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VandenBerg, Alysia L.

Cell and Molecular Biology Program

2004

Ph. D.

C. albicans is a medically important opportunistic pathogen as the fourth leading cause of nosocomial infection in the United States. A rather unique feature of the lifestyle of C. albicans is that it undergoes a dynamic budded-to-hyphal-form transition. This morphological transition is imperative for virulence, as mutants that cannot switch between the budded and hyphal forms are avirulent. The Cdc42p GTPase regulates cell polarity in many organisms. In Candida albicans, Cdc42p performs functions essential for mitotic growth and functions in the budded-to-hyphal-form transition. With the aim of studying the hypha-specific functions of Cdc42p, several cdc42 point mutations were introduced into the genome as the sole copy of cdc42. These mutations were originally identified in S. cerevisiae using a genetic screen to isolate cdc42 mutations that could function as the sole copy of cdc42 but were unable to form pseudo hyphae in that organism's morphological transition. When the cdc42-S261, cdc42-E100G, and cdc42-S158T mutations were introduced into C. albicans, they demonstrated specific defects in the budded-to-hyphal-form transition, while maintaining all of the normal functions of Cdc42p in budded cells, aside from a temperature-sensitive actin polarity defect in the cdc42-E100G mutant. Each of the mutants had a reduced ability to damage endothelial cells, a predictor of virulence. When grown in hypha-inducing conditions, the mutants were deficient in the production of hypha-specific transcripts HWP1, ECE1, and SAP6. Each of these genes is heavily dependent upon the Efg1p transcription factor for their transcription, leading to the proposition that Cdc42p-dependent signaling impinged upon the cAMP/PKAlEfg1p signaling pathway. This pathway is known to function in the budded-to-hyphal-form transition and in virulence. The cdc42-S261 and cdc42-E100G mutants' hyphal formation defect was suppressed by overexpression of the phosphorylation-mimicked Efg1-T206Ep, but not the Efg1 p-T206Ap. Addition of exogenous cAMP was unable to stimulate the budded-to-hyphal-form transition. These results suggested that these two cdc42 mutations might have a deficiency in signaling to the Efg1 p transcription factor and indicate a novel role for Cdc42p in the Efg1 p-dependent signaling pathway.