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A sterol-regulatory element binding protein is required for cell polarity, hypoxia adaptation, azole drug resistance, and virulence in Aspergillus fumigatus.

Willger SD, Puttikamonkul S, Kim KH, Burritt JB, Grahl N, Metzler LJ, Barbuch R, Bard M, Lawrence CB, Cramer RA - PLoS Pathog. (2008)

Bottom Line: At the site of microbial infections, the significant influx of immune effector cells and the necrosis of tissue by the invading pathogen generate hypoxic microenvironments in which both the pathogen and host cells must survive.Loss of SrbA results in a mutant strain of the fungus that is incapable of growth in a hypoxic environment and consequently incapable of causing disease in two distinct murine models of invasive pulmonary aspergillosis (IPA).Significantly, the SrbA mutant was highly susceptible to fluconazole and voriconazole.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Molecular Biology, Montana State University, Bozeman, MT, USA.

ABSTRACT
At the site of microbial infections, the significant influx of immune effector cells and the necrosis of tissue by the invading pathogen generate hypoxic microenvironments in which both the pathogen and host cells must survive. Currently, whether hypoxia adaptation is an important virulence attribute of opportunistic pathogenic molds is unknown. Here we report the characterization of a sterol-regulatory element binding protein, SrbA, in the opportunistic pathogenic mold, Aspergillus fumigatus. Loss of SrbA results in a mutant strain of the fungus that is incapable of growth in a hypoxic environment and consequently incapable of causing disease in two distinct murine models of invasive pulmonary aspergillosis (IPA). Transcriptional profiling revealed 87 genes that are affected by loss of SrbA function. Annotation of these genes implicated SrbA in maintaining sterol biosynthesis and hyphal morphology. Further examination of the SrbA mutant consequently revealed that SrbA plays a critical role in ergosterol biosynthesis, resistance to the azole class of antifungal drugs, and in maintenance of cell polarity in A. fumigatus. Significantly, the SrbA mutant was highly susceptible to fluconazole and voriconazole. Thus, these findings present a new function of SREBP proteins in filamentous fungi, and demonstrate for the first time that hypoxia adaptation is likely an important virulence attribute of pathogenic molds.

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Related in: MedlinePlus

Loss of SrbA does not affect susceptibility to conidia killing by RAW264.7 cells.RAW264.7 cells (macrophages) were infected with a total of 1.25×106 freshly harvested A. fumigatus conidia of strains CEA10, SDW1, and SDW2 to obtain a conidia∶macrophage ratio of 5∶1. Conidia and macrophages were incubated together for 6 hours. After 6 hours, conidia were collected from the macrophages and plated onto glucose minimal media. Shown is the percent of recovered conidia after 6 hours incubation of two biological replicates. No significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01).
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ppat-1000200-g013: Loss of SrbA does not affect susceptibility to conidia killing by RAW264.7 cells.RAW264.7 cells (macrophages) were infected with a total of 1.25×106 freshly harvested A. fumigatus conidia of strains CEA10, SDW1, and SDW2 to obtain a conidia∶macrophage ratio of 5∶1. Conidia and macrophages were incubated together for 6 hours. After 6 hours, conidia were collected from the macrophages and plated onto glucose minimal media. Shown is the percent of recovered conidia after 6 hours incubation of two biological replicates. No significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01).

Mentions: One possible mechanism that could explain the virulence defect of strain SDW1 is an increased susceptibility to oxidative stress as suggested by transcriptional profiling and altered conidia morphology. We examined the growth of CEA10, SDW1, and SDW2 in the presence of 1 mM and 2.5 mM hydrogen peroxide on glucose minimal media. After 48 hours, we observed no detectable difference in growth morphology or colony diameter. In addition, we next examined the ability of RAW264.7 macrophage-like cells to kill SDW1 conidia (Figure 13). As presented in figure 13, no significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01). We conclude that increased susceptibility to oxidative stress and macrophage killing is not responsible for the virulence defect observed in the absence of SrbA.


