Effect of cilofungin (LY121019), a fungal cell wall synthesis inhibitor, on interactions of Candida albicans with human neutrophils

Fungal cell wall synthesis is essential for viability, requiring the activity of genes involved in environmental sensing, precursor synthesis, transport, secretion, and assembly. This multitude of potential targets, the availability of known agents targeting this pathway, and the unique nature of fungal cell wall synthesis make this pathway an appealing target for drug discovery. Here we describe the adaptation of an assay monitoring cell wall synthesis for high-throughput screening. The assay requires fungal cell growth, in the presence of the test compound, for 3 h before the cells are subjected to osmotic shock in the presence of a dye that stains DNA. Miniaturization of the assay to a 384-well plate format and removing a mechanical transfer led to subtle changes in the assay characteristics. Validation of the assay with a library of known pharmacologically active agents has identified a number of different classes of compounds that are active in this assay, causing aberrant cell wall morphology and in many cases the inhibition of fungal cell growth.

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CsiA is a bacterial cell wall synthesis inhibitor contributing to DNA translocation through the cell envelope

Molecular Microbiology ◽

10.1111/j.1365-2958.2009.06683.x ◽

2009 ◽

Vol 72 (3) ◽

pp. 779-794 ◽

Cited By ~ 8

Author(s):

Régis Stentz ◽

Udo Wegmann ◽

Mary Parker ◽

Roy Bongaerts ◽

Laurie Lesaint ◽

...

Keyword(s):

Cell Wall ◽

Bacterial Cell ◽

Cell Envelope ◽

Bacterial Cell Wall ◽

Cell Wall Synthesis ◽

Dna Translocation ◽

Synthesis Inhibitor ◽

Cell Wall Synthesis Inhibitor

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Analysis of the 3H8 antigen of Candida albicans reveals new aspects of the organization of fungal cell wall proteins

FEMS Yeast Research ◽

10.1093/femsyr/foy035 ◽

2018 ◽

Vol 18 (4) ◽

Author(s):

Rafael Sentandreu ◽

Antonio Caminero ◽

Itzel Rentería ◽

Claudia León-Ramirez ◽

Luis González-de-la-Vara ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Fungal Cell Wall ◽

Cell Wall Proteins ◽

Fungal Cell

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The Viscoelastic Properties of the Fungal Cell Wall Allow Traffic of AmBisome as Intact Liposome Vesicles

mBio ◽

10.1128/mbio.02383-17 ◽

2018 ◽

Vol 9 (1) ◽

Cited By ~ 55

Author(s):

Louise Walker ◽

Prashant Sood ◽

Megan D. Lenardon ◽

Gillian Milne ◽

Jon Olson ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Cell Membrane ◽

Amphotericin B ◽

Mode Of Action ◽

Viscoelastic Properties ◽

Fungal Cell Wall ◽

Fungal Cell ◽

Pass Through ◽

Cell Wall Porosity

ABSTRACT The fungal cell wall is a critically important structure that represents a permeability barrier and protective shield. We probed Candida albicans and Cryptococcus neoformans with liposomes containing amphotericin B (AmBisome), with or without 15-nm colloidal gold particles. The liposomes have a diameter of 60 to 80 nm, and yet their mode of action requires them to penetrate the fungal cell wall to deliver amphotericin B to the cell membrane, where it binds to ergosterol. Surprisingly, using cryofixation techniques with electron microscopy, we observed that the liposomes remained intact during transit through the cell wall of both yeast species, even though the predicted porosity of the cell wall (pore size, ~5.8 nm) is theoretically too small to allow these liposomes to pass through intact. C. albicans mutants with altered cell wall thickness and composition were similar in both their in vitro AmBisome susceptibility and the ability of liposomes to penetrate the cell wall. AmBisome exposed to ergosterol-deficient C. albicans failed to penetrate beyond the mannoprotein-rich outer cell wall layer. Melanization of C. neoformans and the absence of amphotericin B in the liposomes were also associated with a significant reduction in liposome penetration. Therefore, AmBisome can reach cell membranes intact, implying that fungal cell wall viscoelastic properties are permissive to vesicular structures. The fact that AmBisome can transit through chemically diverse cell wall matrices when these liposomes are larger than the theoretical cell wall porosity suggests that the wall is capable of rapid remodeling, which may also be the mechanism for release of extracellular vesicles. IMPORTANCE AmBisome is a broad-spectrum fungicidal antifungal agent in which the hydrophobic polyene antibiotic amphotericin B is packaged within a 60- to 80-nm liposome. The mode of action involves perturbation of the fungal cell membrane by selectively binding to ergosterol, thereby disrupting membrane function. We report that the AmBisome liposome transits through the cell walls of both Candida albicans and Cryptococcus neoformans intact, despite the fact that the liposome is larger than the theoretical cell wall porosity. This implies that the cell wall has deformable, viscoelastic properties that are permissive to transwall vesicular traffic. These observations help explain the low toxicity of AmBisome, which can deliver its payload directly to the cell membrane without unloading the polyene in the cell wall. In addition, these findings suggest that extracellular vesicles may also be able to pass through the cell wall to deliver soluble and membrane-bound effectors and other molecules to the extracellular space.

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Iron alters the cell wall composition and intracellular lactate to affect Candida albicans susceptibility to antifungals and host immune response

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.013413 ◽

2020 ◽

Vol 295 (29) ◽

pp. 10032-10044 ◽

Cited By ~ 3

Author(s):

Aparna Tripathi ◽

Elisabetta Liverani ◽

Alexander Y. Tsygankov ◽

Sumant Puri

Keyword(s):

Immune Response ◽

Cell Wall ◽

Candida Albicans ◽

Signaling Pathways ◽

Host Immune Response ◽

Fungal Cell Wall ◽

Mitogen Activated Protein Kinase ◽

Fungal Cell ◽

High Iron ◽

Cell Wall Architecture

Fungal pathogen Candida albicans has a complex cell wall consisting of an outer layer of mannans and an inner layer of β-glucans and chitin. The fungal cell wall is the primary target for antifungals and is recognized by host immune cells. Environmental conditions such as carbon sources, pH, temperature, and oxygen tension can modulate the fungal cell wall architecture. Cellular signaling pathways, including the mitogen-activated protein kinase (MAPK) pathways, are responsible for sensing environmental cues and mediating cell wall alterations. Although iron has recently been shown to affect β-1,3-glucan exposure on the cell wall, we report here that iron changes the composition of all major C. albicans cell wall components. Specifically, high iron decreased the levels of mannans (including phosphomannans) and chitin; and increased β-1,3-glucan levels. These changes increased the resistance of C. albicans to cell wall-perturbing antifungals. Moreover, high iron cells exhibited adequate mitochondrial functioning; leading to a reduction in accumulation of lactate that signals through the transcription factor Crz1 to induce β-1,3-glucan masking in C. albicans. We show here that iron-induced changes in β-1,3-glucan exposure are lactate-dependent; and high iron causes β-1,3-glucan exposure by preventing lactate-induced, Crz1-mediated inhibition of activation of the fungal MAPK Cek1. Furthermore, despite exhibiting enhanced antifungal resistance, high iron C. albicans cells had reduced survival upon phagocytosis by macrophages. Our results underscore the role of iron as an environmental signal in multiple signaling pathways that alter cell wall architecture in C. albicans, thereby affecting its survival upon exposure to antifungals and host immune response.

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