The Experts below are selected from a list of 1197 Experts worldwide ranked by ideXlab platform
Arturo Casadevall - One of the best experts on this subject based on the ideXlab platform.
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A synthetic glycan array containing: Cryptococcus neoformans Glucuronoxylomannan capsular polysaccharide fragments allows the mapping of protective epitopes
Chemical Science, 2020Co-Authors: Lorenzo Guazzelli, Arturo Casadevall, Conor J. Crawford, Rebecca Ulc, Anthony Bowen, Orla Mccabe, Anne J. Jedlicka, Maggie P. Wear, Stefan OscarsonAbstract:A convergent synthetic strategy to Cryptococcus neoformans Glucuronoxylomannan (GXM) capsular polysaccharide part structures was developed based on di-, tri-, tetra-, penta- and hexasaccharide thioglycoside building blocks. The approach permitted the synthesis of a library of spacer-containing serotype A and D related GXM oligosaccharide structures, ranging from di- to octadecasaccharides. Ten deprotected GXM compounds (mono- to decasaccharide) were printed onto microarray plates and screened with seventeen mouse monoclonal antibodies (mAbs) to GXM. For the first time a GXM oligosaccharide structure (a serotype A decasaccharide), capable of being recognized by neutralizing forms of these GXM-specific mAbs, has been identified, offering insight into the binding epitopes of a range of protective monoclonal antibodies and furthering our efforts to develop semi-synthetic conjugate vaccine candidates against C. neoformans.
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Differentiation of Naturally Produced Extracellular Membrane Vesicles from Lipid Aggregation by Glucuronoxylomannan Immunogold Transmission Electron Microscopy in Bacillus subtilis.
Bio-protocol, 2015Co-Authors: Lisa M. Brown, Geoff Perumal, Arturo CasadevallAbstract:Recently, membrane vesicle (MV) production was described in Gram-positive bacteria, which harbor a variety of components such as toxins, antibiotic resistance proteins, proteases, DNA, and immune modulators. Free lipids have the ability to form micelles, thus it is important to rule out spontaneous association of lipids into vesicle-like structures and rather, that MVs are produced naturally by a metabolically active cell. Here, we describe a protocol utilizing the polysaccharide, Glucuronoxylomannan (GXM) from Cryptococcus neoformans (C. neoformans) as a marker to differentiate naturally produced MVs from vesicles that form spontaneously in the Gram-positive model organism, Bacillus subtilis (B. subtilis). MVs are purified from bacterial cultures grown in the presence of GXM; MVs naturally produced by cells would not contain GXM in the lumen whereas vesicular structures forming in the media could encapsulate GXM and this can be visualized via immunogold transmission electron microscopy.
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Cryptococcus neoformans Glucuronoxylomannan fractions of different molecular masses are functionally distinct
Future microbiology, 2014Co-Authors: Priscila C. Albuquerque, Arturo Casadevall, Susana Frases, Fernanda L. Fonseca, Marcelo T. Bozza, Fabianno F. Dutra, Marcio L. RodriguesAbstract:ABSTRACT: Aims: Glucuronoxylomannan (GXM) is the major polysaccharide component of Cryptococcus neoformans. We evaluated in this study whether GXM fractions of different molecular masses were functionally distinct. Materials & methods: GXM samples isolated from C. neoformans cultures were fractionated to generate polysaccharide preparations differing in molecular mass. These fractions were used in experiments focused on the association of GXM with cell wall components of C. neoformans, as well as on the interaction of the polysaccharide with host cells. Results & conclusion: GXM fractions of variable molecular masses bound to the surface of a C. neoformans acapsular mutant in a punctate pattern that is in contrast to the usual annular pattern of surface coating observed when GXM samples containing the full molecular mass range were used. The polysaccharide samples were also significantly different in their ability to stimulate cytokine production by host cells. Our findings indicate that GXM fractions ar...
