Epitope Shaving Promotes Fungal Immune Evasion

ABSTRACT The cell wall provides a major physical interface between fungal pathogens and their mammalian host. This extracellular armor is critical for fungal cell homeostasis and survival. Fungus-specific cell wall moieties, such as β-1,3-glucan, are recognized as pathogen-associated molecular patterns (PAMPs) that activate immune-mediated clearance mechanisms. We have reported that the opportunistic human fungal pathogen Candida albicans masks β-1,3-glucan following exposure to lactate, hypoxia, or iron depletion. However, the precise mechanism(s) by which C. albicans masks β-1,3-glucan has remained obscure. Here, we identify a secreted exoglucanase, Xog1, that is induced in response to lactate or hypoxia. Xog1 functions downstream of the lactate-induced β-glucan “masking” pathway to promote β-1,3-glucan “shaving.” Inactivation of XOG1 blocks most but not all β-1,3-glucan masking in response to lactate, suggesting that other activities contribute to this phenomenon. Nevertheless, XOG1 deletion attenuates the lactate-induced reductions in phagocytosis and cytokine stimulation normally observed for wild-type cells. We also demonstrate that the pharmacological inhibition of exoglucanases undermines β-glucan shaving, enhances the immune visibility of the fungus, and attenuates its virulence. Our study establishes a new mechanism underlying environmentally induced PAMP remodeling that can be manipulated pharmacologically to influence immune recognition and infection outcomes. IMPORTANCE The immune system plays a critical role in protecting us against potentially fatal fungal infections. However, some fungal pathogens have evolved evasion strategies that reduce the efficacy of our immune defenses. Previously, we reported that the fungal pathogen Candida albicans exploits specific host-derived signals (such as lactate and hypoxia) to trigger an immune evasion strategy that involves reducing the exposure of β-glucan at its cell surface. Here, we show that this phenomenon is mediated by the induction of a major secreted exoglucanase (Xog1) by the fungus in response to these host signals. Inactivating XOG1-mediated “shaving” of cell surface-exposed β-glucan enhances immune responses against the fungus. Furthermore, inhibiting exoglucanase activity pharmacologically attenuates C. albicans virulence. In addition to revealing the mechanism underlying a key immune evasion strategy in a major fungal pathogen of humans, our work highlights the potential therapeutic value of drugs that block fungal immune evasion.

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Hypoxia Promotes Immune Evasion by Triggering β-Glucan Masking on theCandida albicansCell Surface via Mitochondrial and cAMP-Protein Kinase A Signaling

mBio ◽

10.1128/mbio.01318-18 ◽

2018 ◽

Vol 9 (6) ◽

Cited By ~ 38

Author(s):

Arnab Pradhan ◽

Gabriela M. Avelar ◽

Judith M. Bain ◽

Delma S. Childers ◽

Daniel E. Larcombe ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Protein Kinase ◽

