Cch1 mediates calcium entry in Cryptococcus neoformans and is essential in low-calcium environments

The ability of Cryptococcus neoformans to grow at the mammalian body temperature (37 degrees C to 39 degrees C) is a well-established virulence factor. Growth of C. neoformans at this physiological temperature requires calcineurin, a Ca2+/calmodulin-dependent protein phosphatase. When cytosolic calcium concentrations are low (similar to 50 to 100 nM), calcineurin is inactive and becomes active only when cytosolic calcium concentrations rise (similar to 1 to 10 mu M) through the activation of calcium channels. In this study we analyzed the function of Cch1 in C. neoformans and found that Cch1 is a Ca2+-permeable channel that mediates calcium entry in C. neoformans. Analysis of the Cch1 protein sequence revealed differences in the voltage sensor (S4 regions), suggesting that Cch1 may have diminished voltage sensitivity or possibly an alternative gating mechanism. The inability of the cch1 mutant to grow under conditions of limited extracellular calcium concentrations ([Ca2+](extracellular), similar to 100 nM) suggested that Cch1 was required for calcium uptake in low-calcium environments. These results are consistent with the role of ScCch1 in mediating high-affinity calcium uptake in Saccharomyces cerevisiae. Although the growth defect of the cch1 mutant under conditions of limited [Ca2+](extracellular) (similar to 100 nM) became more severe with increasing temperature (25 degrees C to 38.5 degrees), this temperature sensitivity was not observed when the cch1 mutant was grown on rich medium ([Ca2+](extracellular), similar to 0.140 mM). Accordingly, the cch1 mutant strain displayed only attenuated virulence when tested in the mouse inhalation model of cryptococcosis, further suggesting that C. neoformans may have a limited requirement for Cch1 and that this requirement appears to include ion stress tolerance.