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Melanie Wellington - One of the best experts on this subject based on the ideXlab platform.
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a genome wide screen of deletion mutants in the filamentous saccharomyces cerevisiae background identifies Ergosterol as a direct trigger of macrophage pyroptosis
Mbio, 2018Co-Authors: Kristy Koselny, Nebibe Mutlu, Annabel Y. Minard, Anuj Kumar, Damian J. Krysan, Melanie WellingtonAbstract:ABSTRACT Phagocytic cells such as macrophages play an important role in the host defense mechanisms mounted in response to the common human fungal pathogen Candida albicans. In vitro, C. albicans triggers macrophage NLRP3-Casp1/11-mediated pyroptosis, an inflammatory programmed cell death pathway. Here, we provide evidence that Casp1/11-dependent pyroptosis occurs in the kidney of infected mice during the early stages of infection. We have also used a genome-wide screen of nonessential Σ1278b Saccharomyces cerevisiae genes to identify genes required for yeast-triggered macrophage pyroptosis. The set of genes identified by this screen was enriched for those with functions in lipid and sterol homeostasis and trafficking. These observations led us to discover that cell surface localization and/or total levels of Ergosterol correlate with the ability of S. cerevisiae, C. albicans, and Cryptococcus neoformans to trigger pyroptosis. Since the mammalian sterol cholesterol triggers NLRP3-mediated pyroptosis, we hypothesized that Ergosterol may also do so. Consistent with that hypothesis, Ergosterol-containing liposomes but not Ergosterol-free liposomes induce pyroptosis. Cell wall mannoproteins directly bind Ergosterol, and we found that Dan1, an Ergosterol receptor mannoprotein, as well as specific mannosyltransferases, is required for pyroptosis, suggesting that cell wall-associated Ergosterol may mediate the process. Taken together, these data indicate that Ergosterol, like mammalian cholesterol, plays a direct role in yeast-mediated pyroptosis. IMPORTANCE Innate immune cells such as macrophages are key components of the host response to the human fungal pathogen Candida albicans. Macrophages undergo pyroptosis, an inflammatory, programmed cell death, in response to some species of pathogenic yeast. Prior to the work described in this report, yeast-triggered pyroptosis has been observed only in vitro; here, we show that pyroptosis occurs in the initial stages of murine kidney infection, suggesting that it plays an important role in the initial response of the innate immune system to invasive yeast infection. We also show that a key component of the fungal plasma membrane, Ergosterol, directly triggers pyroptosis. Ergosterol is also present in the fungal cell wall, most likely associated with mannoproteins, and is increased in hyphal cells compared to yeast cells. Our data indicate that specific mannoproteins are required for pyroptosis. This is consistent with a potential mechanism whereby Ergosterol present in the outer mannoprotein layer of the cell wall is accessible to the macrophage-mediated process. Taken together, our data provide the first evidence that Ergosterol plays a direct role in the host-pathogen interactions of fungi.