Immune Evasion ◽

Fungal Pathogen ◽

Wall Structure ◽

Adaptation To Hypoxia ◽

Low Oxygen ◽

Content Type ◽

Kinase A

ABSTRACTOrganisms must adapt to changes in oxygen tension if they are to exploit the energetic benefits of reducing oxygen while minimizing the potentially damaging effects of oxidation. Consequently, organisms in all eukaryotic kingdoms display robust adaptation to hypoxia (low oxygen levels). This is particularly important for fungal pathogens that colonize hypoxic niches in the host. We show that adaptation to hypoxia in the major fungal pathogen of humansCandida albicansincludes changes in cell wall structure and reduced exposure, at the cell surface, of β-glucan, a key pathogen-associated molecular pattern (PAMP). This leads to reduced phagocytosis by murine bone marrow-derived macrophages and decreased production of IL-10, RANTES, and TNF-α by peripheral blood mononuclear cells, suggesting that hypoxia-induced β-glucan masking has a significant effect uponC. albicans-host interactions. We show that hypoxia-induced β-glucan masking is dependent upon both mitochondrial and cAMP-protein kinase A (PKA) signaling. The decrease in β-glucan exposure is blocked by mutations that affect mitochondrial functionality (goa1Δ andupc2Δ) or that decrease production of hydrogen peroxide in the inner membrane space (sod1Δ). Furthermore, β-glucan masking is enhanced by mutations that elevate mitochondrial reactive oxygen species (aox1Δ). The β-glucan masking defects displayed bygoa1Δ andupc2Δ cells are suppressed by exogenous dibutyryl-cAMP. Also, mutations that inactivate cAMP synthesis (cyr1Δ) or PKA (tpk1Δtpk2Δ) block the masking phenotype. Our data suggest thatC. albicansresponds to hypoxic niches by inducing β-glucan masking via a mitochondrial cAMP-PKA signaling pathway, thereby modulating local immune responses and promoting fungal colonization.IMPORTANCEAnimal, plant, and fungal cells occupy environments that impose changes in oxygen tension. Consequently, many species have evolved mechanisms that permit robust adaptation to these changes. The fungal pathogenCandida albicanscan colonize hypoxic (low oxygen) niches in its human host, such as the lower gastrointestinal tract and inflamed tissues, but to colonize its host, the fungus must also evade local immune defenses. We reveal, for the first time, a defined link between hypoxic adaptation and immune evasion inC. albicans. As this pathogen adapts to hypoxia, it undergoes changes in cell wall structure that include masking of β-glucan at its cell surface, and it becomes better able to evade phagocytosis by innate immune cells. We also define the signaling mechanisms that mediate hypoxia-induced β-glucan masking, showing that they are dependent on mitochondrial signaling and the cAMP-protein kinase pathway. Therefore, hypoxia appears to trigger immune evasion in this fungal pathogen.

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AP-2-Dependent Endocytic Recycling of the Chitin Synthase Chs3 Regulates Polarized Growth inCandida albicans

mBio ◽

10.1128/mbio.02421-18 ◽

2019 ◽

Vol 10 (2) ◽

Cited By ~ 4

Author(s):

H. C. Knafler ◽

I. I. Smaczynska-de Rooij ◽

L. A. Walker ◽

K. K. Lee ◽

N. A. R. Gow ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Cell Surface ◽

Hyphal Growth ◽

Specific Cell ◽

Polarized Growth ◽

Endocytic Recycling ◽

Content Type ◽

Wall Deposition ◽

Cell Wall Deposition

ABSTRACTThe human fungal pathogenCandida albicansis known to require endocytosis to enable its adaptation to diverse niches and to maintain its highly polarized hyphal growth phase. While studies have identified changes in transcription leading to the synthesis and secretion of new proteins to facilitate hyphal growth, effective maintenance of hyphae also requires concomitant removal or relocalization of other cell surface molecules. The key molecules which must be removed from the cell surface, and the mechanisms behind this, have, however, remained elusive. In this study, we show that the AP-2 endocytic adaptor complex is required for the internalization of the major cell wall biosynthesis enzyme Chs3. We demonstrate that this interaction is mediated by the AP-2 mu subunit (Apm4) YXXΦ binding domain. We also show that in the absence of Chs3 recycling via AP-2, cells have abnormal cell wall composition, defective polarized cell wall deposition, and morphological defects. The study also highlights key distinctions between endocytic requirements of growth at yeast buds compared to that at hyphal tips and different requirements of AP-2 in maintaining the polarity of mannosylated proteins and ergosterol at hyphal tips. Together, our findings highlight the importance of correct cell wall deposition in cell shape maintenance and polarized growth and the key regulatory role of endocytic recycling via the AP-2 complex.IMPORTANCECandida albicansis a human commensal yeast that can cause significant morbidity and mortality in immunocompromised individuals. Within humans,C. albicanscan adopt different morphologies as yeast or filamentous hyphae and can occupy different niches with distinct temperatures, pHs, CO2levels, and nutrient availability. Both morphological switching and growth in different environments require cell surface remodelling, which involves both the addition of newly synthesized proteins as well as the removal of other proteins. In our study, we demonstrate the importance of an adaptor complex AP-2 in internalizing and recycling a specific cell surface enzyme to maintain effective polarized hyphal growth. Defects in formation of the complex or in its ability to interact directly with cargo inhibit enzyme uptake and lead to defective cell walls and aberrant hyphal morphology. Our data indicate that the AP-2 adaptor plays a central role in regulating cell surface composition inCandida.

