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Role of the Candida albicans MNN1 gene family in cell wall structure and virulence.

Bates S, Hall RA, Cheetham J, Netea MG, MacCallum DM, Brown AJ, Odds FC, Gow NA - BMC Res Notes (2013)

Bottom Line: However, no gross changes in cell wall composition or N-glycosylation were identified in this mutant, although an extension of phosphomannan chain length was apparent.Although the cell wall defects associated with the mnn14Δ mutant were subtle, this mutant displayed a severe attenuation of virulence in a murine infection model.Mnn14 plays a distinct role from other members of the MNN1 family, demonstrating that specific N-glycan outer chain epitopes are required in the host-pathogen interaction and virulence.

View Article: PubMed Central - HTML - PubMed

Affiliation: College of Life and Environmental Sciences, University of Exeter, Exeter, EX4 4QD, UK. s.bates@ex.ac.uk

ABSTRACT

Background: The Candida albicans cell wall is the first point of contact with the host, and its outer surface is heavily enriched in mannoproteins modified through the addition of N- and O-mannan. Previous work, using mutants with gross defects in glycosylation, has clearly identified the importance of mannan in the host-pathogen interaction, immune recognition and virulence. Here we report the first analysis of the MNN1 gene family, which contains six members predicted to act as α-1,3 mannosyltransferases in the terminal stages of glycosylation.

Findings: We generated single mutants in all members of the C. albicans MNN1 gene family, and disruption of MNN14 led to both in vitro and in vivo defects. Null mutants in other members of the family demonstrated no phenotypic defects, suggesting that these members may display functional redundancy. The mnn14Δ mutant displayed hypersensitivity to agents associated with cell wall and glycosylation defects, suggesting an altered cell wall structure. However, no gross changes in cell wall composition or N-glycosylation were identified in this mutant, although an extension of phosphomannan chain length was apparent. Although the cell wall defects associated with the mnn14Δ mutant were subtle, this mutant displayed a severe attenuation of virulence in a murine infection model.

Conclusion: Mnn14 plays a distinct role from other members of the MNN1 family, demonstrating that specific N-glycan outer chain epitopes are required in the host-pathogen interaction and virulence.

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Biofilm formation of MNN1 family mutants. Biofilm formation was assessed in 96 well plates using a tetrazolium salt (XTT) reduction assay following growth in SC medium for 24 h (closed bars) and 48 h (open bars).
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Figure 5: Biofilm formation of MNN1 family mutants. Biofilm formation was assessed in 96 well plates using a tetrazolium salt (XTT) reduction assay following growth in SC medium for 24 h (closed bars) and 48 h (open bars).

Mentions: Defects in the initiation of O-glycosylation or treatment with tunicamycin, which inhibits N-glycosylation, have previously been shown to impair biofilm formation [28,29]. We therefore screened the MNN1 family mutants for defects in biofilm formation, using a 96 well microplate model and XTT reduction assay to measure metabolic activity of the cells within the biofilm. As predicted the och1∆ mutant [7], which has a gross defect in N-linked glycosylation, displayed a severe biofilm defect demonstrating a 90% reduction in biofilm formation at 48 h, therefore confirming the importance of N-mannan in biofilm formation. In addition the mnt1Δ-mnt2Δ double mutant [11], which lacks the 4 terminal mannose residues in O-mannan, also displayed a clear biofilm defect (80% reduction). This indicates that the extension of O-mannan, in addition to the initiation of its synthesis, is also important for biofilm formation. However, none of the MNN1 family single mutants displayed a significant defect in biofilm formation (Figure 5). This lack of a phenotypic defect may be due to functional redundancy in the MNN1 family. Alternatively, it may suggest that only gross N-glycosylation defects impact upon biofilm formation rather than its being dependent on specific epitopes. Indeed, the pmr1Δ mutant, which displays both O- and N-linked glycosylation defects, but to a lesser extent than the individual defects seen in the och1Δ or mnt1Δ-mnt2Δ mutants, only displays a subtle biofilm defect (40% reduction at 48 h). Therefore, biofilm formation is dependent both on correct O-glycosylation and the overall level of N-glycosylation, presumably through their impact on cellular interactions.


