scholarly journals Interface of Candida albicans Biofilm Matrix-Associated Drug Resistance and Cell Wall Integrity Regulation

2011 ◽  
Vol 10 (12) ◽  
pp. 1660-1669 ◽  
Author(s):  
Jeniel E. Nett ◽  
Hiram Sanchez ◽  
Michael T. Cain ◽  
Kelly M. Ross ◽  
David R. Andes
Keyword(s):  
Drug Resistance ◽  
Cell Wall ◽  
Candida Albicans ◽  
Antifungal Drugs ◽  
Cell Wall Integrity ◽  
Candidate Gene Approach ◽  
Content Type ◽  
Resistant Phenotype ◽  
Cell Wall Integrity Pathway ◽  
Biofilm Matrix

ABSTRACTCandida albicansfrequently infects medical devices by growing as a biofilm, i.e., a community of adherent organisms entrenched in an extracellular matrix. During biofilm growth,Candidaspp. acquire the ability to resist high concentrations of antifungal drugs. One recently recognized biofilm resistance mechanism involves drug sequestration by matrix β-1,3 glucan. Using a candidate gene approach, we investigated potentialC. albicansβ-1,3-glucan regulators, based on their homology toSaccharomyces cerevisiae, includingSMI1and protein kinase C (PKC) pathway components. We identified a role for theSMI1in biofilm matrix glucan production and development of the associated drug resistance phenotype. This pathway appears to act through transcription factor Rlmp and glucan synthase Fks1p. The phenotypes of these mutant biofilms mimicked those of thesmi1Δ/smi1Δ biofilm, and overexpression ofFKS1in thesmi1Δ/smi1Δ mutant restored the biofilm resistant phenotype. However, control of this pathway is distinct from that of the upstream PKC pathway because thepkc1Δ/pkc1Δ,bck1Δ/bck1Δ,mkk2Δ/mkk2Δ, andmkc1Δ/mkc1Δ biofilms retained the resistant phenotype of the parent strain. In addition, resistance to cell-perturbing agents and gene expression data do not support a significant role for the cell wall integrity pathway during the biofilm formation. Here we show that Smi1p functions in conjunction with Rlm1p and Fks1p to produce drug-sequestering biofilm β-glucan. Our work provides new insight into how theC. albicansbiofilm matrix production and drug resistance pathways intersect with the planktonic cell wall integrity pathway. This novel connection helps explain how pathogens in a multicellular biofilm community are protected from anti-infective therapy.

scholarly journals Global Screening of Genomic and Transcriptomic Factors Associated with Phenotype Differences between Multidrug-Resistant and -Susceptible Candida haemulonii Strains

mSystems ◽  
2019 ◽  
Vol 4 (6) ◽  
Author(s):  
Hao Zhang ◽  
Yifei Niu ◽  
Jingwen Tan ◽  
Weixia Liu ◽  
Ming-an Sun ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Cell Wall ◽  
Multidrug Resistance ◽  
Multidrug Resistant ◽  
Antifungal Drugs ◽  
Cell Wall Integrity ◽  
Close Relative ◽  
Content Type ◽  
Candida Haemulonii ◽  
Resistant Strains

ABSTRACT Candida haemulonii, a close relative of Candida auris, is an emerging pathogen which frequently shows multidrug resistance especially to triazoles, the most used antifungal drugs. The mechanisms of drug resistance in C. haemulonii, however, are largely unknown. Here, we sequenced and annotated the genomes of two reference strains from the C. haemulonii complex, compared the phenotypes, genomes, and transcriptomes of a triazole-susceptible and two triazole-resistant C. haemulonii strains, and identified triazole susceptibility, morphology, fitness, and the major genetic and gene expression differences between the strains. A multidrug efflux gene, CDR1, was recurrently found to be upregulated for expression in triazole-resistant strains. Blocking the activity of Cdr1 increased the susceptibility to triazoles strikingly. Comparative transcriptome analysis also demonstrated impaired cell wall integrity, filamentous growth, and iron homeostasis in triazole-resistant strains. Finally, we also identified a zinc-binding MHR family transcription regulator gene that mutated in triazole-resistant strains spontaneously, contributing to the changes of morphology and, possibly, cell wall integrity between the strains. The study provided important clues for future studies exploring the mechanisms of multidrug resistance and related phenotypic differences of C. haemulonii strains. IMPORTANCE A comprehensive, multi-omic survey was performed to disclose the genetic backgrounds and differences between multidrug-resistant and -susceptible C. haemulonii strains. Genes were identified with mutations or significant expression differences in multidrug-resistant compared to multidrug-susceptible strains, which were mainly involved in multidrug resistance, stress fitness, and morphology. The Cdr1-encoding gene, C. haemulonii 2486 (CH_2486), was expressed at a significantly increased level in multidrug-resistant strains. Functional inhibition experiments further implicated potential roles of CH_2486 in drug resistance. A gene spontaneously mutated in resistant strains, CH_4347, was experimentally validated to influence the morphology of spores, possibly by controlling cell wall integrity.