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Melanin Externalization in Candida albicans Depends on Cell Wall Chitin Structures

Eukaryotic Cell ◽

10.1128/ec.00051-10 ◽

2010 ◽

Vol 9 (9) ◽

pp. 1329-1342 ◽

Cited By ~ 55

Author(s):

Claire A. Walker ◽

Beatriz L. Gómez ◽

Héctor M. Mora-Montes ◽

Kevin S. Mackenzie ◽

Carol A. Munro ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Fungal Pathogen ◽

Cell Walls ◽

Chitin Synthesis ◽

Melanin Production ◽

Content Type ◽

Encoding Gene ◽

Chitinase Genes

ABSTRACT The fungal pathogen Candida albicans produces dark-pigmented melanin after 3 to 4 days of incubation in medium containing l-3,4-dihydroxyphenylalanine (l-DOPA) as a substrate. Expression profiling of C. albicans revealed very few genes significantly up- or downregulated by growth in l-DOPA. We were unable to determine a possible role for melanin in the virulence of C. albicans. However, we showed that melanin was externalized from the fungal cells in the form of electron-dense melanosomes that were free or often loosely bound to the cell wall exterior. Melanin production was boosted by the addition of N-acetylglucosamine to the medium, indicating a possible association between melanin production and chitin synthesis. Melanin externalization was blocked in a mutant specifically disrupted in the chitin synthase-encoding gene CHS2. Melanosomes remained within the outermost cell wall layers in chs3Δ and chs2Δ chs3Δ mutants but were fully externalized in chs8Δ and chs2Δ chs8Δ mutants. All the CHS mutants synthesized dark pigment at equivalent rates from mixed membrane fractions in vitro, suggesting it was the form of chitin structure produced by the enzymes, not the enzymes themselves, that was involved in the melanin externalization process. Mutants with single and double disruptions of the chitinase genes CHT2 and CHT3 and the chitin pathway regulator ECM33 also showed impaired melanin externalization. We hypothesize that the chitin product of Chs3 forms a scaffold essential for normal externalization of melanosomes, while the Chs8 chitin product, probably produced in cell walls in greater quantity in the absence of CHS2, impedes externalization.

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Remasking of Candida albicans β-Glucan in Response to Environmental pH Is Regulated by Quorum Sensing

mBio ◽

10.1128/mbio.02347-19 ◽

2019 ◽

Vol 10 (5) ◽

Cited By ~ 7

Author(s):

Fabien Cottier ◽

Sarah Sherrington ◽

Sarah Cockerill ◽

Valentina del Olmo Toledo ◽

Stephen Kissane ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Quorum Sensing ◽