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The Journal of Immunology Mouse Genetic Background Is a Major Determinant of Isotype-Related Differences for Antibody-Mediated Protective Efficacy against Cryptococcus neoformans 1,2
2013Co-Authors: Johanna Rivera, Arturo CasadevallAbstract:The protective efficacy of mAbs to Cryptococcus neoformans Glucuronoxylomannan depends on Ab isotype. Previous studies in A/JCr and C57BL/6J mice showed relative protective efficacy of IgG1, IgG2a � � IgG3. However, we now report that in C57BL/ 6J � 129/Sv mice, IgG3 is protective while IgG1 is not protective, with neither isotype being protective in 129/Sv mice. IgG1, IgG2a, and IgG3 had different effects on IFN- � expression in infected C57BL/6J � 129/Sv mice. IgG1-treated C57BL/6J � 129/Sv mice had significantly more pulmonary eosinophilia than IgG2a- and IgG3-treated C57BL/6J � 129/Sv mice. C. neoforman
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A Paracoccidioides brasiliensis glycan shares serologic and functional properties with cryptococcal Glucuronoxylomannan
Fungal genetics and biology : FG & B, 2012Co-Authors: Priscila C. Albuquerque, Kildare Miranda, Arturo Casadevall, Fernanda L. Fonseca, Radames J. B. Cordero, Roberta Peres Da Silva, Caroline L. Ramos, Rosana Puccia, Joshua D. Nosanchuk, Leonardo NimrichterAbstract:The cell wall of the yeast form of the dimorphic fungus Paracoccidioides brasiliensis is enriched with α1,3-glucans. In Cryptococcus neoformans, α1,3-glucans interact with Glucuronoxylomannan (GXM), a heteropolysaccharide that is essential for fungal virulence. In this study, we investigated the occurrence of P. brasiliensis glycans sharing properties with cryptococcal GXM. Protein database searches in P. brasiliensis revealed the presence of sequences homologous to those coding for enzymes involved in the synthesis of GXM and capsular architecture in C. neoformans. In addition, monoclonal antibodies (mAbs) raised to cryptococcal GXM bound to P. brasiliensis cells. Using protocols that were previously established for extraction and analysis of C. neoformans GXM, we recovered a P. brasiliensis glycan fraction composed of mannose and galactose, in addition to small amounts of glucose, xylose and rhamnose. In comparison with the C. neoformans GXM, the P. brasiliensis glycan fraction components had smaller molecular dimensions. The P. brasiliensis components, nevertheless, reacted with different GXM-binding mAbs. Extracellular vesicle fractions of P. brasiliensis also reacted with a GXM-binding mAb, suggesting that the polysaccharide-like molecule is exported to the extracellular space in secretory vesicles. An acapsular mutant of C. neoformans incorporated molecules from the P. brasiliensis extract onto the cell wall, resulting in the formation of surface networks that resembled the cryptococcal capsule. Coating the C. neoformans acapsular mutant with the P. brasiliensis glycan fraction resulted in protection against phagocytosis by murine macrophages. These results suggest that P. brasiliensis and C. neoformans share metabolic pathways required for the synthesis of similar polysaccharides and that P. brasiliensis yeast cell walls have molecules that mimic certain aspects of C. neoformans GXM. These findings are important because they provide additional evidence for the sharing of antigenically similar components across phylogenetically distant fungal species. Since GXM has been shown to be important for the pathogenesis of C. neoformans and to elicit protective antibodies, the finding of similar molecules in P. brasiliensis raises the possibility that these glycans play similar functions in paracoccidiomycosis.
Marcio L. Rodrigues - One of the best experts on this subject based on the ideXlab platform.
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A Glucuronoxylomannan-like glycan produced by Trichosporon mucoides
Fungal genetics and biology : FG & B, 2018Co-Authors: Ana Claudia G. Zimbres, Leonardo Nimrichter, Fernanda L. Fonseca, Priscila C. Albuquerque, Luna S. Joffe, Taiane N. Souza, Stefânia De Oliveira Frazão, Patrícia Albuquerque, Marcio L. RodriguesAbstract:Abstract Trichosporon asahii shares with Cryptococcus species the ability to produce Glucuronoxylomannan (GXM), an immunomodulatory fungal polysaccharide. The ability of other opportunistic species of Trichosporon to produce GXM-like polysaccharides is unknown. In this study, we observed that T. mucoides was less pathogenic than T. asahii in an infection model of Galleria mellonella and asked whether this difference was related to the characteristics of GXM-like molecules. Compositional analysis of samples obtained from both pathogens indicated that the components of GXM (mannose, xylose and glucuronic acid) were, in fact, detected in T. mucoides and T. asahii glycans. The identification of the T. mucoides glycan as a GXM-like molecule was confirmed by its reactivity with a monoclonal antibody raised to cryptococcal GXM and incorporation of the glycan into the cell surface of an acapsular mutant of C. neoformans. T. mucoides and T. asahii glycans differed in molecular dimensions. The antibody to cryptococcal GXM recognized T. mucoides yeast forms less efficiently than T. asahii cells. Experiments with animal cells revealed that the T. mucoides glycan manifested antiphagocytic properties. Comparative phagocytosis assays revealed that T. mucoides and T. asahii were similarly recognized by macrophages. However, fungal association with the phagocytes did not depend on the typical receptors of cryptococcal GXM, as concluded from assays using macrophages obtained from Tlr2−/− and Cd14−/− knockout mice. These results add T. mucoides to the list of fungal pathogens producing GXM-like glycans, but also indicate a high functional diversity of this major fungal immunogen.
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Cryptococcus neoformans Glucuronoxylomannan fractions of different molecular masses are functionally distinct
Future microbiology, 2014Co-Authors: Priscila C. Albuquerque, Arturo Casadevall, Susana Frases, Fernanda L. Fonseca, Marcelo T. Bozza, Fabianno F. Dutra, Marcio L. RodriguesAbstract:ABSTRACT: Aims: Glucuronoxylomannan (GXM) is the major polysaccharide component of Cryptococcus neoformans. We evaluated in this study whether GXM fractions of different molecular masses were functionally distinct. Materials & methods: GXM samples isolated from C. neoformans cultures were fractionated to generate polysaccharide preparations differing in molecular mass. These fractions were used in experiments focused on the association of GXM with cell wall components of C. neoformans, as well as on the interaction of the polysaccharide with host cells. Results & conclusion: GXM fractions of variable molecular masses bound to the surface of a C. neoformans acapsular mutant in a punctate pattern that is in contrast to the usual annular pattern of surface coating observed when GXM samples containing the full molecular mass range were used. The polysaccharide samples were also significantly different in their ability to stimulate cytokine production by host cells. Our findings indicate that GXM fractions ar...
