scholarly journals Multiple Alternative Carbon Pathways Combine To Promote Candida albicans Stress Resistance, Immune Interactions, and Virulence

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Robert B. Williams ◽  
Michael C. Lorenz
Keyword(s):  
Amino Acids ◽  
Cell Wall ◽  
Candida Albicans ◽  
Stress Resistance ◽  
Innate Immune System ◽  
Carbon Sources ◽  
Innate Immune ◽  
Content Type ◽  
Multiple Alternative ◽  
Carbon Pathways

ABSTRACT The phagocytic cells of the innate immune system are an essential first line of antimicrobial defense, and yet Candida albicans, one of the most problematic fungal pathogens, is capable of resisting the stresses imposed by the macrophage phagosome, eventually resulting in the destruction of the phagocyte. C. albicans rapidly adapts to the phagosome by upregulating multiple alternative carbon utilization pathways, particularly those for amino acids, carboxylic acids, and N-acetylglucosamine (GlcNAc). Here, we report that C. albicans recognizes these carbon sources both as crucial nutrients and as independent signals in its environment. Even in the presence of glucose, each carbon source promotes increased resistance to a unique profile of stressors; lactate promotes increased resistance to osmotic and cell wall stresses, amino acids increased resistance to oxidative and nitrosative stresses, and GlcNAc increased resistance to oxidative stress and caspofungin, while all three alternative carbon sources have been shown to induce resistance to fluconazole. Moreover, we show mutants incapable of utilizing these carbon sources, in particular, strains engineered to be defective in all three pathways, are significantly attenuated in both macrophage and mouse models, with additive effects observed as multiple carbon pathways are eliminated, suggesting that C. albicans simultaneously utilizes multiple carbon sources within the macrophage phagosome and during disseminated candidiasis. Taking the data together, we propose that, in addition to providing energy to the pathogen within host environments, alternative carbon sources serve as niche-specific priming signals that allow C. albicans to recognize microenvironments within the host and to prepare for stresses associated with that niche, thus promoting host adaptation and virulence. IMPORTANCE Candida albicans is a fungal pathogen and a significant cause of morbidity and mortality, particularly in people with defects, sometimes minor ones, in innate immunity. The phagocytes of the innate immune system, particularly macrophages and neutrophils, generally restrict this organism to its normal commensal niches, but C. albicans shows a robust and multifaceted response to these cell types. Inside macrophages, a key component of this response is the activation of multiple pathways for the utilization of alternative carbon sources, particularly amino acids, carboxylic acids, and N-acetylglucosamine. These carbon sources are key sources of energy and biomass but also independently promote stress resistance, induce cell wall alterations, and affect C. albicans interactions with macrophages. Engineered strains incapable of utilizing these alternative carbon pathways are attenuated in infection models. These data suggest that C. albicans recognizes nutrient composition as an indicator of specific host environments and tailors its responses accordingly.

scholarly journals β-(1,3)-Glucan Unmasking in Some Candida albicans Mutants Correlates with Increases in Cell Wall Surface Roughness and Decreases in Cell Wall Elasticity

2016 ◽  
Vol 85 (1) ◽  
Author(s):  
Sahar Hasim ◽  
David P. Allison ◽  
Scott T. Retterer ◽  
Alex Hopke ◽  
Robert T. Wheeler ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Immune System ◽  
Innate Immune System ◽  
Fungal Infections ◽  
Innate Immune ◽  
Surface Topology ◽  
Wild Type ◽  
Content Type ◽  
Cell Wall Elasticity

