Human salivary histatin 5 causes disordered volume regulation and cell cycle arrest in Candida albicans

Human salivary histatin 5 (Hst 5) is a nonimmune salivary protein with antifungal activity against an important human pathogen, Candida albicans. The candidacidal activity of histatins appears to be a distinctive multistep mechanism involving depletion of the C. albicans intracellular ATP content as a result of nonlytic ATP efflux. Hst 5 caused a loss of cell viability concomitant with a decrease in cellular volume as determined both by a classical candidacidal assay with exogenous Hst 5 and by using a genetically engineered C. albicans strain expressing Hst 5. Preincubation of C. albicans cells with pharmacological inhibitors of anion transport provided complete or substantial protection from Hst 5-induced killing and volume reduction of cells. Moreover, intracellular expression of Hst 5 resulted in a reduction in the population mean cell volume that was accompanied by an increase in the percentage of unbudded cells and C. albicans cells in the G(1) phase. Following expression of Hst 5, the smallest cells sorted by fluorescence-activated cell sorting from the total population did not replicate and were exclusively in the G(1) phase. Cells with intracellularly expressed Hst 5 had greatly reduced G(1) cyclin transcript levels, indicating that they arrested in the G(1) phase before the onset of Start. Our data demonstrate that a key determinant in the mechanism of Hst 5 toxicity in C. albicans cells is the disruption of regulatory circuits for cell volume homeostasis that is closely coupled with loss of intracellular ATP. This novel process of fungicidal activity by a human salivary protein has highlighted potential interactions of Hst 5 with volume regulatory mechanisms and the process of yeast cell cycle control.