Blanc SVSE 3 - Sciences de la vie, de la santé et des écosystèmes : Microbiologie, immunologie, infectiosité

HYDROPHOBIN OF Aspergillus fumigatus CONIDIA: STRUCTURAL ANALYSIS AND IMMUNE RESPONSE – HYDROPHOBIN

Deciphering the role of fungal hydrophobin proteins in fungal biology, public health and therapy

Despite we breathe thousands of air-borne fungal spores, hydrophobin surface proteins of spores prevents immune aggression against these inhaled spores. Therefore, air-borne fungal spores do not activate continuously the host innate immune cells and induce detrimental inflammatory responses following their inhalation. The project explores the role of hydrophobins in fungal life cycle and its therapeutic potential.

Molecular and immunological basis for inert nature of fungal hydrophobins and potential applications

We recently show that the surface layer composed of hydrophobins regularly arranged in a rodlet configuration on the dormant spores masks their recognition by the immune system and hence prevents immune response to inhaled air-borne conidia (2009, Nature 460: 1117-21).The present ANR application is the continuation of these studies, wherein we would like to investigate (1) the structure and the self-assembly of the conidial hydrophobin RodAp into rodlets, (2) analyze the molecular basis for the immunologically inertness of the rodlet layer, (3) identify the fungal and host factors that degrade hydrophobin after phagocytosis and during the course of spore-propagation and (4) explore the putative therapeutic use of hydrophobin.

Genetic engineering of hydrophobin and Aspergillus fumigatus conidia. Structural analysis of hydrophobins by using biophysical techniques including nuclear magnetic resonance, circular dichroism, Fourier-transform infrared spectroscopy, electron and atomic force microscopies. Studying the immunogenicity of hydrophobins (wild and mutant) and hydrophobin-coated therapeutic proteins by using human dendritic cells, T cells and in vivo experimental model of hemophilia A.

Aspergillus proteases secreted during the growth are responsible for the degradation of the spore-surface rodlet layer
The structure of the hydrophobin is solved by using doubly labelled (15Nand 13C) RodA protein
Established the in vitro and in vivo system to determine the immunogenicity of recombinant hydrophobins and hydrophobin-mutant conidia
Established a collaboration with Dr. Margaret Sunde, University of Sydney, Discipline of Pharmacology, NSW 2006, Australia, for the structural analysis of hydrophoibins.

Perceiving the structure and the self-assembly of hydrophobins and the molecular basis for the immunologically inertness of the rodlet layer may lead to a better understanding of the release of fungal immunogenic and allergic molecules and their impact on allergic and infectious syndromes. It can be expected that agents that inhibit the degradation of hydrophobin may also block germination and lead to the development of a new generation of antifungals. Alternatively, the immunologically inert nature of hydrophobin could be used to mask the immunogenicity of therapeutic proteins such as coagulant Factor VIII

A commentary/editorial on hydrophobin structure, immunogenicity and perspective applications (PLoS Pathog. 2012, 8:e1002700)
An editorial on chemokine axis as a therapeutic target to enhance the recruitment of regulatory T cells, the immunosuppressor cells to treat organ-specific autoimmune and inflammatory diseases (Immunotherapy 2012, 4: 9-12)
Dissemination of preliminary results on structural analysis of hydrophobin protein (A poster in International Conference on Magnetic Resonance of Biological Systems (ICMRBS), Lyon, France, 19-24 August 2012).

The air we breathe is filled with thousands of fungal spores (conidia). They originate from more than a hundred fungal species mainly belonging to the genus Cladosporium, Penicillium, Alternaria and Aspergillus. Although these spores are reservoirs of antigens and allergens, it was an enigma until recently why airborne fungal microflora do not activate continuously the host innate immune cells and induce detrimental inflammatory responses following their inhalation. We showed recently using several fungal inhabitants of the air-borne microflora including the human opportunistic pathogen Aspergillus fumigatus and various in vitro and in vivo assays with human and murine cells that the hydrophobic rodlet layer on the conidial surface immunologically silences air-borne fungal spores. Accordingly, the hydrophobin RodAp (the protein forming the rodlet layer) did not induce maturation and activation of murine and human immune cells. In contrast, the removal of this surface hydrophobin layer using biochemical and genetic tools resulted in conidial morphotypes that induced immune activation. Although fungal hydrophobins have been known for over 30 years, our publication (Aimanianda et al.,2009, Nature 460: 1117-21 ) is the first to report the immunological inert nature of this surface-protein on the air-borne fungal spores.The present ANR application is the continuation of these studies, wherein we would like to investigate (1) the structure and the self-assembly of the conidial hydrophobin RodAp into rodlets, (2) analyze the molecular basis for the immunologically inertness of the rodlet layer, (3) identify the fungal and host factors that degrade hydrophobin after phagocytosis and during the course of spore-propagation and (4) explore the putative therapeutic use of hydrophobin. This project is centered on A. fumigatus, the mold causing the most serious and often lethal pulmonary infections in the population of industrialized countries. Perceiving the structure and the self-assembly of hydrophobins and the molecular basis for the immunologically inertness of the rodlet layer may lead to a better understanding of the release of fungal molecules and their impact on allergic and infectious syndromes. Other applications can also be foreseen. Since conidial germination is correlated with hydrophobin degradation, it can be expected that agents that inhibit the degradation of hydrophobin may also block germination and lead to the development of a new generation of antifungals. Alternatively, the immunologically inert nature of hydrophobin could be used to mask the immunogenicity of therapeutic proteins such as Factor VIII (FVIII). Production of immunoglobulins (Abs) in patients against a protein-drug remains a major problem since these Abs can inhibit the functions of the administered therapeutic protein. Coating of FVIII by hydrophobin may reduce the immunogenicity of this therapeutic protein.

Project coordinator

Monsieur Jagadeesh Bayry (INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE - DELEGATION PARIS VI) – jagadeesh.bayry@crc.jussieu.fr

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

INSERM INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE - DELEGATION PARIS VI
IP INSTITUT PASTEUR
IP INSTITUT PASTEUR

Help of the ANR 360,000 euros
Beginning and duration of the scientific project: - 36 Months

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