ABSTRACT Candida albicans is among the most common human fungal pathogens, causing a broad range of infections, including life-threatening systemic infections. The cell wall of C. albicans is the interface between the fungus and the innate immune system. The cell wall is composed of an outer layer enriched in mannosylated glycoproteins (mannan) and an inner layer enriched in β-(1,3)-glucan and chitin. Detection of C. albicans by Dectin-1, a C-type signaling lectin specific for β-(1,3)-glucan, is important for the innate immune system to recognize systemic fungal infections. Increased exposure of β-(1,3)-glucan to the immune system occurs when the mannan layer is altered or removed in a process called unmasking. Nanoscale changes to the cell wall during unmasking were explored in live cells with atomic force microscopy (AFM). Two mutants, the cho1Δ/Δ and kre5Δ/Δ mutants, were selected as representatives that exhibit modest and strong unmasking, respectively. Comparisons of the cho1Δ/Δ and kre5Δ/Δ mutants to the wild type reveal morphological changes in their cell walls that correlate with decreases in cell wall elasticity. In addition, AFM tips functionalized with Dectin-1 revealed that the forces of binding of Dectin-1 to all of the strains were similar, but the frequency of binding was highest for the kre5Δ/Δ mutant, decreased for the cho1Δ/Δ mutant, and rare for the wild type. These data show that nanoscale changes in surface topology are correlated with increased Dectin-1 adhesion and decreased cell wall elasticity. AFM, using tips functionalized with immunologically relevant molecules, can map epitopes of the cell wall and increase our understanding of pathogen recognition by the immune system.

scholarly journals Growth of Candida albicans Cells on the Physiologically Relevant Carbon Source Lactate Affects Their Recognition and Phagocytosis by Immune Cells

2012 ◽  
Vol 81 (1) ◽  
pp. 238-248 ◽  
Author(s):  
Iuliana V. Ene ◽  
Shih-Chin Cheng ◽  
Mihai G. Netea ◽  
Alistair J. P. Brown
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Immune System ◽  
Carbon Source ◽  
Immune Responses ◽  
Immune Cells ◽  
Carbon Sources ◽  
Innate Immune ◽  
Phagocytic Cells ◽  
Content Type

Candida albicansis a normal resident of the human gastrointestinal and urogenital tracts and also a prevalent fungal pathogen. During both commensalism and infection, it must match the immunological defenses of its host while adapting to environmental cues and the local nutrient status.C. albicansregularly colonizes glucose-poor niches, thereby depending upon alternative carbon sources for growth. However, most studies of host immune responses toC. albicanshave been performed on fungal cells grown on glucose, and the extent to which alternative physiologically relevant carbon sources impact innate immune responses has not been studied. The fungal cell wall is decorated with multifarious pathogen-associated molecular patterns and is the main target for recognition by host innate immune cells. Cell wall architecture is both robust and dynamic, and it is dramatically influenced by growth conditions. We found that growth ofC. albicanscells on lactate, a nonfermentative carbon source available in numerous anatomical niches, modulates their interactions with immune cells and the resultant cytokine profile. Notably, lactate-grownC. albicansstimulated interleukin-10 (IL-10) production while decreasing IL-17 levels, rendering these cells less visible to the immune system than were glucose-grown cells. This trend was observed in clinicalC. albicansisolates from different host niches and from different epidemiological clades. In addition, lactate-grownC. albicanscells were taken up by macrophages less efficiently, but they were more efficient at killing and escaping these phagocytic cells. Our data indicate that carbon source has a major impact upon theC. albicansinteraction with the innate immune system.

scholarly journals N-Acetylglucosamine Metabolism Promotes Survival of Candida albicans in the Phagosome

mSphere ◽  
2017 ◽  
Vol 2 (5) ◽  
Author(s):  
Elisa M. Vesely ◽  
Robert B. Williams ◽  
James B. Konopka ◽  
Michael C. Lorenz
Keyword(s):  
Amino Acids ◽  
Candida Albicans ◽  
Carboxylic Acids ◽  
Immune Cells ◽  
Carbon Sources ◽  
Innate Immune ◽  
Innate Immune Cells ◽  
Content Type ◽  
Macrophage Phagocytosis ◽  
Ph Neutralization

