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Abstract
Candida albicans is an opportunistic fungal pathogen of humans, and one of the leading causative agents of death and disease in immunocompromised individuals. The ability to switch between the yeast and filamentous morphologies is required for C. albicans ' virulence. However, fungal dimorphism is not the sole virulence determinant, other factors such as the induction and expression of transcription factors, adhesions, lipases, proteases, and heat shock proteins may play a large role in the switch form commensal microbe to pathogen. Similarly, detoxification of toxic compounds also is crucial in protecting cells from host antimicrobial defenses.
The purpose of the studies in chapters two and three is to characterize the role of Glyoxalase 3 (Glx3) through the generation of a glx3Δ/glx3Δ mutant. Phenotypic characterization of Glx3 includes: solving the 3D protein structure, measuring the methylglyoxal-specific enzymatic activity of Glx3, comparing the effect of endogenous and exogenous methylglyoxal on wild type and glx3Δ/glx3Δ growth rate, and the role of this protein in protecting C. albicans during environmental stresses, including high temperature, and oxidative stress.
The results of our studies show that Glx3 in C. albicans appears to be essential for detoxification of methylglyoxal, especially when grown in media with glycerol as the sole carbon source. Additionally, the glx3Δ/glx3Δ mutant strain showed a greater sensitivity to higher temperatures compared to the wild type and a decrease in oxidative stress tolerance and also there is evidence that Glx3 plays a role in modification of farnesol to epoxy farnesol.
In chapter four we investigate the histone biotinylation and the role of biotin in stimulating germ tube formation in C. albicans. Biotin is known to be required for cell growth and fatty acid metabolism. In addition, we have discovered that biotin is used to modify histones in C. albicans. Biotinylation was detected by Western blot using a monoclonal anti-biotin HRP-conjugated antibody as well as with qTOF and LC/MS/MS. During this study we observed that three histones, H2A, H2B, and H4, were biotinylated at many lysine residues in an apparently non-site specific manner. The function of histone biotinylation in C. albicans is still unknown but, because C. albicans is a natural biotin auxotroph, a storage reservoir for biotin is an attractive hypothesis.
