Aspergillus nidulans galactofuranose biosynthesis affects antifungal drug sensitivity

• Published in: Fungal genetics and biology 2012-12, Vol.49 (12), p.1033-1043
• Main Authors: Alam, Md Kausar, El-Ganiny, Amira M., Afroz, Sharmin, Sanders, David A.R., Liu, Juxin, Kaminskyj, Susan G.W.
• Format: Article
• Language: English
• Subjects: alcA promoter; ammonium nitrate; Antifungal Agents - pharmacology; aspergillosis; Aspergillus fumigatus; Aspergillus nidulans; Aspergillus nidulans - cytology; Aspergillus nidulans - drug effects; Aspergillus nidulans - growth & development; Aspergillus nidulans - metabolism; biosynthesis; Biosynthetic Pathways - genetics; cell walls; Culture Media - chemistry; Echinocandins - pharmacology; Fungal Proteins - genetics; Fungal Proteins - metabolism; fungi; Galactofuranose; Galactose - analogs & derivatives; Galactose - biosynthesis; Gene Expression; Gene Expression Profiling; genes; glucose; humans; hyphae; Hyphae - cytology; Hyphae - drug effects; Hyphae - growth & development; Hyphae - metabolism; itraconazole; Itraconazole - pharmacology; lipids; Lipopeptides; Microbial Sensitivity Tests; monoclonal antibodies; morphogenesis; phenotype; Promoter Regions, Genetic; proteins; quantitative polymerase chain reaction; Real-Time Polymerase Chain Reaction; sporulation; threonine; UDP-galactofuranose transporter; UDP-galactopyranose mutase; UDP-glucose/galactose-4-epimerase; vigor
• ISSN: 1087-1845; 1096-0937
• DOI: 10.1016/j.fgb.2012.08.010

[Display omitted] ► Galactofuranose (Galf) biosynthesis is required for wildtype hyphal morphology. ► Galf biosynthesis genes were placed under alcA(p)- and niiA(p)-regulation. ► Under alcA(p)- and niiA(p)-repression regulated strain hyphae resembled gene deletion strains. ► Wild type and niiA(p)-expressed hyphae have Galf; deletion and repressed strains lack Galf. ► Caspofungin sensitivity, but not that of Itraconazole, correlated with wall Galf content. The cell wall is essential for fungal survival in natural environments. Many fungal wall carbohydrates are absent from humans, so they are a promising source of antifungal drug targets. Galactofuranose (Galf) is a sugar that decorates certain carbohydrates and lipids. It comprises about 5% of the Aspergillus fumigatus cell wall, and may play a role in systemic aspergillosis. We are studying Aspergillus wall formation in the tractable model system, A. nidulans. Previously we showed single-gene deletions of three sequential A. nidulans Galf biosynthesis proteins each caused similar hyphal morphogenesis defects and 500-fold reduced colony growth and sporulation. Here, we generated ugeA, ugmA and ugtA strains controlled by the alcA(p) or niiA(p) regulatable promoters. For repression and expression, alcA(p)-regulated strains were grown on complete medium with glucose or threonine, whereas niiA(p)-regulated strains were grown on minimal medium with ammonium or nitrate. Expression was assessed by qPCR and colony phenotype. The alcA(p) and niiA(p) strains produced similar effects: colonies resembling wild type for gene expression, and resembling deletion strains for gene repression. Galf immunolocalization using the L10 monoclonal antibody showed that ugmA deletion and repression phenotypes correlated with loss of hyphal wall Galf. None of the gene manipulations affected itraconazole sensitivity, as expected. Deletion of any of ugmA, ugeA, ugtA, their repression by alcA(p) or niiA(p), OR, ugmA overexpression by alcA(p), increased sensitivity to Caspofungin. Strains with alcA(p)-mediated overexpression of ugeA and ugtA had lower caspofungin sensitivity. Galf appears to play an important role in A. nidulans growth and vigor.