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A Genome-Wide Screen of Deletion Mutants in the Filamentous Saccharomyces cerevisiae Background Identifies Ergosterol as a Direct Trigger of Macrophage Pyroptosis
American Society for Microbiology, 2018Co-Authors: Kristy Koselny, Nebibe Mutlu, Annabel Y. Minard, Anuj Kumar, Damian J. Krysan, Melanie WellingtonAbstract:Phagocytic cells such as macrophages play an important role in the host defense mechanisms mounted in response to the common human fungal pathogen Candida albicans. In vitro, C. albicans triggers macrophage NLRP3-Casp1/11-mediated pyroptosis, an inflammatory programmed cell death pathway. Here, we provide evidence that Casp1/11-dependent pyroptosis occurs in the kidney of infected mice during the early stages of infection. We have also used a genome-wide screen of nonessential Σ1278b Saccharomyces cerevisiae genes to identify genes required for yeast-triggered macrophage pyroptosis. The set of genes identified by this screen was enriched for those with functions in lipid and sterol homeostasis and trafficking. These observations led us to discover that cell surface localization and/or total levels of Ergosterol correlate with the ability of S. cerevisiae, C. albicans, and Cryptococcus neoformans to trigger pyroptosis. Since the mammalian sterol cholesterol triggers NLRP3-mediated pyroptosis, we hypothesized that Ergosterol may also do so. Consistent with that hypothesis, Ergosterol-containing liposomes but not Ergosterol-free liposomes induce pyroptosis. Cell wall mannoproteins directly bind Ergosterol, and we found that Dan1, an Ergosterol receptor mannoprotein, as well as specific mannosyltransferases, is required for pyroptosis, suggesting that cell wall-associated Ergosterol may mediate the process. Taken together, these data indicate that Ergosterol, like mammalian cholesterol, plays a direct role in yeast-mediated pyroptosis.Innate immune cells such as macrophages are key components of the host response to the human fungal pathogen Candida albicans. Macrophages undergo pyroptosis, an inflammatory, programmed cell death, in response to some species of pathogenic yeast. Prior to the work described in this report, yeast-triggered pyroptosis has been observed only in vitro; here, we show that pyroptosis occurs in the initial stages of murine kidney infection, suggesting that it plays an important role in the initial response of the innate immune system to invasive yeast infection. We also show that a key component of the fungal plasma membrane, Ergosterol, directly triggers pyroptosis. Ergosterol is also present in the fungal cell wall, most likely associated with mannoproteins, and is increased in hyphal cells compared to yeast cells. Our data indicate that specific mannoproteins are required for pyroptosis. This is consistent with a potential mechanism whereby Ergosterol present in the outer mannoprotein layer of the cell wall is accessible to the macrophage-mediated process. Taken together, our data provide the first evidence that Ergosterol plays a direct role in the host-pathogen interactions of fungi
Helio K. Takahashi - One of the best experts on this subject based on the ideXlab platform.
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membrane microdomain components of histoplasma capsulatum yeast forms and their role in alveolar macrophage infectivity
Biochimica et Biophysica Acta, 2012Co-Authors: Loriane Tagliari, Tanil G Lacerda, Marcos S. Toledo, Erika Suzuki, Anita H. Straus, Helio K. TakahashiAbstract:Abstract Analysis of membrane lipids of Histoplasma capsulatum showed that ~40% of fungal Ergosterol is present in membrane microdomain fractions resistant to treatment with non-ionic detergent at 4 °C. Specific proteins were also enriched in these fractions, particularly Pma1p a yeast microdomain protein marker (a plasma membrane proton ATPase), a 30 kDa laminin-binding protein, and a 50 kDa protein recognized by anti-α5-integrin antibody. To better understand the role of Ergosterol-dependent microdomains in fungal biology and pathogenicity, H. capsulatum yeast forms were treated with a sterol chelator, methyl-beta-cyclodextrin (mβCD). Removal of Ergosterol by mβCD incubation led to disorganization of Ergosterol-enriched microdomains containing Pma1p and the 30 kDa protein, resulting in displacement of these proteins from detergent-insoluble to -soluble fractions in sucrose density gradient ultracentrifugation. mβCD treatment did not displace/remove the 50 kDa α5-integrin-like protein nor had effect on the organization of glycosphingolipids present in the detergent-resistant fractions. Ergosterol-enriched membrane microdomains were also shown to be important for infectivity of alveolar macrophages; after treatment of yeasts with mβCD, macrophage infectivity was reduced by 45%. These findings suggest the existence of two populations of detergent-resistant membrane microdomains in H. capsulatum yeast forms: (i) Ergosterol-independent microdomains rich in integrin-like proteins and glycosphingolipids, possibly involved in signal transduction; (ii) Ergosterol-enriched microdomains containing Pma1p and the 30 kDa laminin-binding protein; Ergosterol and/or the 30 kDa protein may be involved in macrophage infectivity.
