Candida albicans Ferric Reductases Are Differentially Regulated in Response to Distinct Forms of Iron Limitation by the Rim101 and CBF Transcription Factors

Abstract

Iron is an essential nutrient that is severely limited in the mammalian host. Candida albicans encodes a family of 15 putative ferric reductases, which are required for iron acquisition and utilization. Despite the central role of ferric reductases in iron acquisition and mobilization, relatively little is known about the regulatory networks that govern ferric reductase gene expression in C. albicans. Here we have demonstrated the differential regulation of two ferric reductases, FRE2 and FRP1, in response to distinct iron-limited environments. FRE2 and FRP1 are both induced in alkaline-pH environments directly by the Rim101 transcription factor. However, FRP1 but not FRE2 is also induced by iron chelation. We have identified a CCAAT motif as the critical regulatory sequence for chelatormediated induction and have found that the CCAAT binding factor (CBF) is essential for FRP1 expression in iron-limited environments. We found that a hap5⌬/hap5⌬ mutant, which disrupts the core DNA binding activity of CBF, is unable to grow under iron-limited conditions. C. albicans encodes three CBF-dependent transcription factors, and we identified the Hap43 protein as the CBF-dependent transcription factor required for iron-limited responses. These studies provide key insights into the regulation of ferric reductase gene expression in the fungal pathogen C. albicans.

Cap2-HAP Complex Is a Critical Transcriptional Regulator That Has Dual but Contrasting Roles in Regulation of Iron Homeostasis in Candida albicans

“…Third, we showed that Cap2 could be efficiently coimmunoprecipitated with Hap5 from cell extracts prepared from cultures grown under iron-deprivation conditions. Although previous studies indicated an interaction of Hap4-like proteins with Hap2-Hap3-Hap5 proteins from C. albicans (19,27), our study is the first biochemical demonstration of Cap2-HAP interaction in cell extracts under iron-deprivation conditions. A recent yeast two-hybrid study also showed interaction between H. polymorpha Hap4b and Hap5 (21).…”

“…First, CAP2 is required for growth of C. albicans in iron-limited medium in vitro and for virulence in the mouse model. The requirement of CAP2, also called HAP43, for growth under iron-deprivation conditions was independently identified in other studies (19,20,61). Our mutational analyses showed that both bZIP and HAP4L domains are required for Cap2 function.…”

“…Complex-There are previous reports that Cap2 or its orthologs likely function in a complex with Hap2-Hap3-Hap5 (19,21,29). In a previous study, three HAP4-like genes and two HAP3-like genes were identified in the C. albicans genome (27).…”

“…15 and 16). In addition, the regulation of iron homeostasis in fungal genomes, including C. albicans, is dependent on the HAP regulatory complex (17)(18)(19)(20)(21). In S. cerevisiae, the HAP complex components Hap2, Hap3, and Hap5, are homologous to mammalian NF-YA, NF-YB, and NF-YC subunits, respectively, of the trimeric CCAAT box binding regulator NF-Y (22).…”

“…Interestingly, the Hap5-TAP occupancy was completely abolished in the hap32⌬ background. The Ϫ84 to Ϫ281 sequence contained the 5Ј-CCAAT-3Ј box at Ϫ138, shown previously to be important for activation of the FRP1-lacZ reporter (19).…”