UDP Glucuronate Decarboxylase and Synthesis of Capsular Polysaccharide in Cryptococcus neoformans

1982 ◽  
Vol 152 (2) ◽  
pp. 932-934
Author(s):  
Eric S. Jacobson ◽  
William R. Payne
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Decarboxylase Activity

UDP glucuronate decarboxylase activity was comparable in encapsulated and non-encapsulated strains of Cryptococcus neoformans , required NAD ( K a = 0.2 mM), and was inhibited by NADH ( K i = 0.1 mM) and UDP xylose.

scholarly journals The capsule ofCryptococcus neoformansmodulates phagosomal pH through its acid-base properties

2018 ◽  
Author(s):  
Carlos M. De Leon-Rodriguez ◽  
Man Shun Fu ◽  
M. Osman Corbali ◽  
Radames J.B. Cordero ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Glucuronic Acid ◽  
Pathogenic Fungus ◽  
Weak Acid ◽  
Phagocytic Cells ◽  
Acid Base ◽  
Encapsulated Cells ◽  
Human Pathogenic Fungus ◽  
Base Properties

AbstractPhagosomal acidification is a critical cellular mechanism for the inhibition and killing of ingested microbes by phagocytic cells. The acidic environment activates microbicidal proteins and creates an unfavorable environment for the growth of many microbes. Consequently, numerous pathogenic microbes have developed strategies for countering phagosomal acidification through various mechanisms that include interference with phagosome maturation. The human pathogenic fungusCryptococcus neoformansresides in acidic phagosome after macrophage ingestion that actually provides a favorable environment for replication since the fungus replicates faster at acidic pH. We hypothesized that the glucuronic acid residues in the capsular polysaccharide had the capacity to affect phagosome acidity through their acid-base properties. A ratiometric fluorescence comparison of imaged phagosomes containingC. neoformansto those containing beads showed that the latter were significantly more acidic. Similarly, phagosomes containing non-encapsulatedC. neoformanscells were more acidic than those containing encapsulated cells. Acid-base titrations of isolatedC. neoformanspolysaccharide revealed that it behaves as a weak acid with maximal buffering capacity around pH 4-5. We interpret these results as indicating that the glucuronic acid residues in theC. neoformanscapsular polysaccharide can buffer phagosomal acidification. Interference with phagosomal acidification represents a new function for the cryptococcal capsule in virulence and suggests the importance of considering the acid-base properties of microbial capsules in the host-microbe interaction for other microbes with charged residues in their capsules.ImportanceCryptococcus neoformansis the causative agent of cryptococcosis, a devastating fungal disease that affects thousands of individuals worldwide. This fungus has the capacity to survive inside phagocytic cells, which contributes to persistence of infection and dissemination. One of the major mechanisms of host phagocytes is to acidify the phagosomal compartment after ingestion of microbes. This study shows that the capsule ofC. neoformanscan interfere with full phagosomal acidification by serving as a buffer.

scholarly journals Monoclonal Antibodies Reactive with Immunorecessive Epitopes of Glucuronoxylomannan, the Major Capsular Polysaccharide of Cryptococcus neoformans

2003 ◽  
Vol 10 (5) ◽  
pp. 903-909 ◽  
Author(s):  
Suzanne Brandt ◽  
Peter Thorkildson ◽  
Thomas R. Kozel
Keyword(s):  
Monoclonal Antibody ◽  
Immune Response ◽  
Monoclonal Antibodies ◽  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Passive Immunization ◽  
The Other ◽  
Serotype A ◽  
Humoral Immune ◽  
New Information