Role of the Candida albicans MNN1 gene family in cell wall structure and virulence.

Bates S, Hall RA, Cheetham J, Netea MG, MacCallum DM, Brown AJ, Odds FC, Gow NA - BMC Res Notes (2013)

Biofilm formation of MNN1 family mutants. Biofilm formation was assessed in 96 well plates using a tetrazolium salt (XTT) reduction assay following growth in SC medium for 24 h (closed bars) and 48 h (open bars).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750861&req=5

Figure 5: Biofilm formation of MNN1 family mutants. Biofilm formation was assessed in 96 well plates using a tetrazolium salt (XTT) reduction assay following growth in SC medium for 24 h (closed bars) and 48 h (open bars).
Mentions: Defects in the initiation of O-glycosylation or treatment with tunicamycin, which inhibits N-glycosylation, have previously been shown to impair biofilm formation [28,29]. We therefore screened the MNN1 family mutants for defects in biofilm formation, using a 96 well microplate model and XTT reduction assay to measure metabolic activity of the cells within the biofilm. As predicted the och1∆ mutant [7], which has a gross defect in N-linked glycosylation, displayed a severe biofilm defect demonstrating a 90% reduction in biofilm formation at 48 h, therefore confirming the importance of N-mannan in biofilm formation. In addition the mnt1Δ-mnt2Δ double mutant [11], which lacks the 4 terminal mannose residues in O-mannan, also displayed a clear biofilm defect (80% reduction). This indicates that the extension of O-mannan, in addition to the initiation of its synthesis, is also important for biofilm formation. However, none of the MNN1 family single mutants displayed a significant defect in biofilm formation (Figure 5). This lack of a phenotypic defect may be due to functional redundancy in the MNN1 family. Alternatively, it may suggest that only gross N-glycosylation defects impact upon biofilm formation rather than its being dependent on specific epitopes. Indeed, the pmr1Δ mutant, which displays both O- and N-linked glycosylation defects, but to a lesser extent than the individual defects seen in the och1Δ or mnt1Δ-mnt2Δ mutants, only displays a subtle biofilm defect (40% reduction at 48 h). Therefore, biofilm formation is dependent both on correct O-glycosylation and the overall level of N-glycosylation, presumably through their impact on cellular interactions.

Bottom Line: However, no gross changes in cell wall composition or N-glycosylation were identified in this mutant, although an extension of phosphomannan chain length was apparent.Although the cell wall defects associated with the mnn14Δ mutant were subtle, this mutant displayed a severe attenuation of virulence in a murine infection model.Mnn14 plays a distinct role from other members of the MNN1 family, demonstrating that specific N-glycan outer chain epitopes are required in the host-pathogen interaction and virulence.

View Article: PubMed Central - HTML - PubMed

Affiliation: College of Life and Environmental Sciences, University of Exeter, Exeter, EX4 4QD, UK. s.bates@ex.ac.uk

ABSTRACT

Background: The Candida albicans cell wall is the first point of contact with the host, and its outer surface is heavily enriched in mannoproteins modified through the addition of N- and O-mannan. Previous work, using mutants with gross defects in glycosylation, has clearly identified the importance of mannan in the host-pathogen interaction, immune recognition and virulence. Here we report the first analysis of the MNN1 gene family, which contains six members predicted to act as α-1,3 mannosyltransferases in the terminal stages of glycosylation.

Findings: We generated single mutants in all members of the C. albicans MNN1 gene family, and disruption of MNN14 led to both in vitro and in vivo defects. Null mutants in other members of the family demonstrated no phenotypic defects, suggesting that these members may display functional redundancy. The mnn14Δ mutant displayed hypersensitivity to agents associated with cell wall and glycosylation defects, suggesting an altered cell wall structure. However, no gross changes in cell wall composition or N-glycosylation were identified in this mutant, although an extension of phosphomannan chain length was apparent. Although the cell wall defects associated with the mnn14Δ mutant were subtle, this mutant displayed a severe attenuation of virulence in a murine infection model.

Conclusion: Mnn14 plays a distinct role from other members of the MNN1 family, demonstrating that specific N-glycan outer chain epitopes are required in the host-pathogen interaction and virulence.

Show MeSH
Related in: MedlinePlus