Control of β-glucan exposure by the endo-1,3-glucanase Eng1 in Candida albicans modulates virulence

Candida albicans is a major opportunistic pathogen of humans. It can grow as morphologically distinct yeast, pseudohyphae and hyphae, and the ability to switch reversibly among different forms is critical for its virulence. The relationship between morphogenesis and innate immune recognition is not quite clear. Dectin-1 is a major C-type lectin receptor that recognizes β-glucan in the fungal cell wall. C. albicans β-glucan is usually masked by the outer mannan layer of the cell wall. Whether and how β-glucan masking is differentially regulated during hyphal morphogenesis is not fully understood. Here we show that the endo-1,3-glucanase Eng1 is differentially expressed in yeast, and together with Yeast Wall Protein 1 (Ywp1), regulates β-glucan exposure and Dectin-1-dependent immune activation of macrophage by yeast cells. ENG1 deletion results in enhanced Dectin-1 binding at the septa of yeast cells; while eng1 ywp1 yeast cells show strong overall Dectin-1 binding similar to hyphae of wild-type and eng1 mutants. Correlatively, hyphae of wild-type and eng1 induced similar levels of cytokines in macrophage. ENG1 expression and Eng1-mediated β-glucan trimming are also regulated by antifungal drugs, lactate and N-acetylglucosamine. Deletion of ENG1 modulates virulence in the mouse model of hematogenously disseminated candidiasis in a Dectin-1-dependent manner. The eng1 mutant exhibited attenuated lethality in male mice, but enhanced lethality in female mice, which was associated with a stronger renal immune response and lower fungal burden. Thus, Eng1-regulated β-glucan exposure in yeast cells modulates the balance between immune protection and immunopathogenesis during disseminated candidiasis.

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Fluconazole and Lipopeptide Surfactin Interplay During Candida albicans Plasma Membrane and Cell Wall Remodeling Increases Fungal Immune System Exposure

Pharmaceutics ◽

10.3390/pharmaceutics12040314 ◽

2020 ◽

Vol 12 (4) ◽

pp. 314 ◽

Cited By ~ 3

Author(s):

Jakub Suchodolski ◽

Daria Derkacz ◽

Jakub Muraszko ◽

Jarosław J. Panek ◽

Aneta Jezierska ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Plasma Membrane ◽

Immune System ◽

Antifungal Drugs ◽

Stable Complex ◽

Host Immune System ◽

Chitin Synthesis ◽

Fungal Cell ◽

In Silico Studies

Recognizing the β-glucan component of the Candida albicans cell wall is a necessary step involved in host immune system recognition. Compounds that result in exposed β-glucan recognizable to the immune system could be valuable antifungal drugs. Antifungal development is especially important because fungi are becoming increasingly drug resistant. This study demonstrates that lipopeptide, surfactin, unmasks β-glucan when the C. albicans cells lack ergosterol. This observation also holds when ergosterol is depleted by fluconazole. Surfactin does not enhance the effects of local chitin accumulation in the presence of fluconazole. Expression of the CHS3 gene, encoding a gene product resulting in 80% of cellular chitin, is downregulated. C. albicans exposure to fluconazole changes the composition and structure of the fungal plasma membrane. At the same time, the fungal cell wall is altered and remodeled in a way that makes the fungi susceptible to surfactin. In silico studies show that surfactin can form a complex with β-glucan. Surfactin forms a less stable complex with chitin, which in combination with lowering chitin synthesis, could be a second anti-fungal mechanism of action of this lipopeptide.

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Azole Resistance by Loss of Function of the Sterol Δ5,6-Desaturase Gene (ERG3) in Candida albicans Does Not Necessarily Decrease Virulence

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05720-11 ◽

2012 ◽

Vol 56 (4) ◽

pp. 1960-1968 ◽

Cited By ~ 53

Author(s):

L. A. Vale-Silva ◽

A. T. Coste ◽

F. Ischer ◽

J. E. Parker ◽

S. L. Kelly ◽

...

