Browsing by Author "Du Plessis, Michelle"

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    Carnitine requires choline to exert physiological effects in saccharomyces cerevisiae
    (Frontiers Media, 2018-07-02) Du Plessis, Michelle; Franken, Jaco; Bauer, Florian; De Biase, Daniela
    L-Carnitine is a key metabolite in the energy metabolism of eukaryotic cells, functioning as a shuttling molecule for activated acyl-residues between cellular compartments. In higher eukaryotes this function is essential, and defects in carnitine metabolism has severe effects on fatty acid and carbon metabolism. Carnitine supplementation has been associated with an array of mostly beneficial impacts in higher eukaryotic cells, including stress protection and regulation of redox metabolism in diseased cells. Some of these phenotypes have no obvious link to the carnitine shuttle, and suggest that carnitine has as yet unknown shuttle-independent functions. The existence of shuttle-independent functions has also been suggested in Saccharomyces cerevisiae, including a beneficial effect during hydrogen peroxide stress and a detrimental impact when carnitine is co-supplemented with the reducing agent dithiothreitol (DTT). Here we used these two distinct yeast phenotypes to screen for potential genetic factors that suppress the shuttle independent physiological effects of carnitine. Two deletion strains, Δcho2 and Δopi3, coding for enzymes that catalyze the sequential conversion of phosphatidylethanolamine to phosphatidylcholine were identified for suppressing the phenotypic effects of carnitine. Additional characterisation indicated that the suppression cannot be explained by differences in phospholipid homeostasis. The phenotypes could be reinstated by addition of extracellular choline, but show that the requirement for choline is not based on some overlapping function or the structural similarities of the two molecules. This is the first study to suggest a molecular link between a specific metabolite and carnitine-dependent, but shuttle-independent phenotypes in eukaryotes.
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    The origins of fears in a selected group of middle childhood South African children
    (Stellenbosch : University of Stellenbosch, 2006-03) Du Plessis, Michelle; Loxton, H. S.; University of Stellenbosch. Faculty of Arts and Social Sciences. Dept. of Psychology.
    The aims of the present study were to inquire into the origins of middle childhood South African children’s fears. In order to do this as precisely as possible it was determined to what extent Rachman’s (1977, 1991) three pathways played a role in the experience of the selected sample’s fears; in intensifying their fears; and finally in the actual onset of their fears. The results were examined across the independent variables age, gender, socio-economic status (SES), and culture. In addition, Rachman’s (1977; 1991) hypothesis that stronger, more intense fears originate through Conditioning experiences, while less intense, everyday fears originate through the indirect pathways, namely Modeling and Negative Information/Instruction was tested. The participants consisted of a sample of 660 children aged between 10 and 14 years, attending grades 5 and 7, representing the three main cultural groups found in the Western Cape, South Africa, and residing in low, middle and high socio-economic residential areas. Participants completed a short biographical questionnaire and the Fear Option List (FOL). Data analysis revealed that the majority of participants reported Modeling experiences in connection to their greatest fear, followed by Information and Conditioning experiences. Significant gender, SES, and cultural differences were found. The majority of participants reported that Information experiences were responsible for intensifying their greatest fear, and thus playing a role in maintaining it, followed by Modeling and Conditioning experiences. Significant gender, SES, and cultural differences were found. With regard to the actual onset of participants fears, the majority of participants reported that they had no clear idea of how their fear began, followed by Information, Conditioning, and Modeling. Only significant SES and cultural differences were found. The first half of Rachman’s (1977; 1991) hypothesis was confirmed in that stronger, more intense fears were found to be strongly related to Conditioning experiences. However, the second part of his hypothesis that less intense, everyday fears are likely to originate through the indirect pathways was not confirmed. The implications of the present study’s findings within a South African context are discussed, as well as the limitations of the study and suggestions for future research.
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    The role of carnitine in eukaryotic cells : Using yeast as a model
    (Stellenbosch : Stellenbosch University, 2015-12) Du Plessis, Michelle; Bauer, Florian; Franken, Jaco; Loos, Ben; Stellenbosch University. Faculty of Agrisciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
    ENGLISH ABSTRACT: Previous studies in yeast in this laboratory have found carnitine to be both protective against oxidative stress induced by hydrogen peroxide and to increase the detrimental effect of dithiothreitol. These phenotypes were found to be independent of the role of carnitine within the carnitine shuttle. A screen for suppressor mutations for these carnitine-dependent phenotypes identified, among others, Δcho2 and Δopi3. Cho2p and Opi3p catalyse the sequential methylation reactions in the formation of phosphatidylcholine from phosphatidylethanolamine. Therefore, this study aimed to investigate the relationship between choline, phosphatidylcholine and the carnitine phenotypes. Liquid growth assays of Δcho2 and Δopi3 cultures revealed that addition of choline can restore the protective effects of carnitine against hydrogen peroxide. The connection between the cellular phospholipid composition and the carnitine-dependent shuttleindependent phenotypes was also investigated. Analysis of the lipid composition of cells by LCMS showed that Δcho2 and Δopi3 had a largely different lipid composition compared with the wild type, most notably, a reduction in phosphatidylcholine and an increase in triacylglycerol content were observed for both mutants. These changes were reversed by supplementation with choline. However, no effects on the lipid composition of cells in response to carnitine treatment were observed, either when supplemented alone or in combination with DTT and hydrogen peroxide. Carnitine has also been investigated in mammalian systems for its potential to protect cells from oxidative stress, an effect which would be of benefit in various neurodegenerative disorders. Several studies have documented the positive effects of carnitine against oxidative stress in mammalian cells however the mechanism behind this action remains unknown. It is therefore thought that, provided similar effects for carnitine can be shown in mammalian cells as was observed in yeast, it would be beneficial to use yeast as a model system for the study of the molecular changes induced by carnitine. In view of this, the effects of carnitine on toxicity induced by oxidative stress in mammalian neural cells were compared to that which has been observed in yeast. For this purpose the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay, a measure of reductive capacity of cells, was used. However, no effects for carnitine were observed in the MTT assay in combination with either dithiothreitol or paraquat.