scholarly journals Position of O-Acetylation within the Capsular Repeat Unit Impacts the Biological Properties of Pneumococcal Serotypes 33A and 33F

2017 ◽  
Vol 85 (7) ◽  
Author(s):  
Brady L. Spencer ◽  
Jamil S. Saad ◽  
Anukul T. Shenoy ◽  
Carlos J. Orihuela ◽  
Moon H. Nahm
Keyword(s):  
Cell Wall ◽  
Repeat Unit ◽  
Biological Properties ◽  
Immune Recognition ◽  
Content Type ◽  
Pneumococcal Serotype ◽  
Membrane Bound ◽  
Biochemical Analyses ◽  
Opsonophagocytic Killing ◽  
Small Chemical

ABSTRACT Streptococcus pneumoniae (pneumococcus) produces many capsule types that differ in their abilities to evade host immune recognition. To explain these serotype-dependent protective capacities, many studies have investigated capsular thickness or the interaction of the capsule with complement proteins, but the effects of small chemical modifications of the capsule on its function have not been studied. One small chemical modification found frequently among pneumococcal capsules is O-acetylation. Pneumococcal serotype 33A has two membrane-bound O-acetyltransferase genes, wciG and wcjE. A 33A wcjE-deficient variant, 33F, occurs naturally and is increasing in prevalence in the wake of widespread conjugate vaccine use, but no wciG-deficient variants have been reported. To study the biological consequence of the loss of O-acetylation, we created wciG-deficient variants in both serotypes 33A and 33F, which we named 33X1 (ΔwciG) and 33X2 (ΔwciG ΔwcjE). Serotypes 33X1 and 33X2 express novel capsule types based on serological and biochemical analyses. We found that loss of WcjE-mediated O-acetylation appears not to affect cell wall shielding, since serotypes 33A and 33F exhibit comparable nonspecific opsonophagocytic killing, biofilm production, and adhesion to nasopharyngeal cells, though serotype 33F survived short-term drying better than serotype 33A. Loss of WciG-mediated O-acetylation in serotypes 33X1 and 33X2, however, resulted in a phenotype resembling that of nonencapsulated strains: increased cell wall accessibility, increased nonspecific opsonophagocytic killing, enhanced biofilm formation, and increased adhesion to nasopharyngeal cells. We conclude that WciG-mediated, but not WcjE-mediated, O-acetylation is important for producing protective capsules in 33A and that small chemical changes to the capsule can drastically affect its biological properties.

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 Recognition and Blocking of Innate Immunity Cells by Candida albicans Chitin

2011 ◽  
Vol 79 (5) ◽  
pp. 1961-1970 ◽  
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.

scholarly journals Novel Biological Functions of the NsdC Transcription Factor in Aspergillus fumigatus

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Patrícia Alves de Castro ◽  
Clara Valero ◽  
Jéssica Chiaratto ◽  
Ana Cristina Colabardini ◽  
Lakhansing Pardeshi ◽  
...  
Keyword(s):  
Cell Wall ◽  
Transcription Factor ◽  
Aspergillus Fumigatus ◽  
Sexual Reproduction ◽  
Rna Polymerase Ii ◽  
Vegetative Growth ◽  
Wall Structure ◽  
Immune Recognition ◽  
Biological Functions ◽  
Content Type

ABSTRACT The fungal zinc finger transcription factor NsdC is named after, and is best known for, its essential role in sexual reproduction (never in sexual development). In previous studies with Aspergillus nidulans, it was also shown to have roles in promotion of vegetative growth and suppression of asexual conidiation. In this study, the function of the nsdC homologue in the opportunistic human pathogen A. fumigatus was investigated. NsdC was again found to be essential for sexual development, with deletion of the nsdC gene in both MAT1-1 and MAT1-2 mating partners of a cross leading to complete loss of fertility. However, a functional copy of nsdC in one mating partner was sufficient to allow sexual reproduction. Deletion of nsdC also led to decreased vegetative growth and allowed conidiation in liquid cultures, again consistent with previous findings. However, NsdC in A. fumigatus was shown to have additional biological functions including response to calcium stress, correct organization of cell wall structure, and response to the cell wall stressors. Furthermore, virulence and host immune recognition were affected. Gene expression studies involving chromatin immunoprecipitation (ChIP) of RNA polymerase II (PolII) coupled to next-generation sequencing (Seq) revealed that deletion of nsdC resulted in changes in expression of over 620 genes under basal growth conditions. This demonstrated that this transcription factor mediates the activity of a wide variety of signaling and metabolic pathways and indicates that despite the naming of the gene, the promotion of sexual reproduction is just one among multiple roles of NsdC. IMPORTANCE Aspergillus fumigatus is an opportunistic human fungal pathogen and the main causal agent of invasive aspergillosis, a life-threatening infection especially in immunocompromised patients. A. fumigatus can undergo both asexual and sexual reproductive cycles, and the regulation of both cycles involves several genes and pathways. Here, we have characterized one of these genetic determinants, the NsdC transcription factor, which was initially identified in a screen of transcription factor null mutants showing sensitivity when exposed to high concentrations of calcium. In addition to its known essential roles in sexual reproduction and control of growth rate and asexual reproduction, we have shown in the present study that A. fumigatus NsdC transcription factor has additional previously unrecognized biological functions including calcium tolerance, cell wall stress response, and correct cell wall organization and functions in virulence and host immune recognition. Our results indicate that NsdC can play novel additional biological functions not directly related to its role played during sexual and asexual processes.