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Glucuronoxylomannan from Cryptococcus neoformans Down-regulates the Enzyme 6-Phosphofructo-1-kinase of Macrophages
The Journal of biological chemistry, 2011Co-Authors: Juliana Grechi, Leonardo Nimrichter, Marcio L. Rodrigues, Monica M. Marinho-carvalho, Patricia Zancan, Leonardo P. Cinelli, Andre M. O. Gomes, Mauro Sola-pennaAbstract:The encapsulated yeast Cryptococcus neoformans is the causative agent of cryptococosis, an opportunistic life-threatening infection. C. neoformans is coated by a polysaccharide capsule mainly composed of Glucuronoxylomannan (GXM). GXM is considered a key virulence factor of this pathogen. The present work aimed at evaluating the effects of GXM on the key glycolytic enzyme, 6-phosphofructo-1-kinase (PFK). GXM inhibited PFK activity in cultured murine macrophages in both dose- and time-dependent manners, which occurred in parallel to cell viability decrease. The polysaccharide also inhibited purified PFK, promoting a decrease on the enzyme affinity for its substrates. In macrophages GXM and PFK partially co-localized, suggesting that internalized polysaccharide directly may interact with this enzyme. The mechanism of PFK inhibition involved dissociation of tetramers into weakly active dimers, as revealed by fluorescence spectroscopy. Allosteric modulators of the enzyme able to stabilize its tetrameric conformation attenuated the inhibition promoted by GXM. Altogether, our results suggest that the mechanism of GXM-induced cell death involves the inhibition of the glycolytic flux.
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Glucuronoxylomannan fromCryptococcus neoformans Down-regulates the Enzyme 6-Phosphofructo-1-kinase of
2011Co-Authors: Juliana Grechi, Leonardo Nimrichter, Marcio L. Rodrigues, Monica M. Marinho-carvalho, Patricia Zancan, Leonardo P. Cinelli, Andre M. O. Gomes, Mauro Sola-pennaAbstract:is coated by a polysaccharide capsulemainly composed of Glucuronoxylomannan (GXM). GXM isconsidered a key virulence factor of this pathogen. The presentwork aimed at evaluating the effects of GXM on the key glyco-lytic enzyme, 6-phosphofructo-1-kinase (PFK). GXM inhibitedPFKactivityinculturedmurinemacrophagesinbothdose-andtime-dependentmanners,whichoccurredinparalleltocellvia-bilitydecrease.ThepolysaccharidealsoinhibitedpurifiedPFK,promoting a decrease on the enzyme affinity for its substrates.In macrophages GXM and PFK partially co-localized, suggest-ing that internalized polysaccharide directly may interact withthis enzyme. The mechanism of PFK inhibition involved disso-ciation of tetramers into weakly active dimers, as revealed byfluorescencespectroscopy.Allostericmodulatorsoftheenzymeable to stabilize its tetrameric conformation attenuated theinhibition promoted by GXM. Altogether, our results suggestthat the mechanism of GXM-induced cell death involves theinhibition of the glycolytic flux.
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Immunomodulatory Effects of Serotype B Glucuronoxylomannan from Cryptococcus gattii Correlate with Polysaccharide Diameter
Infection and immunity, 2010Co-Authors: Fernanda L. Fonseca, Leonardo Nimrichter, Arturo Casadevall, Susana Frases, Radames J. B. Cordero, Lilian L. Nohara, Igor C. Almeida, Marcio L. RodriguesAbstract:Glucuronoxylomannan (GXM), the major capsular component in the Cryptococcus complex, interacts with the immune system in multiple ways, which include the activation of Toll-like receptors (TLRs) and the modulation of nitric oxide (NO) production by phagocytes. In this study, we analyzed several structural parameters of GXM samples from Cryptococcus neoformans (serotypes A and D) and Cryptococcus gattii (serotypes B and C) and correlated them with the production of NO by phagocytes and the activation of TLRs. GXM fractions were differentially recognized by TLR2/TLR1 (TLR2/1) and TLR2/6 heterodimers expressed on TLR-transfected HEK293A cells. Higher NF-kappaB luciferase reporter activity induced by GXM was observed in cells expressing TLR2/1 than in cells transfected with TLR2/6 constructs. A serotype B GXM from C. gattii was the most effective polysaccharide fraction activating the TLR-mediated response. This serotype B polysaccharide, which was also highly efficient at eliciting the production of NO by macrophages, was similar to the other GXM samples in monosaccharide composition, zeta potential, and electrophoretic mobility. However, immunofluorescence with four different monoclonal antibodies and dynamic light-scattering analysis revealed that the serotype B GXM showed particularities in serological reactivity and had the smallest effective diameter among the GXM samples analyzed in this study. Fractionation of additional serotype B GXMs, followed by exposure of these fractions to macrophages, revealed a correlation between NO production and reduced effective diameters. Our results demonstrate a great functional diversity in GXM samples from different isolates and establish their abilities to differentially activate cellular responses. We propose that serological properties as well as physical chemical parameters, such as the diameter of polysaccharide molecules, may potentially influence the inflammatory response against Cryptococcus spp. and may contribute to the differences in granulomatous inflammation between cryptococcal species.
