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The Role of RodA-Conserved Cysteine Residues in the Aspergillus fumigatus Conidial Surface Organization.

ABSTRACT: Immune inertness of Aspergillusfumigatus conidia is attributed to its surface rodlet-layer made up of RodAp, characterized by eight conserved cysteine residues forming four disulfide bonds. Earlier, we showed that the conserved cysteine residue point (ccrp) mutations result in conidia devoid of the rodlet layer. Here, we extended our study comparing the surface organization and immunoreactivity of conidia carrying ccrp-mutations with the RODA deletion mutant (∆rodA). Western blot analysis using anti-RodAp antibodies indicated the absence of RodAp in the cytoplasm of ccrp-mutant conidia. Immunolabeling revealed differential reactivity to conidial surface glucans, the ccrp-mutant conidia preferentially binding to α-(1,3)-glucan, ∆rodA conidia selectively bound to β-(1,3)-glucan; the parental strain conidia showed negative labeling. However, permeability of ccrp-mutants and ∆rodA was similar to the parental strain conidia. Proteomic analyses of the conidial surface exposed proteins of the ccrp-mutants showed more similarities with the parental strain, but were significantly different from the ∆rodA. Ccrp-mutant conidia were less immunostimulatory compared to ∆rodA conidia. Our data suggest that (i) the conserved cysteine residues are essential for the trafficking of RodAp and the organization of the rodlet layer on the conidial surface, and (ii) targeted point mutation could be an alternative approach to study the role of fungal cell-wall genes in host-fungal interaction.

SUBMITTER: Valsecchi I

PROVIDER: S-EPMC7558875 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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The Role of RodA-Conserved Cysteine Residues in the Aspergillus fumigatus Conidial Surface Organization.

Valsecchi Isabel I Stephen-Victor Emmanuel E Wong Sarah Sze Wah SSW Karnam Anupama A Sunde Margaret M Guijarro J Iñaki JI Rodríguez de Francisco Borja B Krüger Thomas T Kniemeyer Olaf O Brown Gordon D GD Willment Janet A JA Latgé Jean-Paul JP Brakhage Axel A AA Bayry Jagadeesh J Aimanianda Vishukumar V

Journal of fungi (Basel, Switzerland) 20200826 3

Immune inertness of Aspergillus fumigatus conidia is attributed to its surface rodlet-layer made up of RodAp, characterized by eight conserved cysteine residues forming four disulfide bonds. Earlier, we showed that the conserved cysteine residue point (ccrp) mutations result in conidia devoid of the rodlet layer. Here, we extended our study comparing the surface organization and immunoreactivity of conidia carrying ccrp-mutations with the RODA deletion mutant (∆rod ...[more]

PMID: 32859091

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Project description:The hydrophobic layer of Aspergillus conidia, composed of RodA, plays a crucial role in conidia transfer and immune evasion. It self-assembles into hydrophobic rodlets through intramolecular disulfide bonds. However, the secretory process of RodA and its regulatory elements remain unknown. Since protein disulfide isomerase (PDI) is essential for the secretion of many disulfide-bonded proteins, we investigated whether PDI is also involved in RodA secretion and assembly. By gene knockout and phenotypic analysis, we found that Pdi1, one of the four PDI-related proteins of Aspergillus fumigatus, determines the hydrophobicity and integrity of the rodlet layer of the conidia. Preservation of the thioredoxin-active domain of Pdi1 was sufficient to maintain conidial hydrophobicity, suggesting that Pdi1 mediates RodA assembly through its disulfide isomerase activity. In the absence of Pdi1, the disulfide mismatch of RodA in conidia may prevent its delivery from the inner to the outer layer of the cell wall for rodlet assembly. This was demonstrated using a strain expressing a key cysteine-mutated RodA. The dormant conidia of the Pdi1-deficient strain (Δpdi) elicited an immune response, suggesting that the defective conidia surface in the absence of Pdi1 exposes internal immunogenic sources. In conclusion, Pdi1 ensures the correct folding of RodA in the inner layer of conidia, facilitating its secretion into the outer layer of the cell wall and allowing self-assembly of the hydrophobic layer. This study has identified a regulatory element for conidia rodlet assembly.IMPORTANCEAspergillus fumigatus is the major cause of invasive aspergillosis, which is mainly transmitted by the inhalation of conidia. The spread of conidia is largely dependent on their hydrophobicity, which is primarily attributed to the self-assembly of the hydrophobic protein RodA on the cell wall. However, the mechanisms underlying RodA secretion and transport to the outermost layer of the cell wall are still unclear. Our study identified a critical role for Pdi1, a fungal protein disulfide isomerase found in regulating RodA secretion and assembly. Inhibition of Pdi1 prevents the formation of correct S-S bonds in the inner RodA, creating a barrier to RodA delivery and resulting in a defective hydrophobic layer. Our findings provided insight into the formation of the conidial hydrophobic layer and suggested potential drug targets to inhibit A. fumigatus infections by limiting conidial dispersal and altering their immune inertia.

Dataset Information

The Role of RodA-Conserved Cysteine Residues in the Aspergillus fumigatus Conidial Surface Organization.

Publications

The Role of RodA-Conserved Cysteine Residues in the <i>Aspergillus fumigatus</i> Conidial Surface Organization.

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Dataset Information

The Role of RodA-Conserved Cysteine Residues in the Aspergillus fumigatus Conidial Surface Organization.

Publications

The Role of RodA-Conserved Cysteine Residues in the &lt;i&gt;Aspergillus fumigatus&lt;/i&gt; Conidial Surface Organization.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

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The Role of RodA-Conserved Cysteine Residues in the <i>Aspergillus fumigatus</i> Conidial Surface Organization.