scholarly journals Highly Dynamic and Specific Phosphatidylinositol 4,5-Bisphosphate, Septin, and Cell Wall Integrity Pathway Responses Correlate with Caspofungin Activity against Candida albicans

2016 ◽  
Vol 60 (6) ◽  
pp. 3591-3600 ◽  
Author(s):  
Hassan Badrane ◽  
M. Hong Nguyen ◽  
Cornelius J. Clancy
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Fluorescent Protein ◽  
Cell Wall Integrity ◽  
Ph Domain ◽  
Calcofluor White ◽  
Content Type ◽  
Pathway Activation ◽  
Cell Wall Integrity Pathway ◽  
Green Fluorescent

Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] activates the yeast cell wall integrity pathway.Candida albicansexposure to caspofungin results in the rapid redistribution of PI(4,5)P2and septins to plasma membrane foci and subsequent fungicidal effects. We studiedC. albicansPI(4,5)P2and septin dynamics and protein kinase C (PKC)-Mkc1 cell wall integrity pathway activation following exposure to caspofungin and other drugs. PI(4,5)P2and septins were visualized by live imaging ofC. albicanscells coexpressing green fluorescent protein (GFP)-pleckstrin homology (PH) domain and red fluorescent protein-Cdc10p, respectively. PI(4,5)P2was also visualized in GFP-PH domain-expressingC. albicans mkc1mutants. Mkc1p phosphorylation was measured as a marker of PKC-Mkc1 pathway activation. Fungicidal activity was assessed using 20-h time-kill assays. Caspofungin immediately induced PI(4,5)P2and Cdc10p colocalization to aberrant foci, a process that was highly dynamic over 3 h. PI(4,5)P2levels increased in a dose-response manner at caspofungin concentrations of ≤4× MIC and progressively decreased at concentrations of ≥8× MIC. Caspofungin exposure resulted in broad-based mother-daughter bud necks and arrested septum-like structures, in which PI(4,5)P2and Cdc10 colocalized. PKC-Mkc1 pathway activation was maximal within 10 min, peaked in response to caspofungin at 4× MIC, and declined at higher concentrations. The caspofungin-induced PI(4,5)P2redistribution remained apparent inmkc1mutants. Caspofungin exerted dose-dependent killing and paradoxical effects at ≤4× and ≥8× MIC, respectively. Fluconazole, amphotericin B, calcofluor white, and H2O2did not impact the PI(4,5)P2or Cdc10p distribution like caspofungin did. Caspofungin exerts rapid PI(4,5)P2-septin and PKC-Mkc1 responses that correlate with the extent ofC. albicanskilling, and the responses are not induced by other antifungal agents. PI(4,5)P2-septin regulation is crucial in early caspofungin responses and PKC-Mkc1 activation.

scholarly journals Role of Matrix β-1,3 Glucan in Antifungal Resistance of Non-albicans Candida Biofilms

2013 ◽  
Vol 57 (4) ◽  
pp. 1918-1920 ◽  
Author(s):  
K. F. Mitchell ◽  
H. T. Taff ◽  
M. A. Cuevas ◽  
E. L. Reinicke ◽  
H. Sanchez ◽  
...  
Keyword(s):  
Health Care ◽  
Drug Resistance ◽  
Candida Albicans ◽  
Care Setting ◽  
Antifungal Susceptibility ◽  
Antifungal Drug ◽  
Content Type ◽  
The Matrix ◽  
Biofilm Matrix

