Calcium signaling components in the human pathogen<i>Cryptococcus neoformans</i>

Kmetzsch, Lívia; Staats, Charley Christian; Rodrigues, Márcio L.; Schrank, Augusto; Vainstein, Marilene Henning

doi:10.4161/cib.4.2.14271

Cited by 10 publications

(9 citation statements)

References 23 publications

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“…3). This value, which was in agreement with previous studies using similar models (Kmetzsch et al, 2011a,b,c, 2010), was about threefold higher than the percentage of Tlr2 −/− macrophages infected by C. neoformans . Blocking of fungal chitooligomers with WGA before incubation with BMDM caused a decrease of approximately 50% in the efficacy of association of C. neoformans with macrophages obtained from WT mice.…”

Section: Resultssupporting

confidence: 93%

“…Fungal suspensions were prepared in DMEM at a ratio of 10 yeasts per host cell. Interactions between fungal and host cells occurred at 37 °C with 5% CO 2 for 12 h. Incubation time was based on previous phagocytosis studies developed in our laboratory showing that 60–70% of the phagocytes become infected and efficiently internalize C. neoformans after overnight incubations in the absence of opsonins (Kmetzsch et al, 2011a,b,c, 2010). In some systems, the medium of interaction was supplemented with the trisaccharide (GlcNAc) 3 at 10 μg/ml (Sigma; Richmond, VA).…”

Section: Methodsmentioning

confidence: 99%

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Binding of the wheat germ lectin to Cryptococcus neoformans chitooligomers affects multiple mechanisms required for fungal pathogenesis

Fonseca

Guimarães

Kmetzsch

et al. 2013

Fungal Genetics and Biology

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The principal capsular component of Cryptococcus neoformans, glucuronoxylomannan (GXM), interacts with surface glycans, including chitin-like oligomers. Although the role of GXM in cryptococcal infection has been well explored, there is no information on how chitooligomers affect fungal pathogenesis. In this study, surface chitooligomers of C. neoformans were blocked through the use of the wheat germ lectin (WGA) and the effects on animal pathogenesis, interaction with host cells, fungal growth and capsule formation were analyzed. Treatment of C. neoformans cells with WGA followed by infection of mice delayed mortality relative to animals infected with untreated fungal cells. This observation was associated with reduced brain colonization by lectin-treated cryptococci. Blocking chitooligomers also rendered yeast cells less efficient in their ability to associate with phagocytes. WGA did not affect fungal viability, but inhibited GXM release to the extracellular space and capsule formation. In WGA-treated yeast cells, genes that are involved in capsule formation and GXM traffic had their transcription levels decreased in comparison with untreated cells. Our results suggest that cellular pathways required for capsule formation and pathogenic mechanisms are affected by blocking chitin-derived structures at the cell surface of C. neoformans. Targeting chitooligomers with specific ligands may reveal new therapeutic alternatives to control cryptococcosis.

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Section: Resultssupporting

confidence: 93%

Section: Methodsmentioning

confidence: 99%

Binding of the wheat germ lectin to Cryptococcus neoformans chitooligomers affects multiple mechanisms required for fungal pathogenesis

Fonseca

Guimarães

Kmetzsch

et al. 2013

Fungal Genetics and Biology

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“…These genes were individually inactivated in the C. gattii genome. The cloning reaction was performed with the BP clonase (Life Technologies, Carlsbad, CA, USA) and the 5 0 and 3 0 target gene flanks, as well the pDONRHYG vector [42,49,50]. The 5 0 and 3 0 target gene flanks were PCR-amplified (Table S1) and purified from agarose gels (Illustra GFX PCR DNA and Gel Band Purification Kit; GE Healthcare, Uppsala, Sweden).…”

Section: Disruption and Complementation Of The Ure Genesmentioning

confidence: 99%

“…The 5 0 and 3 0 target gene flanks were PCR-amplified (Table S1) and purified from agarose gels (Illustra GFX PCR DNA and Gel Band Purification Kit; GE Healthcare, Uppsala, Sweden). The cloning reaction was performed with the BP clonase (Life Technologies, Carlsbad, CA, USA) and the 5 0 and 3 0 target gene flanks, as well the pDONRHYG vector [42,49,50]. The product of this reaction was transformed into the E. coli OmniMAX 2-T1 strain.…”