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membrane microdomain components of histoplasma capsulatum yeast forms and their role in alveolar macrophage infectivity
Biochimica et Biophysica Acta, 2012Co-Authors: Loriane Tagliari, Tanil G Lacerda, Marcos S. Toledo, Erika Suzuki, Anita H. Straus, Helio K. TakahashiAbstract:Abstract Analysis of membrane lipids of Histoplasma capsulatum showed that ~40% of fungal Ergosterol is present in membrane microdomain fractions resistant to treatment with non-ionic detergent at 4 °C. Specific proteins were also enriched in these fractions, particularly Pma1p a yeast microdomain protein marker (a plasma membrane proton ATPase), a 30 kDa laminin-binding protein, and a 50 kDa protein recognized by anti-α5-integrin antibody. To better understand the role of Ergosterol-dependent microdomains in fungal biology and pathogenicity, H. capsulatum yeast forms were treated with a sterol chelator, methyl-beta-cyclodextrin (mβCD). Removal of Ergosterol by mβCD incubation led to disorganization of Ergosterol-enriched microdomains containing Pma1p and the 30 kDa protein, resulting in displacement of these proteins from detergent-insoluble to -soluble fractions in sucrose density gradient ultracentrifugation. mβCD treatment did not displace/remove the 50 kDa α5-integrin-like protein nor had effect on the organization of glycosphingolipids present in the detergent-resistant fractions. Ergosterol-enriched membrane microdomains were also shown to be important for infectivity of alveolar macrophages; after treatment of yeasts with mβCD, macrophage infectivity was reduced by 45%. These findings suggest the existence of two populations of detergent-resistant membrane microdomains in H. capsulatum yeast forms: (i) Ergosterol-independent microdomains rich in integrin-like proteins and glycosphingolipids, possibly involved in signal transduction; (ii) Ergosterol-enriched microdomains containing Pma1p and the 30 kDa laminin-binding protein; Ergosterol and/or the 30 kDa protein may be involved in macrophage infectivity.
Jeremy C Smith - One of the best experts on this subject based on the ideXlab platform.
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differential effects of cholesterol Ergosterol and lanosterol on a dipalmitoyl phosphatidylcholine membrane a molecular dynamics simulation study
Journal of Physical Chemistry B, 2007Co-Authors: Zoe Cournia, Matthias G Ullmann, Jeremy C SmithAbstract:Lipid raft/domain formation may arise as a result of the effects of specific sterols on the physical properties of membranes. Here, using molecular dynamics simulation, we examine the effects of three closely-related sterols, Ergosterol, cholesterol, and lanosterol, at a biologically relevant concentration (40 mol %) on the structural properties of a model dipalmitoyl phosphatidylcholine (DPPC) membrane at 309 and 323 K. All three sterols are found to order the DPPC acyl tails and condense the membrane relative to the DPPC liquid-phase membrane, but each one does this to a significantly different degree. The smooth alpha-face of Ergosterol, together with the presence of tail unsaturation in this sterol, leads to closer interaction of Ergosterol with the lipids and closer packing of the lipids with each other, so Ergosterol has a higher condensing effect on the membrane, as reflected by the area per lipid. Moreover, Ergosterol induces a higher proportion of trans lipid conformers, a thicker membrane, and higher lipid order parameters and is aligned more closely with the membrane normal. Ergosterol also positions itself closer to the bilayer/water interface. In contrast, the rough alpha-face of lanosterol leads to a less close interaction of the steroid ring system with the phospholipid acyl chains, and so lanosterol orders, straightens, and packs the lipid acyl chains less well and is less closely aligned with the membrane normal. Furthermore, lanosterol lies closer to the relatively disordered membrane center than do the other sterols. The behavior of cholesterol in all the above respects is intermediate between that of lanosterol and Ergosterol. The findings here may explain why Ergosterol is the most efficient of the three sterols at promoting the liquid-ordered phase and lipid domain formation and may also furnish part of the explanation as to why cholesterol is evolutionarily preferred over lanosterol in higher-vertebrate plasma membranes.