Immune Responses ◽

Female Reproductive Tract ◽

Innate Immune ◽

Immune Recognition ◽

Innate Immune Responses ◽

Content Type ◽

Cell Wall Remodeling

ABSTRACT Candida albicans is a commensal yeast of the human gut which is tolerated by the immune system but has the potential to become an opportunistic pathogen. One way in which C. albicans achieves this duality is through concealing or exposing cell wall pathogen-associated molecular patterns (PAMPs) in response to host-derived environment cues (pH, hypoxia, and lactate). This cell wall remodeling allows C. albicans to evade or hyperactivate the host’s innate immune responses, leading to disease. Previously, we showed that adaptation of C. albicans to acidic environments, conditions encountered during colonization of the female reproductive tract, induces significant cell wall remodeling resulting in the exposure of two key fungal PAMPs (β-glucan and chitin). Here, we report that this pH-dependent cell wall remodeling is time dependent, with the initial change in pH driving cell wall unmasking, which is then remasked at later time points. Remasking of β-glucan was mediated via the cell density-dependent fungal quorum sensing molecule farnesol, while chitin remasking was mediated via a small, heat-stable, nonproteinaceous secreted molecule(s). Transcript profiling identified a core set of 42 genes significantly regulated by pH over time and identified the transcription factor Efg1 as a regulator of chitin exposure through regulation of CHT2. This dynamic cell wall remodeling influenced innate immune recognition of C. albicans, suggesting that during infection, C. albicans can manipulate the host innate immune responses. IMPORTANCE Candida albicans is part of the microbiota of the skin and gastrointestinal and reproductive tracts of humans and has coevolved with us for millennia. During that period, C. albicans has developed strategies to modulate the host’s innate immune responses, by regulating the exposure of key epitopes on the fungal cell surface. Here, we report that exposing C. albicans to an acidic environment, similar to the one of the stomach or vagina, increases the detection of the yeast by macrophages. However, this effect is transitory, as C. albicans is able to remask these epitopes (glucan and chitin). We found that glucan remasking is controlled by the production of farnesol, a molecule secreted by C. albicans in response to high cell densities. However, chitin-remasking mechanisms remain to be identified. By understanding the relationship between environmental sensing and modulation of the host-pathogen interaction, new opportunities for the development of innovative antifungal strategies are possible.

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Staurosporine Induces Filamentation in the Human Fungal Pathogen Candida albicans via Signaling through Cyr1 and Protein Kinase A

mSphere ◽

10.1128/msphere.00056-17 ◽

2017 ◽

Vol 2 (2) ◽

Cited By ~ 12

Author(s):

Jinglin L. Xie ◽

Teresa R. O’Meara ◽

Elizabeth J. Polvi ◽

Nicole Robbins ◽

Leah E. Cowen

Keyword(s):

Candida Albicans ◽

Small Molecules ◽

Fungal Pathogen ◽

Fungal Pathogens ◽

Bloodstream Infections ◽

Filamentous Growth ◽

Genetic Pathways ◽

Content Type ◽

Virulence Trait ◽

The Impact

ABSTRACT The impact of fungal pathogens on human health is devastating. One of the most pervasive fungal pathogens is Candida albicans, which kills ~40% of people suffering from bloodstream infections. Treatment of these infections is extremely difficult, as fungi are closely related to humans, and there are limited drugs that kill the fungus without host toxicity. The capacity of C. albicans to transition between yeast and filamentous forms is a key virulence trait. Thus, understanding the genetic pathways that regulate morphogenesis could provide novel therapeutic targets to treat C. albicans infections. Here, we establish the small molecule staurosporine as an inducer of filamentous growth. We unveil distinct regulatory circuitry required for staurosporine-induced filamentation that appears to be unique to this filament-inducing cue. Thus, this work highlights the fact that small molecules, such as staurosporine, can improve our understanding of the pathways required for key virulence programs, which may lead to the development of novel therapeutics. Protein kinases are key regulators of signal transduction pathways that participate in diverse cellular processes. In fungal pathogens, kinases regulate signaling pathways that govern drug resistance, stress adaptation, and pathogenesis. The impact of kinases on the fungal regulatory circuitry has recently garnered considerable attention in the opportunistic fungal pathogen Candida albicans, which is a leading cause of human morbidity and mortality. Complex regulatory circuitry governs the C. albicans morphogenetic transition between yeast and filamentous growth, which is a key virulence trait. Here, we report that staurosporine, a promiscuous kinase inhibitor that abrogates fungal drug resistance, also influences C. albicans morphogenesis by inducing filamentation in the absence of any other inducing cue. We further establish that staurosporine exerts its effect via the adenylyl cyclase Cyr1 and the cyclic AMP (cAMP)-dependent protein kinase A (PKA). Strikingly, filamentation induced by staurosporine does not require the known upstream regulators of Cyr1, Ras1 or Pkc1, or effectors downstream of PKA, including Efg1. We further demonstrate that Cyr1 is capable of activating PKA to enable filamentation in response to staurosporine through a mechanism that does not require degradation of the transcriptional repressor Nrg1. We establish that staurosporine-induced filamentation is accompanied by a defect in septin ring formation, implicating cell cycle kinases as potential staurosporine targets underpinning this cellular response. Thus, we establish staurosporine as a chemical probe to elucidate the architecture of cellular signaling governing fungal morphogenesis and highlight the existence of novel circuitry through which the Cyr1 and PKA govern a key virulence trait. IMPORTANCE The impact of fungal pathogens on human health is devastating. One of the most pervasive fungal pathogens is Candida albicans, which kills ~40% of people suffering from bloodstream infections. Treatment of these infections is extremely difficult, as fungi are closely related to humans, and there are limited drugs that kill the fungus without host toxicity. The capacity of C. albicans to transition between yeast and filamentous forms is a key virulence trait. Thus, understanding the genetic pathways that regulate morphogenesis could provide novel therapeutic targets to treat C. albicans infections. Here, we establish the small molecule staurosporine as an inducer of filamentous growth. We unveil distinct regulatory circuitry required for staurosporine-induced filamentation that appears to be unique to this filament-inducing cue. Thus, this work highlights the fact that small molecules, such as staurosporine, can improve our understanding of the pathways required for key virulence programs, which may lead to the development of novel therapeutics.