A sterol-regulatory element binding protein is required for cell polarity, hypoxia adaptation, azole drug resistance, and virulence in Aspergillus fumigatus.

Willger SD, Puttikamonkul S, Kim KH, Burritt JB, Grahl N, Metzler LJ, Barbuch R, Bard M, Lawrence CB, Cramer RA - PLoS Pathog. (2008)

Loss of SrbA does not affect susceptibility to conidia killing by RAW264.7 cells.RAW264.7 cells (macrophages) were infected with a total of 1.25×106 freshly harvested A. fumigatus conidia of strains CEA10, SDW1, and SDW2 to obtain a conidia∶macrophage ratio of 5∶1. Conidia and macrophages were incubated together for 6 hours. After 6 hours, conidia were collected from the macrophages and plated onto glucose minimal media. Shown is the percent of recovered conidia after 6 hours incubation of two biological replicates. No significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2572145&req=5

ppat-1000200-g013: Loss of SrbA does not affect susceptibility to conidia killing by RAW264.7 cells.RAW264.7 cells (macrophages) were infected with a total of 1.25×106 freshly harvested A. fumigatus conidia of strains CEA10, SDW1, and SDW2 to obtain a conidia∶macrophage ratio of 5∶1. Conidia and macrophages were incubated together for 6 hours. After 6 hours, conidia were collected from the macrophages and plated onto glucose minimal media. Shown is the percent of recovered conidia after 6 hours incubation of two biological replicates. No significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01).
Mentions: One possible mechanism that could explain the virulence defect of strain SDW1 is an increased susceptibility to oxidative stress as suggested by transcriptional profiling and altered conidia morphology. We examined the growth of CEA10, SDW1, and SDW2 in the presence of 1 mM and 2.5 mM hydrogen peroxide on glucose minimal media. After 48 hours, we observed no detectable difference in growth morphology or colony diameter. In addition, we next examined the ability of RAW264.7 macrophage-like cells to kill SDW1 conidia (Figure 13). As presented in figure 13, no significant difference in conidia killing was observed between CEA10, SDW1, and SDW2 (P>0.01). We conclude that increased susceptibility to oxidative stress and macrophage killing is not responsible for the virulence defect observed in the absence of SrbA.

Bottom Line: At the site of microbial infections, the significant influx of immune effector cells and the necrosis of tissue by the invading pathogen generate hypoxic microenvironments in which both the pathogen and host cells must survive.Loss of SrbA results in a mutant strain of the fungus that is incapable of growth in a hypoxic environment and consequently incapable of causing disease in two distinct murine models of invasive pulmonary aspergillosis (IPA).Significantly, the SrbA mutant was highly susceptible to fluconazole and voriconazole.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Molecular Biology, Montana State University, Bozeman, MT, USA.

ABSTRACT
At the site of microbial infections, the significant influx of immune effector cells and the necrosis of tissue by the invading pathogen generate hypoxic microenvironments in which both the pathogen and host cells must survive. Currently, whether hypoxia adaptation is an important virulence attribute of opportunistic pathogenic molds is unknown. Here we report the characterization of a sterol-regulatory element binding protein, SrbA, in the opportunistic pathogenic mold, Aspergillus fumigatus. Loss of SrbA results in a mutant strain of the fungus that is incapable of growth in a hypoxic environment and consequently incapable of causing disease in two distinct murine models of invasive pulmonary aspergillosis (IPA). Transcriptional profiling revealed 87 genes that are affected by loss of SrbA function. Annotation of these genes implicated SrbA in maintaining sterol biosynthesis and hyphal morphology. Further examination of the SrbA mutant consequently revealed that SrbA plays a critical role in ergosterol biosynthesis, resistance to the azole class of antifungal drugs, and in maintenance of cell polarity in A. fumigatus. Significantly, the SrbA mutant was highly susceptible to fluconazole and voriconazole. Thus, these findings present a new function of SREBP proteins in filamentous fungi, and demonstrate for the first time that hypoxia adaptation is likely an important virulence attribute of pathogenic molds.

Show MeSH
Related in: MedlinePlus