Farnesol-Induced Apoptosis in Candida albicans Is Mediated by Cdr1-p Extrusion and Depletion of Intracellular Glutathione

Jingsong Zhu,Dominique Sanglard,Bastiaan P Krom,Chaidan Intapa,Mark E Shirtliff,Brian M Peters,Mary Ann Jabra-Rizk,Clinton C Dawson,Kirsten Nielsen

doi:10.1371/journal.pone.0028830

Abstract

Farnesol is a key derivative in the sterol biosynthesis pathway in eukaryotic cells previously identified as a quorum sensing molecule in the human fungal pathogen Candida albicans. Recently, we demonstrated that above threshold concentrations, farnesol is capable of triggering apoptosis in C. albicans. However, the exact mechanism of farnesol cytotoxicity is not fully elucidated. Lipophilic compounds such as farnesol are known to conjugate with glutathione, an antioxidant crucial for cellular detoxification against damaging compounds. Glutathione conjugates act as substrates for ATP-dependent ABC transporters and are extruded from the cell. To that end, this current study was undertaken to validate the hypothesis that farnesol conjugation with intracellular glutathione coupled with Cdr1p-mediated extrusion of glutathione conjugates, results in total glutathione depletion, oxidative stress and ultimately fungal cell death. The combined findings demonstrated a significant decrease in intracellular glutathione levels concomitant with up-regulation of CDR1 and decreased cell viability. However, addition of exogenous reduced glutathione maintained intracellular glutathione levels and enhanced viability. In contrast, farnesol toxicity was decreased in a mutant lacking CDR1, whereas it was increased in a CDR1-overexpressing strain. Further, gene expression studies demonstrated significant up-regulation of the SOD genes, primary enzymes responsible for defense against oxidative stress, with no changes in expression in CDR1. This is the first study describing the involvement of Cdr1p-mediated glutathione efflux as a mechanism preceding the farnesol-induced apoptotic process in C. albicans. Understanding of the mechanisms underlying farnesol-cytotoxicity in C. albicans may lead to the development of this redox-cycling agent as an alternative antifungal agent.

Highlights

Candida albicans is the most important human fungal pathogen causing diseases varying from superficial mucosal infections to lifethreatening systemic disorders [1,2]
Several key physiological roles have been described for the quorum-sensing molecule farnesol in C. albicans cell cycle
Since farnesol is described to be increasingly secreted by the fungus in aging cultures, it is conceivable to speculate that through the regulation of farnesol production, C. albicans cells may have developed a mechanism of programmed cell death with evolutionary advantages [35]

Summary

Introduction

Candida albicans is the most important human fungal pathogen causing diseases varying from superficial mucosal infections to lifethreatening systemic disorders [1,2]. Farnesol, synthesized from farnesyl pyrophosphate, an intermediate in the sterol biosynthetic pathway, was identified as a quorum sensing molecule secreted by C. albicans, able to prevent yeast-to-hyphal conversion and biofilm formation [6,7,8,9]. Through assessment of classical apoptotic markers and global proteomic analysis we demonstrated that above threshold concentrations, farnesol induces apoptosis in C. albicans and in human oral tumor cells [10,11]. Among the apoptotic markers identified were the activation of intracellular caspases, a class of proteolytic enzymes which when activated, convey an apoptotic signal and induce apoptosis [10,11,12]. To date, the underlying mechanism of farnesol cytotoxicity in eukaryotic cells is yet to be fully elucidated

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