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Candida albicans Quorum Sensing Molecules Stimulate Mouse Macrophage Migration

Infection and Immunity ◽

10.1128/iai.00886-15 ◽

2015 ◽

Vol 83 (10) ◽

pp. 3857-3864 ◽

Cited By ~ 17

Author(s):

Jessica C. Hargarten ◽

Tyler C. Moore ◽

Thomas M. Petro ◽

Kenneth W. Nickerson ◽

Audrey L. Atkin

Keyword(s):

Candida Albicans ◽

Fungal Pathogens ◽

Physiological Mechanism ◽

Fold Increase ◽

Immune Recognition ◽

Macrophage Migration ◽

Content Type ◽

Transplant Patients

The polymorphic commensal fungusCandida albicanscauses life-threatening disease via bloodstream and intra-abdominal infections in immunocompromised and transplant patients. Although host immune evasion is a common strategy used by successful human fungal pathogens,C. albicansprovokes recognition by host immune cells less capable of destroying it. To accomplish this,C. albicanswhite cells secrete a low-molecular-weight chemoattractive stimulant(s) of macrophages, a phagocyte that they are able to survive within and eventually escape from.C. albicansopaque cells do not secrete this chemoattractive stimulant(s). We report here a physiological mechanism that contributes to the differences in the interaction ofC. albicanswhite and opaque cells with macrophages.E,E-Farnesol, which is secreted by white cells only, is a potent stimulator of macrophage chemokinesis, whose activity is enhanced by yeast cell wall components and aromatic alcohols.E,E-farnesol results in up to an 8.5-fold increase in macrophage migrationin vitroand promotes a 3-fold increase in the peritoneal infiltration of macrophagesin vivo. Therefore, modulation of farnesol secretion to stimulate host immune recognition by macrophages may help explain why this commensal is such a successful pathogen.

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A CRISPR Interference Platform for Efficient Genetic Repression in Candida albicans

mSphere ◽

10.1128/msphere.00002-19 ◽

2019 ◽

Vol 4 (1) ◽

Cited By ~ 16

Author(s):

Lauren Wensing ◽

Jehoshua Sharma ◽

Deeva Uthayakumar ◽

Yannic Proteau ◽

Alejandro Chavez ◽

...

