The Hyphal-Associated Adhesin and Invasin Als3 of Candida albicans Mediates Iron Acquisition from Host Ferritin

Aaron P Mitchell,Ricardo S Almeida,John E Edwards,Michael Laue,Sascha Brunke,Sascha Thewes,Bernhard Hube,Antje Albrecht,Scott G Filler

doi:10.1371/journal.ppat.1000217

Abstract

Iron sequestration by host iron-binding proteins is an important mechanism of resistance to microbial infections. Inside oral epithelial cells, iron is stored within ferritin, and is therefore not usually accessible to pathogenic microbes. We observed that the ferritin concentration within oral epithelial cells was directly related to their susceptibility to damage by the human pathogenic fungus, Candida albicans. Thus, we hypothesized that host ferritin is used as an iron source by this organism. We found that C. albicans was able to grow on agar at physiological pH with ferritin as the sole source of iron, while the baker's yeast Saccharomyces cerevisiae could not. A screen of C. albicans mutants lacking components of each of the three known iron acquisition systems revealed that only the reductive pathway is involved in iron utilization from ferritin by this fungus. Additionally, C. albicans hyphae, but not yeast cells, bound ferritin, and this binding was crucial for iron acquisition from ferritin. Transcriptional profiling of wild-type and hyphal-defective C. albicans strains suggested that the C. albicans invasin-like protein Als3 is required for ferritin binding. Hyphae of an Δals3 null mutant had a strongly reduced ability to bind ferritin and these mutant cells grew poorly on agar plates with ferritin as the sole source of iron. Heterologous expression of Als3, but not Als1 or Als5, two closely related members of the Als protein family, allowed S. cerevisiae to bind ferritin. Immunocytochemical localization of ferritin in epithelial cells infected with C. albicans showed ferritin surrounding invading hyphae of the wild-type, but not the Δals3 mutant strain. This mutant was also unable to damage epithelial cells in vitro. Therefore, C. albicans can exploit iron from ferritin via morphology dependent binding through Als3, suggesting that this single protein has multiple virulence attributes.

Highlights

Iron is an essential element for virtually all organisms, ranging from microbes to multicellular animals
To elucidate which iron sources are exploited during growth on and invasion of oral epithelial cells and to determine how the availability of iron influences fungal-host cell interactions, we incubated oral epithelial cell monolayers in the presence of additional free iron or the iron chelator bathophenanthrolindisulphonic acid (BPS)
Through immunocytochemical localization of ferritin within epithelial cells, we found that addition of BPS caused a dramatic decrease in cellular ferritin within 24 hours of incubation (Figure 1A), in comparison to non-treated cells (Figure 1B)

Summary

Introduction

Iron is an essential element for virtually all organisms, ranging from microbes to multicellular animals. Successful microbial pathogens have developed multiple iron acquisition and uptake systems (reviewed in [2,3]). These systems include enzymes for reduction and oxidization of iron ions (Fe2+ or Fe3+), high-affinity permeases for iron transport, chelators (siderophores) and uptake systems for siderophores. With the exception of ferritin, each of these proteins has been reported to serve as an iron source for some pathogenic microbes These iron sources are exploited via direct binding, degradation, and/or uptake [4,5,6,7,8,9,10,11,12,13]

Methods

Results

Conclusion