ABSTRACT Cryptococcus neoformans is surrounded by an antiphagocytic capsule whose primary constituent is glucuronoxylomannan (GXM). An epitope shared by GXM serotypes A, B, C, and D is immunodominant when mice are immunized with serotype A GXM. In contrast, an epitope shared only by serotypes A and D is immunodominant when mice are immunized with serotype D. Hybridomas secreting antibodies reactive with subdominant epitopes were identified through a positive-negative screening procedure in which antibody-secreting colonies were characterized by reactivity with both the immunizing polysaccharide and GXMs from each of the four major serotypes. In this manner, a monoclonal antibody (MAb) that was reactive with an epitope shared only by serotypes A and B was identified and designated F10F5. Such an epitope has not been described previously. Immunization of mice with de-O-acetylated serotype A GXM generated a hybridoma that secreted an antibody, designated F12D2, that was reactive with all four serotypes. Unlike previously described monoclonal and polyclonal panspecific antibodies, the reactivity of MAb F12D2 was not altered by de-O-acetylation of GXM. These results indicate that there are at least two panspecific GXM epitopes; one epitope is dependent on O acetylation for antibody reactivity, and the other is independent of O acetylation. This study identifies strategies for production of MAbs that are reactive with subdominant or cryptic GXM epitopes and provides new information regarding the antigenic makeup and the humoral immune response to GXM, an essential virulence factor that is a target for active and passive immunization.

SPECIFIC DEGRADATION OF CRYPTOCOCCUS NEOFORMANS 3723 CAPSULAR POLYSACCHARIDE BY A MICROBIAL ENZYME: I. ISOLATION, PARTIAL PURIFICATION, AND PROPERTIES OF THE ENZYME

1961 ◽  
Vol 7 (1) ◽  
pp. 53-60 ◽  
Author(s):  
Hans H. Gadebusch ◽  
John D. Johnson
Keyword(s):  
Cryptococcus Neoformans ◽  
Capsular Polysaccharide ◽  
Enzyme Reaction ◽  
Partial Purification ◽  
Intracellular Enzyme ◽  
Purification And Properties ◽  
Microbial Enzyme ◽  
Specific Degradation ◽  
Kinetics Of ◽  
Enzyme I

A partially purified intracellular enzyme from a species of Alcaligenes is described which specifically initiates the degradation of the he heteropolysaccharide of Cryptococcus neoformans, isolate 3723, The enzyme is active in the presence of serum and can be inactivated by heating at 45 °C for 10 minutes, The kinetics of the enzyme reaction are similar to those of other enzymes. Recovery and identification of the four known monosaccharides from enzymatic hydrolyzates suggest the presence of a number of other enzymes in these preparations.

scholarly journals Immunoglobulin G3 blocking antibodies to the fungal pathogen Cryptococcus neoformans.

1996 ◽  
Vol 183 (4) ◽  
pp. 1905-1909 ◽  
Author(s):  
G Nussbaum ◽  
R Yuan ◽  
A Casadevall ◽  
M D Scharff
Keyword(s):  
Cryptococcus Neoformans ◽  
Host Defense ◽  
Fungal Pathogen ◽  
Capsular Polysaccharide ◽  
Protective Efficacy ◽  
Antibody Therapy ◽  
Antibody Isotype ◽  
Cryptococcal Infection ◽  
Epitope Specificity ◽  
Blocking Antibodies

Vaccination and infection can elicit protective and nonprotective antibodies to the fungus Cryptococcus neoformans in mice. The effect of nonprotective antibodies on host defense is unknown. In this study we used mixtures of protective and nonprotective monoclonal antibodies (mAbs) to determine if nonprotective mAbs blocked the activity of the protective mAbs. Antibody isotype and epitope specificity are important in determining the ability to prolong survival in mice given a lethal C. neoformans infection. Three different nonprotective immunoglobulin (Ig) G23 mAbs to cryptococcal capsular polysaccharide were used to study the interaction between the IgG3 isotype and protective IgG1 and IgG2a mAbs in murine cryptococcal infection. One IgG3 mAb reduced the protective efficacy of an IgG1 with identical epitope specificity. A second IgG3 mAb with different epitope specificity also reduced the protection provided by the IgG1 mAb. The protective efficacy of an IgG2a mAb was also dramatically decreased by still another IgG3 mAb. To our knowledge this is the first report of blocking antibodies to a fungal pathogen. The results have important implications for the development of vaccines and passive antibody therapy against C. neoformans.