ABSTRACTCandidabiofilm infections pose an increasing threat in the health care setting due to the drug resistance associated with this lifestyle. Several mechanisms underlie the resistance phenomenon. InCandida albicans, one mechanism involves drug impedance by the biofilm matrix linked to β-1,3 glucan. Here, we show this is important for otherCandidaspp. We identified β-1,3 glucan in the matrix, found that the matrix sequesters antifungal drug, and enhanced antifungal susceptibility with matrix β-1,3 glucan hydrolysis.

scholarly journals Aspergillus fumigatus G-Protein Coupled Receptors GprM and GprJ Are Important for the Regulation of the Cell Wall Integrity Pathway, Secondary Metabolite Production, and Virulence

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Aílton Pereira da Costa Filho ◽  
Guilherme Thomaz Pereira Brancini ◽  
Patrícia Alves de Castro ◽  
Clara Valero ◽  
Jaire Alves Ferreira Filho ◽  
...  
Keyword(s):  
Cell Wall ◽  
G Protein ◽  
Galleria Mellonella ◽  
G Protein Coupled Receptors ◽  
Cell Wall Integrity ◽  
Fungal Development ◽  
Content Type ◽  
Cell Wall Integrity Pathway ◽  
Mycotoxin Biosynthesis ◽  
G Protein Coupled

ABSTRACT G-protein coupled receptors (GPCRs) are extracellular signaling receptors that sense environmental cues. Fungi sense their environment primarily through GPCR-mediated signaling pathways, which, in turn, regulate fungal development, metabolism, virulence, and mycotoxin biosynthesis. Aspergillus fumigatus is an important human pathogen that causes aspergillosis, a heterogeneous group of diseases that present a wide range of clinical manifestations. Here, we investigate in detail the role of the GPCRs GprM and GprJ in growth and gene expression. GprM and GprJ are important for melanin production and the regulation of the cell wall integrity (CWI) pathway. Overexpression of gprM and gprJ causes a 20 and 50% reduction in growth rate compared to the wild-type (WT) strain and increases sensitivity to cell wall-damaging agents. Phosphorylation of the CWI protein kinase MpkA is increased in the ΔgprM and ΔgprJ strains and decreased in the overexpression mutants compared to the WT strain. Furthermore, differences in cell wall polysaccharide concentrations and organization were observed in these strains. Transcriptome sequencing suggests that GprM and GprJ negatively regulate genes encoding secondary metabolites (SMs). Mass spectrometry analysis confirmed that the production of fumagillin, pyripyropene, fumigaclavine C, fumiquinazoline, and fumitremorgin is reduced in the ΔgprM and ΔgprJ strains, at least partially through the activation of MpkA. Overexpression of grpM also resulted in the regulation of many transcription factors, with AsgA predicted to function downstream of GprM and MpkA signaling. Finally, we show that the ΔgprM and ΔgprJ mutants are reduced in virulence in the Galleria mellonella insect model of invasive aspergillosis. IMPORTANCE A. fumigatus is the main etiological agent of invasive pulmonary aspergillosis, a life-threatening fungal disease that occurs in severely immunocompromised humans. Withstanding the host environment is essential for A. fumigatus virulence, and sensing of extracellular cues occurs primarily through G-protein coupled receptors (GPCRs) that activate signal transduction pathways, which, in turn, regulate fungal development, metabolism, virulence, and mycotoxin biosynthesis. The A. fumigatus genome encodes 15 putative classical GPCRs, with only three having been functionally characterized to date. In this work, we show that the two GPCRs GprM and GprJ regulate the phosphorylation of the mitogen-activated protein kinase MpkA and thus control the regulation of the cell wall integrity pathway. GprM and GprJ are also involved in the regulation of the production of the secondary metabolites fumagillin, pyripyropene, fumigaclavine C, fumiquinazoline, melanin, and fumitremorgin, and this regulation partially occurs through the activation of MpkA. Furthermore, GprM and GprJ are important for virulence in the insect model Galleria mellonella. This work therefore functionally characterizes two GPCRs and shows how they regulate several intracellular pathways that have been shown to be crucial for A. fumigatus virulence.

scholarly journals Curcumin Targets Cell Wall Integrity via Calcineurin-Mediated Signaling in Candida albicans

