The molecular mechanism of Nystatin action is dependent on the membrane biophysical properties and lipid composition

Abstract

Nystatin (Nys) is a pore forming broad-spectrum and efficient antifungal drug with significant toxicity in mammalian organisms. In order to develop a non-toxic and more effective Nys formulation, its molecular mechanism of action at the cell membrane needs to be better understood. It is widely accepted that Nys activity and toxicity depend on the presence and type of membrane sterols. Taking advantage of multiple biophysical methodologies, we now show that the formation and stabilization of Nys aqueous pores, which are associated with Nys cytotoxicity, occur in the absence of membrane sterols. Our results suggest that the Nys mechanism of action is driven by the presence of highly ordered membrane domains capable of stabilizing the Nys oligomers. Moreover, Nys pore formation is accompanied by strong Nys-induced membrane reorganization that depends on membrane lipid composition and seems to underlie the Nys cytotoxic effect. Accordingly, in membranes enriched in a gel-phase forming phospholipid, Nys incorporates within the phospholipid-enriched gel domains, where it forms pores able to expand the gel domains. In contrast, in membranes enriched in gel domain forming sphingolipids, Nys-induced pore formation occurs through the destabilization of the gel phase. These results show that the Nys mechanism of action is complex and not only dependent on membrane sterols, and provide further insight into the molecular details governing Nys activity and toxicity.

TOR complex 2–regulated protein kinase Ypk1 controls sterol distribution by inhibiting StARkin domain–containing proteins located at plasma membrane–endoplasmic reticulum contact sites

“…This difference is due to a modification in the distribution of ergosterol rather than in the level of cellular ergosterol, as Ysp2 AAA and WT Ysp2 cells contain the same amount of total ergosterol ( Figure 4B ). The change in PM ergosterol homeostasis was confirmed by showing that cells expressing Ysp2 AAA are more resistant than cells expressing WT Ysp2 to nystatin, another pore-forming polyene antifungal that functions, in part, by interacting with ergosterol in membranes ( Dos Santos et al, 2017 ; Figure 4C ). Therefore, the role of Ypk1 phosphorylation is to negatively regulate Ysp2 function.…”

The Verticillium dahliae Sho1‐MAPK pathway regulates melanin biosynthesis and is required for cotton infection

“…S13 and S14), suggesting that VdSho1 was not involved in responses to high osmotic stress and cell wall integrity in V. dahliae. Furthermore, VdSho1 sensing of the osmotic stress was determined using the pharmacological agent nystatin, which inhibits the Sho1-mediated signalling and reduce osmotic stress intracellularly by enhancing membrane permeability (Reiser et al, 2003;dos Santos et al, 2017). In the VdSho1 complemented transformants, the growth of ΔSho1 strain was not restricted in response to nystatin ( Fig.…”

Yeast Sphingolipid-Enriched Domains and Membrane Compartments in the Absence of Mannosyldiinositolphosphorylceramide

“…The present work shows how the headgroup structure of complex sphingolipids can be an important determinant in the organization of plasma membrane lipids and proteins in S. cerevisiae. Recently, we showed that the polyene antifungal nystatin forms active pores in membranes lacking any sterol, but containing gel domains [12], challenging the classical model which describes the molecular mode of action of nystatin through an ergosterol-dependent mechanism. Moreover, the action of other antifungal drugs has been shown to involve an interplay with sphingolipids that is conditioned by their headgroup structure.…”

“…DPH steady-state fluorescence anisotropy, r , is a well-established parameter to report alterations in the global fluidity of the plasma membrane [11,58,59]. The fluorescence anisotropy of DPH changes linearly with composition (and temperature) along the gel/ liquid disordered (l d ) and l d / liquid ordered (l o ) phase coexistence region, i.e., responds linearly to the fraction of each phase, in several binary and ternary lipid systems [12,54,58], including in the 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/ergosterol binary system [60]. Moreover, in the POPC/phytoceramide (the yeast sphingolipid backbone) system, the fluorescence anisotropy and intensity of DPH behave similarly to typical gel/ l d systems in the region of coexistence of l d and a phytoceramide-rich gel phase where the hydrophobic packing is comparable to the one found for the gel domains in yeast [48].…”

“…These domains found in the plasma membrane of Saccharomyces cerevisiae differ from the prototypical mammalian lipid raft both in composition, given they are ergosterol-depleted, and biophysically, as they are highly rigid gel domains [11]. Hence, the study of these domains is particularly important, as they may explain fundamental differences between fungal and mammalian membranes [9], providing potential targets that can be explored concerning antifungal drug development [12].…”