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differential effects of cholesterol Ergosterol and lanosterol on a dipalmitoyl phosphatidylcholine membrane a molecular dynamics simulation study
Journal of Physical Chemistry B, 2007Co-Authors: Zoe Cournia, Matthias G Ullmann, Jeremy C SmithAbstract:Lipid raft/domain formation may arise as a result of the effects of specific sterols on the physical properties of membranes. Here, using molecular dynamics simulation, we examine the effects of three closely-related sterols, Ergosterol, cholesterol, and lanosterol, at a biologically relevant concentration (40 mol %) on the structural properties of a model dipalmitoyl phosphatidylcholine (DPPC) membrane at 309 and 323 K. All three sterols are found to order the DPPC acyl tails and condense the membrane relative to the DPPC liquid-phase membrane, but each one does this to a significantly different degree. The smooth α-face of Ergosterol, together with the presence of tail unsaturation in this sterol, leads to closer interaction of Ergosterol with the lipids and closer packing of the lipids with each other, so Ergosterol has a higher condensing effect on the membrane, as reflected by the area per lipid. Moreover, Ergosterol induces a higher proportion of trans lipid conformers, a thicker membrane, and highe...
Eric Chauvet - One of the best experts on this subject based on the ideXlab platform.
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Effect of Culture Conditions on Ergosterol as an Indicator of Biomass in the Aquatic Hyphomycetes
Applied and Environmental Microbiology, 2001Co-Authors: Jean-yves Charcosset, Eric ChauvetAbstract:Ergosterol is a membrane component specific to fungi that can be used to estimate fungal biomass using appropriate factors of conversion. Our objectives were to determine the limits of use of Ergosterol content as a measure of biomass for aquatic hyphomycetes, and to evaluate a previously established Ergosterol-to-biomass conversion factor. We varied inoculum quality, growth medium, and degree of shaking of four aquatic hyphomycete species. In cultures inoculated with homogenized mycelium, we found a significant effect of shaking condition and culture age on Ergosterol content. In liquid cultures with defined medium, Ergosterol content reached 10 to 11 mg/mg of mycelium (dry mass) and varied by factors of 2.2 during exponential growth and 1.3 during stationary phase. The increase in Ergosterol content during exponential phase could be attributed, at least in part, to rapid depletion of glucose. Oxygen availability to internal hyphae within the mycelial mass is also responsible for the differences found between culture conditions. Ergosterol concentration ranged from 0.8 to 1.6 mg/mg in static cultures inoculated with agar plugs. Ergosterol content varied by a factor of 4 in two media of different richnesses. For different combinations of these parameters, strong (r2 5 0.83 to 0.98) and highly significant (P ! 0.001) linear relationships between Ergosterol and mycelial dry mass (up to 110 mg) were observed. Overall, the Ergosterol content varied by a factor of 14 (0.8 to 11 mg/g). These results suggest that care must be taken when the Ergosterol content is used to compare data generated in different field environments.
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Ergosterol-to-Biomass Conversion Factors for Aquatic Hyphomycetes.
Applied and Environmental Microbiology, 1993Co-Authors: Mark O. Gessner, Eric ChauvetAbstract:Fourteen strains of aquatic hyphomycete species that are common on decaying leaves in running waters were grown in liquid culture and analyzed for total Ergosterol contents. Media included an aqueous extract from senescent alder leaves, a malt extract broth, and a glucose-mineral salt solution. Concentrations of Ergosterol in fungal mycelium ranged from 2.3 to 11.5 mg/g of dry mass. The overall average was 5.5 mg/g. Differences among both species and growth media were highly significant but followed no systematic pattern. Stationary-phase mycelium had Ergosterol contents 10 to 12% lower or higher than mycelium harvested during the growth phase, but these differences were only significant for one of four species examined. Availability of plant sterols in the growth medium had no clear effect on Ergosterol concentrations in two species tested. To convert Ergosterol contents determined in field samples to biomass values of aquatic hyphomycetes, a general multiplicative factor of 182 is proposed. More accurate estimates would be obtained with species-specific factors. Using these in combination with estimates of the proportion of the dominant species in a naturally established community on leaves resulted in biomass estimates that were typically 20% lower than those obtained with the general conversion factor. Improvements of estimates with species-specific factors may be limited, however, by intraspecific variability in fungal Ergosterol content.