Thomas R. Kozel - One of the best experts on this subject based on the ideXlab platform.
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vivo clearance of Glucuronoxylomannan, the major capsular polysaccharide of Cryptococcus neoformans: a critical role for tissue macrophages
2016Co-Authors: Matthew M. Grinsell, Jim E. Cutler, Yongmoon Han, Leanne A C. Weinhold, Thomas R. KozelAbstract:Cryptococcus neoformans produces a life-threatening meningitis in patients who are im-munocompromised by AIDS. A striking feature of cryptococcosis in AIDS is high serum levels of the major capsular polysaccharide, Glucuronoxylomannan (GXM). Soluble GXM has numerous biologic activities that may contribute to the pathogenesis of infection. The objective of the study was to further understand in vivo processing of GXM. Mice were injected intravenously with GXM, and the tissue distribution was determined. A macrophage suicide technique that used liposome-encapsulated dichloromethylene diphosphonate deter-mined the role of macrophages. GXM was cleared from serum with a half-life of 24–48 h but was retained for an indefinite period in tissues rich in cells of the mononuclear phagocyte system. Ablation of macrophages decreased GXM in the liver and spleen and increased serum GXM. The results identify a key role for macrophages in the clearance of GXM from serum and identify macrophages as a long-term reservoir for storage. Cryptococcus neoformans is an encapsulated yeast that may produce a life-threatening meningitis in immunocompromised persons, particularly those with AIDS. The major componen
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Microbial immune suppression mediated by direct engagement of inhibitory Fc receptor.
The Journal of Immunology, 2006Co-Authors: Claudia Monari, Arturo Casadevall, Francesco Bistoni, Thomas R. Kozel, Eva Pericolini, Francesca Paganelli, Stefano Perito, Anna VecchiarelliAbstract:A microbial polysaccharide (Glucuronoxylomannan (GXM)) exerts potent immunosuppression by direct engagement to immunoinhibitory receptor FcgammaRIIB. Activation of FcgammaRIIB by GXM leads to the recruitment and phosphorylation of SHIP that prevents IkappaBalpha activation. The FcgammaRIIB blockade inhibits GXM-induced IL-10 production and induces TNF-alpha secretion. GXM quenches LPS-induced TNF-alpha release via FcgammaRIIB. The addition of mAb to GXM reverses GXM-induced immunosuppression by shifting recognition from FcgammaRIIB to FcgammaRIIA. These findings indicate a novel mechanism by which microbial products can impair immune function through direct stimulation of an inhibitory receptor. Furthermore, our observations provide a new mechanism for the ability of specific Ab to reverse the immune inhibitory effects of certain microbial products.
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cryptococcus neoformans capsular Glucuronoxylomannan induces expression of fas ligand in macrophages
Journal of Immunology, 2005Co-Authors: Claudia Monari, Arturo Casadevall, Thomas R. Kozel, Eva Pericolini, Giovanni Bistoni, Anna VecchiarelliAbstract:The major component of capsular material of Cryptococcus neoformans is glucuronoxylomannnan (GXM), a polysaccharide that exhibits potent immunosuppressive properties in vitro and in vivo. The results reported here show that 1) soluble purified GXM induces a prompt, long-lasting, and potent up-regulation of Fas ligand (FasL) on macrophages, 2) the up-regulation of FasL is related to induced synthesis and increased mobilization to the cellular surface, 3) this effect is largely mediated by interaction between GXM and TLR4, 4) FasL up-regulation occurs exclusively in GXM-loaded macrophages, 5) macrophages that show up-regulation of FasL induce apoptosis of activated T cells expressing Fas and Jurkat cells that constitutively express Fas, and 6) anti-Fas Abs rescue T cells from apoptosis induced by GXM. Collectively our results reveal novel aspects of the immunoregulatory properties of GXM and suggest that this nontoxic soluble compound could be used to dampen the immune response, to promote or accelerate the death receptor, and to fix FasL expression in a TLR/ligand-dependent manner. In the present study, we delineate potential new therapeutic applications for GXM that exploit death receptors as key molecular targets in regulating cell-mediated cytotoxicity, immune homeostasis, and the immunopathology of diseases.
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Glucuronoxylomannan, a Microbial Compound, Regulates Expression of Costimulatory Molecules and Production of Cytokines in Macrophages
The Journal of infectious diseases, 2004Co-Authors: Claudia Monari, Arturo Casadevall, Francesco Bistoni, Thomas R. Kozel, Eva Pericolini, Donatella Pietrella, Anna VecchiarelliAbstract:Glucuronoxylomannan (GXM) is a microbial compound that can modulate the immune response. We investigated (1) the receptors involved in uptake of GXM on monocyte-derived macrophages (MDMs) from healthy donors, (2) the effects of GXM on expression of specific receptors, (3) the effects of GXM mediated by pattern-recognition receptors, and (4) GXM modulation of MDM accessory and secretory functions. Cellular receptors involved in uptake of GXM included Fc gamma RII, CD18, Toll-like receptor (TLR) 4, and CD14. Some biological functions of MDMs were profoundly affected by treatment with GXM, resulting in (1) increased expression of CD40 and CD86 via perturbation of TLR4, (2) decreased expression of major histocompatibility complex class II, (3) induction of interleukin-10 but not of tumor necrosis factor-alpha, and (4) decreased lipopolysaccharide (LPS)-induced production of cytokines. GXM represents an attractive compound to limit inflammatory processes and induce an LPS-tolerant state.
