A live-cell ergosterol reporter for visualization of the effects of fluconazole on the human fungal pathogen Candida albicans

Abstract

ABSTRACTErgosterol, an essential plasma membrane amphipathic lipid, is a major component of the fungal plasma membrane. Most fungal pathogens are susceptible to azole drugs that target ergosterol biosynthesis, and resistance/tolerance to azoles is increasingly problematic. Candida albicans is the most prevalent etiology of candidiasis, and, in this fungal pathogen, ergosterol-rich sub-domains are likely to include sphingolipids, as well as specific membrane proteins, such as multidrug transporters. To investigate the dynamics of ergosterol during the cell cycle and whether drug treatment affects these dynamics in this opportunistic pathogen, we adapted the D4H (domain 4 of the perfringolysin O bacterial toxin) reporter for studying sterol-rich membrane domains. We show that D4H provides a direct readout for the cellular effects of fluconazole and that highly polarized ergosterol is not critical for budding or filamentous growth.IMPORTANCEErgosterol is a critical membrane lipid in fungi. In Candida albicans, this essential plasma membrane amphipathic lipid is important for interactions with host cells, in particular, host immune responses. Here, we use a live-cell reporter for specifically visualizing ergosterol and show that apical enrichment of this sterol is not critical for budding and filamentous growth in this human fungal pathogen. Our results highlight that this live-cell reporter is likely to be a useful tool in the analyses of azole resistance and tolerance mechanisms, including alterations in drug targets and upregulation of efflux activities.

Keywords

• ergosterol
• antifungal agents
• Candida albicans
• fungal growth
• lipid signaling