ABSTRACT Candida albicans is the most important medically relevant fungal pathogen, with disseminated candidiasis being the fourth most common hospital-associated bloodstream infection. Macrophages and neutrophils are innate immune cells that play a key role in host defense by phagocytosing and destroying C. albicans cells. To survive this attack by macrophages, C. albicans generates energy by utilizing alternative carbon sources that are available in the phagosome. Interestingly, metabolism of amino acids and carboxylic acids by C. albicans raises the pH of the phagosome and thereby blocks the acidification of the phagosome, which is needed to initiate antimicrobial attack. In this work, we demonstrate that metabolism of a third type of carbon source, the amino sugar GlcNAc, also induces pH neutralization and survival of C. albicans upon phagocytosis. This mechanism is genetically and physiologically distinct from the previously described mechanisms of pH neutralization, indicating that the robust metabolic plasticity of C. albicans ensures survival upon macrophage phagocytosis. Phagocytosis by innate immune cells is one of the most effective barriers against the multiplication and dissemination of microbes within the mammalian host. Candida albicans, a pathogenic yeast, has robust mechanisms that allow survival upon macrophage phagocytosis. C. albicans survives in part because it can utilize the alternative carbon sources available in the phagosome, including carboxylic acids and amino acids. Furthermore, metabolism of these compounds raises the pH of the extracellular environment, which combats the acidification and maturation of the phagolysosome. In this study, we demonstrate that metabolism by C. albicans of an additional carbon source, N-acetylglucosamine (GlcNAc), facilitates neutralization of the phagosome by a novel mechanism. Catabolism of GlcNAc raised the ambient pH through release of ammonia, which is distinct from growth on carboxylic acids but similar to growth on amino acids. However, the effect of GlcNAc metabolism on pH was genetically distinct from the neutralization induced by catabolism of amino acids, as mutation of STP2 or ATO5 did not impair the effects of GlcNAc. In contrast, mutants lacking the dedicated GlcNAc transporter gene NGT1 or the enzymes responsible for catabolism of GlcNAc were defective in altering the pH of the phagosome. This correlated with reduced survival following phagocytosis and decreased ability to damage macrophages. Thus, GlcNAc metabolism represents the third genetically independent mechanism that C. albicans utilizes to combat the rapid acidification of the phagolysosome, allowing for cells to escape and propagate infection. IMPORTANCE Candida albicans is the most important medically relevant fungal pathogen, with disseminated candidiasis being the fourth most common hospital-associated bloodstream infection. Macrophages and neutrophils are innate immune cells that play a key role in host defense by phagocytosing and destroying C. albicans cells. To survive this attack by macrophages, C. albicans generates energy by utilizing alternative carbon sources that are available in the phagosome. Interestingly, metabolism of amino acids and carboxylic acids by C. albicans raises the pH of the phagosome and thereby blocks the acidification of the phagosome, which is needed to initiate antimicrobial attack. In this work, we demonstrate that metabolism of a third type of carbon source, the amino sugar GlcNAc, also induces pH neutralization and survival of C. albicans upon phagocytosis. This mechanism is genetically and physiologically distinct from the previously described mechanisms of pH neutralization, indicating that the robust metabolic plasticity of C. albicans ensures survival upon macrophage phagocytosis.

scholarly journals Masking of β(1-3)-Glucan in the Cell Wall of Candida albicans from Detection by Innate Immune Cells Depends on Phosphatidylserine

2014 ◽  
Vol 82 (10) ◽  
pp. 4405-4413 ◽  
Author(s):  
Sarah E. Davis ◽  
Alex Hopke ◽  
Steven C. Minkin ◽  
Anthony E. Montedonico ◽  
Robert T. Wheeler ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
De Novo ◽  
Invasive Disease ◽  
Innate Immune ◽  
Dependent Manner ◽  
Content Type ◽  
Immune Effector ◽  
Host Interactions ◽  
Innate Immune Effector

ABSTRACTThe virulence ofCandida albicansin a mouse model of invasive candidiasis is dependent on the phospholipids phosphatidylserine (PS) and phosphatidylethanolamine (PE). Disruption of the PS synthase geneCHO1(i.e.,cho1Δ/Δ) eliminates PS and blocks thede novopathway for PE biosynthesis. In addition, thecho1Δ/Δ mutant's ability to cause invasive disease is severely compromised. Thecho1Δ/Δ mutant also exhibits cell wall defects, and in this study, it was determined that loss of PS results in decreased masking of cell wall β(1-3)-glucan from the immune system. In wild-typeC. albicans, the outer mannan layer of the wall masks the inner layer of β(1-3)-glucan from exposure and detection by innate immune effector molecules like the C-type signaling lectin Dectin-1, which is found on macrophages, neutrophils, and dendritic cells. Thecho1Δ/Δ mutant exhibits increases in exposure of β(1-3)-glucan, which leads to greater binding by Dectin-1 in both yeast and hyphal forms. The unmasking of β(1-3)-glucan also results in increased elicitation of TNF-α from macrophages in a Dectin-1-dependent manner. The role of phospholipids in fungal pathogenesis is an emerging field, and this is the first study showing that loss of PS inC. albicansresults in decreased masking of β(1-3)-glucan, which may contribute to our understanding of fungus-host interactions.

