Evaluation of pseudouridine monophosphate glycosidase and snR10c Candida albicans mutants

Candida albicans is an opportunistic human fungal pathogen capable of causing both superficial and disseminated infections. C. albicans bloodstream infections continue to have high mortality rates despite existing antifungal treatments. Therefore, it is important to continually work toward finding new drug targets. snR10c is a C. albicans specific H/ACA box snoRNA involved in the conversion of uridine into pseudouridine, the most abundant RNA modification. We isolated 4 mutants of snR10c that altered the binding pocket residues. We found two of four mutant isolates also contained an additional mutation. Isolate 1, which was heterozygous for this additional mutation had severe growth defects under temperature- and drug-induced stress. Isolate 1 also had filamentation defects and had significantly attenuated virulence compared to wildtype. This could suggest that snR10c also has role outside of pseudouridylation. The study also set out to investigate pseudouridine degradation, a metabolic process that does not occur in humans, but is predicted to occur in C. albicans. The second aim of this study was to determine if knockout of Pug1, Tna12, and/or Gap2 changes C. albicans virulence in the Galleria mellonella model system. We found minor growth and filamentation defects in the strains pug1, pug1tna12, and pug1gap2. I also found tna12 had increased virulence compared to wildtype. Our data suggests that targeting pseudouridine degradation alone may not be a suitable therapeutic target. The third aim of this study was to design a PUG1-TurboID construct for downstream use in identifying protein-protein interactions with Pug1. This construct has been designed and will be integrated into the C. albicans genome to identify novel interactors. My data addresses under described aspects of RNA metabolism in C. albicans and has identified likely distinctions between C. albicans RNA metabolism and that of mammals.