Deacetylation of Fungal Exopolysaccharide Mediates Adhesion and Biofilm Formation
Authors: Lee Mark J. , Geller Alexander M. , Bamford Natalie C. , Liu Hong , Gravelat Fabrice N. , Snarr Brendan D. , Le Mauff François , Chabot Joseé , Ralph Benjamin , Ostapska Hanna , Lehoux Mélanie , Cerone Robert P. , Baptista Stephanie D. , Vinogradov Evgeny , Stajich Jason E. , Filler Scott G. , Howell P. Lynne , Sheppard Donald C. 
Published: April 2016

The mold
Aspergillus fumigatus
causes invasive infection in immunocompromised patients. Recently, galactosaminogalactan (GAG), an exopolysaccharide composed of galactose and
N
-acetylgalactosamine (GalNAc), was identified as a virulence factor required for biofilm formation. The molecular mechanisms underlying GAG biosynthesis and GAG-mediated biofilm formation were unknown. We identified a cluster of five coregulated genes that were dysregulated in GAG-deficient mutants and whose gene products share functional similarity with proteins that mediate the synthesis of the bacterial biofilm exopolysaccharide poly-(β1-6)-
N
-acetyl-
d
-glucosamine (PNAG). Bioinformatic analyses suggested that the GAG cluster gene
agd3
encodes a protein containing a deacetylase domain. Because deacetylation of
N
-acetylglucosamine residues is critical for the function of PNAG, we investigated the role of GAG deacetylation in fungal biofilm formation. Agd3 was found to mediate deacetylation of GalNAc residues within GAG and render the polysaccharide polycationic. As with PNAG, deacetylation is required for the adherence of GAG to hyphae and for biofilm formation. Growth of the Δ
agd3
mutant in the presence of culture supernatants of the GAG-deficient Δ
uge3
mutant rescued the biofilm defect of the Δ
agd3
mutant and restored the adhesive properties of GAG, suggesting that deacetylation is an extracellular process. The GAG biosynthetic gene cluster is present in the genomes of members of the
Pezizomycotina
subphylum of the
Ascomycota
including a number of plant-pathogenic fungi and a single basidiomycete species,
Trichosporon asahii
, likely a result of recent horizontal gene transfer. The current study demonstrates that the production of cationic, deacetylated exopolysaccharides is a strategy used by both fungi and bacteria for biofilm formation.

IMPORTANCE
This study sheds light on the biosynthetic pathways governing the synthesis of galactosaminogalactan (GAG), which plays a key role in
A. fumigatus
virulence and biofilm formation. We find that bacteria and fungi use similar strategies to synthesize adhesive biofilm exopolysaccharides. The presence of orthologs of the GAG biosynthetic gene clusters in multiple fungi suggests that this exopolysaccharide may also be important in the virulence of other fungal pathogens. Further, these studies establish a molecular mechanism of adhesion in which GAG interacts via charge-charge interactions to bind to both fungal hyphae and other substrates. Finally, the importance of deacetylation in the synthesis of functional GAG and the extracellular localization of this process suggest that inhibition of deacetylation may be an attractive target for the development of novel antifungal therapies.