Keyword(s):

Candida Albicans ◽

Human Disease ◽

Transcriptional Repression ◽

Fungal Pathogen ◽

Fungal Pathogens ◽

Fungal Infections ◽

Content Type ◽

Guide Rna ◽

Crispr Interference ◽

Functional Genomic Analysis

ABSTRACT Fungal pathogens are emerging as an important cause of human disease, and Candida albicans is among the most common causative agents of fungal infections. Studying this fungal pathogen is of the utmost importance and necessitates the development of molecular technologies to perform comprehensive genetic and functional genomic analysis. Here, we designed and developed a novel clustered regularly interspaced short palindromic repeat interference (CRISPRi) system for targeted genetic repression in C. albicans. We engineered a nuclease-dead Cas9 (dCas9) construct that, paired with a guide RNA targeted to the promoter of an endogenous gene, is capable of targeting that gene for transcriptional repression. We further optimized a favorable promoter locus to achieve repression and demonstrated that fusion of dCas9 to an Mxi1 repressor domain was able to further enhance transcriptional repression. Finally, we demonstrated the application of this CRISPRi system through genetic repression of the essential molecular chaperone HSP90. This is the first demonstration of a functional CRISPRi repression system in C. albicans, and this valuable technology will enable many future applications in this critical fungal pathogen. IMPORTANCE Fungal pathogens are an increasingly important cause of human disease and mortality, and Candida albicans is among the most common causes of fungal disease. Studying this important fungal pathogen requires a comprehensive genetic toolkit to establish how different genetic factors play roles in the biology and virulence of this pathogen. Here, we developed a CRISPR-based genetic regulation platform to achieve targeted repression of C. albicans genes. This CRISPR interference (CRISPRi) technology exploits a nuclease-dead Cas9 protein (dCas9) fused to transcriptional repressors. The dCas9 fusion proteins pair with a guide RNA to target genetic promoter regions and to repress expression from these genes. We demonstrated the functionality of this system for repression in C. albicans and show that we can apply this technology to repress essential genes. Taking the results together, this work presents a new technology for efficient genetic repression in C. albicans, with important applications for genetic analysis in this fungal pathogen.

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Recognition and Blocking of Innate Immunity Cells by Candida albicans Chitin

Infection and Immunity ◽

10.1128/iai.01282-10 ◽

2011 ◽

Vol 79 (5) ◽

pp. 1961-1970 ◽

Cited By ~ 111

Author(s):

Héctor M. Mora-Montes ◽

Mihai G. Netea ◽

Gerben Ferwerda ◽

Megan D. Lenardon ◽

Gordon D. Brown ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Cell Walls ◽

Mononuclear Cells ◽

Cytokine Production ◽

Immune Cell ◽

Human Peripheral Blood ◽

Immune Recognition ◽

Cell Wall Polysaccharide ◽

Content Type

ABSTRACTChitin is a skeletal cell wall polysaccharide of the inner cell wall of fungal pathogens. As yet, little about its role during fungus-host immune cell interactions is known. We show here that ultrapurified chitin fromCandida albicanscell walls did not stimulate cytokine production directly but blocked the recognition ofC. albicansby human peripheral blood mononuclear cells (PBMCs) and murine macrophages, leading to significant reductions in cytokine production. Chitin did not affect the induction of cytokines stimulated by bacterial cells or lipopolysaccharide (LPS), indicating that blocking was not due to steric masking of specific receptors. Toll-like receptor 2 (TLR2), TLR4, and Mincle (the macrophage-inducible C-type lectin) were not required for interactions with chitin. Dectin-1 was required for immune blocking but did not bind chitin directly. Cytokine stimulation was significantly reduced upon stimulation of PBMCs with heat-killed chitin-deficientC. albicanscells but not with live cells. Therefore, chitin is normally not exposed to cells of the innate immune system but is capable of influencing immune recognition by blocking dectin-1-mediated engagement with fungal cell walls.

