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Deletion of the ?-(1,3)-glucan synthase genes induces a restructuring of the conidial cell wall responsible for the avirulence of Aspergillus fumigatus.

ABSTRACT: ?-(1,3)-Glucan is a major component of the cell wall of Aspergillus fumigatus, an opportunistic human fungal pathogen. There are three genes (AGS1, AGS2 and AGS3) controlling the biosynthesis of ?-(1,3)-glucan in this fungal species. Deletion of all the three AGS genes resulted in a triple mutant that was devoid of ?-(1,3)-glucan in its cell wall; however, its growth and germination was identical to that of the parental strain in vitro. In the experimental murine aspergillosis model, this mutant was less pathogenic than the parental strain. The AGS deletion resulted in an extensive structural modification of the conidial cell wall, especially conidial surface where the rodlet layer was covered by an amorphous glycoprotein matrix. This surface modification was responsible for viability reduction of conidia in vivo, which explains decrease in the virulence of triple ags? mutant.

SUBMITTER: Beauvais A

PROVIDER: S-EPMC3828178 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Deletion of the α-(1,3)-glucan synthase genes induces a restructuring of the conidial cell wall responsible for the avirulence of Aspergillus fumigatus.

Beauvais Anne A Bozza Silvia S Kniemeyer Olaf O Formosa Cécile C Balloy Viviane V Henry Christine C Roberson Robert W RW Dague Etienne E Chignard Michel M Brakhage Axel A AA Romani Luigina L Latgé Jean-Paul JP

PLoS pathogens 20131114 11

α-(1,3)-Glucan is a major component of the cell wall of Aspergillus fumigatus, an opportunistic human fungal pathogen. There are three genes (AGS1, AGS2 and AGS3) controlling the biosynthesis of α-(1,3)-glucan in this fungal species. Deletion of all the three AGS genes resulted in a triple mutant that was devoid of α-(1,3)-glucan in its cell wall; however, its growth and germination was identical to that of the parental strain in vitro. In the experimental murine aspergillosis model, this mutant ...[more]

PMID: 24244155

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Project description:Aerial conidia of Ascomycetes and Basidiomycetes are coated with an amphipathic hydrophobin layer commonly called rodlet layer, that renders their surface hydrophobic and resist wetting, thus facilitating effective dispersal in the air. Fungal hydrophobins are small secreted proteins with specific hydropathy patterns and eight cysteine residues ordered in a particular manner which define the two well characterized classes I and II of hydrophobins and a third intermediate class (Kershaw and Talbot, 1998. Littlejohn et al, 2012; Jensen et al, 2010). Wessels 1994). Class I assemblies are generally water insoluble, have an amyloid structure and form the patterned rodlet layer. Class II are smaller proteins than class I and are soluble in aqueous ethanol mixtures of SDS. The intermediate class is a rather heterogenous tote class which would gather non-class I and non-class II hydrophobins without a defined structure. In addition to the role of the rodlet in the fungal life during conidiogenesis and conidial dispersal in the air, it was shown that rodlets can have a role in fungal – host interactions and favour the pathogenic behavior of the fungal pathogens in humans, plant and insect hosts (Talbot et al., 1996, Aimanianda et al, 2009; Zhang et al, 2011). In Aspergillusfumigatus rodlets immunosilence the conidium and their loss from the conidial surface initiates the host immune response (Aimanianda et al., 2009). Our study is focused on the Rod A protein which forms the rodlet structure on the surface of the conidium of A. fumigatus. In spite of their prominent morphological and biological role, the molecular basis for rodlet self-assembly and desagregation in this fungal species has not been understood yet. Point mutations in the rodA gene have helped understanding the mechanism which drives hydrophobin proteins to assemble into highly ordered structures anchored to the conidium surface. Moreover, the biochemical events responsible for the lysis of the rodlets have been investigated.

Project description:?-(1,3)-Glucan, the major fungal cell wall component, ramifies through ?-(1,6)-glycosidic linkages, which facilitates its binding with other cell wall components contributing to proper cell wall assembly. Using Saccharomyces cerevisiae as a model, we developed a protocol to quantify ?-(1,6)-branching on ?-(1,3)-glucan. Permeabilized S. cerevisiae and radiolabeled substrate UDP-(14C)glucose allowed us to determine branching kinetics. A screening aimed at identifying deletion mutants with reduced branching among them revealed only two, the bgl2? and gas1? mutants, showing 15% and 70% reductions in the branching, respectively, compared to the wild-type strain. Interestingly, a recombinant Gas1p introduced ?-(1,6)-branching on the ?-(1,3)-oligomers following its ?-(1,3)-elongase activity. Sequential elongation and branching activity of Gas1p occurred on linear ?-(1,3)-oligomers as well as Bgl2p-catalyzed products [short ?-(1,3)-oligomers linked by a linear ?-(1,6)-linkage]. The double S. cerevisiae gas1? bgl2? mutant showed a drastically sick phenotype. An ScGas1p ortholog, Gel4p from Aspergillus fumigatus, also showed dual ?-(1,3)-glucan elongating and branching activity. Both ScGas1p and A. fumigatus Gel4p sequences are endowed with a carbohydrate binding module (CBM), CBM43, which was required for the dual ?-(1,3)-glucan elongating and branching activity. Our report unravels the ?-(1,3)-glucan branching mechanism, a phenomenon occurring during construction of the cell wall which is essential for fungal life.IMPORTANCE The fungal cell wall is essential for growth, morphogenesis, protection, and survival. In spite of being essential, cell wall biogenesis, especially the core ?-(1,3)-glucan ramification, is poorly understood; the ramified ?-(1,3)-glucan interconnects other cell wall components. Once linear ?-(1,3)-glucan is synthesized by plasma membrane-bound glucan synthase, the subsequent event is its branching event in the cell wall space. Using Saccharomyces cerevisiae as a model, we identified GH72 and GH17 family glycosyltransferases, Gas1p and Bgl2p, respectively, involved in the ?-(1,3)-glucan branching. The sick phenotype of the double Scgas1? bgl2? mutant suggested that ?-(1,3)-glucan branching is essential. In addition to ScGas1p, GH72 family ScGas2p and Aspergillus fumigatus Gel4p, having CBM43 in their sequences, showed dual ?-(1,3)-glucan elongating and branching activity. Our report identifies the fungal cell wall ?-(1,3)-glucan branching mechanism. The essentiality of ?-(1,3)-glucan branching suggests that enzymes involved in the glucan branching could be exploited as antifungal targets.

Dataset Information

Deletion of the ?-(1,3)-glucan synthase genes induces a restructuring of the conidial cell wall responsible for the avirulence of Aspergillus fumigatus.

Publications

Deletion of the α-(1,3)-glucan synthase genes induces a restructuring of the conidial cell wall responsible for the avirulence of Aspergillus fumigatus.

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