Purpose: Aspergillus fumigatus is the main fungal pathogen found in cystic fibrosis (CF) airways, though its role in disease progression remains unclear. As the first point of contact for A. fumigatus following inhalation, the mucus barrier provides an important site for host-conidia interactions. Previous studies have suggested that A. fumigatus may alter the composition of airway mucus by altering mucin gene expression (1), and growth of A. fumigatus in mucin-based medium has been shown to upregulate the expression and secretion of specific proteases (2). Although these findings suggest a potential role for the mucus barrier during A. fumigatus infection,many aspects of this interplay are yet to be explored. We aimed to investigate this further by studying the effects of A. fumigatus on MUC5B and MUC5AC, the major gel-forming mucins found in airway mucus. Methods: The effects of A. fumigatus culture filtrates on the size of purified mucins were studied using agarose gel electrophoresis and sedimentation analysis, and recombinant mucin domains were used to identify proteolytic cleavage sites. Microrheology studies were also performed to measure the effects of A. fumigatus proteases on mucus rheological properties. Ongoing work using a panel of >200 A. fumigatus isolates is looking at the potential role of various signalling molecules and secreted products in regulating mucin degradation. Results: Our data show that A. fumigatus is able to degrade MUC5B and MUC5AC, with proteolytic activity observed in culture filtrates collected after 12 hours in liquid culture. Degradation of mucins resulted in a decrease in size and was accompanied by a reduction in mucus viscosity. Furthermore, cleavage was observed in both the N- and C-terminal regions of MUC5B. Knockout studies highlighted a potential role for PalH/PacC signalling in regulating mucin degradation, and have allowed us to identify putative mucin-degrading proteases including alkaline serine protease 1 (Alp1). Conclusion: These findings provide a valuable insight into the effects of A. fumigatus on the mucus barrier, and its potential involvement in A. fumigatus infections. Our data suggest that A. fumigatus may degrade mucins either as a nutrient source, or to alter mucus barrier properties in order to facilitate airway colonisation. This may represent an important mechanism of A. fumigatus pathogenesis, which could provide novel therapeutic targets for treating aspergillosis in patients such as those with cystic fibrosis. References (1) Oguma T, Asano K, Tomomatsu K, Kodama M, Fukunaga K, Shiomi T, Ohmori N, Ueda S, Takihara T, Shiraishi Y, Sayama K, Kagawa S, Natori Y, Lilly CM, Satoh K, Makimura K and Ishizaka A (2011). Induction of mucin and MUC5AC expression by the protease activity of aspergillus fumigatus in airway epithelial cells. The Journal of Immunology 187. 999-1005. (2) Farnell E, Rousseau K, Thornton DJ, Bowyer P and Herrick SE (2012). Expression and secretion of Aspergillus fumigatus proteases are regulated in response to different protein substrates. Fungal Biology 116. 1003-1012.
Full conference title:
The 8th Advances Against Aspergillus, Lisbon Conference Center, Lisbon, Portugal
- AAA 8th (2018)