Author:
François Le Mauff 1,2, Joshua Kerkaert 3, Ira Lacdao 1,2, Marc Yacoub 4, Benjamin Wucher 5, Fabrice Gravelat 1,2, Pierre-Guy Millette 1,2,*, Mario Vergas 6, Lynne P. Howell 6,7, Carey Nadell 5, Robert Cramer 3, J. Stajich 4, Shizhu Zhang 8, Donald C. Sheppard 1,2.
Author address:
1 Departments of Microbiology and Immunology, Faculty of Medicine, McGill University, Montréal, Canada 2 Infectious Diseases in Global Health Program, Research Institute of the McGill University Health Centre, Montréal, Canada 3 Department of Microbiology and Immunology, Geisel School of Medicine at Dartmouth, Hanover, NH 03755 USA 4 Department of Microbiology and Plant Pathology, Institute for Integrative Genome Biology, University of California Riverside, Riverside, California, USA 5 Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA 8 Jiangsu Key Laboratory for Microbes and Functional Genomics, Jiangsu Engineering and Technology Research Center for Microbiology, College of Life Sciences, Nanjing Normal University, Nanjing, China
* Current address: Faculty of Medicine, Université de Montréal, Montréal, Canada
Category: Fungal biology and genomics
Abstract:
Objectives: Biofilm formation in Aspergillus fumigatus has been reported to be dependent on the synthesis and secretion of galactosaminogalactan (GAG), a partially deacetylated heteropolymer of α-1,4-N-acetylgalactosamine (GalNAc) and galactose (Gal). GAG biosynthesis is believed to be mediated by the products of a 5-gene cluster located on chromosome 3 (cluster 3). However, the majority of enzymes in this cluster exhibit GalNAc-specific activity, suggesting that other enzymes might contribute to the synthesis and incorporation of Gal into GAG.
Methods: Blast searches identified a cluster of genes on chromosome 4 with similar functions to those found in cluster 3. Mutants deficient in the glycosyltransferases located within each cluster (gtb3, gtb4) were constructed. Biochemical characterization of GAG produced by each mutant was then carried out by gas chromatography coupled to mass spectrometry. Biofilm formation was assessed by crystal violet staining and confocal microscopy.
Results: Deletion of gtb4 resulted in a strain that formed fully adherent biofilms, but produced a GAG which was composed of a partially deacetylated homopolymer of GalNAc uniquely, a polyGalNAc. Deletion of gtb3 was associated with the inability to form an adherent biofilm, and a complete loss of GAG production. However, gene expression studies demonstrated that deletion of gtb3 was also associated with a complete repression of gtb4 expression. Therefore, to study the effects of gtb4 expression in isolation, gtb4 was constitutively expressed in a Δgtb3 background. Analysis of this strain revealed the production of a polymer of galactose and N-acetylgalactosamine which cannot be deacetylated by Agd3, the N-acetylgalactosaminogalactan. Biofilm microscopy demonstrated that polyGalNAc was largely adherent to the hyphal surface, while N-acetylgalactosaminogalactan was found within the biofilm intercellular matrix.
These findings suggest that A. fumigatus GAG is actually comprised of two separate polymers: a polymer of partially deacetylated GalNAc, produced by the gene products of cluster 3, which mediates adherent biofilm formation (partially deacetylated polyGalNAc), and a polymer composed largely of Gal with random insertion of low levels of GalNAc, produced by the gene products of cluster 4, which is dispensable for adhesion and largely found within the extracellular matrix (N-acetylgalactosaminogalactan).
Conclusion: This study highlights that GAG is actually comprised of two polymers with similar yet distinct compositions and functions. Further studies are required to determine the role that each of these two polymers plays in the multiple functions that have been described for GAG.
Abstract Number: 55
Conference Year: 2024
Conference abstracts, posters & presentations
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Title
Author
Year
Number
Poster
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v
Vasireddy Teja, Bibhuti Saha Hod, Soumendranath Haldar (IN)
2024
81
n/a
-
v
Rose-Anne Lavergne (FR)
2024
79
n/a
-
v
Rutuja H. Patil1, Tereza Juříková1, Andrea Palyzová1, and Vladimír Havlíček1,2
2024
78
n/a
-
v
Rui Xu1-2,, Chuqin Huang1 , Rong Yan1, Yanyan Lou1-2, Bo Zhang1, Yingyi Pan1, Yingying Liu1, Sini Huang1, Zhiyuan Zhang1, Antonin Tidu2, Thorsten Heinekamp3, Philippe Bulet4, Franck Martin2, , Axel Brakhage3, Li Zi1, Samuel Liegeois1-2, Dominique Ferrandon* 1-2
2024
75
n/a
-
v
Dominika Luptáková1, Tereza Hřivnová Juříková 1, Miloš Petřík2, Andrea Palyzová1, Vladimír Havlíček1
2024
74
n/a
-
v
Amer Ali Abd El-Hafeez1, Gabriele Sass2, David A. Stevens2,3, Vladimír Havlíček4,5, Ioly Kotta-Loizou6,7, Joe Hsu1
2024
73
n/a