Section: Disruption and Complementation Of The Ure Genesmentioning

confidence: 99%

Cryptococcus gattii urease as a virulence factor and the relevance of enzymatic activity in cryptococcosis pathogenesis

et al. 2015

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Ureases (EC 3.5.1.5) are Ni(2+) -dependent metalloenzymes produced by plants, fungi and bacteria that hydrolyze urea to produce ammonia and CO2 . The insertion of nickel atoms into the apo-urease is better characterized in bacteria, and requires at least three accessory proteins: UreD, UreF, and UreG. Our group has demonstrated that ureases possess ureolytic activity-independent biological properties that could contribute to the pathogenicity of urease-producing microorganisms. The presence of urease in pathogenic bacteria strongly correlates with pathogenesis in some human diseases. Some medically important fungi also produce urease, including Cryptococcus neoformans and Cryptococcus gattii. C. gattii is an etiological agent of cryptococcosis, most often affecting immunocompetent individuals. The cryptococcal urease might play an important role in pathogenesis. It has been proposed that ammonia produced via urease action might damage the host endothelium, which would enable yeast transmigration towards the central nervous system. To analyze the role of urease as a virulence factor in C. gattii, we constructed knockout mutants for the structural urease-coding gene URE1 and for genes that code the accessory proteins Ure4 and Ure6. All knockout mutants showed reduced multiplication within macrophages. In intranasally infected mice, the ure1Δ (lacking urease protein) and ure4Δ (enzymatically inactive apo-urease) mutants caused reduced blood burdens and a delayed time of death, whereas the ure6Δ (enzymatically inactive apo-urease) mutant showed time and dose dependency with regard to fungal burden. Our results suggest that C. gattii urease plays an important role in virulence, in part possibly through enzyme activity-independent mechanism(s).

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“…In the same way, calcium-dependent activation of calcineurin can be modeled by addition of high amounts of exogenous calcium, but sensitivity of cna1 α mutants to conditions of high carbon dioxide, alkaline pH and high concentrations of cations suggest that these environmental signals are the physiological conditions responsible for the activation of intracellular calcium stores. In addition, calcium signaling through calmodulin and the phosphatase calcineurin are required for mating and virulence of C. neoformans [71–73]. An increased level of calcium is sensed by calmodulin, which activates the conserved serine/threonine phosphatase calcineurin (Fig.…”

Section: Calcium-calcineurin Signaling Pathwaymentioning

confidence: 99%

‘Popping the Clutch’: Novel Mechanisms Regulating Sexual Development in Cryptococcus neoformans

Park

Williamson

2011

Mycopathologia

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Sexual reproduction in fungal pathogens such as Cryptococcus provides natural selection and adaptation of the organisms to environmental conditions by allowing beneficial mutations to spread. However, successful mating in these fungi requires a time-critical induction of signaling pheromones when appropriate partners become available. Recently, it has been shown that the fungus uses the transcriptional equivalent of the racing technique: ‘popping the clutch’-- pushing in the clutch pedal, putting the car in gear, revving with the gas pedal and then dropping the clutch pedal to accelerate rapidly. In the same way, Cryptococcus during vegetative growth constitutively matches a high rate of pheromone synthesis with a high rate of degradation to produce repressed levels of transcript. Then, when mating is required, the fungus drops the degradative machinery, resulting in a rapid induction of the pheromone. Pairing with this novel regulatory cycle are a host of mitogen-activated protein (MAP) kinase cascades, cyclic AMP (cAMP) dependent, and calcium-calcineurin signaling pathways that maintain these high rates of pheromone synthesis and prime downstream pathways for an effective mating response. The intersection of a number of virulence-associated traits with sexual development such as the synthesis of an immune-disruptive laccase as well as a protective polysaccharide capsule makes these rapid regulatory strategies a formidable foe in the battle against human disease.

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Calcium signaling components in the human pathogenCryptococcus neoformans

Cited by 10 publications

References 23 publications

Binding of the wheat germ lectin to Cryptococcus neoformans chitooligomers affects multiple mechanisms required for fungal pathogenesis

Binding of the wheat germ lectin to Cryptococcus neoformans chitooligomers affects multiple mechanisms required for fungal pathogenesis

Cryptococcus gattii urease as a virulence factor and the relevance of enzymatic activity in cryptococcosis pathogenesis

‘Popping the Clutch’: Novel Mechanisms Regulating Sexual Development in Cryptococcus neoformans

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