Molecular characterization of AfuFleA, an L-fucose-specific lectin from Aspergillus fumigatus.

Aspergillus fumigatus causes life-threatening infections in immunocompromised patients. We have found that extracts of mycelial mats of A. fumigatus contain a potent hemagglutinin. To clarify the characteristics of this factor, the hemagglutinin was purified from late-stage cultures and characterized at the molecular level. The hemagglutinin is a 32-kilodalton protein that shows activity as an L-fucose lectin. The gene encoding this protein, AfufleA, was identified from a genomic DNA library utilizing consensus primers designed for amino acid sequences obtained from peptides following limited trypsin proteolysis. An open reading frame was found that consists of 942 nucleotides encoding 314 amino acids with a deduced molecular mass of 34,498 and contains all seven of trypsin-digested peptide sequences; four short introns, 49-63 bp, were also identified. AfufleA shares homology with a fucose-specific lectin produced by the orange peel mushroom, Aleuria aurantia. The role of AfufleA fucose-specific lectin is not clear, but this lectin may enhance attachment of fungal spores to mammalian cell membranes and contribute to the pathogenicity of A. fumigatus.

Fungal cell wall septation and cytokinesis are inhibited by bleomycins.

When the essential and distinctive cell walls of either pathogenic or nonpathogenic fungi break, cytoplasmic membranes rupture and fungi die. This fungicidal activity was discovered previously on nonproliferating Saccharomyces cerevisiae cells treated briefly with the oxidative tool and anticancer drug family of bleomycins. The present studies investigated effects of bleomycin on growing fungal organisms. These included the medically important Aspergillus fumigatus and Cryptococcus neoformans, as well as the emerging human pathogen and fungal model, S. cerevisiae. Bleomycin had its highest potency against A. fumigatus. Scanning electron microscopy and thin-section transmission electron microscopy were used to study morphological growth characteristics. Killing and growth inhibition were also measured. Long, thin, and segmented hyphae were observed when A. fumigatus was grown without bleomycin but were never observed when the mold was grown with the drug. Bleomycin arrested conidial germination, hyphal development, and the progression and completion of cell wall septation. Similarly, the drug inhibited the construction of yeast cell wall septa, preventing cytokinesis and progression in the cell division cycle of S. cerevisiae. Even when cytoplasms of mother and daughter cells separated, septation and cell division did not necessarily occur. Bizarre cell configurations, abnormally thickened cell walls at mother-daughter necks, abnormal polarized growth, large undivided cells, fragmented cells, and empty cell ghosts were also produced. This is the first report of a fungicidal agent that arrests fungal growth and development, septum formation, and cytokinesis and that also preferentially localizes to cell walls and alters isolated cell walls as well as intact cell walls on nongrowing cells.