Biochemical characterization of recombinant Candida albicans mannosyltransferases Mnt1, Mnt2 and Mnt5 reveals new functions in O- and N-mannan biosynthesis

Díaz-Jiménez, Diana F.; Mora-Montes, Héctor M.; Hernández‐Cervantes, Arturo; Luna-Arias, Juan Pedro; Gow, Neil A. R.; Flores-Carreón, Arturo

doi:10.1016/j.bbrc.2012.01.131

Cited by 43 publications

(54 citation statements)

References 31 publications

(43 reference statements)

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“…Loss of cell viability was confirmed by an absence of growth on SC plates after 48 h incubation at 28°C. For β-elimination assays, cell samples were treated as previously described (Díaz-Jiménez et al, 2012). Briefly, cells were suspended in 10 ml of NaOH 0.1 N and incubated at room temperature during 18 h with gently shaking.…”

Section: Methodsmentioning

confidence: 99%

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Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

et al. 2016

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The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans, Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rarely causes candidiasis, Candida guilliermondii. We disrupted C. guilliermondii PMR1 and found that loss of Pmr1 affected cell growth and morphology, biofilm formation, susceptibility to cell wall perturbing agents, mannan levels, and the wall composition and organization. Despite the significant increment in the amount of β1,3-glucan exposed at the wall surface, this positively influenced only the ability of the mutant to stimulate IL-10 production by human monocytes, suggesting that recognition of both mannan and β1,3-glucan, is required to stimulate strong levels of pro-inflammatory cytokines. Accordingly, our results indicate C. guilliermondii sensing by monocytes was critically dependent on the recognition of N-linked mannans and β1,3-glucan, as reported in other Candida species. In addition, chemical remotion of cell wall O-linked mannans was found to positively influence the recognition of C. guilliermondii by human monocytes, suggesting that O-linked mannans mask other cell wall components from immune cells. This observation contrasts with that reported in C. albicans. Finally, mice infected with C. guilliermondii pmr1Δ null mutant cells had significantly lower fungal burdens compared to animals challenged with the parental strain. Accordingly, the null mutant showed inability to kill larvae in the Galleria mellonella infection model. This study thus demonstrates that mannans are relevant for the C. guilliermondii-host interaction, with an atypical role for O-linked mannans.

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Section: Methodsmentioning

confidence: 99%

“…Cells were also β-eliminated as described (Díaz-Jiménez et al, 2012). In both cases, cells were centrifuged, and the supernatant were freeze-dried and used to quantify the sugar content according to a reported the phenol-sulfuric-acid protocol (Dubois et al, 1956).…”

Section: Methodsmentioning

confidence: 99%

Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

et al. 2016

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“…Phosphodiester linkages are also included. Mannan synthesis is regulated by several specific glycosyl transferases [15][16][17][18] . It is also an immunochemical determinant of Candida spp 19 .…”

Section: Introductionmentioning

confidence: 99%

β-mannosyl linkages inhibit CAWS arteritis by negatively regulating dectin-2-dependent signaling in spleen and dendritic cells

Hirata

Ishibashi

Sato

et al. 2013

Immunopharmacology and Immunotoxicology

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“…However, mannan structure differs significantly between fungal species, and this has important consequences for host-fungus interactions, and in the future development of vaccines and diagnostics. For example, S. cerevisiae O -mannan is comprised of two or more α1,2-mannose units, which are capped by several α1,3-mannose units [21]–[23], while the O -mannan in C. albicans is comprised solely of α1,2-mannose [11], [24]–[25]. In addition, the N -mannan extracted from C. albicans has a higher molecular weight than that extracted from S. cerevisiae , includes β1,2-mannan in both the acid-labile and acid-stable fractions as well as higher levels of phosphomannan [26].…”

Section: Introductionmentioning

confidence: 99%

The Mnn2 Mannosyltransferase Family Modulates Mannoprotein Fibril Length, Immune Recognition and Virulence of Candida albicans

et al. 2013

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The fungal cell wall is the first point of interaction between an invading fungal pathogen and the host immune system. The outer layer of the cell wall is comprised of GPI anchored proteins, which are post-translationally modified by both N- and O-linked glycans. These glycans are important pathogen associated molecular patterns (PAMPs) recognised by the innate immune system. Glycan synthesis is mediated by a series of glycosyl transferases, located in the endoplasmic reticulum and Golgi apparatus. Mnn2 is responsible for the addition of the initial α1,2-mannose residue onto the α1,6-mannose backbone, forming the N-mannan outer chain branches. In Candida albicans, the MNN2 gene family is comprised of six members (MNN2, MNN21, MNN22, MNN23, MNN24 and MNN26). Using a series of single, double, triple, quintuple and sextuple mutants, we show, for the first time, that addition of α1,2-mannose is required for stabilisation of the α1,6-mannose backbone and hence regulates mannan fibril length. Sequential deletion of members of the MNN2 gene family resulted in the synthesis of lower molecular weight, less complex and more uniform N-glycans, with the sextuple mutant displaying only un-substituted α1,6-mannose. TEM images confirmed that the sextuple mutant was completely devoid of the outer mannan fibril layer, while deletion of two MNN2 orthologues resulted in short mannan fibrils. These changes in cell wall architecture correlated with decreased proinflammatory cytokine induction from monocytes and a decrease in fungal virulence in two animal models. Therefore, α1,2-mannose of N-mannan is important for both immune recognition and virulence of C. albicans.

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Biochemical characterization of recombinant Candida albicans mannosyltransferases Mnt1, Mnt2 and Mnt5 reveals new functions in O- and N-mannan biosynthesis

Abstract: Highlights► Mnt5 adds the second residues of lateral branches of the N-linked mannan outer chain. ► Mnt1 and Mnt2 participate in the addition of the 4° and 5° residues of O-linked mannan. ► Candida albicansO-linked mannans can be extended further than five mannose residues.

Cited by 43 publications

References 31 publications

Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

β-mannosyl linkages inhibit CAWS arteritis by negatively regulating dectin-2-dependent signaling in spleen and dendritic cells

The Mnn2 Mannosyltransferase Family Modulates Mannoprotein Fibril Length, Immune Recognition and Virulence of Candida albicans

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