Keyword(s):

Candida Albicans ◽

Premature Stop Codon ◽

Antifungal Drugs ◽

Ergosterol Biosynthesis ◽

Azole Resistance ◽

Wild Type ◽

Disseminated Candidiasis ◽

Double Base

ABSTRACTThe inactivation ofERG3, a gene encoding sterol Δ5,6-desaturase (essential for ergosterol biosynthesis), is a known mechanism ofin vitroresistance to azole antifungal drugs in the human pathogenCandida albicans. ERG3inactivation typically results in loss of filamentation and attenuated virulence in animal models of disseminated candidiasis. In this work, we identified aC. albicansclinical isolate (VSY2) with high-level resistance to azole drugsin vitroand an absence of ergosterol but normal filamentation. Sequencing ofERG3in VSY2 revealed a double base deletion leading to a premature stop codon and thus a nonfunctional enzyme. The reversion of the double base deletion in the mutant allele (erg3-1) restored ergosterol biosynthesis and full fluconazole susceptibility in VSY2, confirming thatERG3inactivation was the mechanism of azole resistance. Additionally, the replacement of bothERG3alleles byerg3-1in the wild-type strain SC5314 led to the absence of ergosterol and to fluconazole resistance without affecting filamentation. In a mouse model of disseminated candidiasis, the clinicalERG3mutant VSY2 produced kidney fungal burdens and mouse survival comparable to those obtained with the wild-type control. Interestingly, while VSY2 was resistant to fluconazole bothin vitroandin vivo, theERG3-derived mutant of SC5314 was resistant onlyin vitroand was less virulent than the wild type. This suggests that VSY2 compensated for thein vivofitness defect ofERG3inactivation by a still unknown mechanism(s). Taken together, our results provide evidence that contrary to previous reports inactivation ofERG3does not necessarily affect filamentation and virulence.

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Crosstalk between calcineurin and the cell wall integrity pathways prevents chitin overexpression in Candida albicans

Journal of Cell Science ◽

10.1242/jcs.258889 ◽

2021 ◽

Author(s):

Alessandra da Silva Dantas ◽

Filomena Nogueira ◽

Keunsook K. Lee ◽

Louise A. Walker ◽

Matt Edmondson ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Antifungal Drugs ◽

Cell Wall Integrity ◽

Yeast Cells ◽

Outer Cell ◽

Chitin Synthesis ◽

Simultaneous Activation ◽

Glucan Synthesis ◽

Calcineurin Pathway

Echinocandins such as caspofungin are front line antifungal drugs that compromise β-1,3 glucan synthesis in the cell wall. Recent reports have shown that fungal cells can resist killing by caspofungin by up-regulation of chitin synthesis, thereby sustaining cell wall integrity. When echinocandins are removed, the chitin content of cells quickly returns to basal levels, suggesting that there is a fitness cost associated with having elevated levels of chitin in the cell wall. We show here that simultaneous activation of the calcineurin and CWI pathways generates a sub-population of Candida albicans yeast cells that have supra-normal chitin levels interspersed throughout the inner and outer cell wall, and that these cells are non-viable, perhaps due to loss of wall elasticity required for cell expansion and growth. Mutations in the Ca2+-calcineurin pathway prevented the formation of these non-viable super high chitin cells by negatively regulating chitin synthesis driven by the CWI pathway. The Ca2+-calcineurin pathway may therefore act as an attenuator that prevents the overproduction of chitin by coordinating both chitin upregulation and negative regulation of the CWI signaling pathway.

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Molecular Analysis of the Candida albicans Homolog of Saccharomyces cerevisiae MNN9, Required for Glycosylation of Cell Wall Mannoproteins

Journal of Bacteriology ◽

10.1128/jb.181.24.7439-7448.1999 ◽

1999 ◽

Vol 181 (24) ◽

pp. 7439-7448 ◽

Cited By ~ 37

Author(s):

