Director: Jean-Paul Latgé
Aspergillus fumigatus is a thermophilic, saprophytic filamentous fungus that sporulates abundantly in nature. Pulmonary diseases consecutive to the inhalation of airborne conidia of A. fumigatus are often fatal among immunocompromised patients. Indeed, A. fumigatus has become today the most important fungal aerial pathogen in developed countries. The host-pathogen interactions are specially analysed to understand the early stages of the establishment of A. fumigatus in the lung parenchyma. On the applied side, new serological diagnostic methods are currently investigated since a better management of invasive aspergillosis patients is based on a better diagnostic system for this nosocomial mycosis. The first research theme of the laboratory is medically orientated and is focused on the study of invasive aspergillosis (IA). The other research area is more fundamental and concerns the study of the fungal cell wall biosynthesis that is a key morphogenetic event in the fungal life. In addition, such studies should identify targets for the development of new antifungal drugs that are urgently needed for the treatment of fungal diseases.
I - Immunity and Aspergillus infections
The interactions between the alveolar macrophage (AM) that is the main phagocytic cells of the lung environment and the conidia of A. fumigatus have been studied. Killing of the conidia in the immunocompetent host is due to reactive oxidant intermediates associated to the acidification of the phagolysosome. Activation of the MAP kinases ERK and p38 and translocation of NFkB were associated to the phagocytosis of conidia by AM. Glucocorticoïds are responsible for a reduction of reactive oxidant species and an inhibition of the phosphorylation of ERK and p38. These cellular events that resulted in an intracellular germination of conidia in the AM, explain a posteriori why glucocorticoïds trigger the development of IA among immunocompromised patients (Granet, Dubourdeau).
In accordance with the phagocytosis data, analysis of virulence factors of A. fumigatus has been now focused on the study of mutants affected in cell wall permeability and sensitivity to oxidative stress. However, mutants of A. fumigatus deficient in catalases, that are supposed to counteract the host reactive oxidant species, are as pathogenic as parental strains in an experimental murine aspergillosis model. These results associated to the analysis of other A. fumigatus mutants obtained by reverse genetics suggest that the virulence of A. fumigatus is polygenic (Paris). To approach this problem, and to identify genes concomitanly expressed during infection, a transcriptome analysis has been started. Such large scale investigation studies are now feasible since the sequence the genome of A. fumigatus (11 mb, 11000 genes) that has been sequenced is now annotated by an international consortium to which we participate (Beauvais, Debeaupuis, Henry, Mouyna, Sarfati, Lamarre, collaboration PT2, J.Y. Coppée).
Our unit has played a major role in the chemical and immunological characterization of the galactomannan of A. fumigatus that is at the origin of the development of the only commercial kit for the diagnosis of IA. This kit is a sandwich ELISA based on a monoclonal antibody detecting circulating beta1-5 galactofuran. New A. fumigatus molecules bearing this epitope have been now characterized : (i) glycoproteins with terminal galactofuranose at the non-reducing end of the mannan core and (ii) a lipophosphogalactomannan (Debeaupuis, Fontaine). Quantification of anti-A. fumigatus antibodies using recombinant protein antigens has been shown recently to be useful to identify patients at risk for IA (Sarfati).
II - Structure and biosynthesis of the cell wall of A. fumigatus
Major polysaccharides of the cell wall of A. fumigatus are alpha and beta(1-3) glucans and chitin. Biosynthesis of beta(1-3) glucans is under the control of FKS1, a unique essential membrane protein. In contrast, chitin and alpha(1,3) glucan are synthesized by families of genes that have different functions. Synthesis of chitin is under the control of a chitin synthase gene family composed of 7 members. Disruption of only 2 members of this family lead to phenotype with reduced growth and altered conidial permeability. Three AGS genes involved in the biosynthesis of alpha(1-3) glucans have been identified and only 2 of the AGS mutants are characterized by a reduced growth and altered conidiation (Beauvais).
In the periplasmic space, neosynthesized beta(1-3) glucans are modified and associated to the other cell wall polysaccharides (chitin, galactomannan and beta1-3, 1-4 glucan) to produce the rigid three-dimensional network characteristic of the cell wall. Such organization requires the presence of active glucanosyltransferases in cell wall space. During the search for such activities, it was shown for the first time that glycosyltransferases bound to the membrane by a glycosylphosphatidyl inositol (GPI) anchor play a major role in the biosynthesis of the yeast and mold cell wall (Mouyna). Chemical structure of the GPI anchor and its biosynthesis in A. fumigatus have been elucidated (Fontaine). Five families of GPI-anchored proteins, common to both filamentous fungi and yeast have been identified following a proteome and comparative genomic analysis of GPI-anchored proteins of S. cerevisiae and A. fumigatus and are now studied using a combination of biochemistry and molecular biology techniques (Chabane, Mouyna).
Three Dimensional schematic representation of the Aspergillus fumigatus cell wall.
Keywords: Aspergillus, lung, aspergillosis, cell wall, glucan, alveolar macrophage