scholarly journals Epitope Shaving Promotes Fungal Immune Evasion

mBio ◽  
2020 ◽  
Vol 11 (4) ◽  
Author(s):  
Delma S. Childers ◽  
Gabriela Mol Avelar ◽  
Judith M. Bain ◽  
Arnab Pradhan ◽  
Daniel E. Larcombe ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Cell Surface ◽  
Immune Evasion ◽  
Fungal Pathogen ◽  
Fungal Pathogens ◽  
Specific Cell ◽  
Immune Recognition ◽  
Content Type ◽  
Evasion Strategy

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.

scholarly journals Glucan Unmasking Identifies Regulators of Temperature-Induced Translatome Reprogramming in C. neoformans

mSphere ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Amanda L. M. Bloom ◽  
David Goich ◽  
Corey M. Knowles ◽  
John C. Panepinto
Keyword(s):  
Cell Wall ◽  
Cryptococcus Neoformans ◽  
Temperature Stress ◽  
Fungal Pathogen ◽  
Cell Walls ◽  
Mapk Pathway ◽  
Regulation Of Transcription ◽  
Immune Recognition ◽  
Content Type ◽  
Signaling Modules

ABSTRACT The cell walls of fungi are critical for cellular structure and rigidity but also serve as a major communicator to alert the cell to the changing environment. In response to stresses encountered in human hosts, pathogenic fungi remodel their cell walls. Masking the β-1,3-glucan component of the cell wall is critical to escape detection by innate immune cells. We previously demonstrated that β-1,3-glucan is unmasked in response to host temperature stress when translatome reprogramming is defective in Cryptococcus neoformans. Here, we used β-1,3-glucan unmasking as an output to identify signaling modules involved both in masking and in translatome reprogramming in response to host temperature stress. We reveal that the high-osmolarity glycerol (HOG) mitogen-activated protein kinase (MAPK) pathway is involved in translatome reprogramming and that mutants in this pathway display moderate unmasking when grown at 37°C. Additionally, we show that mutants of the cell wall integrity (CWI)/Mpk1 MAPK pathway extensively unmask β-1,3-glucan. While the CWI pathway does not impact translatome reprogramming, our data suggest that it may play a role in the posttranslational regulation of transcription factors that govern masking. IMPORTANCE Cryptococcus neoformans is a fungal pathogen that causes devastating morbidity and mortality in immunocompromised individuals. It possesses several virulence factors that aid in its evasion from the host immune system, including a large polysaccharide capsule that cloaks the antigenic cell wall. Studies investigating how the cell wall is remodeled to keep this pathogen disguised in response to stress have been limited. We previously found that host temperature stress results in translatome reprogramming that is necessary for keeping the highly antigenic β-(1, 3)-glucan component masked. Our data reveal signaling modules that trigger these responses and suggest the points of regulation at which these pathways act in achieving masking. Understanding these mechanisms may allow for therapeutic manipulation that may promote the immune recognition and clearance of this fungal pathogen.

scholarly journals Proteome Analysis Reveals the Conidial Surface Protein CcpA Essential for Virulence of the Pathogenic FungusAspergillus fumigatus

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Vera Voltersen ◽  
Matthew G. Blango ◽  
Sahra Herrmann ◽  
Franziska Schmidt ◽  
Thorsten Heinekamp ◽  
...  
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Surface Structure ◽  
Fungal Spores ◽  
Fungal Spore ◽  
Invasive Infection ◽  
Immune Recognition ◽  
Wild Type ◽  
Content Type ◽  
Conidial Surface