Susan B. Southard ◽

Charles A. Specht ◽

Chitra Mishra ◽

Joan Chen-Weiner ◽

Phillips W. Robbins

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Wall ◽

Candida Albicans ◽

Gene Family ◽

Molecular Analysis ◽

Transmembrane Domain ◽

Antifungal Drugs ◽

Northern Analysis ◽

Fungal Cell ◽

Poor Growth

ABSTRACT The fungal cell wall has generated interest as a potential target for developing antifungal drugs, and the genes encoding glucan and chitin in fungal pathogens have been studied to this end. Mannoproteins, the third major component of the cell wall, contain mannose in either O- or N-glycosidic linkages. Here we describe the molecular analysis of the Candida albicans homolog ofSaccharomyces cerevisiae MNN9, a gene required for the synthesis of N-linked outer-chain mannan in yeast, and the phenotypes associated with its disruption. CaMNN9 has significant homology with S. cerevisiae MNN9, including a putative N-terminal transmembrane domain, and represents a member of a similar gene family in Candida. CaMNN9 resides on chromosome 3 and is expressed at similar levels in both yeast and hyphal cells. Disruption of both copies of CaMNN9 leads to phenotypic effects characteristic of cell wall defects including poor growth in liquid media and on solid media, formation of aggregates in liquid culture, osmotic sensitivity, aberrant hyphal formation, and increased sensitivity to lysis after treatment with β-1,3-glucanase. Like all members of the S. cerevisiae MNN9 gene family theCamnn9Δ strain is resistant to sodium orthovanadate and sensitive to hygromycin B. Analysis of cell wall-associated carbohydrates showed the Camnn9Δ strain to contain half the amount of mannan present in cell walls derived from the wild-type parent strain. Reverse transcription-PCR and Northern analysis of the expression of MNN9 gene family members CaVAN1and CaANP1 in the Camnn9Δ strain showed that transcription of those genes is not affected in the absence ofCaMNN9 transcription. Our results suggest that, while the role MNN9 plays in glycosylation in bothCandida and Saccharomyces is conserved, loss ofMNN9 function in C. albicans leads to phenotypes that are inconsistent with the pathogenicity of the organism and thus identify CaMnn9p as a potential drug target.

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(R)-(+)-β-Citronellol and (S)-(−)-β-Citronellol in Combination with Amphotericin B against Candida Spp.

International Journal of Molecular Sciences ◽

10.3390/ijms21051785 ◽

2020 ◽

Vol 21 (5) ◽

pp. 1785 ◽

Cited By ~ 4

Author(s):

Daniele Silva ◽

Hermes Diniz-Neto ◽

Laísa Cordeiro ◽

Maria Silva-Neta ◽

Shellygton Silva ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Minimum Inhibitory Concentration ◽

Amphotericin B ◽

Mechanism Of Action ◽

Inhibitory Concentration ◽

Association Studies ◽

Antifungal Drugs ◽

Candida Spp ◽

Fungal Cell

The enantiomers (R)-(+)-β-citronellol and (S)-(−)-β-citronellol are present in many medicinal plants, but little is understood about their bioactivity against Candida yeasts. This study aimed to evaluate the behavior of positive and negative enantiomers of β-citronellol on strains of Candida albicans and C. tropicalis involved in candidemia. The minimum inhibitory concentration (MIC) and minimum fungicide concentration (MFC) were determined. The evaluation of growth kinetics, mechanism of action, and association studies with Amphotericin B (AB) using the checkerboard method was also performed. R-(+)-β-citronellol and S-(−)-β-citronellol presented a MIC50% of 64 µg/mL and a MFC50% of 256 µg/mL for C. albicans strains. For C. tropicalis, the isomers exhibited a MIC50% of 256 µg/mL and a MFC50% of 1024 µg/mL. In the mechanism of action assay, both substances displayed an effect on the fungal membrane but not on the fungal cell wall. Synergism and indifference were observed in the association of R-(+)-β-citronellol and AB, while the association between S-(−)-β-citronellol and AB displayed synergism, additivity, and indifference. In conclusion, both isomers of β-citronellol presented a similar profile of antifungal activity. Hence, they can be contemplated in the development of new antifungal drugs providing that further research is conducted about their pharmacology and toxicity.

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Elevated Chitin Content Reduces the Susceptibility of Candida Species to Caspofungin

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01486-12 ◽

2012 ◽

Vol 57 (1) ◽

pp. 146-154 ◽

Cited By ~ 98

Author(s):