scholarly journals Altered Dynamics of Candida albicans Phagocytosis by Macrophages and PMNs When Both Phagocyte Subsets Are Present

mBio ◽  
2013 ◽  
Vol 4 (6) ◽  
Author(s):  
Fiona M. Rudkin ◽  
Judith M. Bain ◽  
Catriona Walls ◽  
Leanne E. Lewis ◽  
Neil A. R. Gow ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Immune System ◽  
Innate Immune System ◽  
Video Microscopy ◽  
Innate Immune ◽  
Live Cell ◽  
Yeast Cells ◽  
Polymorphonuclear Cells ◽  
Fungal Cell ◽  
Content Type

ABSTRACT An important first line of defense against Candida albicans infections is the killing of fungal cells by professional phagocytes of the innate immune system, such as polymorphonuclear cells (PMNs) and macrophages. In this study, we employed live-cell video microscopy coupled with dynamic image analysis tools to provide insights into the complexity of C. albicans phagocytosis when macrophages and PMNs were incubated with C. albicans alone and when both phagocyte subsets were present. When C. albicans cells were incubated with only one phagocyte subtype, PMNs had a lower overall phagocytic capacity than macrophages, despite engulfing fungal cells at a higher rate once fungal cells were bound to the phagocyte surface. PMNs were more susceptible to C. albicans-mediated killing than macrophages, irrespective of the number of C. albicans cells ingested. In contrast, when both phagocyte subsets were studied in coculture, the two cell types phagocytosed and cleared C. albicans at equal rates and were equally susceptible to killing by the fungus. The increase in macrophage susceptibility to C. albicans-mediated killing was a consequence of macrophages taking up a higher proportion of hyphal cells under these conditions. In the presence of both PMNs and macrophages, C. albicans yeast cells were predominantly cleared by PMNs, which migrated at a greater speed toward fungal cells and engulfed bound cells more rapidly. These observations demonstrate that the phagocytosis of fungal pathogens depends on, and is modified by, the specific phagocyte subsets present at the site of infection. IMPORTANCE Extensive work investigating fungal cell phagocytosis by macrophages and PMNs of the innate immune system has been carried out. These studies have been informative but have examined this phenomenon only when one phagocyte subset is present. The current study employed live-cell video microscopy to break down C. albicans phagocytosis into its component parts and examine the effect of a single phagocyte subset, versus a mixed phagocyte population, on these individual stages. Through this approach, we identified that the rate of fungal cell engulfment and rate of phagocyte killing altered significantly when both macrophages and PMNs were incubated in coculture with C. albicans compared to the rate of either phagocyte subset incubated alone with the fungus. This research highlights the significance of studying pathogen-host cell interactions with a combination of phagocytes in order to gain a greater understanding of the interactions that occur between cells of the host immune system in response to fungal invasion.

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

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
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.

scholarly journals Phosphoric Metabolites Link Phosphate Import and Polysaccharide Biosynthesis for Candida albicans Cell Wall Maintenance

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Ning-Ning Liu ◽  
Maikel Acosta-Zaldívar ◽  
Wanjun Qi ◽  
Joann Diray-Arce ◽  
Louise A. Walker ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Stress Resistance ◽  
Wall Stress ◽  
Phosphate Starvation ◽  
Nucleotide Sugar ◽  
Content Type ◽  
Cell Wall Stress ◽  
Chitin Synthases ◽  
Nucleotide Sugars