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Differences in outcome of the interaction between Cryptococcus neoformans Glucuronoxylomannan and human monocytes and neutrophils
European journal of immunology, 2003Co-Authors: Claudia Monari, Arturo Casadevall, Francesco Bistoni, Cinzia Retini, Thomas R. Kozel, Dale Netski, Anna VecchiarelliAbstract:Disseminated infections by the opportunistic yeast Cryptococcus neoformans are characterized by accumulation in tissues of Glucuronoxylomannan (GXM), the major component of the capsular polysaccharide. We investigated binding, uptake, and disposal of GXM by peripheral blood neutrophils and monocytes, and the effect of GXM uptake on phagocytic cell function. GXM was efficiently bound and internalized by both types of phagocytic cells, with maximal loading at 50 μg/ml, a GXM concentration found in serum and cerebrospinal fluid of some cryptococcosis patients. However, substantial differences were noted in the kinetics for uptake by macrophages and neutrophils. Whereas neutrophils rapidly ingested limited amounts of GXM and then expelled or degraded it after 1 h of incubation, macrophages demonstrated continuous intracellular accumulation for up to 1 week of incubation. Accumulation of GXM by neutrophils was accompanied by reduced anticryptococcal activity, suggesting one more mechanism for virulence enhancement by the major capsular component of C. neoformans.
Leonardo Nimrichter - One of the best experts on this subject based on the ideXlab platform.
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A Glucuronoxylomannan-like glycan produced by Trichosporon mucoides
Fungal genetics and biology : FG & B, 2018Co-Authors: Ana Claudia G. Zimbres, Leonardo Nimrichter, Fernanda L. Fonseca, Priscila C. Albuquerque, Luna S. Joffe, Taiane N. Souza, Stefânia De Oliveira Frazão, Patrícia Albuquerque, Marcio L. RodriguesAbstract:Abstract Trichosporon asahii shares with Cryptococcus species the ability to produce Glucuronoxylomannan (GXM), an immunomodulatory fungal polysaccharide. The ability of other opportunistic species of Trichosporon to produce GXM-like polysaccharides is unknown. In this study, we observed that T. mucoides was less pathogenic than T. asahii in an infection model of Galleria mellonella and asked whether this difference was related to the characteristics of GXM-like molecules. Compositional analysis of samples obtained from both pathogens indicated that the components of GXM (mannose, xylose and glucuronic acid) were, in fact, detected in T. mucoides and T. asahii glycans. The identification of the T. mucoides glycan as a GXM-like molecule was confirmed by its reactivity with a monoclonal antibody raised to cryptococcal GXM and incorporation of the glycan into the cell surface of an acapsular mutant of C. neoformans. T. mucoides and T. asahii glycans differed in molecular dimensions. The antibody to cryptococcal GXM recognized T. mucoides yeast forms less efficiently than T. asahii cells. Experiments with animal cells revealed that the T. mucoides glycan manifested antiphagocytic properties. Comparative phagocytosis assays revealed that T. mucoides and T. asahii were similarly recognized by macrophages. However, fungal association with the phagocytes did not depend on the typical receptors of cryptococcal GXM, as concluded from assays using macrophages obtained from Tlr2−/− and Cd14−/− knockout mice. These results add T. mucoides to the list of fungal pathogens producing GXM-like glycans, but also indicate a high functional diversity of this major fungal immunogen.
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A Paracoccidioides brasiliensis glycan shares serologic and functional properties with cryptococcal Glucuronoxylomannan
Fungal genetics and biology : FG & B, 2012Co-Authors: Priscila C. Albuquerque, Kildare Miranda, Arturo Casadevall, Fernanda L. Fonseca, Radames J. B. Cordero, Roberta Peres Da Silva, Caroline L. Ramos, Rosana Puccia, Joshua D. Nosanchuk, Leonardo NimrichterAbstract:The cell wall of the yeast form of the dimorphic fungus Paracoccidioides brasiliensis is enriched with α1,3-glucans. In Cryptococcus neoformans, α1,3-glucans interact with Glucuronoxylomannan (GXM), a heteropolysaccharide that is essential for fungal virulence. In this study, we investigated the occurrence of P. brasiliensis glycans sharing properties with cryptococcal GXM. Protein database searches in P. brasiliensis revealed the presence of sequences homologous to those coding for enzymes involved in the synthesis of GXM and capsular architecture in C. neoformans. In addition, monoclonal antibodies (mAbs) raised to cryptococcal GXM bound to P. brasiliensis cells. Using protocols that were previously established for extraction and analysis of C. neoformans GXM, we recovered a P. brasiliensis glycan fraction composed of mannose and galactose, in addition to small amounts of glucose, xylose and rhamnose. In comparison with the C. neoformans GXM, the P. brasiliensis glycan fraction components had smaller molecular dimensions. The P. brasiliensis components, nevertheless, reacted with different GXM-binding mAbs. Extracellular vesicle fractions of P. brasiliensis also reacted with a GXM-binding mAb, suggesting that the polysaccharide-like molecule is exported to the extracellular space in secretory vesicles. An acapsular mutant of C. neoformans incorporated molecules from the P. brasiliensis extract onto the cell wall, resulting in the formation of surface networks that resembled the cryptococcal capsule. Coating the C. neoformans acapsular mutant with the P. brasiliensis glycan fraction resulted in protection against phagocytosis by murine macrophages. These results suggest that P. brasiliensis and C. neoformans share metabolic pathways required for the synthesis of similar polysaccharides and that P. brasiliensis yeast cell walls have molecules that mimic certain aspects of C. neoformans GXM. These findings are important because they provide additional evidence for the sharing of antigenically similar components across phylogenetically distant fungal species. Since GXM has been shown to be important for the pathogenesis of C. neoformans and to elicit protective antibodies, the finding of similar molecules in P. brasiliensis raises the possibility that these glycans play similar functions in paracoccidiomycosis.