2013 ◽  
Vol 58 (1) ◽  
pp. 167-175 ◽  
Author(s):  
Awanish Kumar ◽  
Sanjiveeni Dhamgaye ◽  
Indresh Kumar Maurya ◽  
Ashutosh Singh ◽  
Monika Sharma ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Critical Role ◽  
Pathogenic Fungi ◽  
Cell Wall Integrity ◽  
Ergosterol Biosynthesis ◽  
Ros Generation ◽  
Calcofluor White ◽  
Content Type ◽  
Cell Wall Damage

ABSTRACTCurcumin (CUR) shows antifungal activity against a range of pathogenic fungi, includingCandida albicans. The reported mechanisms of action of CUR include reactive oxygen species (ROS) generation, defects in the ergosterol biosynthesis pathway, decrease in hyphal development, and modulation of multidrug efflux pumps. Reportedly, each of these pathways is independently linked to the cell wall machinery inC. albicans, but surprisingly, CUR has not been previously implicated in cell wall damage. In the present study, we performed transcriptional profiling to identify the yet-unidentified targets of CUR inC. albicans. We found that, among 348 CUR-affected genes, 51 were upregulated and 297 were downregulated. Interestingly, most of the cell wall integrity pathway genes were downregulated. The possibility of the cell wall playing a critical role in the mechanism of CUR required further validation; therefore, we performed specific experiments to establish if there was any link between the two. The fractional inhibitory concentration index values of 0.24 to 0.37 show that CUR interacts synergistically with cell wall-perturbing (CWP) agents (caspofungin, calcofluor white, Congo red, and SDS). Furthermore, we could observe cell wall damage and membrane permeabilization by CUR alone, as well as synergistically with CWP agents. We also found hypersusceptibility in calcineurin and mitogen-activated protein (MAP) kinase pathway mutants against CUR, which confirmed that CUR also targets cell wall biosynthesis inC. albicans. Together, these data provide strong evidence that CUR disrupts cell wall integrity inC. albicans. This new information on the mechanistic action of CUR could be employed in improving treatment strategies and in combinatorial drug therapy.

scholarly journals Nanoscale Effects of Caspofungin against Two Yeast Species, Saccharomyces cerevisiae and Candida albicans

2013 ◽  
Vol 57 (8) ◽  
pp. 3498-3506 ◽  
Author(s):  
C. Formosa ◽  
M. Schiavone ◽  
H. Martin-Yken ◽  
J. M. François ◽  
R. E. Duval ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Candida Albicans ◽  
Yeast Species ◽  
Young Modulus ◽  
Antifungal Drugs ◽  
Molecular Organization ◽  
Response To Treatment ◽  
Content Type ◽  
High Doses

ABSTRACTSaccharomyces cerevisiaeandCandida albicansare model yeasts for biotechnology and human health, respectively. We used atomic force microscopy (AFM) to explore the effects of caspofungin, an antifungal drug used in hospitals, on these two species. Our nanoscale investigation revealed similar, but also different, behaviors of the two yeasts in response to treatment with the drug. While administration of caspofungin induced deep cell wall remodeling in both yeast species, as evidenced by a dramatic increase in chitin and decrease in β-glucan content, changes in cell wall composition were more pronounced withC. albicanscells. Notably, the increase of chitin was proportional to the increase in the caspofungin dose. In addition, the Young modulus of the cell was three times lower forC. albicanscells than forS. cerevisiaecells and increased proportionally with the increase of chitin, suggesting differences in the molecular organization of the cell wall between the two yeast species. Also, at a low dose of caspofungin (i.e., 0.5× MIC), the cell surface ofC. albicansexhibited a morphology that was reminiscent of cells expressing adhesion proteins. Interestingly, this morphology was lost at high doses of the drug (i.e., 4× MIC). However, the treatment ofS. cerevisiaecells with high doses of caspofungin resulted in impairment of cytokinesis. Altogether, the use of AFM for investigating the effects of antifungal drugs is relevant in nanomedicine, as it should help in understanding their mechanisms of action on fungal cells, as well as unraveling unexpected effects on cell division and fungal adhesion.

