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

Himanshu K. Prasad,Rana Pratap Singh,Krishnamurthy Natarajan,Neha Agarwal,Ishani Sinha

doi:10.1074/jbc.m111.233569

Himanshu K. Prasad, Rana Pratap Singh + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m111.233569

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Abstract

Iron homeostasis is highly regulated in organisms across evolutionary time scale as iron is essential for various cellular processes. In a computational screen, we identified the Yap/bZIP domain family in Candida clade genomes. Cap2/Hap43 is essential for C. albicans growth under iron-deprivation conditions and for virulence in mouse. Cap2 has an amino-terminal bipartite domain comprising a fungal-specific Hap4-like domain and a bZIP domain. Our mutational analyses showed that both the bZIP and Hap4-like domains perform critical and independent functions for growth under iron-deprivation conditions. Transcriptome analysis conducted under iron-deprivation conditions identified about 16% of the C. albicans ORFs that were differentially regulated in a Cap2-dependent manner. Microarray data also suggested that Cap2 is required to mobilize iron through multiple mechanisms; chiefly by activation of genes in three iron uptake pathways and repression of iron utilizing and iron storage genes. The expression of HAP2, HAP32, and HAP5, core components of the HAP regulatory complex was induced in a Cap2-dependent manner indicating a feed-forward loop. In a feed-back loop, Cap2 repressed the expression of Sfu1, a negative regulator of iron uptake genes. Cap2 was coimmunoprecipitated with Hap5 from cell extracts prepared from iron-deprivation conditions indicating an in vivo association. ChIP assays demonstrated Hap32-dependent recruitment of Hap5 to the promoters of FRP1 (Cap2-induced) and ACO1 (Cap2-repressed). Together our data indicates that the Cap2-HAP complex functions both as a positive and a negative regulator to maintain iron homeostasis in C. albicans.

Highlights

Fungal genomes [1, 2]
C. albicans Cap2 Is a HAP4L-bZIP Bipartite Domain Protein— We identified the complete repertoire of bZIP domain containing sequences from S. cerevisiae, Candida glabrata, and 10 Candida clade species (Fig. 1A, and supplemental Fig. S1)
We demonstrate that the Cap2-HAP complex is the central and a critical regulator of iron homeostasis in C. albicans

Summary

Introduction

Fungal genomes [1, 2]. the lessons from the S. cerevisiae transcriptional programs would have to be re-evaluated as new genomes are sequenced and annotated. Our mutational analyses showed that the CAP2/HAP43 gene was required for growth of C. albicans under iron deprivation conditions (Fig. 2A). RT-PCR analysis showed that iron deprivation led to up-regulation of CAP2 mRNA in a time-dependent manner, with a maximum induction of 5.7-fold after 3 h of BPS addition (Fig. 4A).

Results

Conclusion