Loriane Tagliari - One of the best experts on this subject based on the ideXlab platform.
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membrane microdomain components of histoplasma capsulatum yeast forms and their role in alveolar macrophage infectivity
Biochimica et Biophysica Acta, 2012Co-Authors: Loriane Tagliari, Tanil G Lacerda, Marcos S. Toledo, Erika Suzuki, Anita H. Straus, Helio K. TakahashiAbstract:Abstract Analysis of membrane lipids of Histoplasma capsulatum showed that ~40% of fungal Ergosterol is present in membrane microdomain fractions resistant to treatment with non-ionic detergent at 4 °C. Specific proteins were also enriched in these fractions, particularly Pma1p a yeast microdomain protein marker (a plasma membrane proton ATPase), a 30 kDa laminin-binding protein, and a 50 kDa protein recognized by anti-α5-integrin antibody. To better understand the role of Ergosterol-dependent microdomains in fungal biology and pathogenicity, H. capsulatum yeast forms were treated with a sterol chelator, methyl-beta-cyclodextrin (mβCD). Removal of Ergosterol by mβCD incubation led to disorganization of Ergosterol-enriched microdomains containing Pma1p and the 30 kDa protein, resulting in displacement of these proteins from detergent-insoluble to -soluble fractions in sucrose density gradient ultracentrifugation. mβCD treatment did not displace/remove the 50 kDa α5-integrin-like protein nor had effect on the organization of glycosphingolipids present in the detergent-resistant fractions. Ergosterol-enriched membrane microdomains were also shown to be important for infectivity of alveolar macrophages; after treatment of yeasts with mβCD, macrophage infectivity was reduced by 45%. These findings suggest the existence of two populations of detergent-resistant membrane microdomains in H. capsulatum yeast forms: (i) Ergosterol-independent microdomains rich in integrin-like proteins and glycosphingolipids, possibly involved in signal transduction; (ii) Ergosterol-enriched microdomains containing Pma1p and the 30 kDa laminin-binding protein; Ergosterol and/or the 30 kDa protein may be involved in macrophage infectivity.
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membrane microdomain components of histoplasma capsulatum yeast forms and their role in alveolar macrophage infectivity
Biochimica et Biophysica Acta, 2012Co-Authors: Loriane Tagliari, Tanil G Lacerda, Marcos S. Toledo, Erika Suzuki, Anita H. Straus, Helio K. TakahashiAbstract:Abstract Analysis of membrane lipids of Histoplasma capsulatum showed that ~40% of fungal Ergosterol is present in membrane microdomain fractions resistant to treatment with non-ionic detergent at 4 °C. Specific proteins were also enriched in these fractions, particularly Pma1p a yeast microdomain protein marker (a plasma membrane proton ATPase), a 30 kDa laminin-binding protein, and a 50 kDa protein recognized by anti-α5-integrin antibody. To better understand the role of Ergosterol-dependent microdomains in fungal biology and pathogenicity, H. capsulatum yeast forms were treated with a sterol chelator, methyl-beta-cyclodextrin (mβCD). Removal of Ergosterol by mβCD incubation led to disorganization of Ergosterol-enriched microdomains containing Pma1p and the 30 kDa protein, resulting in displacement of these proteins from detergent-insoluble to -soluble fractions in sucrose density gradient ultracentrifugation. mβCD treatment did not displace/remove the 50 kDa α5-integrin-like protein nor had effect on the organization of glycosphingolipids present in the detergent-resistant fractions. Ergosterol-enriched membrane microdomains were also shown to be important for infectivity of alveolar macrophages; after treatment of yeasts with mβCD, macrophage infectivity was reduced by 45%. These findings suggest the existence of two populations of detergent-resistant membrane microdomains in H. capsulatum yeast forms: (i) Ergosterol-independent microdomains rich in integrin-like proteins and glycosphingolipids, possibly involved in signal transduction; (ii) Ergosterol-enriched microdomains containing Pma1p and the 30 kDa laminin-binding protein; Ergosterol and/or the 30 kDa protein may be involved in macrophage infectivity.