ABSTRACT The Candida albicans high-affinity phosphate transporter Pho84 is required for normal Target of Rapamycin (TOR) signaling, oxidative stress resistance, and virulence of this fungal pathogen. It also contributes to C. albicans’ tolerance of two antifungal drug classes, polyenes and echinocandins. Echinocandins inhibit biosynthesis of a major cell wall component, beta-1,3-glucan. Cells lacking Pho84 were hypersensitive to other forms of cell wall stress beyond echinocandin exposure, while their cell wall integrity signaling response was weak. Metabolomics experiments showed that levels of phosphoric intermediates, including nucleotides like ATP and nucleotide sugars, were low in pho84 mutant compared to wild-type cells recovering from phosphate starvation. Nonphosphoric precursors like nucleobases and nucleosides were elevated. Outer cell wall phosphomannan biosynthesis requires a nucleotide sugar, GDP-mannose. The nucleotide sugar UDP-glucose is the substrate of enzymes that synthesize two major structural cell wall polysaccharides, beta-1,3- and beta-1,6-glucan. Another nucleotide sugar, UDP-N-acetylglucosamine, is the substrate of chitin synthases which produce a stabilizing component of the intercellular septum and of lateral cell walls. Lack of Pho84 activity, and phosphate starvation, potentiated pharmacological or genetic perturbation of these enzymes. We posit that low substrate concentrations of beta-d-glucan- and chitin synthases, together with pharmacologic inhibition of their activity, diminish enzymatic reaction rates as well as the yield of their cell wall-stabilizing products. Phosphate import is not conserved between fungal and human cells, and humans do not synthesize beta-d-glucans or chitin. Hence, inhibiting these processes simultaneously could yield potent antifungal effects with low toxicity to humans. IMPORTANCE Candida species cause hundreds of thousands of invasive infections with high mortality each year. Developing novel antifungal agents is challenging due to the many similarities between fungal and human cells. Maintaining phosphate balance is essential for all organisms but is achieved completely differently by fungi and humans. A protein that imports phosphate into fungal cells, Pho84, is not present in humans and is required for normal cell wall stress resistance and cell wall integrity signaling in C. albicans. Nucleotide sugars, which are phosphate-containing building block molecules for construction of the cell wall, are diminished in cells lacking Pho84. Cell wall-constructing enzymes may be slowed by lack of these building blocks, in addition to being inhibited by drugs. Combined targeting of Pho84 and cell wall-constructing enzymes may provide a strategy for antifungal therapy by which two sequential steps of cell wall maintenance are blocked for greater potency.

scholarly journals Mutations in Alternative Carbon Utilization Pathways in Candida albicans Attenuate Virulence and Confer Pleiotropic Phenotypes

2006 ◽  
Vol 6 (2) ◽  
pp. 280-290 ◽  
Author(s):  
Melissa A. Ramírez ◽  
Michael C. Lorenz
Keyword(s):  
Candida Albicans ◽  
Carbon Metabolism ◽  
Innate Immune System ◽  
Glyoxylate Cycle ◽  
Transcriptional Profiling ◽  
Carbon Sources ◽  
Innate Immune ◽  
Carbon Utilization ◽  
The Glyoxylate Cycle

ABSTRACT The interaction between Candida albicans and cells of the innate immune system is a key determinant of disease progression. Transcriptional profiling has revealed that C. albicans has a complex response to phagocytosis, much of which is similar to carbon starvation. This suggests that nutrient limitation is a significant stress in vivo, and we have shown that glyoxylate cycle mutants are less virulent in mice. To examine whether other aspects of carbon metabolism are important in vivo during an infection, we have constructed strains lacking FOX2 and FBP1, which encode key components of fatty acid β-oxidation and gluconeogenesis, respectively. As expected, fox2Δ mutants failed to utilize several fatty acids as carbon sources. Surprisingly, however, these mutants also failed to grow in the presence of several other carbon sources, whose assimilation is independent of β-oxidation, including ethanol and citric acid. Mutants lacking the glyoxylate enzyme ICL1 also had more severe carbon utilization phenotypes than were expected. These results suggest that the regulation of alternative carbon metabolism in C. albicans is significantly different from that in other fungi. In vivo, fox2Δ mutants show a moderate but significant reduction in virulence in a mouse model of disseminated candidiasis, while disruption of the glyoxylate cycle or gluconeogenesis confers a severe attenuation in this model. These data indicate that C. albicans often encounters carbon-poor conditions during growth in the host and that the ability to efficiently utilize multiple nonfermentable carbon sources is a virulence determinant. Consistent with this in vivo requirement, C. albicans uniquely regulates carbon metabolism in a more integrated manner than in Saccharomyces cerevisiae, such that defects in one part of the machinery have wider impacts than expected. These aspects of alternative carbon metabolism may then be useful as targets for therapeutic intervention.

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