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Glucuronoxylomannan from Cryptococcus neoformans Down-regulates the Enzyme 6-Phosphofructo-1-kinase of Macrophages
The Journal of biological chemistry, 2011Co-Authors: Juliana Grechi, Leonardo Nimrichter, Marcio L. Rodrigues, Monica M. Marinho-carvalho, Patricia Zancan, Leonardo P. Cinelli, Andre M. O. Gomes, Mauro Sola-pennaAbstract:The encapsulated yeast Cryptococcus neoformans is the causative agent of cryptococosis, an opportunistic life-threatening infection. C. neoformans is coated by a polysaccharide capsule mainly composed of Glucuronoxylomannan (GXM). GXM is considered a key virulence factor of this pathogen. The present work aimed at evaluating the effects of GXM on the key glycolytic enzyme, 6-phosphofructo-1-kinase (PFK). GXM inhibited PFK activity in cultured murine macrophages in both dose- and time-dependent manners, which occurred in parallel to cell viability decrease. The polysaccharide also inhibited purified PFK, promoting a decrease on the enzyme affinity for its substrates. In macrophages GXM and PFK partially co-localized, suggesting that internalized polysaccharide directly may interact with this enzyme. The mechanism of PFK inhibition involved dissociation of tetramers into weakly active dimers, as revealed by fluorescence spectroscopy. Allosteric modulators of the enzyme able to stabilize its tetrameric conformation attenuated the inhibition promoted by GXM. Altogether, our results suggest that the mechanism of GXM-induced cell death involves the inhibition of the glycolytic flux.
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Glucuronoxylomannan fromCryptococcus neoformans Down-regulates the Enzyme 6-Phosphofructo-1-kinase of
2011Co-Authors: Juliana Grechi, Leonardo Nimrichter, Marcio L. Rodrigues, Monica M. Marinho-carvalho, Patricia Zancan, Leonardo P. Cinelli, Andre M. O. Gomes, Mauro Sola-pennaAbstract:is coated by a polysaccharide capsulemainly composed of Glucuronoxylomannan (GXM). GXM isconsidered a key virulence factor of this pathogen. The presentwork aimed at evaluating the effects of GXM on the key glyco-lytic enzyme, 6-phosphofructo-1-kinase (PFK). GXM inhibitedPFKactivityinculturedmurinemacrophagesinbothdose-andtime-dependentmanners,whichoccurredinparalleltocellvia-bilitydecrease.ThepolysaccharidealsoinhibitedpurifiedPFK,promoting a decrease on the enzyme affinity for its substrates.In macrophages GXM and PFK partially co-localized, suggest-ing that internalized polysaccharide directly may interact withthis enzyme. The mechanism of PFK inhibition involved disso-ciation of tetramers into weakly active dimers, as revealed byfluorescencespectroscopy.Allostericmodulatorsoftheenzymeable to stabilize its tetrameric conformation attenuated theinhibition promoted by GXM. Altogether, our results suggestthat the mechanism of GXM-induced cell death involves theinhibition of the glycolytic flux.
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Immunomodulatory Effects of Serotype B Glucuronoxylomannan from Cryptococcus gattii Correlate with Polysaccharide Diameter
Infection and immunity, 2010Co-Authors: Fernanda L. Fonseca, Leonardo Nimrichter, Arturo Casadevall, Susana Frases, Radames J. B. Cordero, Lilian L. Nohara, Igor C. Almeida, Marcio L. RodriguesAbstract:Glucuronoxylomannan (GXM), the major capsular component in the Cryptococcus complex, interacts with the immune system in multiple ways, which include the activation of Toll-like receptors (TLRs) and the modulation of nitric oxide (NO) production by phagocytes. In this study, we analyzed several structural parameters of GXM samples from Cryptococcus neoformans (serotypes A and D) and Cryptococcus gattii (serotypes B and C) and correlated them with the production of NO by phagocytes and the activation of TLRs. GXM fractions were differentially recognized by TLR2/TLR1 (TLR2/1) and TLR2/6 heterodimers expressed on TLR-transfected HEK293A cells. Higher NF-kappaB luciferase reporter activity induced by GXM was observed in cells expressing TLR2/1 than in cells transfected with TLR2/6 constructs. A serotype B GXM from C. gattii was the most effective polysaccharide fraction activating the TLR-mediated response. This serotype B polysaccharide, which was also highly efficient at eliciting the production of NO by macrophages, was similar to the other GXM samples in monosaccharide composition, zeta potential, and electrophoretic mobility. However, immunofluorescence with four different monoclonal antibodies and dynamic light-scattering analysis revealed that the serotype B GXM showed particularities in serological reactivity and had the smallest effective diameter among the GXM samples analyzed in this study. Fractionation of additional serotype B GXMs, followed by exposure of these fractions to macrophages, revealed a correlation between NO production and reduced effective diameters. Our results demonstrate a great functional diversity in GXM samples from different isolates and establish their abilities to differentially activate cellular responses. We propose that serological properties as well as physical chemical parameters, such as the diameter of polysaccharide molecules, may potentially influence the inflammatory response against Cryptococcus spp. and may contribute to the differences in granulomatous inflammation between cryptococcal species.