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

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.

scholarly journals Mitochondrial Sorting and Assembly Machinery Subunit Sam37 in Candida albicans: Insight into the Roles of Mitochondria in Fitness, Cell Wall Integrity, and Virulence

2012 ◽  
Vol 11 (4) ◽  
pp. 532-544 ◽  
Author(s):  
Yue Qu ◽  
Branka Jelicic ◽  
Filomena Pettolino ◽  
Andrew Perry ◽  
Tricia L. Lo ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Mitochondrial Function ◽  
Drug Tolerance ◽  
Cell Wall Integrity ◽  
Mitochondrial Outer Membrane ◽  
Wall Structure ◽  
Content Type ◽  
Mitochondrial Functions ◽  
Insight Into

ABSTRACT Recent studies indicate that mitochondrial functions impinge on cell wall integrity, drug tolerance, and virulence of human fungal pathogens. However, the mechanistic aspects of these processes are poorly understood. We focused on the mitochondrial outer membrane SAM ( S orting and A ssembly M achinery) complex subunit Sam37 in Candida albicans . Inactivation of SAM37 in C. albicans leads to a large reduction in fitness, a phenotype not conserved with the model yeast Saccharomyces cerevisiae . Our data indicate that slow growth of the sam37ΔΔ mutant results from mitochondrial DNA loss, a new function for Sam37 in C. albicans , and from reduced activity of the essential SAM complex subunit Sam35. The sam37ΔΔ mutant was hypersensitive to drugs that target the cell wall and displayed altered cell wall structure, supporting a role for Sam37 in cell wall integrity in C. albicans . The sensitivity of the mutant to membrane-targeting antifungals was not significantly altered. The sam37ΔΔ mutant was avirulent in the mouse model, and bioinformatics showed that the fungal Sam37 proteins are distant from their animal counterparts and could thus represent potential drug targets. Our study provides the first direct evidence for a link between mitochondrial function and cell wall integrity in C. albicans and is further relevant for understanding mitochondrial function in fitness, antifungal drug tolerance, and virulence of this major pathogen. Beyond the relevance to fungal pathogenesis, this work also provides new insight into the mitochondrial and cellular roles of the SAM complex in fungi.

scholarly journals Endoplasmic Reticulum α-Glycosidases of Candida albicans Are Required for N Glycosylation, Cell Wall Integrity, and Normal Host-Fungus Interaction

2007 ◽  
Vol 6 (12) ◽  
pp. 2184-2193 ◽  
Author(s):  
Héctor M. Mora-Montes ◽  
Steven Bates ◽  
Mihai G. Netea ◽  
Diana F. Díaz-Jiménez ◽  
Everardo López-Romero ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Wall ◽  
Candida Albicans ◽  
Systemic Infection ◽  
Cell Wall Integrity ◽  
Yeast Cells ◽  
Chain Elongation ◽  
Cell Wall Integrity Pathway ◽  
Oligosaccharide Processing ◽  
Host Fungus

ABSTRACT The cell surface of Candida albicans is enriched in highly glycosylated mannoproteins that are involved in the interaction with the host tissues. N glycosylation is a posttranslational modification that is initiated in the endoplasmic reticulum (ER), where the Glc3Man9GlcNAc2 N-glycan is processed by α-glucosidases I and II and α1,2-mannosidase to generate Man8GlcNAc2. This N-oligosaccharide is then elaborated in the Golgi to form N-glycans with highly branched outer chains rich in mannose. In Saccharomyces cerevisiae, CWH41, ROT2, and MNS1 encode for α-glucosidase I, α-glucosidase II catalytic subunit, and α1,2-mannosidase, respectively. We disrupted the C. albicans CWH41, ROT2, and MNS1 homologs to determine the importance of N-oligosaccharide processing on the N-glycan outer-chain elongation and the host-fungus interaction. Yeast cells of Cacwh41Δ, Carot2Δ, and Camns1Δ null mutants tended to aggregate, displayed reduced growth rates, had a lower content of cell wall phosphomannan and other changes in cell wall composition, underglycosylated β-N-acetylhexosaminidase, and had a constitutively activated PKC-Mkc1 cell wall integrity pathway. They were also attenuated in virulence in a murine model of systemic infection and stimulated an altered pro- and anti-inflammatory cytokine profile from human monocytes. Therefore, N-oligosaccharide processing by ER glycosidases is required for cell wall integrity and for host-fungus interactions.

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