nutrient studies suggested that multiple G. gracilis nitrate reductase and glutamine synthetase isoforms are present and differentially regulated via transcriptional, post-trancriptional, translational and posttranslational mechanisms. Furthermore, the insensitivity of these nitrogen metabolic enzymes to ammonium inhibition and the ability to alter the GS1:GS2 activity ratio possibly represents adaptive strategies developed by G. gracilis to survive nitrogen limitation. Immunocytochemical investigations confirmed the presence of multiple nitrate reductase and glutamine synthetase isoforms. The enzymes were successfully localised to the cell wall, chloroplast and cytosol of G. gracilis. A novel finding was the immuno-localisation of glutamine synthetase to intracellular starch granules. Overall, findings in the current study have suggested multiple roles for these metabolic enzymes that include nitrogen assimilation/transport, cell wall biosynthesis and senescence. This study led to the development of a model of the metabolic changes that occur in nitrogen replete and deplete G. gracilis and provides a firm foundation for future studies of the nitrogen stress response in G. gracilis. Characterisation of the G. gracilis nitrogen stress response may ultimately revive mariculture of this commercially important alga in South Africa.en_ZA
dc.identifier.apacitationReddy, A. F. (2012). <i>Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis</i>. (Thesis). University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology. Retrieved from http://hdl.handle.net/11427/6878en_ZA
dc.identifier.chicagocitationReddy, Amelia Francine. <i>"Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis."</i> Thesis., University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology, 2012. http://hdl.handle.net/11427/6878en_ZA
dc.identifier.citationReddy, A. 2012. Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis. University of Cape Town.en_ZA
dc.identifier.ris TY - Thesis / Dissertation AU - Reddy, Amelia Francine AB - Gracilaria gracilis occurs naturally in Saldanha Bay, and was important to the South African export industry as it is a source of two important types of agar, namely food grade and sugar reactive agar. However, a number of abiotic and biotic factors, such as nutrient limitation, has virtually destroyed the G. gracilis resource. An understanding of the physiological response of the alga to nutrient limitation will aid in re-establishing and sustaining G. gracilis populations. However, modelling algal physiology and growth in response to abiotic stresses such as nutrient limitation requires an understanding of the underlying metabolic processes. The present study aimed to address this by investigating nitrogen metabolism and the mechanisms regulating nitrogen metabolism in G. gracilis. This was achieved by profiling changes in gene and protein expression, and activity of two major nitrogen metabolic enzymes, nitrate reductase and glutamine synthetase. Long term culture of G. gracilis in nitrogen replete and lacking conditions indicated that nutrient limitation causes a reduction in intracellular nitrogen and nitrogen protein stores such as phycoerythrin. When various sources were introduced to the culture medium to replenish nitrogen starved G. gracilis, changes in nitrate reductase and glutamine synthetase mRNA, protein and activity seemed to be dependent on the nutrient history of the cells, intracellular and extracellular nitrogen concentrations, metabolites of nitrogen assimilation and other metabolic processes such as carbon metabolism and photosynthesis. Nutrient studies suggested that multiple G. gracilis nitrate reductase and glutamine synthetase isoforms are present and differentially regulated via transcriptional, post-trancriptional, translational and posttranslational mechanisms. Furthermore, the insensitivity of these nitrogen metabolic enzymes to ammonium inhibition and the ability to alter the GS1:GS2 activity ratio possibly represents adaptive strategies developed by G. gracilis to survive nitrogen limitation. Immunocytochemical investigations confirmed the presence of multiple nitrate reductase and glutamine synthetase isoforms. The enzymes were successfully localised to the cell wall, chloroplast and cytosol of G. gracilis. A novel finding was the immuno-localisation of glutamine synthetase to intracellular starch granules. Overall, findings in the current study have suggested multiple roles for these metabolic enzymes that include nitrogen assimilation/transport, cell wall biosynthesis and senescence. This study led to the development of a model of the metabolic changes that occur in nitrogen replete and deplete G. gracilis and provides a firm foundation for future studies of the nitrogen stress response in G. gracilis. Characterisation of the G. gracilis nitrogen stress response may ultimately revive mariculture of this commercially important alga in South Africa. DA - 2012 DB - OpenUCT DP - University of Cape Town LK - https://open.uct.ac.za PB - University of Cape Town PY - 2012 T1 - Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis TI - Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis UR - http://hdl.handle.net/11427/6878 ER - en_ZA
dc.identifier.urihttp://hdl.handle.net/11427/6878
dc.identifier.vancouvercitationReddy AF. Characterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilis. [Thesis]. University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology, 2012 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/6878en_ZA
dc.language.isoengen_ZA
dc.publisher.departmentDepartment of Molecular and Cell Biologyen_ZA
dc.publisher.facultyFaculty of Scienceen_ZA
dc.publisher.institutionUniversity of Cape Town
dc.titleCharacterisation of the effect of stress on nitrogen metabolism in the commercially important agarophyte, Gracilaria gracilisen_ZA
dc.typeDoctoral Thesis
dc.type.qualificationlevelDoctoral
dc.type.qualificationnamePhDen_ZA
uct.type.filetypeText
uct.type.filetypeImage
uct.type.publicationResearchen_ZA
uct.type.resourceThesisen_ZA
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