Louise A. Walker ◽

Neil A. R. Gow ◽

Carol A. Munro

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Point Mutations ◽

Antifungal Drugs ◽

Cell Wall Polysaccharide ◽

Chitin Synthesis ◽

Fungal Cell ◽

Content Type ◽

Chitin Content ◽

Compensatory Increase

ABSTRACTThe echinocandin antifungal drugs inhibit synthesis of the major fungal cell wall polysaccharide β(1,3)-glucan. Echinocandins have good efficacy againstCandida albicansbut reduced activity against otherCandidaspecies, in particularCandida parapsilosisandCandida guilliermondii. Treatment ofCandida albicanswith a sub-MIC level of caspofungin has been reported to cause a compensatory increase in chitin content and to select for sporadic echinocandin-resistantFKS1point mutants that also have elevated cell wall chitin. Here we show that elevated chitin in response to caspofungin is a common response in variousCandidaspecies. Activation of chitin synthesis was observed in isolates ofC. albicans,Candida tropicalis,C. parapsilosis, andC. guilliermondiiand in some isolates ofCandida kruseiin response to caspofungin treatment. However,Candida glabrataisolates demonstrated no exposure-induced change in chitin content. Furthermore, isolates ofC. albicans,C. krusei,C. parapsilosis, andC. guilliermondiiwhich were stimulated to have higher chitin levels via activation of the calcineurin and protein kinase C (PKC) signaling pathways had reduced susceptibility to caspofungin. Isolates containing point mutations in theFKS1gene generally had higher chitin levels and did not demonstrate a further compensatory increase in chitin content in response to caspofungin treatment. These results highlight the potential of increased chitin synthesis as a potential mechanism of tolerance to caspofungin for the major pathogenicCandidaspecies.

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Revisiting the Vital Drivers and Mechanisms of β-Glucan Masking in Human Fungal Pathogen, Candida albicans

Pathogens ◽

10.3390/pathogens10080942 ◽

2021 ◽

Vol 10 (8) ◽

pp. 942

Author(s):

Saif Hameed ◽

Sandeep Hans ◽

Shweta Singh ◽

Ruby Dhiman ◽

Ross Monasky ◽

...

Keyword(s):

Candida Albicans ◽

Fungal Pathogens ◽

Pathogenic Fungi ◽

Antifungal Drugs ◽

Immune Recognition ◽

Fungal Cell ◽

Distinctive Features ◽

Host Immune Responses ◽

Human Fungal Pathogen ◽

Buried Layers

Among the several human fungal pathogens, Candida genus represents one of the most implicated in the clinical scenario. There exist several distinctive features that govern the establishment of Candida infections in addition to their capacity to adapt to multiple stress conditions inside humans which also include evasion of host immune responses. The complex fungal cell wall of the prevalent pathogen, Candida albicans, is one of the main targets of antifungal drugs and recognized by host immune cells. The wall consists of tiered arrangement of an outer thin but dense covering of mannan and inner buried layers of β-glucan and chitin. However, the pathogenic fungi adopt strategies to evade immune recognition by masking these molecules. This capacity to camouflage the immunogenic polysaccharide β-glucan from the host is a key virulence factor of C. albicans. The present review is an attempt to collate various underlying factors and mechanisms involved in Candida β-glucan masking from the available pool of knowledge and provide a comprehensive understanding. This will further improve therapeutic approaches to candidiasis by identifying new antifungal targets that blocks fungal immune evasion.

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Efficacy of Novel Schiff base Derivatives as Antifungal Compounds in Combination with Approved Drugs Against Candida Albicans

Medicinal Chemistry ◽

10.2174/1573406415666181203115957 ◽

2019 ◽

Vol 15 (6) ◽

pp. 648-658 ◽

Cited By ~ 1

Author(s):

Manzoor Ahmad Malik ◽

Shabir Ahmad Lone ◽

Parveez Gull ◽

Ovas Ahmad Dar ◽

Mohmmad Younus Wani ◽

...

Keyword(s):

Schiff Base ◽

Candida Albicans ◽

Antifungal Activity ◽

Fungal Infections ◽

Total Sterol ◽

Antifungal Drugs ◽

New Drugs ◽

Ergosterol Biosynthesis ◽

Dependent Manner ◽

Schiff Base Derivatives

Background: The increasing incidence of fungal infections, especially caused by Candida albicans, and their increasing drug resistance has drastically increased in recent years. Therefore, not only new drugs but also alternative treatment strategies are promptly required. Methods: We previously reported on the synergistic interaction of some azole and non-azole compounds with fluconazole for combination antifungal therapy. In this study, we synthesized some non-azole Schiff-base derivatives and evaluated their antifungal activity profile alone and in combination with the most commonly used antifungal drugs- fluconazole (FLC) and amphotericin B (AmB) against four drug susceptible, three FLC resistant and three AmB resistant clinically isolated Candida albicans strains. To further analyze the mechanism of antifungal action of these compounds, we quantified total sterol contents in FLC-susceptible and resistant C. albicans isolates. Results: A pyrimidine ring-containing derivative SB5 showed the most potent antifungal activity against all the tested strains. After combining these compounds with FLC and AmB, 76% combinations were either synergistic or additive while as the rest of the combinations were indifferent. Interestingly, none of the combinations was antagonistic, either with FLC or AmB. Results interpreted from fractional inhibitory concentration index (FICI) and isobolograms revealed 4-10-fold reduction in MIC values for synergistic combinations. These compounds also inhibit ergosterol biosynthesis in a concentration-dependent manner, supported by the results from docking studies. Conclusion: The results of the studies conducted advocate the potential of these compounds as new antifungal drugs. However, further studies are required to understand the other mechanisms and in vivo efficacy and toxicity of these compounds.