Fernanda L. Fonseca - One of the best experts on this subject based on the ideXlab platform.
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A Glucuronoxylomannan-like glycan produced by Trichosporon mucoides
Fungal genetics and biology : FG & B, 2018Co-Authors: Ana Claudia G. Zimbres, Leonardo Nimrichter, Fernanda L. Fonseca, Priscila C. Albuquerque, Luna S. Joffe, Taiane N. Souza, Stefânia De Oliveira Frazão, Patrícia Albuquerque, Marcio L. RodriguesAbstract:Abstract Trichosporon asahii shares with Cryptococcus species the ability to produce Glucuronoxylomannan (GXM), an immunomodulatory fungal polysaccharide. The ability of other opportunistic species of Trichosporon to produce GXM-like polysaccharides is unknown. In this study, we observed that T. mucoides was less pathogenic than T. asahii in an infection model of Galleria mellonella and asked whether this difference was related to the characteristics of GXM-like molecules. Compositional analysis of samples obtained from both pathogens indicated that the components of GXM (mannose, xylose and glucuronic acid) were, in fact, detected in T. mucoides and T. asahii glycans. The identification of the T. mucoides glycan as a GXM-like molecule was confirmed by its reactivity with a monoclonal antibody raised to cryptococcal GXM and incorporation of the glycan into the cell surface of an acapsular mutant of C. neoformans. T. mucoides and T. asahii glycans differed in molecular dimensions. The antibody to cryptococcal GXM recognized T. mucoides yeast forms less efficiently than T. asahii cells. Experiments with animal cells revealed that the T. mucoides glycan manifested antiphagocytic properties. Comparative phagocytosis assays revealed that T. mucoides and T. asahii were similarly recognized by macrophages. However, fungal association with the phagocytes did not depend on the typical receptors of cryptococcal GXM, as concluded from assays using macrophages obtained from Tlr2−/− and Cd14−/− knockout mice. These results add T. mucoides to the list of fungal pathogens producing GXM-like glycans, but also indicate a high functional diversity of this major fungal immunogen.
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Cryptococcus neoformans Glucuronoxylomannan fractions of different molecular masses are functionally distinct
Future microbiology, 2014Co-Authors: Priscila C. Albuquerque, Arturo Casadevall, Susana Frases, Fernanda L. Fonseca, Marcelo T. Bozza, Fabianno F. Dutra, Marcio L. RodriguesAbstract:ABSTRACT: Aims: Glucuronoxylomannan (GXM) is the major polysaccharide component of Cryptococcus neoformans. We evaluated in this study whether GXM fractions of different molecular masses were functionally distinct. Materials & methods: GXM samples isolated from C. neoformans cultures were fractionated to generate polysaccharide preparations differing in molecular mass. These fractions were used in experiments focused on the association of GXM with cell wall components of C. neoformans, as well as on the interaction of the polysaccharide with host cells. Results & conclusion: GXM fractions of variable molecular masses bound to the surface of a C. neoformans acapsular mutant in a punctate pattern that is in contrast to the usual annular pattern of surface coating observed when GXM samples containing the full molecular mass range were used. The polysaccharide samples were also significantly different in their ability to stimulate cytokine production by host cells. Our findings indicate that GXM fractions ar...