scholarly journals Lipophilic Dye Staining of Cryptococcus neoformans Extracellular Vesicles and Capsule

2009 ◽  
Vol 8 (9) ◽  
pp. 1373-1380 ◽  
Author(s):  
André Moraes Nicola ◽  
Susana Frases ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Extracellular Vesicles ◽  
Capsular Polysaccharide ◽  
Complex Structure ◽  
Spectral Characteristics ◽  
Acid Dyes ◽  
Whole Cells ◽  
Direct Binding

ABSTRACT Cryptococcus neoformans is an encapsulated yeast that causes systemic mycosis in immunosuppressed individuals. Recent studies have determined that this fungus produces vesicles that are released to the extracellular environment both in vivo and in vitro. These vesicles contain assorted cargo that includes several molecules associated with virulence and implicated in host-pathogen interactions, such as capsular polysaccharides, laccase, urease, and other proteins. To date, visualization of extracellular vesicles has relied on transmission electron microscopy, a time-consuming technique. In this work we report the use of fluorescent membrane tracers to stain lipophilic structures in cryptococcal culture supernatants and capsules. Two dialkylcarbocyanine probes with different spectral characteristics were used to visualize purified vesicles by fluorescence microscopy and flow cytometry. Dual staining of vesicles with dialkylcarbocyanine and RNA-selective nucleic acid dyes suggested that a fraction of the vesicle population carried RNA. Use of these dyes to stain whole cells, however, was hampered by their possible direct binding to capsular polysaccharide. A fluorescent phospholipid was used as additional membrane tracer to stain whole cells, revealing punctate structures on the edge of the capsule which are consistent with vesicular trafficking. Lipophilic dyes provide new tools for the study of fungal extracellular vesicles and their content. The finding of hydrophobic regions in the capsule of C. neoformans adds to the growing evidence for a structurally complex structure composed of polysaccharide and nonpolysaccharide components.

scholarly journals The Vacuolar Ca2+ Exchanger Vcx1 Is Involved in Calcineurin-Dependent Ca2+ Tolerance and Virulence in Cryptococcus neoformans

2010 ◽  
Vol 9 (11) ◽  
pp. 1798-1805 ◽  
Author(s):  
Lívia Kmetzsch ◽  
Charley Christian Staats ◽  
Elisa Simon ◽  
Fernanda L. Fonseca ◽  
Débora L. de Oliveira ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Cyclosporine A ◽  
Capsular Polysaccharide ◽  
Cytosolic Calcium ◽  
Content Type ◽  
Human Body Temperature ◽  
Life Threatening ◽  
High Concentrations ◽  
Capsule Size ◽  
Virulence Attribute

ABSTRACT Cryptococcus neoformans is an encapsulated yeast that causes a life-threatening meningoencephalitis in immunocompromised individuals. The ability to survive and proliferate at the human body temperature is an essential virulence attribute of this pathogen. This trait is controlled in part by the Ca2+-calcineurin pathway, which senses and utilizes cytosolic calcium for signaling. In the present study, the identification of the C. neoformans gene VCX1, which encodes a vacuolar calcium exchanger, is reported. The VCX1 knockout results in hypersensitivity to the calcineurin inhibitor cyclosporine A at 35°C, but not at 30°C. Furthermore, high concentrations of CaCl2 lead to growth inhibition of the vcx1 mutant strain only in the presence of cyclosporine A, indicating that Vcx1 acts in parallel with calcineurin. The loss of VCX1 does not influence cell wall integrity or capsule size but decreases secretion of the major capsular polysaccharide glucuronoxylomannan (GXM) in culture supernatants.Vcx1 also influences C. neoformans phagocytosis by murine macrophages and is required for full virulence in mice. Analysis of cellular distribution by confocal microscopy confirmed the vacuolar localization of Vcx1 in C. neoformans cells.

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