Squalene lipotoxicity in a lipid droplet-less yeast mutant is linked to plasma membrane dysfunction.

Squalene is a naturally occurring triterpene with wide industrial applications. Due to limited natural resources, production of this valuable lipid in yeast is of high commercial relevance. Typically low levels of squalene in yeast can be significantly increased by specific cultivation conditions or genetic modifications. Under normal conditions, excess squalene is stored in lipid droplets (LD), while in a Saccharomyces cerevisiae mutant unable to form LD it is distributed to cellular membranes. We present here the evidence that squalene accumulation in this LD-less mutant treated with squalene monooxygenase inhibitor terbinafine induces growth defects and loss of viability. We show that plasma membrane malfunction is involved in squalene toxicity. We have found that subinhibitory concentrations of terbinafine increased the sensitivity of LD-less mutant to several membrane-active substances. Furthermore, squalene accumulation in terbinafine-treated LD-less cells disturbed the maintenance of membrane potential and increased plasma membrane permeability to rhodamine 6G. LD-less cells treated with terbinafine showed also high sensitivity to osmotic stress. To confirm the causal relationship between squalene accumulation, loss of viability and impaired plasma membrane functions we treated LD-less cells simultaneously with terbinafine and squalene synthase inhibitor zaragozic acid. Reduction of squalene levels by zaragozic acid improved cell growth and viability and decreased plasma membrane permeability to rhodamine 6G in terbinafine-treated LD-less cells. Our results support the hypothesis that plasma membrane malfunction is involved in the mechanisms of squalene lipotoxicity in yeast cells with defective lipid storage.

Variations in yeast plasma-membrane lipid composition affect killing activity of three families of insect antifungal peptides.

Naturally occurring antimicrobial peptides and their synthetic analogues are promising candidates for new antifungal drugs. We focused on three groups of peptides isolated from the venom of bees and their synthetic analogues (lasioglossins, halictines and hylanines), which all rapidly permeabilised the plasma membrane. We compared peptides' potency against six pathogenic Candida species (C. albicans, C. glabrata, C. parapsilosis, C. tropicalis, C. krusei and C. dubliniensis) and the non-pathogenic model yeast Saccharomyces cerevisiae. Their activity was independent of the presence of the multidrug-resistant pumps of C. glabrata but was influenced by the lipid composition of cell plasma membranes. Although the direct interaction of the peptides with ergosterol was negligible in comparison with amphotericin B, the diminished ergosterol content after terbinafine pretreatment resulted in an increased resistance of C. glabrata to the peptides. The tested peptides strongly interacted with phosphatidylglycerol, phosphatidic acid and cardiolipin and partly with phosphatidylinositol and phosphatidylethanolamine. The interactions between predominantly anionic phospholipids and cationic peptides indicated a mainly electrostatic binding of peptides to the membranes. The results obtained also pointed to a considerable role of the components of lipid rafts (composed from sphingolipids and ergosterol) in the interaction of yeast cells with the peptides.