<i>KRE</i> genes are required for β‐1,6‐glucan synthesis, maintenance of capsule architecture and cell wall protein anchoring in <i>Cryptococcus neoformans</i>

Gilbert, Nicole M.; Donlin, Maureen J.; Gerik, Kimberly J.; Specht, Charles A.; Djordjevic, Julianne T.; Wilson, Christabel F.; Sorrell, Tania C.; Lodge, Jennifer K.

doi:10.1111/j.1365-2958.2010.07119.x

Cited by 95 publications

(87 citation statements)

References 66 publications

(103 reference statements)

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“…carinii cell wall preparations contain ␤-1,6 glucans and can incorporate UDP-glucose into ␤-1,6 polysaccharide. To determine the presence of ␤-1,6 glucans in P. carinii, we isolated a ␤-1,6 glucan-enriched cell wall fraction and implemented a competitive dot blot assay (39) to verify its ␤-1,6 glucan content, using a polyclonal ␤-1,6 glucan antibody we generated. Similar to what others have shown with ␤-1,6 glucans derived from C. neoformans (39), isolated P. carinii ␤-1,6 preparations immobilized on the membrane bound the ␤-1,6 glucan antibody.…”

Section: Resultsmentioning

confidence: 99%

“…To determine the presence of ␤-1,6 glucans in P. carinii, we isolated a ␤-1,6 glucan-enriched cell wall fraction and implemented a competitive dot blot assay (39) to verify its ␤-1,6 glucan content, using a polyclonal ␤-1,6 glucan antibody we generated. Similar to what others have shown with ␤-1,6 glucans derived from C. neoformans (39), isolated P. carinii ␤-1,6 preparations immobilized on the membrane bound the ␤-1,6 glucan antibody. Furthermore, the soluble ␤-1,6 glucan pustulan was the single polysaccharide that could significantly compete with the P. carinii ␤-1,6 glucan immobilized on the membrane (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…More recently, another glucan component of the fungal and yeast cell walls, termed ␤-1,6 glucan, has been shown both to possess relevant cell wall structural activity and to promote host inflammatory responses and organism virulence (11,39). ␤-1,6 Glucan linkages are thought to be involved in anchoring glycophosphatidylinositol (GPI)-linked membrane-associated proteins to ␤-1,3 glucans (66, 67).…”

Section: Discussionmentioning

confidence: 99%

“…To verify the ␤-1,6 glucan contents of the carbohydrates contained in the P. carinii cell wall isolates, competitive dot blot assays were performed (39). Serial dilutions of P. carinii cell wall preparations (CWP) isolated as described above were spotted (2 l) onto Hybond-C nitrocellulose (Amersham) and air dried.…”

Section: Methodsmentioning

confidence: 99%

“…The ␤-1,6 glucan antibody used in the dot blot assay, as well as in the inhibition enzyme immunoassay (IEIA) described below, was generated in rabbits (Bethyl Laboratories) as detailed above. The anti-␤-1,6 glucan antibody working solution (1:5,000 in TBST) was mixed with various soluble competing carbohydrates, including diluent alone (no competing carbohydrate) or pustulan (soluble ␤-1,6 glucan), laminarin (soluble ␤-1,3 glucans), or yeast mannan (soluble mannoproteins), and rocked at 25°C for 1 h prior to addition of the mixture to the spotted membrane (39). Each competing carbohydrate was present at a concentration of 5 g/ml.…”

Section: Methodsmentioning

confidence: 99%

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Evidence for Proinflammatory β-1,6 Glucans in the Pneumocystis carinii Cell Wall