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Ploidy determines the consequences of antifungal-induced mutagenesis in Candida albicans, a human fungal pathogen

10.1101/686881 ◽

2019 ◽

Cited By ~ 1

Author(s):

Ognenka Avramovska ◽

Meleah A. Hickman

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Environmental Stress ◽

Genome Size ◽

Fungal Pathogen ◽

Fungal Pathogens ◽

Antifungal Drugs ◽

Relevant Model ◽

Human Fungal Pathogen ◽

Induced Mutagenesis

AbstractOrganismal ploidy state and environmental stress impact the mutational spectrum and the mutational rate. The human fungal pathogen Candida albicans, serves as a clinically relevant model for studying the interaction between eukaryotic ploidy and stress-induced mutagenesis. In this study, we compared the rates and types of genome perturbations in diploid and tetraploid C. albicans following exposure to two classes of antifungal drugs, azoles and echinocandins. We measured mutations at three different scales: point mutation, loss-of-heterozygosity (LOH), and genome size changes in cells treated with fluconazole and caspofungin. We find that caspofungin induced higher rates of mutation than fluconazole, likely an indirect result from the stress associated with cell wall perturbations rather than an inherent genotoxicity. Furthermore, we found disproportionately elevated rates of LOH and genome size changes in response to both antifungals in tetraploid C. albicans compared to diploid C. albicans, suggesting that the magnitude of stress-induced mutagenesis results from an interaction between ploidy state and the environment. These results have both clinical and evolutionary implications for how fungal pathogens generate mutations in response to antifungal drug stress, and may facilitate the emergence of antifungal resistance.

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Candida albicans Hypha Formation and Mannan Masking of β-Glucan Inhibit Macrophage Phagosome Maturation

mBio ◽

10.1128/mbio.01874-14 ◽

2014 ◽

Vol 5 (6) ◽

Cited By ~ 70

Author(s):

Judith M. Bain ◽

Johanna Louw ◽

Leanne E. Lewis ◽

Blessing Okai ◽

Catriona A. Walls ◽

...

Keyword(s):

Cell Wall ◽

Candida Albicans ◽

Fungal Pathogen ◽

Fluorescent Protein ◽

Cell Wall Composition ◽

Live Cell ◽

Actin Dynamics ◽

Phagosome Maturation ◽

Content Type ◽

Life Threatening

ABSTRACTCandida albicansis a major life-threatening human fungal pathogen in the immunocompromised host. Host defense against systemicCandidainfection relies heavily on the capacity of professional phagocytes of the innate immune system to ingest and destroy fungal cells. A number of pathogens, includingC. albicans, have evolved mechanisms that attenuate the efficiency of phagosome-mediated inactivation, promoting their survival and replication within the host. Here we visualize host-pathogen interactions using live-cell imaging and show that viable, but not heat- or UV-killedC. albicanscells profoundly delay phagosome maturation in macrophage cell lines and primary macrophages. The ability ofC. albicansto delay phagosome maturation is dependent on cell wall composition and fungal morphology. Loss of cell wallO-mannan is associated with enhanced acquisition of phagosome maturation markers, distinct changes in Rab GTPase acquisition by the maturing phagosome, impaired hyphal growth within macrophage phagosomes, profound changes in macrophage actin dynamics, and ultimately a reduced ability of fungal cells to escape from macrophage phagosomes. The loss of cell wallO-mannan leads to exposure of β-glucan in the inner cell wall, facilitating recognition by Dectin-1, which is associated with enhanced phagosome maturation.IMPORTANCEInnate cells engulf and destroy invading organisms by phagocytosis, which is essential for the elimination of fungal cells to protect against systemic life-threatening infections. Yet comparatively little is known about what controls the maturation of phagosomes following ingestion of fungal cells. We used live-cell microscopy and fluorescent protein reporter macrophages to understand howC. albicansviability, filamentous growth, and cell wall composition affect phagosome maturation and the survival of the pathogen within host macrophages. We have demonstrated that cell wall glycosylation and yeast-hypha morphogenesis are required for disruption of host processes that function to inactivate pathogens, leading to survival and escape of this fungal pathogen from within host phagocytes. The methods employed here are applicable to study interactions of other pathogens with phagocytic cells to dissect how specific microbial features impact different stages of phagosome maturation and the survival of the pathogen or host.

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