Lactoferrin perturbs lipid rafts and requires integrity of Pma1p-lipid rafts association to exert its antifungal activity against Saccharomyces cerevisiae

Abstract

Lactoferrin (Lf) is a bioactive milk-derived protein with remarkable wide-spectrum antifungal activity. To deepen our understanding of the molecular mechanisms underlying Lf cytotoxicity, the role of plasma membrane ergosterol-and sphingolipid-rich lipid rafts and their association with the proton pump Pma1p was explored. Pma1p was previously identified as a Lf-binding protein. Results showed that bovine Lf (bLf) perturbs ergosterol-rich lipid rafts organization by inducing intracellular accumulation of ergosterol. Using yeast mutant strains lacking lipid rafts-associated proteins or enzymes involved in the synthesis of ergosterol and sphingolipids, we found that perturbations in the composition of these membrane domains increase resistance to bLf-induced yeast cell death. Also, when Pma1p-lipid rafts association is compromised in the Pma1-10 mutant and in the absence of the Pma1p-binding protein Ast1p, the bLf killing activity is impaired. Altogether, results showed that the perturbation of lipid rafts and the inhibition of both Pma1p and V-ATPase activities mediate the antifungal activity of bLf. Since it is suggested that the combination of conventional antifungals with lipid rafts-disrupting compounds is a powerful antifungal approach, our data will help to pave the way for the use of bLf alone or in combination for the treatment/eradication of clinically and agronomically relevant yeast pathogens/fungi.

Emerging insights on the role of V‐ATPase in human diseases: Therapeutic challenges and opportunities

“…In agreement with this, BafA1 and ammonium chloride, two lysosomotropic agents that inhibit V‐ATPase activity, almost completely reduced the entry of a SARS‐CoV‐2 pseudovirion in cells expressing its receptor 320 . Our studies demonstrating the ability of lactoferrin to inhibit V‐ATPase add this natural protein to the list of anti‐viral agents, which in particular can be explored in the treatment of infections caused by SARS‐CoV‐2 82,102,322 . These examples further highlight the power of modulating V‐ATPase activity for antiviral purposes.…”

Cyclo(Pro-Tyr) elicits conserved cellular damage in fungi by targeting the [H+]ATPase Pma1 in plasma membrane domains

“…Based on these studies and our experimental evidence, we propose that failure of Pma1 to oligomerize may trigger the internalization of membrane sections containing incorrectly folded proteins and, consequently, trigger the remaining physiological alterations associated with the disruption of membrane homeostasis. Thus, our findings support the rationale of using Pma1 as a target for designing novel antifungal molecules 39–41 . Methotrexate and fluconazole are two molecules that deplete Pma1 in S. cerevisiae , compromising ion transport and the influx of nutrients 29,42 .…”

A comprehensive review on lactoferrin: a natural multifunctional glycoprotein

“…48 A recent study revealed that Lf binding protein Pma1p, a proton pump which is inhibited by Lf through interruption of the membrane ergosterol and sphingolipid rich lipid rafts connected with Pma1p, promotes the intracellular accumulation of ergosterol, and it also reduces the V-ATPase activity to stimulate antifungal activity. 49 Furthermore, hLf prevents oral candidiasis caused by C. albicans, via upregulation of anti-inflammatory cytokines…”

“…48 A recent study revealed that Lf binding protein Pma1p, a proton pump which is inhibited by Lf through interruption of the membrane ergosterol and sphingolipid rich lipid rafts connected with Pma1p, promotes the intracellular accumulation of ergosterol, and it also reduces the V-ATPase activity to stimulate antifungal activity. 49 Furthermore, hLf prevents oral candidiasis caused by C. albicans , via upregulation of anti-inflammatory cytokines and downregulation of virulent candida genes. 50 The antifungal drug like fluconazole combined with Lf from various species significantly enhanced the inhibitory effect and decreased the minimum inhibitory concentration (MIC) against Candida species, Saccharomyces cerevisiae , and Cryptococcus neoformans .…”