Functional characterization of the Aspergillus fumigatusPHO80 homologue

Phosphate is an ion that is essential for fungal growth. The systems for inorganic phosphate (P_i) acquisition in eukaryotic cells (PHO) have been characterized as a low-affinity (that assures a supply of P_i at normal or high external P_i concentrations) and a high-affinity (activated in response to P_i starvation). Here, as an initial step to understand the PHO pathway in Aspergillus fumigatus, we characterized the PHO80 homologue, PhoB^PHO80. We show that the ΔphoB^PHO80 mutant has a polar growth defect (i.e., a delayed germ tube emergence) and, by phenotypic and phosphate uptake analyses, establish a link between PhoB^PHO80, calcineurin and calcium metabolism. Microarray hybridizations carried out with RNA obtained from wild-type and ΔphoB^PHO80 mutant cells identify Afu4g03610 (phoD^PHO84), Afu7g06350 (phoE^PHO89), Afu4g06020 (phoC^PHO81), and Afu2g09040 (vacuolar transporter Vtc4) as more expressed both in the ΔphoB^PHO80 mutant background and under phosphate-limiting conditions of 0.1 mM P_i. Epifluorescence microscopy revealed accumulation of poly-phosphate in ΔphoB^PHO80 vacuoles, which was independent of extracellular phosphate concentration. Surprisingly, a phoD^PHO84 deletion mutant is indistinguishable phenotypically from the corresponding wild-type strain. mRNA analyses suggest that protein kinase A absence supports the expression of PHO genes in A. fumigatus. Furthermore, ΔphoB^PHO80 and ΔphoD^PHO84 mutant are fully virulent in a murine low dose model for invasive aspergillosis.