rdi1? vs WT during macrophage co-culture. Cryptococcus neoformans H99

Rho-GDP dissociation inhibitors (RDI) are repressors of Rho-type monomeric GTPases that allow for precise control of their target processes, e.g. cytoskeletal arrangement, vesicle trafficking, and polarized growth. In the human pathogenic yeast Cryptococcus neoformans, maintenance of normal cell morphology is vital for pathogenicity. We identified and deleted the gene encoding an RDI homolog in the human fungal pathogen Cryptococcus neoformans and investigated its impact on pathogenicity in animal models of cryptococcosis. Rdi1 deletion resulted in altered vacuole size in tissue culture medium, with corresponding alterations in expression of vesicle trafficking related genes. The rdi1? mutant strain showed reduced intracellular survival in macrophages, and severe attenuation of virulence in murine models of cryptococcosis. This reduction in virulence of the rdi1 mutant occurs in the absence of major defects in growth, morphology, or classical virulence-associated phenotypes. Keywords: mutant response, macrophage co-culture Overall design: WT and rdi1? total RNA were compared in a dye-swap experiment. One culture of each representing a population of each strain were incubated with macrophages at a ratio of 10:1 (yeast:macrophage). The co-cultures were incubated for 1hour at 37°C, 5% CO2 in tissue culture medium.