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Title: RNA synthesis in Candida albicans
Author: Peters, David William
ISNI:       0000 0001 3485 1370
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 1985
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The thesis describes the results from investigations into RNA synthesis in the dimorphic fungus Candida albicans. Methods were described, and evaluated, for the preparation of protoplasts and nuclei - two in vitro systems that were used to study RNA synthesis. It was found that most radiolabelled precursor was incorporated into low molecular weight RNA by nuclei. In contrast, the precursor was associated with higher molecular weight RNA species from protoplasts, from which the nuclei were prepared. Contamination by RNases was a serious problem in the preparation of this in vitro system from protoplasts. The RNA polymerases were purified from yeast and part-purified from mycelial C. albicans. Three classes of RNA polymerase were resolved by ion exchange chromatography of cell-free extract from yeast, whilst only one was found in mycelia. Some characteristics of RNA polymerases from the yeast were described. All three isozymes had optimal activity in vitro when the media contained mono and divalent ion concentrations that were similar to those reported for Saccharomyces cerevisiae - a yeast often compared with C. albicans. In addition, the three isozymes from C. albicans had similar Km values for CTP to that of these enzymes from other eukaryotes. RNA polymerases I and III from yeast C. albicans showed similar sensitivities to α-amanitin as the corresponding isozymes from S. cerevisiae. RNA polymerase II was far more insensitive to the amatoxin than the corresponding enzyme from higher eukaryotes. A variety of nucleoside analogues were suggested as potential antifungal agents warranting further investigations. These were capable of inhibiting RNA synthesis by C. albicans in vitro and/or in vivo assays. Studies were also made on RNA synthesis during germ tube formation - the initial stage of the yeast-mycelial transformation. It was found that it was necessary to cultivate C, albicans yeast in nutritionally impoverished media and starved for 24 hr to achieve reproducible germ tube formation. The strain of C. albicans used in this research formed germ tubes when incubated in imidazole HC1 buffer containing serum, N-acetyl glucosamine, glucose or glucose plus glutamine at temperatures above 35°C. During germ tube formation there was an increase in the RNA content per unit yeast cell. Both the rate and maximum amount of radiolabelled precursor incorporated into RNA depended upon the conditions used to induce germ tube formation. Some inhibitors of RNA synthesis were capable of inhibiting germ tube formation. It was found that the ratio of high to low molecular weight RNA species changed over the period of germ tube formation. The thesis concludes by evaluating the results of this research and suggests further directions for future research.
Supervisor: Not available Sponsor: Science and Engineering Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QK Botany ; QP Physiology