ABSTRACTAspergillus fumigatusis a common airborne fungal pathogen of humans and a significant source of mortality in immunocompromised individuals. Here, we provide the most extensive cell wall proteome profiling to date ofA. fumigatusresting conidia, the fungal morphotype pertinent to first contact with the host. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), we identified proteins within the conidial cell wall by hydrogen-fluoride (HF)–pyridine extraction and proteins exposed on the surface using a trypsin-shaving approach. One protein, designatedconidialcell wallproteinA(CcpA), was identified by both methods and was found to be nearly as abundant as hydrophobic rodlet layer-forming protein RodA. CcpA, an amphiphilic protein, like RodA, peaks in expression during sporulation on resting conidia. Despite high cell wall abundance, the cell surface structure of ΔccpAresting conidia appeared normal. However, trypsin shaving of ΔccpAconidia revealed novel surface-exposed proteins not detected on conidia of the wild-type strain. Interestingly, the presence of swollen ΔccpAconidia led to higher activation of neutrophils and dendritic cells than was seen with wild-type conidia and caused significantly less damage to epithelial cellsin vitro. In addition, virulence was highly attenuated when cortisone-treated, immunosuppressed mice were infected with ΔccpAconidia. CcpA-specific memory T cell responses were detectable in healthy human donors naturally exposed toA. fumigatusconidia, suggesting a role for CcpA as a structural protein impacting conidial immunogenicity rather than possessing a protein-intrinsic immunosuppressive effect. Together, these data suggest that CcpA serves as a conidial stealth protein by altering the conidial surface structure to minimize innate immune recognition.IMPORTANCEThe mammalian immune system relies on recognition of pathogen surface antigens for targeting and clearance. In the absence of immune evasion strategies, pathogen clearance is rapid. In the case ofAspergillus fumigatus, the successful fungus must avoid phagocytosis in the lung to establish invasive infection. In healthy individuals, fungal spores are cleared by immune cells; however, in immunocompromised patients, clearance mechanisms are impaired. Here, using proteome analyses, we identified CcpA as an important fungal spore protein involved in pathogenesis.A. fumigatuslacking CcpA was more susceptible to immune recognition and prompt eradication and, consequently, exhibited drastically attenuated virulence. In infection studies, CcpA was required for virulence in infected immunocompromised mice, suggesting that it could be used as a possible immunotherapeutic or diagnostic target in the future. In summary, our report adds a protein to the list of those known to be critical to the complex fungal spore surface environment and, more importantly, identifies a protein important for conidial immunogenicity during infection.

scholarly journals Identification and Characterization of a Novel Family of Selective Antifungal Compounds (CANBEFs) That Interfere with Fungal Protein Synthesis

2015 ◽  
Vol 59 (9) ◽  
pp. 5631-5640 ◽  
Author(s):  
Gabriel Mircus ◽  
Nathaniel Albert ◽  
Dafna Ben-Yaakov ◽  
Dodo Chikvashvili ◽  
Yona Shadkchan ◽  
...  
Keyword(s):  
Cell Wall ◽  
Protein Synthesis ◽  
Aspergillus Nidulans ◽  
Galleria Mellonella ◽  
Fungal Infections ◽  
Chemical Compounds ◽  
Fungal Cell ◽  
Content Type ◽  
Mammalian Cell Lines ◽  
Small Chemical

ABSTRACTInvasive mycotic infections have become more common during recent decades, posing an increasing threat to public health. However, despite the growing needs, treatments for invasive fungal infections remain unsatisfactory and are limited to a small number of antifungals. The aim of this study was to identify novel fungal cell wall inhibitors from a library of small chemical compounds using a conditional protein kinase C (PKC)-expressing strain ofAspergillus nidulanssensitive to cell wall-active agents. Eight “hit” compounds affecting cell wall integrity were identified from a screen of 35,000 small chemical compounds. Five shared a common basic molecular structure of 4-chloro-6-arylamino-7-nitro-benzofurazane (CANBEF). The most potent compound, CANBEF-24, was characterized further and was shown to inhibit the growth of pathogenicAspergillus,Candida,Fusarium, andRhizopusisolates at micromolar concentrations but not to affect the growth of mammalian cell lines. CANBEF-24 demonstrated strong synergy in combination with caspofungin, an antifungal that inhibits cell wall biosynthesis. Genetic and biochemical analyses withAspergillus nidulansandSaccharomyces cerevisiaeindicated that CANBEFs selectively inhibit fungal rRNA maturation and protein synthesis, suggesting that their effect on the cell wall is indirect. CANBEFs were nontoxic in insect (Galleria mellonella,Drosophila melanogaster) and mouse models of fungal infection. Preliminary evidence showing no therapeutic benefit in these models suggests that further cycles of optimization are needed for the development of this novel class of compounds for systemic use.

