Whether this phenotype was due to a direct involvement of Hog1p in the regulation of the iron JQEZ5 supplier responsive network or due to indirect effects, such as perturbations of copper metabolism, which may have impaired the functionality of iron uptake proteins was not yet studied. As expected, high levels of extracellular iron increased the formation of intracellular ROS. Thus, we used intracellular ROS levels together with learn more the removal of iron from growth medium as indicators of iron entry into the cells. We detected

increased basal ROS levels in the Δhog1 mutants, as previously reported [36]. These ROS levels were further increased by exposure to 30 μM Fe3+ confirming that iron was taken up by Δhog1 cells. Moreover, iron ions were removed from the growth medium with the same efficiency by Δhog1 as by the reference (DAY286) cells. Thus, Hog1p dependent phenotypes of the C. albicans response to iron were not due to iron uptake

deficiencies, but could be rather due to the involvement of Hog1p in the response to iron availability. This is supported by our data on the transient hyper-phosphorylation of Hog1p during exposure of cells to high iron concentrations. Elevated iron concentrations induced a flocculent phenotype of C. albicans, which was dependent on the presence of both Hog1p and Pbs2p, as well as on protein synthesis. As high iron concentrations led to increased phosphorylation of Hog1p, this could induce the synthesis of proteins of which TNF-alpha inhibitor some mediate cell aggregation. This iron triggered activation of Hog1p is likely not related to oxidative stress, as the potent radical scavenger NAC did not prevent the flocculent phenotype upon exposure to high iron concentrations, while it decreased intracellular ROS levels. For the closely related almost yeast S. cerevisiae, a function of ScHog1p in cell aggregation was reported, in that hyperactive

ScHog1p mutants resulted in increased flocculation [51]. First hints on an involvement of Hog1p in the response of C. albicans to iron came from the observation of the de-repression of several iron uptake genes in the Δhog1 mutant under otherwise repressive conditions [27]. In agreement with these gene expression data, we observed increased MCFOs protein levels and ferric reductase activity in Δhog1 mutants. Furthermore we found that MCFOs were also de-repressed in Δpbs2 mutants, indicating that the HOG1 mediated regulation of MCFOs was dependent on PBS2. Remarkably, induction of these components in RIM was not strictly dependent on Hog1p, as this induction was also observed in the Δhog1 mutant. Thus deletion of HOG1 de-repressed components of the iron uptake system, and this elevated basal level was further enhanced when iron availability was limited. Hog1p was shown to be essential for C. albicans under oxidative stress conditions [30].