Comparison of Sterol Import under Aerobic and Anaerobic Conditions in Three Fungal Species, Candida albicans, Candida glabrata, and Saccharomyces cerevisiae

Abstract

bSterol import has been characterized under various conditions in three distinct fungal species, the model organism Saccharomyces cerevisiae and two human fungal pathogens Candida glabrata and Candida albicans, employing cholesterol, the sterol of higher eukaryotes, as well as its fungal equivalent, ergosterol. Import was confirmed by the detection of esterified cholesterol within the cells. Comparing the three fungal species, we observe sterol import under three different conditions. First, as previously well characterized, we observe sterol import under low oxygen levels in S. cerevisiae and C. glabrata, which is dependent on the transcription factor Upc2 and/or its orthologs or paralogs. Second, we observe sterol import under aerobic conditions exclusively in the two pathogenic fungi C. glabrata and C. albicans. Uptake emerges during post-exponential-growth phases, is independent of the characterized Upc2-pathway and is slower compared to the anaerobic uptake in S. cerevisiae and C. glabrata. Third, we observe under normoxic conditions in C. glabrata that Upc2-dependent sterol import can be induced in the presence of fetal bovine serum together with fluconazole. In summary, C. glabrata imports sterols both in aerobic and anaerobic conditions, and the limited aerobic uptake can be further stimulated by the presence of serum together with fluconazole. S. cerevisiae imports sterols only in anaerobic conditions, demonstrating aerobic sterol exclusion. Finally, C. albicans imports sterols exclusively aerobically in post-exponential-growth phases, independent of Upc2. For the first time, we provide direct evidence of sterol import into the human fungal pathogen C. albicans, which until now was believed to be incapable of active sterol import.

Structural mechanism of ergosterol regulation by fungal sterol transcription factor Upc2

“…Although there are several structural differences between cholesterol and ergosterol, they both efficiently support the growth of S. cerevisiae and C. glabrata incapable of ergosterol biosynthesis 47 . Since the human body under azole treatment provides conditions favouring the stimulation of sterol import from the fungal environment, the ability of C. glabrata to replace ergosterol with a host sterol may be responsible for its elevated azole resistance in vivo 6,47 . The deletion of UPC2 leads to anaerobic inviability and high susceptibility to azole antifungals in S. cerevisiae and in C. albicans 6,14 .…”

“…Since the human body under azole treatment provides conditions favouring the stimulation of sterol import from the fungal environment, the ability of C. glabrata to replace ergosterol with a host sterol may be responsible for its elevated azole resistance in vivo 6,47 . The deletion of UPC2 leads to anaerobic inviability and high susceptibility to azole antifungals in S. cerevisiae and in C. albicans 6,14 . Therefore, the inhibition of Upc2, which subsequently suppresses the adaptive responses of fungal cells to azoles, could be a novel strategy to improve the combined therapy with antifungal agents 16,48 .…”

“…However, conservation of this region across a large number of zinc TFs in many fungal species indicates that the CTD may represent a novel domain 13 . Deletion of UPC2 in pathogenic fungi, C. albicans and C. glabrata, leads to a significant decrease of tolerance to azole drugs and deletion of both UPC2 and ECM22 in S. cerevisiae is lethal in certain yeast strains 6,14,15 . Therefore, the Upc2 pathway may represent a potential co-therapeutic target for enhancing azole activity against these organisms 16 .…”

“…However, many other fungal species, including Saccharomyces cerevisiae and Candida species, lack protein homologues of the mammalian cholesterol regulators and appear to have evolved distinct regulatory mechanism 5 . In S. cerevisiae, Upc2 and Ecm22, two similar zinc finger transcriptional factors, have been implicated in ergosterol regulation by binding to the promoters of ergosterol biosynthetic and sterol uptake genes and activating the transcription on sterol depletion 2,6 . In addition, S. cerevisiae Upc2 is important for adaptation to hypoxia regulating up to one-third of anaerobically expressed genes 7 .…”

The Ins and Outs of Azole Antifungal Drug Resistance: Molecular Mechanisms of Transport

“…To date, this phenomenon is related only to two fungal species -the model organism S. cerevisiae (Andreasen and Stier 1953) and the closely related pathogen C. glabrata (Zavrel et al 2013;Bard et al 2005;Nakayama et al 2007;Nagi et al 2013). Although C. albicans is also capable of sterol import, the rate is insufficient to replace the endogenous ergosterol biosynthesis (Zavrel et al 2013). S. cerevisiae and C. glabrata are the only two described species able to import extracellular sterols in quantities sufficient to replace endogenous ergosterol biosynthesis in the presence of azoles.…”