ABCA1 transporter reduces amphotericin B cytotoxicity in mammalian cells

A Wu,K Wójtowicz,R Luchowski,N Filipczak,T Trombik,M Gagoś,W I Gruszecki,A Szczepaniak,O Raducka-Jaszul,E Grela,W Grudziński,G Chimini,Y Hamon

doi:10.1007/s00018-019-03154-w

Abstract

Amphotericin B (AmB) belongs to a group of polyene antibiotics commonly used in the treatment of systemic mycotic infections. A widely accepted mechanism of action of AmB is based on the formation of an oligomeric pore structure within the plasma membrane (PM) by interaction with membrane sterols. Although AmB binds preferentially to ergosterol, it can also bind to cholesterol in the mammalian PM and cause severe cellular toxicity. The lipid content and its lateral organization at the cell PM appear to be significant for AmB binding. Several ATP-binding cassette (ABC) transporters, including ABCA1, play a crucial role in lipid translocation, cholesterol redistribution and efflux. Here, we demonstrate that cells expressing ABCA1 are more resistant to AmB treatment, while cells lacking ABCA1 expression or expressing non-active ABCA1MM mutant display increased sensitivity. Further, a FLIM analysis of AmB-treated cells reveals a fraction of the antibiotic molecules, characterized by relatively high fluorescence lifetimes (> 6 ns), involved in formation of bulk cholesterol–AmB structures at the surface of ABCA1-expressing cells. Finally, lowering the cellular cholesterol content abolishes resistance of ABCA1-expressing cells to AmB. Therefore, we propose that ABCA1-mediated cholesterol efflux from cells induces formation of bulk cholesterol–AmB structures at the cell surface, preventing AmB cytotoxicity.

Highlights

Amphotericin B (AmB) belongs to a group of lifesaving polyene antibiotics used in the treatment of systemic mycoses, whose number is growing, especially in immunosuppressed patients, and as a response to more and more widely used treatments with broad-spectrum antibacterial antibiotics
We used murine Raw 264.7 macrophages that were treated with the liver X receptor (LXR) agonist GW3965 to induce ABCA1 expression [36, 37]
We propose a new potential mechanism of cell resistance to AmB based on the activity of the cholesterol efflux protein ABCA1

Summary

Introduction

Amphotericin B (AmB) belongs to a group of lifesaving polyene antibiotics used in the treatment of systemic mycoses, whose number is growing, especially in immunosuppressed patients, and as a response to more and more widely used treatments with broad-spectrum antibacterial antibiotics. AmB has been demonstrated to effectively “pull out” sterol molecules from the lipid phase and immobilize them within extramembranous, sterol–AmB hybrid structures [7]. Both mechanisms are relevant from the standpoint of the mode of action of AmB. In sterol-free, liquid-disordered (Ld) membranes, AmB has low orientational and positional stability [6, 9] This suggests that the lipid-ordered phase in model membrane (considered as experimental lipid rafts) favors the formation of stable channels [5]

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