et al. 2015

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Inflammation is a major cause of respiratory impairment during Pneumocystis pneumonia. Studies support a significant role for cell wall ␤-glucans in stimulating inflammatory responses. Fungal ␤-glucans are comprised of D-glucose homopolymers containing ␤-1,3-linked glucose backbones with ␤-1,6-linked glucose side chains. Prior studies in Pneumocystis carinii have characterized ␤-1,3 glucan components of the organism. However, recent investigations in other organisms support important roles for ␤-1,6 glucans, predominantly in mediating host cellular activation. Accordingly, we sought to characterize ␤-1,6 glucans in the cell wall of Pneumocystis and to establish their activity in lung cell inflammation. Immune staining revealed specific ␤-1,6 localization in P. carinii cyst walls. Homology-based cloning facilitated characterization of a functional P. carinii kre6 (Pckre6) ␤-1,6 glucan synthase in Pneumocystis that, when expressed in kre6-deficient Saccharomyces cerevisiae, restored cell wall stability. Recently synthesized ␤-1,6 glucan synthase inhibitors decreased the ability of isolated P. carinii preparations to generate ␤-1,6 carbohydrate. In addition, isolated ␤-1,6 glucan fractions from Pneumocystis elicited vigorous tumor necrosis factor alpha (TNF-␣) responses from macrophages. These inflammatory responses were significantly dampened by inhibition of host cell plasma membrane microdomain function. Together, these studies indicate that ␤-1,6 glucans are present in the P. carinii cell wall and contribute to lung cell inflammatory activation during infection. P neumocystis organisms are opportunistic fungi that produce significant morbidity and mortality in immunocompromised hosts, with infection-related fatalities ranging between 10% and 45% (1). Pneumocystis jirovecii is the species known to infect humans, while Pneumocystis carinii represents the parallel species studied widely in rodents (2). Pneumocystis pneumonia remains a significant cause of mortality during AIDS, despite highly active antiretroviral therapy (3-5). Severe Pneumocystis pneumonia is characterized by intense lung inflammation involving CD8 ϩ cells and neutrophils, impairing gas exchange (6-9).The P. carinii cell walls contain abundant ␤-glucan molecules (10). Fungal cell wall ␤-glucans are homopolymers of D-glucose consisting of ␤-1,3 core chains with variable numbers of ␤-1,6 glucose side chains. The variable inflammatory activities of different glucan preparations have been postulated to be related to the relative amounts and configurations of these two major structures (␤-1,3 versus ␤-1,6) (11). Almost all of the initial studies in fungi have largely focused on unfractionated glucans (10). In fact, all prior studies in P. carinii previously utilized only unfractionated ␤-1,3/␤-1,6 glucans. Interestingly, recent investigations in Saccharomyces cerevisiae indicate major roles for ␤-1,6 glucans in strongly mediating cellular activation and inflammation (11). Our investigations of unfractionated P. carinii ␤-glucans indicate that innate ...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Evidence for Proinflammatory β-1,6 Glucans in the Pneumocystis carinii Cell Wall

et al. 2015

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Synthetic amino acids‐based short amphipathic peptides exhibit antifungal activity by targeting cell membrane disruption

Aaghaz

Sharma

Maurya

et al. 2023

Drug Development Research

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Availability of a limited number of antifungal drugs created a necessity to develop new antifungals with distinct mode of action. Investigation on a new series of peptides led us to identify Boc‐His‐Trp‐His[1‐(4‐tert‐butylphenyl)] (10g) as the most promising inhibitor exhibiting IC50 value of 4.4 µg/mL against Cryptococcus neoformans. Analog 10g exhibit high selectivity to fungal cells and was nonhemolytic and noncytotoxic at its minimum inhibitory concentration. 10g produced fungicidal effect on growing cryptococcal cells and displayed synergistic effect with amphotericin B. Overall cationic character of 10g resulted in interaction with negatively charged fungal membrane while hydrophobicity enhanced penetration inside the cryptococcal cells causing hole(s) formation and disruption to the membrane as evident by the scanning electron microscopy, transmission electron microscopy, and confocal laser scanning microscopy analyses. Flow cytometric investigation revealed rapid death of fungal cells by apopotic pathway.

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Overexpression of a beta‐1,6‐glucanase gene GluM in transgenic rice confers high resistance to rice blast, sheath blight and false smut

Shen

Wang

et al. 2023

Pest Management Science

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BACKGROUND: Frequent fungal diseases tend to lead to severe losses in rice production. As a main component of the fungal cell wall, glucan plays an important role in the growth and development of fungi. Glucanase can inhibit the growth of fungi by breaking glycosidic bonds, and may be a promising target for developing rice varieties with broad-spectrum disease resistance.RESULTS: We transferred a codon-optimized ⊎-1,6-glucanase gene (GluM) from myxobacteria into the japonica rice variety Zhonghua11 (ZH11), and obtained a large number of individual transgenic plants with GluM overexpression. Based on molecular analysis, three single-copy homozygous lines with GluM overexpression were selected for assessment of fungal disease resistance at the T 3 generation. Compared with that of the recipient cultivar ZH11, the area of rice blast lesion in transgenic rice was reduced by 82.71%; that of sheath blight lesion was decreased by 35.76%-43.67%; the sheath blight resistance in the field was enhanced by an average of 0.75 grade over 3 years; and the incidence of diseased panicles due to rice false smut was decreased by 65.79%. More importantly, there was no obvious loss of yield (without a significant effect on agronomic traits). Furthermore, plants overexpressing a ⊎-1,6-glucanase gene showed higher disease resistance than rice plants overexpressing a ⊎-1,3-glucanase gene derived from tobacco.CONCLUSION: The ⊎-1,6-glucanase gene GluM can confer broad-spectrum disease resistance to rice, providing an environmentally friendly alternative way to effectively manage fungal pathogens in rice production.

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KRE genes are required for β‐1,6‐glucan synthesis, maintenance of capsule architecture and cell wall protein anchoring in Cryptococcus neoformans

Cited by 95 publications

References 66 publications

Evidence for Proinflammatory β-1,6 Glucans in the Pneumocystis carinii Cell Wall

Evidence for Proinflammatory β-1,6 Glucans in the Pneumocystis carinii Cell Wall

Synthetic amino acids‐based short amphipathic peptides exhibit antifungal activity by targeting cell membrane disruption

Overexpression of a beta‐1,6‐glucanase gene GluM in transgenic rice confers high resistance to rice blast, sheath blight and false smut

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