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A Paracoccidioides brasiliensis glycan shares serologic and functional properties with cryptococcal Glucuronoxylomannan
Fungal genetics and biology : FG & B, 2012Co-Authors: Priscila C. Albuquerque, Kildare Miranda, Arturo Casadevall, Fernanda L. Fonseca, Radames J. B. Cordero, Roberta Peres Da Silva, Caroline L. Ramos, Rosana Puccia, Joshua D. Nosanchuk, Leonardo NimrichterAbstract:The cell wall of the yeast form of the dimorphic fungus Paracoccidioides brasiliensis is enriched with α1,3-glucans. In Cryptococcus neoformans, α1,3-glucans interact with Glucuronoxylomannan (GXM), a heteropolysaccharide that is essential for fungal virulence. In this study, we investigated the occurrence of P. brasiliensis glycans sharing properties with cryptococcal GXM. Protein database searches in P. brasiliensis revealed the presence of sequences homologous to those coding for enzymes involved in the synthesis of GXM and capsular architecture in C. neoformans. In addition, monoclonal antibodies (mAbs) raised to cryptococcal GXM bound to P. brasiliensis cells. Using protocols that were previously established for extraction and analysis of C. neoformans GXM, we recovered a P. brasiliensis glycan fraction composed of mannose and galactose, in addition to small amounts of glucose, xylose and rhamnose. In comparison with the C. neoformans GXM, the P. brasiliensis glycan fraction components had smaller molecular dimensions. The P. brasiliensis components, nevertheless, reacted with different GXM-binding mAbs. Extracellular vesicle fractions of P. brasiliensis also reacted with a GXM-binding mAb, suggesting that the polysaccharide-like molecule is exported to the extracellular space in secretory vesicles. An acapsular mutant of C. neoformans incorporated molecules from the P. brasiliensis extract onto the cell wall, resulting in the formation of surface networks that resembled the cryptococcal capsule. Coating the C. neoformans acapsular mutant with the P. brasiliensis glycan fraction resulted in protection against phagocytosis by murine macrophages. These results suggest that P. brasiliensis and C. neoformans share metabolic pathways required for the synthesis of similar polysaccharides and that P. brasiliensis yeast cell walls have molecules that mimic certain aspects of C. neoformans GXM. These findings are important because they provide additional evidence for the sharing of antigenically similar components across phylogenetically distant fungal species. Since GXM has been shown to be important for the pathogenesis of C. neoformans and to elicit protective antibodies, the finding of similar molecules in P. brasiliensis raises the possibility that these glycans play similar functions in paracoccidiomycosis.
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Immunomodulatory Effects of Serotype B Glucuronoxylomannan from Cryptococcus gattii Correlate with Polysaccharide Diameter
Infection and immunity, 2010Co-Authors: Fernanda L. Fonseca, Leonardo Nimrichter, Arturo Casadevall, Susana Frases, Radames J. B. Cordero, Lilian L. Nohara, Igor C. Almeida, Marcio L. RodriguesAbstract:Glucuronoxylomannan (GXM), the major capsular component in the Cryptococcus complex, interacts with the immune system in multiple ways, which include the activation of Toll-like receptors (TLRs) and the modulation of nitric oxide (NO) production by phagocytes. In this study, we analyzed several structural parameters of GXM samples from Cryptococcus neoformans (serotypes A and D) and Cryptococcus gattii (serotypes B and C) and correlated them with the production of NO by phagocytes and the activation of TLRs. GXM fractions were differentially recognized by TLR2/TLR1 (TLR2/1) and TLR2/6 heterodimers expressed on TLR-transfected HEK293A cells. Higher NF-kappaB luciferase reporter activity induced by GXM was observed in cells expressing TLR2/1 than in cells transfected with TLR2/6 constructs. A serotype B GXM from C. gattii was the most effective polysaccharide fraction activating the TLR-mediated response. This serotype B polysaccharide, which was also highly efficient at eliciting the production of NO by macrophages, was similar to the other GXM samples in monosaccharide composition, zeta potential, and electrophoretic mobility. However, immunofluorescence with four different monoclonal antibodies and dynamic light-scattering analysis revealed that the serotype B GXM showed particularities in serological reactivity and had the smallest effective diameter among the GXM samples analyzed in this study. Fractionation of additional serotype B GXMs, followed by exposure of these fractions to macrophages, revealed a correlation between NO production and reduced effective diameters. Our results demonstrate a great functional diversity in GXM samples from different isolates and establish their abilities to differentially activate cellular responses. We propose that serological properties as well as physical chemical parameters, such as the diameter of polysaccharide molecules, may potentially influence the inflammatory response against Cryptococcus spp. and may contribute to the differences in granulomatous inflammation between cryptococcal species.
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Structural and functional properties of the Trichosporon asahii Glucuronoxylomannan.
Fungal Genetics and Biology, 2009Co-Authors: Fernanda L. Fonseca, Leonardo Nimrichter, Arturo Casadevall, Susana Frases, Olga Fischman-gompertz, Marcio L. RodriguesAbstract:The virulence attributes of Trichosporon asahii are virtually unknown, despite its growing relevance as causative agent of superficial and invasive diseases in humans. Glucuronoxylomannan (GXM) is a well described virulence factor of pathogenic species in the Cryptococcus genus. GXM is also produced by species of the Trichosporon genus, and both polysaccharides share antigenic determinants, but unlike cryptococcal GXM, relatively little work has been done on trichosporal GXMs. In this study, we analyzed structural and functional aspects of GXM produced by T. asahii and compared them to the properties of the cryptococcal polysaccharide. Trichosporal and cryptococcal GXM shared antigenic reactivity, but the former polysaccharide had smaller effective diameter and negative charge. GXM anchoring to the cell wall was perturbed by dimethylsulfoxide and required interactions of chitin-derived oligomers with the polysaccharide. GXM from T. asahii supernatants are incorporated by acapsular mutants of Cryptococcus neoformans, which renders these cells more resistant to phagocytosis by mouse macrophages. In summary, our results establish that despite similarities in cell wall anchoring, antigenic and antiphagocytic properties, trichosporal and cryptococcal GXMs manifest major structural differences that may directly affect polysaccharide assembly at the fungal surface.