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The newly synthesized thiazole derivatives as potential antifungal compounds against Candida albicans

Applied Microbiology and Biotechnology ◽

10.1007/s00253-021-11477-7 ◽

2021 ◽

Author(s):

Anna Biernasiuk ◽

Anna Berecka-Rycerz ◽

Anna Gumieniczek ◽

Maria Malm ◽

Krzysztof Z. Łączkowski ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Antifungal Activity ◽

Cell Membrane ◽

Mode Of Action ◽

Thiazole Derivatives ◽

Fungal Cell ◽

Erythrocyte Lysis ◽

Low Cytotoxicity ◽

High Antifungal Activity

Abstract Recently, the occurrence of candidiasis has increased dramatically, especially in immunocompromised patients. Additionally, their treatment is often ineffective due to the resistance of yeasts to antimycotics. Therefore, there is a need to search for new antifungals. A series of nine newly synthesized thiazole derivatives containing the cyclopropane system, showing promising activity against Candida spp., has been further investigated. We decided to verify their antifungal activity towards clinical Candida albicans isolated from the oral cavity of patients with hematological malignancies and investigate the mode of action on fungal cell, the effect of combination with the selected antimycotics, toxicity to erythrocytes, and lipophilicity. These studies were performed by the broth microdilution method, test with sorbitol and ergosterol, checkerboard technique, erythrocyte lysis assay, and reversed phase thin-layer chromatography, respectively. All derivatives showed very strong activity (similar and even higher than nystatin) against all C. albicans isolates with minimal inhibitory concentration (MIC) = 0.008–7.81 µg/mL Their mechanism of action may be related to action within the fungal cell wall structure and/or within the cell membrane. The interactions between the derivatives and the selected antimycotics (nystatin, chlorhexidine, and thymol) showed additive effect only in the case of combination some of them and thymol. The erythrocyte lysis assay confirmed the low cytotoxicity of these compounds as compared to nystatin. The high lipophilicity of the derivatives was related with their high antifungal activity. The present studies confirm that the studied thiazole derivatives containing the cyclopropane system appear to be a very promising group of compounds in treatment of infections caused by C. albicans. However, this requires further studies in vivo. Key points • The newly thiazoles showed high antifungal activity and some of them — additive effect in combination with thymol. • Their mode of action may be related with the influence on the structure of the fungal cell wall and/or the cell membrane. • The low cytotoxicity against erythrocytes and high lipophilicity of these derivatives are their additional good properties. Graphical abstract

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CaNdt80 Is Involved in Drug Resistance in Candida albicans by Regulating CDR1

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.12.4505-4512.2004 ◽

2004 ◽

Vol 48 (12) ◽

pp. 4505-4512 ◽

Cited By ~ 66

Author(s):

Chia-Geun Chen ◽

Yun-Liang Yang ◽

Hsin-I Shih ◽

Chia-Li Su ◽

Hsiu-Jung Lo

Keyword(s):

Saccharomyces Cerevisiae ◽

Drug Resistance ◽

Candida Albicans ◽

Type Strain ◽

Antifungal Agents ◽

Wild Type Strain ◽

Efflux Pump ◽

Antifungal Drugs ◽

Galactosidase Activity ◽

Wild Type

ABSTRACT Overexpression of CDR1, an efflux pump, is one of the major mechanisms contributing to drug resistance in Candida albicans. CDR1 p-lacZ was constructed and transformed into a Saccharomyces cerevisiae strain so that the lacZ gene could be used as the reporter to monitor the activity of the CDR1 promoter. Overexpression of CaNDT80, the C. albicans homolog of S. cerevisiae NDT80, increases the β-galactosidase activity of the CDR1 p-lacZ construct in S. cerevisiae. Furthermore, mutations in CaNDT80 abolish the induction of CDR1 expression by antifungal agents in C. albicans. Consistently, the Candt80/Candt80 mutant is also more susceptible to antifungal drugs than the wild-type strain. Thus, the gene for CaNdt80 may be the first gene among the regulatory factors involved in drug resistance in C. albicans whose function has been identified.

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