scholarly journals Methanobactin from Methylocystis sp. Strain SB2 Affects Gene Expression and Methane Monooxygenase Activity in Methylosinus trichosporium OB3b

2015 ◽  
Vol 81 (7) ◽  
pp. 2466-2473 ◽  
Author(s):  
Muhammad Farhan Ul-Haque ◽  
Bhagyalakshmi Kalidass ◽  
Alexey Vorobev ◽  
Bipin S. Baral ◽  
Alan A. DiSpirito ◽  
...  
Keyword(s):  
Gene Expression ◽  
Methane Monooxygenase ◽  
Particulate Methane Monooxygenase ◽  
Methylosinus Trichosporium Ob3b ◽  
Methylosinus Trichosporium ◽  
Monooxygenase Activity ◽  
Content Type ◽  
Soluble Methane Monooxygenase ◽  
A Subunit ◽  
Membrane Bound

ABSTRACTMethanotrophs can express a cytoplasmic (soluble) methane monooxygenase (sMMO) or membrane-bound (particulate) methane monooxygenase (pMMO). Expression of these MMOs is strongly regulated by the availability of copper. Many methanotrophs have been found to synthesize a novel compound, methanobactin (Mb), that is responsible for the uptake of copper, and methanobactin produced byMethylosinus trichosporiumOB3b plays a key role in controlling expression of MMO genes in this strain. As all known forms of methanobactin are structurally similar, it was hypothesized that methanobactin from one methanotroph may alter gene expression in another. WhenMethylosinus trichosporiumOB3b was grown in the presence of 1 μM CuCl2, expression ofmmoX, encoding a subunit of the hydroxylase component of sMMO, was very low.mmoXexpression increased, however, when methanobactin fromMethylocystissp. strain SB2 (SB2-Mb) was added, as did whole-cell sMMO activity, but there was no significant change in the amount of copper associated withM. trichosporiumOB3b. IfM. trichosporiumOB3b was grown in the absence of CuCl2, themmoXexpression level was high but decreased by several orders of magnitude if copper prebound to SB2-Mb (Cu-SB2-Mb) was added, and biomass-associated copper was increased. Exposure ofMethylosinus trichosporiumOB3b to SB2-Mb had no effect on expression ofmbnA, encoding the polypeptide precursor of methanobactin in either the presence or absence of CuCl2.mbnAexpression, however, was reduced when Cu-SB2-Mb was added in both the absence and presence of CuCl2. These data suggest that methanobactin acts as a general signaling molecule in methanotrophs and that methanobactin “piracy” may be commonplace.

scholarly journals The “Vesicular Intelligence” Strategy of Blood Cancers

Genes ◽  
2021 ◽  
Vol 12 (3) ◽  
pp. 416
Author(s):  
Dorian Forte ◽  
Martina Barone ◽  
Francesca Palandri ◽  
Lucia Catani
Keyword(s):  
Clonal Evolution ◽  
Biological Properties ◽  
Hemopoietic Stem Cell ◽  
Target Cells ◽  
Cancer Cell Growth ◽  
Cell Compartment ◽  
Blood Cancer ◽  
Non Invasive ◽  
Membrane Bound ◽  
Immune Potential

Blood cancers are a heterogeneous group of disorders including leukemia, multiple myeloma, and lymphoma. They may derive from the clonal evolution of the hemopoietic stem cell compartment or from the transformation of progenitors with immune potential. Extracellular vesicles (EVs) are membrane-bound nanovesicles which are released by cells into body fluids with a role in intercellular communication in physiology and pathology, including cancer. EV cargos are enriched in nucleic acids, proteins, and lipids, and these molecules can be delivered to target cells to influence their biological properties and modify surrounding or distant targets. In this review, we will describe the “smart strategy” on how blood cancer-derived EVs modulate tumor cell development and maintenance. Moreover, we will also depict the function of microenvironment-derived EVs in blood cancers and discuss how the interplay between tumor and microenvironment affects blood cancer cell growth and spreading, immune response, angiogenesis, thrombogenicity, and drug resistance. The potential of EVs as non-invasive biomarkers will be also discussed. Lastly, we discuss the clinical application viewpoint of EVs in blood cancers. Overall, blood cancers apply a ‘vesicular intelligence’ strategy to spread signals over their microenvironment, promoting the development and/or maintenance of the malignant clone.

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